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Sample records for arrests human monocytic

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

    Science.gov (United States)

    Maniwa, Yasuhisa; Kasukabe, Takashi; Kumakura, Shunichi

    2015-08-01

    Although all-trans retinoic acid (ATRA) is a standard and effective drug used for differentiation therapy in acute promyelocytic leukemia, ATRA-resistant leukemia cells ultimately emerge during this treatment. Therefore, the development of new drugs or effective combination therapy is urgently needed. We demonstrate that the combined treatment of vitamin K2 and cotylenin A synergistically induced monocytic differentiation in HL-60 cells. This combined treatment also synergistically induced NBT-reducing activity and non-specific esterase-positive cells as well as morphological changes to monocyte/macrophage-like cells. Vitamin K2 and cotylenin A cooperatively inhibited the proliferation of HL-60 cells in short-term and long-term cultures. This treatment also induced growth arrest at the G1 phase. Although 5 µg/ml cotylenin A or 5 µM vitamin K2 alone reduced c-MYC gene expression in HL-60 cells to approximately 45% or 80% that of control cells, respectively, the combined treatment almost completely suppressed c-MYC gene expression. We also demonstrated that the combined treatment of vitamin K2 and cotylenin A synergistically induced the expression of cyclin G2, which had a positive effect on the promotion and maintenance of cell cycle arrest. These results suggest that the combination of vitamin K2 and cotylenin A has therapeutic value in the treatment of acute myeloid leukemia. PMID:26046133

  2. Cyclic dinucleotides modulate human T-cell response through monocyte cell death.

    Science.gov (United States)

    Tosolini, Marie; Pont, Frédéric; Verhoeyen, Els; Fournié, Jean-Jacques

    2015-12-01

    Cyclic dinucleotides, a class of microbial messengers, have been recently identified in bacteria, but their activity in humans remains largely unknown. Here, we have studied the function of cyclic dinucleotides in humans. We found that c-di-AMP and cGAMP, two adenosine-based cyclic dinucleotides, activated T lymphocytes in an unusual manner through monocyte cell death. c-di-AMP and cGAMP induced the selective apoptosis of human monocytes, and T lymphocytes were activated by the direct contact with these dying monocytes. The ensuing T-cell response comprised cell-cycle exit, phenotypic maturation into effector memory cells and proliferation arrest, but not cell death. This quiescence was transient since T cells remained fully responsive to further restimulation. Together, our results depict a novel activation pattern for human T lymphocytes: a transient quiescence induced by c-di-AMP- or cGAMP-primed apoptotic monocytes. PMID:26460927

  3. The transcriptional network that controls growth arrest and differentiation in a human myeloid leukemia cell line

    DEFF Research Database (Denmark)

    Suzuki, Harukazu; Forrest, Alistair R R; van Nimwegen, Erik;

    2009-01-01

    Using deep sequencing (deepCAGE), the FANTOM4 study measured the genome-wide dynamics of transcription-start-site usage in the human monocytic cell line THP-1 throughout a time course of growth arrest and differentiation. Modeling the expression dynamics in terms of predicted cis-regulatory sites...

  4. Programming of Human Monocytes by the Uteroplacental Environment

    OpenAIRE

    McIntire, Ramsey H.; Ganacias, Karen G.; Hunt, Joan S.

    2008-01-01

    During human pregnancy, monocytes recruited to the uterus (decidua) are modified to promote immune defense and semiallogeneic pregnancy. The purpose of this study was to identify decidual factors involved in programming of monocytes into decidual macrophages by comparing the surface and secretory phenotypes of resting and interferon-γ (IFN-γ)–activated monocytes, unfractionated decidual cells, purified term decidual macrophages, and monocyte-derived macrophages. Surface markers for antigen pr...

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

    OpenAIRE

    MANIWA, YASUHISA; Kasukabe, Takashi; Kumakura, Shunichi

    2015-01-01

    Although all-trans retinoic acid (ATRA) is a standard and effective drug used for differentiation therapy in acute promyelocytic leukemia, ATRA-resistant leukemia cells ultimately emerge during this treatment. Therefore, the development of new drugs or effective combination therapy is urgently needed. We demonstrate that the combined treatment of vitamin K2 and cotylenin A synergistically induced monocytic differentiation in HL-60 cells. This combined treatment also synergistically induced NB...

  6. Effect of prostaglandin I2 analogs on monocyte chemoattractant protein-1 in human monocyte and macrophage.

    Science.gov (United States)

    Tsai, Ming-Kai; Hsieh, Chong-Chao; Kuo, Hsuan-Fu; Lee, Min-Sheng; Huang, Ming-Yii; Kuo, Chang-Hung; Hung, Chih-Hsing

    2015-08-01

    Chemokines play essential roles during inflammatory responses and in pathogenesis of inflammatory diseases. Monocyte chemotactic protein-1 (MCP-1) is a critical chemokine in the development of atherosclerosis and acute cardiovascular syndromes. MCP-1, by its chemotactic activity, causes diapedesis of monocytes from the lumen to the subendothelial space that leads to atherosclerotic plaque formation. Prostaglandin I2 (PGI2) analogs are used clinically for patients with pulmonary hypertension and have anti-inflammatory effects. However, little is known about the effect of PGI2 analogs on the MCP-1 production in human monocytes and macrophages. We investigated the effects of three conventional (iloprost, beraprost and treprostinil) and one new (ONO-1301) PGI2 analogs, on the expression of MCP-1 expression in human monocytes and macrophages. Human monocyte cell line, THP-1 cell, was treated with PGI2 analogs after LPS stimulation. Supernatants were harvested to measure MCP-1 levels and measured by ELISA. To explore which receptors involved the effects of PGI2 analogs on the expression of MCP-1 expression, IP and EP, PPAR-α and PPAR-γ receptor antagonists were used. Forskolin, a cAMP activator, was used to further confirm the involvement of cAMP on MCP-1 production in human monocytes. Three PGI2 analogs suppressed LPS-induced MCP-1 production in THP-1 cells and THP-1-induced macrophages. Higher concentrations of ONO-1301 also had the suppressive effect. CAY 10449, an IP receptor antagonist, could reverse the effects on MCP-1 production of iloprost on THP-1 cells. Other reported PGI2 receptor antagonists including EP1, EP2, EP4, PPAR-α and PPAR-γ antagonists could not reverse the effect. Forskolin, a cAMP activator, also suppressed MCP-1 production in THP-1 cells. PGI2 analogs suppressed LPS-induced MCP-1 production in human monocytes and macrophages via the IP receptor and cAMP pathway. The new PGI2 analog (ONO-1301) was not better than conventional PGI2 analog in

  7. Lactic acid delays the inflammatory response of human monocytes

    Energy Technology Data Exchange (ETDEWEB)

    Peter, Katrin, E-mail: katrin.peter@ukr.de [Department of Internal Medicine III, University Hospital Regensburg, Franz-Josef-Strauß-Allee 11, 93053 Regensburg (Germany); Rehli, Michael, E-mail: michael.rehli@ukr.de [Department of Internal Medicine III, University Hospital Regensburg, Franz-Josef-Strauß-Allee 11, 93053 Regensburg (Germany); RCI Regensburg Center for Interventional Immunology, University Hospital Regensburg, Franz-Josef-Strauß-Allee 11, 93053 Regensburg (Germany); Singer, Katrin, E-mail: katrin.singer@ukr.de [Department of Internal Medicine III, University Hospital Regensburg, Franz-Josef-Strauß-Allee 11, 93053 Regensburg (Germany); Renner-Sattler, Kathrin, E-mail: kathrin.renner-sattler@ukr.de [Department of Internal Medicine III, University Hospital Regensburg, Franz-Josef-Strauß-Allee 11, 93053 Regensburg (Germany); Kreutz, Marina, E-mail: marina.kreutz@ukr.de [Department of Internal Medicine III, University Hospital Regensburg, Franz-Josef-Strauß-Allee 11, 93053 Regensburg (Germany); RCI Regensburg Center for Interventional Immunology, University Hospital Regensburg, Franz-Josef-Strauß-Allee 11, 93053 Regensburg (Germany)

    2015-02-13

    Lactic acid (LA) accumulates under inflammatory conditions, e.g. in wounds or tumors, and influences local immune cell functions. We previously noted inhibitory effects of LA on glycolysis and TNF secretion of human LPS-stimulated monocytes. Here, we globally analyze the influence of LA on gene expression during monocyte activation. To separate LA-specific from lactate- or pH-effects, monocytes were treated for one or four hours with LPS in the presence of physiological concentrations of LA, sodium lactate (NaL) or acidic pH. Analyses of global gene expression profiles revealed striking effects of LA during the early stimulation phase. Up-regulation of most LPS-induced genes was significantly delayed in the presence of LA, while this inhibitory effect was attenuated in acidified samples and not detected after incubation with NaL. LA targets included genes encoding for important monocyte effector proteins like cytokines (e.g. TNF and IL-23) or chemokines (e.g. CCL2 and CCL7). LA effects were validated for several targets by quantitative RT-PCR and/or ELISA. Further analysis of LPS-signaling pathways revealed that LA delayed the phosphorylation of protein kinase B (AKT) as well as the degradation of IκBα. Consistently, the LPS-induced nuclear accumulation of NFκB was also diminished in response to LA. These results indicate that the broad effect of LA on gene expression and function of human monocytes is at least partially caused by its interference with immediate signal transduction events after activation. This mechanism might contribute to monocyte suppression in the tumor environment. - Highlights: • Lactic acid broadly delays LPS-induced gene expression in human monocytes. • Expression of important monocyte effector molecules is affected by lactic acid. • Interference of lactic acid with TLR signaling causes the delayed gene expression. • The profound effect of lactic acid might contribute to immune suppression in tumors.

  8. Lactic acid delays the inflammatory response of human monocytes

    International Nuclear Information System (INIS)

    Lactic acid (LA) accumulates under inflammatory conditions, e.g. in wounds or tumors, and influences local immune cell functions. We previously noted inhibitory effects of LA on glycolysis and TNF secretion of human LPS-stimulated monocytes. Here, we globally analyze the influence of LA on gene expression during monocyte activation. To separate LA-specific from lactate- or pH-effects, monocytes were treated for one or four hours with LPS in the presence of physiological concentrations of LA, sodium lactate (NaL) or acidic pH. Analyses of global gene expression profiles revealed striking effects of LA during the early stimulation phase. Up-regulation of most LPS-induced genes was significantly delayed in the presence of LA, while this inhibitory effect was attenuated in acidified samples and not detected after incubation with NaL. LA targets included genes encoding for important monocyte effector proteins like cytokines (e.g. TNF and IL-23) or chemokines (e.g. CCL2 and CCL7). LA effects were validated for several targets by quantitative RT-PCR and/or ELISA. Further analysis of LPS-signaling pathways revealed that LA delayed the phosphorylation of protein kinase B (AKT) as well as the degradation of IκBα. Consistently, the LPS-induced nuclear accumulation of NFκB was also diminished in response to LA. These results indicate that the broad effect of LA on gene expression and function of human monocytes is at least partially caused by its interference with immediate signal transduction events after activation. This mechanism might contribute to monocyte suppression in the tumor environment. - Highlights: • Lactic acid broadly delays LPS-induced gene expression in human monocytes. • Expression of important monocyte effector molecules is affected by lactic acid. • Interference of lactic acid with TLR signaling causes the delayed gene expression. • The profound effect of lactic acid might contribute to immune suppression in tumors

  9. Susceptibility and Response of Human Blood Monocyte Subsets to Primary Dengue Virus Infection

    OpenAIRE

    Wong, Kok Loon; Chen, Weiqiang; Balakrishnan, Thavamalar; Toh, Ying Xiu; Fink, Katja; Wong, Siew-Cheng

    2012-01-01

    Human blood monocytes play a central role in dengue infections and form the majority of virus infected cells in the blood. Human blood monocytes are heterogeneous and divided into CD16− and CD16+ subsets. Monocyte subsets play distinct roles during disease, but it is not currently known if monocyte subsets differentially contribute to dengue protection and pathogenesis. Here, we compared the susceptibility and response of the human CD16− and CD16+ blood monocyte subsets to primary dengue viru...

  10. Maturation and demise of human primary monocytes by carbon nanotubes

    International Nuclear Information System (INIS)

    The possibility of exploiting carbon nanotubes (CNT) in biomedical practices requires thorough analysis of the chemical or bulk effects they may exert on the immune system, the complex network that recognizes and eliminates foreign particles. In particular, the phagocytosing ability of cells belonging to the monocyte/macrophage lineage may render these immune cells an ideal toxicological target of pristine CNT, which may form aggregates of size exceeding monocyte/macrophage phagocytosing plasticity. To shed light on this issue, we analyzed the effects that pristine multi-walled CNT (MWCNT) without metal or biological impurities exert on survival and activation of freshly explanted human peripheral blood monocytes, analyzing in parallel the non-phagocytosing lymphocytes, and using graphite as control carbon material. MWCNT (diameter 10–50 nm, length up to 10 μm) exert two different toxic effects on mononuclear leukocytes: a minor apoptogenic effect (on lymphocytes > monocytes), and a major, apoptosis-independent effect that exclusively and deeply affect monocyte homeostasis. Analysis of monocyte number, adhesion, redox equilibrium, and the differentiation markers CD14 and CD11b reveals that MWCNT cause the selective disappearance of phagocytosis-competent monocytes by mechanisms related to the presence of large nanoparticle aggregates, suggesting phenomena of bulk toxicity possibly consisting of frustrated phagocytosis. At the same time, MWCNT stimulate adhesion of the phagocytosis-incompetent monocytes, and their differentiation toward a peculiar maturation asset. These observations point out novel mechanisms of CNT toxicity, renewing concerns that they may impair the innate immune system deranging the inflammatory responses.

  11. Maturation and demise of human primary monocytes by carbon nanotubes

    Energy Technology Data Exchange (ETDEWEB)

    De Nicola, Milena, E-mail: milena.de.nicola@uniroma2.it [University of Rome ' Tor Vergata' , Department of Biology (Italy); Mirabile Gattia, Daniele, E-mail: daniele.mirabile@enea.it [UTTMAT, ENEA-C.R. Casaccia (Italy); Traversa, Enrico, E-mail: Enrico.Traversa@kaust.edu.sa [King Abdullah University of Science and Technology (KAUST), Division of Physical Science and Engineering (Saudi Arabia); Ghibelli, Lina, E-mail: ghibelli@uniroma2.it [University of Rome ' Tor Vergata' , Department of Biology (Italy)

    2013-06-15

    The possibility of exploiting carbon nanotubes (CNT) in biomedical practices requires thorough analysis of the chemical or bulk effects they may exert on the immune system, the complex network that recognizes and eliminates foreign particles. In particular, the phagocytosing ability of cells belonging to the monocyte/macrophage lineage may render these immune cells an ideal toxicological target of pristine CNT, which may form aggregates of size exceeding monocyte/macrophage phagocytosing plasticity. To shed light on this issue, we analyzed the effects that pristine multi-walled CNT (MWCNT) without metal or biological impurities exert on survival and activation of freshly explanted human peripheral blood monocytes, analyzing in parallel the non-phagocytosing lymphocytes, and using graphite as control carbon material. MWCNT (diameter 10-50 nm, length up to 10 {mu}m) exert two different toxic effects on mononuclear leukocytes: a minor apoptogenic effect (on lymphocytes > monocytes), and a major, apoptosis-independent effect that exclusively and deeply affect monocyte homeostasis. Analysis of monocyte number, adhesion, redox equilibrium, and the differentiation markers CD14 and CD11b reveals that MWCNT cause the selective disappearance of phagocytosis-competent monocytes by mechanisms related to the presence of large nanoparticle aggregates, suggesting phenomena of bulk toxicity possibly consisting of frustrated phagocytosis. At the same time, MWCNT stimulate adhesion of the phagocytosis-incompetent monocytes, and their differentiation toward a peculiar maturation asset. These observations point out novel mechanisms of CNT toxicity, renewing concerns that they may impair the innate immune system deranging the inflammatory responses.

  12. Maturation and demise of human primary monocytes by carbon nanotubes

    KAUST Repository

    De Nicola, Milena D.

    2013-05-17

    The possibility of exploiting carbon nanotubes (CNT) in biomedical practices requires thorough analysis of the chemical or bulk effects they may exert on the immune system, the complex network that recognizes and eliminates foreign particles. In particular, the phagocytosing ability of cells belonging to the monocyte/macrophage lineage may render these immune cells an ideal toxicological target of pristine CNT, which may form aggregates of size exceeding monocyte/macrophage phagocytosing plasticity. To shed light on this issue, we analyzed the effects that pristine multi-walled CNT (MWCNT) without metal or biological impurities exert on survival and activation of freshly explanted human peripheral blood monocytes, analyzing in parallel the non-phagocytosing lymphocytes, and using graphite as control carbon material. MWCNT (diameter 10-50 nm, length up to 10 μm) exert two different toxic effects on mononuclear leukocytes: a minor apoptogenic effect (on lymphocytes > monocytes), and a major, apoptosis-independent effect that exclusively and deeply affect monocyte homeostasis. Analysis of monocyte number, adhesion, redox equilibrium, and the differentiation markers CD14 and CD11b reveals that MWCNT cause the selective disappearance of phagocytosis-competent monocytes by mechanisms related to the presence of large nanoparticle aggregates, suggesting phenomena of bulk toxicity possibly consisting of frustrated phagocytosis. At the same time, MWCNT stimulate adhesion of the phagocytosis-incompetent monocytes, and their differentiation toward a peculiar maturation asset. These observations point out novel mechanisms of CNT toxicity, renewing concerns that they may impair the innate immune system deranging the inflammatory responses. © 2013 Springer Science+Business Media Dordrecht.

  13. Primary human monocyte differentiation regulated by Nigella sativa pressed oil

    OpenAIRE

    Mat Mahaya C; Mohamed Azman S; Hamid Shahrul S

    2011-01-01

    Abstract Background Oxidized low density lipoprotein plays an important role in development of foam cells in atherosclerosis. The study was focused on regulation of primary human monocyte growth and CD11b expression in presence of Nigella sativa oil. Methods Primary human monocytes were isolated from whole blood and grown at 37°C and 5% CO2 saturation for five days prior to treatment with Nigella sativa oil. The cells were plated and washed before treatment with ox-LDL (10 μg/ml) as positive ...

  14. No evidence for histamine H4 receptor in human monocytes.

    Science.gov (United States)

    Werner, Kristin; Neumann, Detlef; Buschauer, Armin; Seifert, Roland

    2014-12-01

    The histamine H4 receptor (H4R) is a classic pertussis toxin-sensitive Gi protein-coupled receptor that mediates increases in intracellular calcium concentration ([Ca(2+)]i). The presence of H4R in human eosinophils has been rigorously documented by several independent groups. It has also been suggested that H4R is expressed in human monocytes, but this suggestion hinges in part on H4R antibodies with questionable specificity. This situation prompted us to reinvestigate H4R expression in human monocytes. As positive control, we studied human embryonic kidney 293T cells stably expressing the human H4R (hH4R). In these cells, histamine (HA) and the H4R agonist UR-PI376 (2-cyano-1-[4-(1H-imidazol-4-yl)butyl]-3-[(2-phenylthio)ethyl]guanidine) induced pertussis toxin-sensitive [Ca(2+)]i increases. However, in quantitative real-time polymerase chain reaction studies we failed to detect hH4R mRNA in human monocytes and U937 promonocytes. In human monocytes, ATP and N-formyl-l-methionyl-l-leucyl-l-phenylalanine increased [Ca(2+)]i, but HA, UR-PI376, and 5-methylhistamine (a dual H4R/H2 receptor agonist) did not. In U937 promonocytes and differentiated U937 cells, HA increased [Ca(2+)]i, but this increase was mediated via HA H1 receptor. In conclusion, there is no evidence for the presence of H4R in human monocytes. PMID:25273276

  15. DMPD: LPS induction of gene expression in human monocytes. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 11257452 LPS induction of gene expression in human monocytes. Guha M, Mackman N. Ce...ll Signal. 2001 Feb;13(2):85-94. (.png) (.svg) (.html) (.csml) Show LPS induction of gene expression in human... monocytes. PubmedID 11257452 Title LPS induction of gene expression in human monocytes. Authors Guha M, Ma

  16. Maturation arrest of human oocytes at germinal vesicle stage

    Directory of Open Access Journals (Sweden)

    Zhi Qin Chen

    2010-01-01

    Full Text Available Maturation arrest of human oocytes may occur at various stages of the cell cycle. A total failure of human oocytes to complete meiosis is rarely observed during assisted conception cycles. We describe here a case of infertile couples for whom all oocytes repeatedly failed to mature at germinal vesicle (GV stage during in vitro fertilization/Intra cytoplasmic sperm injection (IVF/ICSI. The patient underwent controlled ovarian stimulation followed by oocyte retrieval and IVF/ICSI. The oocytes were stripped off cumulus cells prior to the ICSI procedure and their maturity status was defined. The oocyte maturation was repeatedly arrested at the GV. Oocyte maturation arrest may be the cause of infertility in this couple. The recognition of oocyte maturation arrest as a specific medical condition may contribute to the characterization of the currently known as "oocyte factor." The cellular and genetic mechanisms causing oocyte maturation arrest should be the subject for further investigation.

  17. [The effects of PEMF on the activation of human monocytes].

    Science.gov (United States)

    Chen, Xiaoying; Han, Xiaoyu; Wang, Qian; Wu, Wenchao; Liu, Xiaojing

    2012-08-01

    The aim of the present study is to investigate the effect of pulsed electromagnetic field (PEMF) on the activation of human monocytes (THP-1). Cultured THP-1 cells were exposed to PEMF stimulation with radiation of 32Hz or 64Hz respectively, using sinusoidal wave, and 1mT, twice a day, 30 minutes each time, with an interval of 8 hours, for 3 days. Those with 0Hz stimulation served as the controls. Monocytes activation was monitored by measuring both the release of monocyte chemoattractant protein-1 (MCP-1) from monocytes and their adhesion to monolayers of human umbilical vein endothelial cells (HUVECs). The adhesion of THP-1 cells to HUVECs was evaluated by cell counting method. The secretion of MCP-1 from THP-1 cells was detected by ELISA and MCP-1 mRNA expression was assessed by real time quantitative RT-PCR. The data showed that exposure to PEMF with above parameters could significantly inhibit the adhesion of THP-1 cells to HUVECs and decrease the MCP-1 mRNA and protein expression. The results demonstrated that exposure to PEMF of 1mT, 32Hz or 64Hz for 3 days could significantly inhibit the activation of THP-1 cells. PMID:23016400

  18. Δ9-Tetrahydrocannabinol (THC) enhances lipopolysaccharide-stimulated tissue factor in human monocytes and monocyte-derived microvesicles

    OpenAIRE

    Williams, Julie C.; Thomas W Klein; Goldberger, Bruce A.; Sleasman, John W.; Mackman, Nigel; Goodenow, Maureen M.

    2015-01-01

    Background Immunomodulatory effects in humans of Δ9−Tetrahydrocannabinol (THC), the psychoactive component of marijuana are controversial. Tissue factor (TF), the activator of the extrinsic coagulation cascade, is increased on circulating activated monocytes and is expressed on microvesicles released from activated monocytes during inflammatory conditions, which perpetuate coagulopathies in a number of diseases. In view of the increased medicinal use of marijuana, effects of THC on human mono...

  19. Decreased Monocyte HLA-DR Expression in Patients After Non-Shockable out-of-Hospital Cardiac Arrest.

    Science.gov (United States)

    Venet, Fabienne; Cour, Martin; Demaret, Julie; Monneret, Guillaume; Argaud, Laurent

    2016-07-01

    Out-of-hospital cardiac arrest (OHCA) constitutes a major health care problem with the development in immediate survivors of a post-cardiac arrest syndrome including systemic inflammatory response as observed in sepsis. As a decreased monocyte HLA-DR expression (mHLA-DR) has been repeatedly described in septic patients in association with an increased risk of death and nosocomial infections, we tested whether this immune alteration could also be observed after OHCA. Fifty-five non-shockable OHCA patients sampled at Day 0 (D0: within 4 h after OHCA), D1 (the next day), and D3: (after 2 additional days) were included. CD4+ lymphocyte count and mHLA-DR were evaluated by flow cytometry. We observed a marked decrease in mHLA-DR as early as D0 in patients compared with normal values. This decrease persisted till D3 and was associated with a moderate decrease in the number of circulating CD4+ lymphocytes. No correlations were identified between mHLA-DR and usual prognostic markers after OHCA. However, overtime evolution in mHLA-DR values appeared different between survivors and non-survivors with a quasisystematic decrease between D1 and D3 in non-survivors versus an increased expression in survivors. In conclusion, this preliminary pilot study describes the occurrence of OHCA-induced immune alterations as illustrated by a decreased mHLA-DR and CD4+ lymphopenia. Furthermore, we show for the first time the differential overtime evolution in mHLA-DR between survivors and non-survivors without association with usual prognostic markers and multiple organ failure. These initial results should now be confirmed in a larger cohort of OHCA patients. PMID:26796574

  20. Normal human monocytes exposed to glioma cells acquire myeloid-derived suppressor cell-like properties

    OpenAIRE

    Rodrigues, Jennifer C.; Gonzalez, Guido C.; Zhang, Lei; Ibrahim, George; Kelly, John J.; Gustafson, Michael P.; Yi LIN; Dietz, Allan B.; Forsyth, Peter A; Yong, V. Wee; Parney, Ian F.

    2009-01-01

    Glioblastoma patients are immunosuppressed, yet glioblastomas are highly infiltrated by monocytes/macrophages. Myeloid-derived suppressor cells (MDSC; immunosuppressive myeloid cells including monocytes) have been identified in other cancers and correlate with tumor burden. We hypothesized that glioblastoma exposure causes normal monocytes to assume an MDSC-like phenotype and that MDSC are increased in glioblastoma patients. Healthy donor human CD14+ monocytes were cultured with human gliobla...

  1. A curated compendium of monocyte transcriptome datasets of relevance to human monocyte immunobiology research.

    Science.gov (United States)

    Rinchai, Darawan; Boughorbel, Sabri; Presnell, Scott; Quinn, Charlie; Chaussabel, Damien

    2016-01-01

    Systems-scale profiling approaches have become widely used in translational research settings. The resulting accumulation of large-scale datasets in public repositories represents a critical opportunity to promote insight and foster knowledge discovery. However, resources that can serve as an interface between biomedical researchers and such vast and heterogeneous dataset collections are needed in order to fulfill this potential. Recently, we have developed an interactive data browsing and visualization web application, the Gene Expression Browser (GXB). This tool can be used to overlay deep molecular phenotyping data with rich contextual information about analytes, samples and studies along with ancillary clinical or immunological profiling data. In this note, we describe a curated compendium of 93 public datasets generated in the context of human monocyte immunological studies, representing a total of 4,516 transcriptome profiles. Datasets were uploaded to an instance of GXB along with study description and sample annotations. Study samples were arranged in different groups. Ranked gene lists were generated based on relevant group comparisons. This resource is publicly available online at http://monocyte.gxbsidra.org/dm3/landing.gsp. PMID:27158452

  2. Follicular dendritic-like cells derived from human monocytes

    Directory of Open Access Journals (Sweden)

    Peters J Hinrich

    2005-09-01

    Full Text Available Abstract Background Follicular dendritic cells (FDCs play a central role in controlling B-cell response maturation, isotype switching and the maintenance of B-cell memory. These functions are based on prolonged preservation of antigen and its presentation in its native form by FDCs. However, when entrapping entire pathogens, FDCs can turn into dangerous long-term reservoirs that may preserve viruses or prions in highly infectious form. Despite various efforts, the ontogeny of FDCs has remained elusive. They have been proposed to derive either from bone marrow stromal cells, myeloid cells or local mesenchymal precursors. Still, differentiating FDCs from their precursors in vitro may allow addressing many unsolved issues associated with the (patho- biology of these important antigen-presenting cells. The aim of our study was to demonstrate that FDC-like cells can be deduced from monocytes, and to develop a protocol in order to quantitatively generate them in vitro. Results Employing highly purified human monocytes as a starter population, low concentrations of Il-4 (25 U/ml and GM-CSF (3 U/ml in combination with Dexamethasone (Dex (0.5 μM in serum-free medium trigger the differentiation into FDC-like cells. After transient de-novo membrane expression of alkaline phosphatase (AP, such cells highly up-regulate surface expression of complement receptor I (CD35. Co-expression of CD68 confirms the monocytic origin of both, APpos and CD35pos cells. The common leukocyte antigen CD45 is strongly down-regulated. Successive stimulation with TNF-α up-regulates adhesion molecules ICAM-1 (CD54 and VCAM (CD106. Importantly, both, APpos as well as APneg FDC-like cells, heterotypically cluster with and emperipolese B cells and exhibit the FDC characteristic ability to entrap functionally preserved antigen for prolonged times. Identical characteristics are found in monocytes which were highly expanded in vitro by higher doses of GM-CSF (25 U/ml in the absence of

  3. Localisation of the monocyte-binding region on human immunoglobulin G.

    Science.gov (United States)

    Woof, J M; Partridge, L J; Jefferis, R; Burton, D R

    1986-03-01

    Earlier studies, which provided indirect evidence for the involvement of the C gamma 2 domain of human immunoglobulin G (IgG) in human immunoglobulin G (IgG) in human monocyte binding, have been extended to further localise the site of interaction on human IgG. A number of IgGs from several different species and fragments of human IgGs were assayed for ability to inhibit the interaction of radio-labelled human IgG and the human monocyte. By comparison of the amino-acid sequences of those IgGs found to exhibit relatively tight, intermediate or weak binding to human monocyte Fc receptors we are able to postulate a possible monocyte-binding site on human IgG. In addition, the results have implications for the applicability of monoclonal antibodies and antisera when used in the presence of human monocytes and possibly macrophages. PMID:3487030

  4. Burkholderia pseudomallei induces IL-23 production in primary human monocytes.

    Science.gov (United States)

    Kulsantiwong, Panthong; Pudla, Matsayapan; Boondit, Jitrada; Wikraiphat, Chanthiwa; Dunachie, Susanna J; Chantratita, Narisara; Utaisincharoen, Pongsak

    2016-06-01

    Burkholderia pseudomallei, a gram-negative intracellular bacterium, is a causative agent of melioidosis. The bacterium has been shown to induce the innate immune response, particularly pro-inflammatory cytokine production in several of both mouse and human cell types. In the present study, we investigate host immune response in B. pseudomallei-infected primary human monocytes. We discover that wild-type B. pseudomallei is able to survive and multiply inside the primary human monocytes. In contrast, B. pseudomallei LPS mutant, a less virulent strain, is susceptible to host killing during bacterial infection. Moreover, microarray result showed that wild-type B. pseudomallei but not B. pseudomallei LPS mutant is able to activate gene expression of IL-23 as demonstrated by the up-regulation of p19 and p40 subunit expression. Consistent with gene expression analysis, the secretion of IL-23 analyzed by ELISA also showed that wild-type B. pseudomallei induces a significantly higher level of IL-23 secretion than that of B. pseudomallei LPS mutant. These results implied that IL-23 may be an important cytokine for the innate immune response during B. pseudomallei infection. The regulation of IL-23 production may drive the different host innate immune responses between patients and may relate to the severity of melioidosis. PMID:26563410

  5. Differential induction from X-irradiated human peripheral blood monocytes to dendritic cells

    International Nuclear Information System (INIS)

    Dendritic cells (DCs) are a type of antigen-presenting cell which plays an essential role in the immune system. To clarify the influences of ionizing radiation on the differentiation to DCs, we focused on human peripheral blood monocytes and investigated whether X-irradiated monocytes can differentiate into DCs. The non-irradiated monocytes and 5 Gy-irradiated monocytes were induced into immature DCs (iDCs) and mature DCs (mDCs) with appropriate cytokine stimulation, and the induced cells from each monocyte expressed each DC-expressing surface antigen such as CD40, CD86 and HLA-DR. However, the expression levels of CD40 and CD86 on the iDCs derived from the 5 Gy-irradiated monocytes were higher than those of iDCs derived from non-irradiated monocytes. Furthermore, the mDCs derived from 5 Gy-irradiated monocytes had significantly less ability to stimulate allogeneic T cells in comparison to the mDCs derived from non-irradiated monocytes. There were no significant differences in the phagocytotic activity of the iDCs and cytokines detected in the supernatants conditioned by the DCs from the non-irradiated and irradiated monocytes. These results suggest that human monocytes which are exposed to ionizing radiation can thus differentiate into DCs, but there is a tendency that X-irradiation leads to an impairment of the function of DCs. (author)

  6. Suppressive effects of liquid crystal compounds on the growth of U937 human leukemic monocyte lymphoma cells

    Directory of Open Access Journals (Sweden)

    Ishikawa Junya

    2012-02-01

    Full Text Available Abstract Background The aim of this study was to evaluate the biological and pharmaceutical activities of 14 amphiphilic liquid-crystalline compounds (LCs, i.e, phenylpyrimidine derivatives possessing D-glucamine and cyanobiphenyl derivatives with a terminal hydroxyl unit. Results The cytotoxic properties of the LCs on the cell growth, cell cycle distribution, and cell signaling pathway of U937 human leukemic monocyte lymphoma cells were assessed by flow cytometry and western blot analysis. Some LCs showed cytostatic effects, suppressing cell growth via S-phase arrest and without apoptosis in U937 cells. To investigate the mechanisms of the LC-induced S-phase arrest, proteins relevant to cell cycle regulation were investigated by western blot analysis. The rate of LC-induced S-phase arrest was congruent with the decreased expression of MCM2, cyclin A, cyclin B, CDK2, phospho-CDK1 and Cdc25C. Observed changes in cell cycle distribution by LC treated might be caused by insufficient preparation for G2/M transition. Considering the structure of the LCs, the rod-like molecules displaying cytotoxicity against U937 cells possessed flexible spacers with no bulky polar group attached via the flexible spacer. Conclusions Our results revealed that some LCs showed cytotoxic properties against non-solid type tumor human leukemic cells via LC-induced S-phase arrest and decreasing expression of several cell cycle related proteins.

  7. Moesin Functions as a Lipopolysaccharide Receptor on Human Monocytes

    Science.gov (United States)

    Tohme, Ziad N.; Amar, Salomon; Van Dyke, Thomas E.

    1999-01-01

    Bacterial endotoxin (lipopolysaccharide [LPS]), a glycolipid found in the outer membranes of gram-negative bacteria, induces the secretion of proinflammatory cytokines such as tumor necrosis factor alpha (TNF-α), interleukin-1 (IL-1), and IL-6 by monocytes/macrophages. The secretion of these biologically active compounds leads to multiple pathological conditions, such as septic shock. There is substantial evidence that chronic exposure to LPS mediates, at least in part, the tissue destruction associated with gram-negative infection. CD14, a 55-kDa protein, has been identified as an LPS receptor. In conjunction with a serum protein, LPS binding protein (LBP), LPS-CD14 interactions mediate many LPS functions in the inflammatory response. However, CD14 lacks a cytoplasmic domain, or any known signal transduction sequence motif, suggesting the existence of another cell surface domain capable of transducing signals. In this paper, we report a second, CD14-independent LPS binding site, which, based on biological activity, appears to be a functional LPS receptor. Cross-linking experiments were performed to identify LPS binding sites. Two molecules were identified: a 55-kDa protein (CD14) and a second, 78-kDa band. Sequencing of the 78-kDa protein by mass spectroscopic analysis revealed 100% homology with moesin (membrane-organizing extension spike protein). Antibody to CD14 induced partial blocking of the LPS response. However, antimoesin monoclonal antibody completely blocked the LPS-induced TNF-α response in human monocytes, without blocking CD14 binding of LPS. Irrelevant isotype controls had no effect. Additional experiments were performed to evaluate the specificity of the antimoesin blocking. Separate experiments evaluated antimoesin effects on monocyte chemotaxis, IL-1 production in response to IL-1 stimulation, and TNF-α secretion in response to Staphylococcus aureus stimulation. Antimoesin blocked only LPS-mediated events. The data suggest that moesin

  8. A case of human monocytic ehrlichiosis in Serbia

    Directory of Open Access Journals (Sweden)

    Arsić Bogdan

    2014-01-01

    Full Text Available Introduction. Ehrlichiosis is a bacterial zoonosis transmitted by hematophagous arthropods - ticks. In humans, it occurs as monocytic, granulocytic, and ewingii ehrlichiosis. Pathological process is based on parasitic presence of Ehrlichia organisms within peripheral blood cells - monocytes and granulocytes. Case Outline. Fifty-two year old patient was admitted to hospital due to high fever of over 40°C that lasted two days, accompanied with chills, muscle aches, malaise, loss of appetite, headache, confusion, breathing difficulties, and mild dry cough. The history suggested tick bite that occurred seven days before the onset of disease. Doxycycline was introduced and administered for 14 days, causing the disease to subside. Indirect immunofluorescence assay was used to analyze three serum samples obtained from this patient for Ehrlichia chaffeensis antibodies, and peripheral blood smear was evaluated for the presence of Ehrlichia and Ehrlichia aggregation into morulae. Conclusion. Ehrlichiosis should be considered in each case where there is a history of tick bite together with the clinical picture (high fever, chills, muscle aches, headache, generalized weakness and malaise, and possible maculopapular rash. The presence of Ehrlichia chaffeensis antibodies was confirmed in a patient with the history of tick bite, appropriate clinical picture and indirect immunofluorescence assay. This confirmed the presence of human monocytotropic ehrlichiosis, a disease that is uncommonly identified in our country.

  9. Interleukin-1α activation and localization in lipopolysaccharide-stimulated human monocytes and macrophages

    DEFF Research Database (Denmark)

    Carlsen, Thomas Gelsing; Kjærsgaard, Pernille; Jørgensen, Trine Lykke; Foldbjerg, Rasmus; Nielsen, Mads Lausen; Guldbæk Poulsen, Thomas Bouet; Zabieglo, Katarzyna; Christiansen, Gunna; Birkelund, Svend

    2015-01-01

    - 1α in inflammation is only partly understood. Results: Human macrophages/monocytes, stimulated with lipopolysaccharide (LPS) were analyzed for production and localization of IL-1α by use of a monoclonal antibody (MAb) generated against IL-1α pro piece. We found that IL-1α propiece was detected...... being a marker for monocytes. Conclusions: Here, we demonstrate, for the first time, a method to visualize and measure the production of IL-1α in both human monocytes and macrophages....

  10. Replication-incompetent herpesvirus vector delivery of an interferon alpha gene inhibits human immunodeficiency virus replication in human monocytes.

    OpenAIRE

    Weir, J P; Elkins, K L

    1993-01-01

    Human monocytes and macrophages are nondividing cells that serve as a major reservoir for human immunodeficiency virus (HIV) at all stages of infection. To investigate viral-mediated gene delivery as a means of inhibiting HIV replication in human monocytes, a replication-incompetent herpes simplex virus vector was developed that expressed human interferon alpha. Monocytes infected with this herpes simplex virus vector and then challenged with HIV showed dramatically reduced cytopathic effects...

  11. High glucose-induced oxidative stress increases transient receptor potential channel expression in human monocytes

    DEFF Research Database (Denmark)

    Wuensch, Tilo; Thilo, Florian; Krueger, Katharina; Scholze, Alexandra; Ristow, Michael; Tepel, Martin

    2010-01-01

    Transient receptor potential (TRP) channel-induced cation influx activates human monocytes, which play an important role in the pathogenesis of atherosclerosis. In the present study, we investigated the effects of high glucose-induced oxidative stress on TRP channel expression in human monocytes....

  12. Momilactone B induces apoptosis and G1 arrest of the cell cycle in human monocytic leukemia U937 cells through downregulation of pRB phosphorylation and induction of the cyclin-dependent kinase inhibitor p21Waf1/Cip1.

    Science.gov (United States)

    Park, Cheol; Jeong, Na Young; Kim, Gi-Young; Han, Min Ho; Chung, Ill-Min; Kim, Wun-Jae; Yoo, Young Hyun; Choi, Yung Hyun

    2014-04-01

    Momilactone B, a terpenoid phytoalexin present in rice bran, has been shown to exhibit several biological activities. The present study was conducted using cultured human leukemia U937 cells to elucidate the possible mechanisms by which momilactone B exerts its anticancer activity, which to date has remained poorly understood. Momilactone B treatment of U937 cells resulted in a dose-dependent inhibition of cell growth and induced apoptotic cell death as detected by chromatin condensation, DNA fragmentation, the cleavage of poly(ADP-ribose) polymerase and Annexin V-FITC staining. Flow cytometric analysis revealed that momilactone B resulted in G1 arrest in cell cycle progression, which was associated with the dephosphorylation of retinoblastoma protein (pRB) and enhanced binding of pRB with the E2F transcription factor family proteins. Treatment with momilactone B also increased the expression of cyclin-dependent kinase (Cdk) inhibitor p21Waf1/Cip1 in a p53-independent manner, without any noticeable changes in G1 cyclins and cyclin-dependent kinases (Cdks), except a slight decrease in cyclin E. Moreover, in vitro kinase assay indicated that momilactone B significantly decreased Cdk4- and Cdk6-associated kinase activities through a notably increased binding of p21 to Cdk4 and Cdk6. Our results demonstrated that momilactone B caused G1 cell cycle arrest and apoptosis in U937 cells through the induction of p21 expression, inhibition of Cdk/cyclin-associated kinase activities, and reduced phosphorylation of pRB, which may be related to anticancer activity. PMID:24503697

  13. Sphingosylphosphorylcholine stimulates human monocyte-derived dendritic cell chemotaxis

    Institute of Scientific and Technical Information of China (English)

    Ha-young LEE; Eun-ha SHIN; Yoe-sik BAE

    2006-01-01

    Aim: To investigate the effects of Sphingosylphosphorylcholine (SPC) on human monocyte-derived dendritic cell (DC) chemotaxis. Methods: Human DC were generated from peripheral blood monocytes by culturing them with granulocyte macrophage-colony stimulating factor and interleukin-4. The effect of SPC on the DC chemotactic migration was measured by chemotaxis assay. Intracellular signaling event involved in the SPC-induced DC chemotaxis was investigated with several inhibitors for specific kinase. The expression of the SPC receptors was examined by reverse transcription polymerase chain reaction. Results: We found that SPC induced chemotactic migration in immature DC (iDC) and mature DC (mDC). In terms of SPC-induced signaling events, mitogen activated protein kinase activation and Akt activation in iDC and mDC were stimulated. SPC-induced chemotaxis was mediated by extracellular signal-regulated protein kinase and phosphoino-sitide-3-kinase, but not by calcium in both iDC and mDC. Although mDC express ovarian cancer G protein-coupled receptor 1, but not G protein-coupled receptor 4, iDC do not express any of these receptors. To examine the involvement of sphin-gosine-1-phosphate (SIP) receptors, we checked the effect of an SIP receptor antagonist (VPC23019) on SPC-induced DC chemotaxis. VPC23019 did not affect SPC-induced DC chemotaxis. Conclusion: The results suggest that SPC may play a role in regulating DC trafficking during phagocytosis and the T cell-stimulating phase, and the unique SPC receptor, which is different from SIP receptors, is involved in SPC-induced chemotaxis.

  14. CD13 mediates phagocytosis in human monocytic cells.

    Science.gov (United States)

    Licona-Limón, Ileana; Garay-Canales, Claudia A; Muñoz-Paleta, Ofelia; Ortega, Enrique

    2015-07-01

    CD13 is a membrane-bound ectopeptidase, highly expressed on monocytes, macrophages, and dendritic cells. CD13 is involved in diverse functions, including degradation of peptide mediators, cellular adhesion, migration, viral endocytosis, signaling, and positive modulation of phagocytosis mediated by FcγRs and other phagocytic receptors. In this work, we explored whether besides acting as an accessory receptor, CD13 by itself is a primary phagocytic receptor. We found that hCD13 mediates efficient phagocytosis of large particles (erythrocytes) modified so as to interact with the cell only through CD13 in human macrophages and THP-1 monocytic cells. The extent of this phagocytosis is comparable with the phagocytosis mediated through the canonical phagocytic receptor FcγRI. Furthermore, we demonstrated that hCD13 expression in the nonphagocytic cell line HEK293 is sufficient to enable these cells to internalize particles bound through hCD13. CD13-mediated phagocytosis is independent of other phagocytic receptors, as it occurs in the absence of FcγRs, CR3, and most phagocytic receptors. Phagocytosis through CD13 is independent of its enzymatic activity but is dependent on actin rearrangement and activation of PI3K and is partially dependent on Syk activation. Moreover, the cross-linking of CD13 with antibodies rapidly induced pSyk in human macrophages. Finally, we observed that antibody-mediated cross-linking of hCD13, expressed in the murine macrophage-like J774 cell line, induces production of ROS. These results demonstrate that CD13 is a fully competent phagocytic receptor capable of mediating internalization of large particles. PMID:25934926

  15. Epigenetic regulation of extracellular-superoxide dismutase in human monocytes.

    Science.gov (United States)

    Kamiya, Tetsuro; Machiura, Masatomo; Makino, Junya; Hara, Hirokazu; Hozumi, Isao; Adachi, Tetsuo

    2013-08-01

    Extracellular-superoxide dismutase (EC-SOD) is a major SOD isozyme mainly present in the vascular wall and plays an important role in normal redox homeostasis. We previously showed the significant reduction or induction of EC-SOD during human monocytic U937 or THP-1 cell differentiation induced by 12-O-tetradecanoylphorbol-13-acetate (TPA), respectively; however, its cell-specific expression and regulation have not been fully elucidated. It has been reported that epigenetic factors, such as DNA methylation and histone modification, are involved in several kinds of gene regulation. In this study, we investigated the involvement of epigenetic factors in EC-SOD expression and determined high levels of DNA methylation within promoter and coding regions of EC-SOD in THP-1 cells compared to those in U937 cells. Moreover, treatment with a DNA methyltransferase inhibitor, 5-azacytidine, significantly induced the expression of EC-SOD in THP-1 cells, indicating the importance of DNA methylation in the suppression of EC-SOD expression; however, the DNA methylation status did not change during THP-1 cell differentiation induced by TPA. On the other hand, we detected histone H3 and H4 acetylation during differentiation. Further, pretreatment with histone acetyltransferase inhibitors, CPTH2 or garcinol, significantly suppressed the TPA-inducible EC-SOD expression. We also determined the epigenetic suppression of EC-SOD in peripheral blood mononuclear cells. Treatment with granulocyte macrophage colony-stimulating factor (GM-CSF)/granulocyte-CSF induced that expression. Overall, these findings provide novel evidence that cell-specific and TPA-inducible EC-SOD expression are regulated by DNA methylation and histone H3 and H4 acetylation in human monocytic cells. PMID:23602908

  16. Peroxidatic activity distinct from myeloperoxidase in human monocytes cultured in vitro and in alveolar macrophages.

    Science.gov (United States)

    Breton-Gorius, J; Vildé, J L; Guichard, J; Vainchenker, W; Basset, F

    1982-01-01

    Human monocytes develop a peroxidatic activity (PA) in rough endoplasmic reticulum (RER) after adherence or after culture in semi-solid medium. This enzyme activity disappears after three days of culture in the majority of macrophages derived from adult monocytes but persists for one week in macrophages derived from neonatal monocytes. The PA is due to an enzyme distinct from myeloperoxidase (MPO), since monocytes from a patient with MPO deficiency develop the same PA as that of normal monocytes after adherence. By its localization and other characteristics, PA of adherent monocytes resembles that of rodent macrophages. We therefore investigated whether human alveolar macrophages exhibit PA, using a sensitive cytochemical method which prevents inhibition by aldehyde in adherent monocytes. In various pathological cases, four types of macrophages could be identified: the majority were peroxidase-negative, a small percentage was of exudate type exhibiting a PA in granules as blood monocytes, while few macrophages were intermediate, possessing only PA in RER i.e. of type resident and a smaller proportion had PA in RER and in granules i.e. exudate-resident macrophages. These findings demonstrate that human macrophages and adherent monocytes may exhibit PA in RER as has been reported for rodent macrophages. The true nature and function of the enzyme responsible for this PA, which is distinct from MPO, remains unknown, but some arguments seem to suggest its role in prostaglandin synthesis. PMID:6283838

  17. Expression and regulation of Schlafen (SLFN family members in primary human monocytes, monocyte-derived dendritic cells and T cells

    Directory of Open Access Journals (Sweden)

    Alexander Puck

    2015-01-01

    Full Text Available Schlafen (SLFN/Slfn family members have been investigated for their involvement in fundamental cellular processes including growth regulation, differentiation and control of viral replication. However, most research has been focused on the characterization of Slfns within the murine system or in human cell lines. Since little is known about SLFNs in primary human immune cells, we set out to analyze the expression and regulation of the six human SLFN genes in monocytes, monocyte-derived dendritic cells (moDCs and T cells. Comparison of SLFN gene expression across these three cell types showed high mRNA expression of SLFN11 in monocytes and moDCs and high SLFN5 expression in T cells, indicating functional importance within these cell types. Differentiation of monocytes to moDCs leads to the gradual upregulation of SLFN12L and SLFN13 while SLFN12 levels were decreased by differentiation stimuli. Stimulation of moDCs via human rhinovirus, lipopolysaccharide, or IFN-α lead to strong upregulation of SLFN gene expression, while peptidoglycan poorly stimulated regulation of both SLFNs and the classical interferon-stimulated gene MxA. T cell activation was found to downregulate the expression of SLFN5, SLFN12 and SLFN12L, which was reversible upon addition of exogenous IFN-α. In conclusion, we demonstrate, that SLFN gene upregulation is mainly dependent on autocrine type I interferon signaling in primary human immune cells. Rapid decrease of SLFN expression levels following T cell receptor stimulation indicates a role of SLFNs in the regulation of human T cell quiescence.

  18. DMPD: Differential responses of human monocytes and macrophages to IL-4 and IL-13. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 10534111 Differential responses of human monocytes and macrophages to IL-4 and IL-1...):575-8. (.png) (.svg) (.html) (.csml) Show Differential responses of human monocytes and macrophages to IL-...4 and IL-13. PubmedID 10534111 Title Differential responses of human monocytes an

  19. Microgravity modifies protein kinase C isoform translocation in the human monocytic cell line U937 and human peripheral blood T-cells

    Science.gov (United States)

    Hatton, Jason P.; Gaubert, Francois; Cazenave, Jean-Pierre; Schmitt, Didier; Hashemi, B. B. (Principal Investigator); Hughes-Fulford, M. (Principal Investigator)

    2002-01-01

    Individual protein kinase C (PKC) isoforms fulfill distinct roles in the regulation of the commitment to differentiation, cell cycle arrest, and apoptosis in both monocytes and T-cells. The human monocyte like cell line U937 and T-cells were exposed to microgravity, during spaceflight and the translocation (a critical step in PKC signaling) of individual isoforms to cell particulate fraction examined. PKC activating phorbol esters induced a rapid translocation of several PKC isoforms to the particulate fraction of U937 monocytes under terrestrial gravity (1 g) conditions in the laboratory. In microgravity, the translocation of PKC beta II, delta, and epsilon in response to phorbol esters was reduced in microgravity compared to 1 g, but was enhanced in weak hypergravity (1.4 g). All isoforms showed a net increase in particulate PKC following phorbol ester stimulation, except PKC delta which showed a net decrease in microgravity. In T-cells, phorbol ester induced translocation of PKC delta was reduced in microgravity, compared to 1 g, while PKC beta II translocation was not significantly different at the two g-levels. These data show that microgravity differentially alters the translocation of individual PKC isoforms in monocytes and T-cells, thus providing a partial explanation for the modifications previously observed in the activation of these cell types under microgravity.

  20. Gamma interferon augments Fc gamma receptor-mediated dengue virus infection of human monocytic cells.

    OpenAIRE

    Kontny, U; Kurane, I; Ennis, F A

    1988-01-01

    It has been reported that anti-dengue antibodies at subneutralizing concentrations augment dengue virus infection of monocytic cells. This is due to the increased uptake of dengue virus in the form of virus-antibody complexes by cells via Fc gamma receptors. We analyzed the effects of recombinant human gamma interferon (rIFN-gamma) on dengue virus infection of human monocytic cells. U937 cells, a human monocytic cell line, were infected with dengue virus in the form of virus-antibody complexe...

  1. High-Affinity Fc Receptor Expression Indicates Relative Immaturity in Human Monocytes.

    Science.gov (United States)

    Clanchy, Felix I L

    2016-05-01

    Within monocyte heterogeneity, subsets represent discrete, well-characterized phenotypes. Although many studies have highlighted differences between subsets, there is evidence that subpopulations represent contiguous stages in a maturational series. As CD14(hi)CD64(hi) monocytes have higher proliferative potential than CD14(hi)CD64(lo) monocytes, the surface marker profile on 4 subsets defined by CD14 and CD64 was measured. The profiles were compared to that of subsets defined by the high-affinity IgE receptor (FcɛRIα), CD16, and CD14; further differences in size, granularity, and buoyancy were measured in subsets delineated by these markers. There was a positive correlation between proliferative monocyte (PM) prevalence and CD64 expression on the classical monocyte subset, and also between PM prevalence and circulating FcɛRIα(+) monocytes. The expression of CD64, the high-affinity IgG receptor, on canonical human monocyte subsets was determined before and after short-term culture, and in response to interleukin (IL)-6, IL-10, macrophage colony-stimulating factor, granulocyte/macrophage colony-stimulating factor and interferon-γ; the influence of these cytokines on monocyte subset transition was also measured. The loss of FcɛRIα expression preceded an increase in CD16 expression in whole blood cultures. These data indicate that high-affinity Fc receptors are expressed on less mature monocytes and that FcɛRIα(+) monocytes are developmentally antecedent to the canonical classical and intermediate monocyte subsets. PMID:26714112

  2. Lymphokines Enhance the Capacity of Human Monocytes to Secrete Reactive Oxygen Intermediates

    OpenAIRE

    Nakagawara, Akira; Desantis, Nancy M.; Nogueira, Nadia; Carl F Nathan

    1982-01-01

    Supernatants from mitogen- or antigen-stimulated human blood mononuclear cells enhanced the capacity of human monocytes or monocyte-derived macrophages (MDM) to release H2O2 or O2̇̄ in response to phorbol myristate acetate or zymosan. The stimulatory effect of lymphokines (LK) lasted ∼5 d, regardless of the time of their addition. However, the magnitude of stimulation depended on whether LK were added to freshly explanted monocytes or to MDM. When LK were added on day 0 of culture, they enhan...

  3. Inhibition of human monocyte chemotaxis and chemiluminescence by Pseudomonas aeruginosa elastase

    DEFF Research Database (Denmark)

    Kharazmi, A; Nielsen, H

    1991-01-01

    The in vitro effect of Pseudomonas aeruginosa elastase on human monocyte function was examined. Mononuclear cells isolated from the peripheral blood of healthy individuals were incubated with various concentrations of elastase, and the chemotactic activity and chemiluminescence response of these ...

  4. Effect of Legionella pneumophila cytotoxic protease on human neutrophil and monocyte function

    DEFF Research Database (Denmark)

    Rechnitzer, C; Kharazmi, A

    1992-01-01

    protease on the chemotactic activity of neutrophils was demonstrated by the continued inhibition of neutrophil chemotaxis when the protease was removed following pre-incubation of the cells. In contrast, the enzyme had no effect on monocyte chemotaxis. The protease inhibited, also in a concentration...... activity of L. pneumophila protease on neutrophil chemotaxis and on the listericidal activity of human neutrophils and monocytes demonstrated in this study provides evidence for a role of this enzyme in the pathogenesis of Legionnaires' disease....

  5. Effect of extracellular serum in the stimulation of intracellular killing of streptococci by human monocytes.

    OpenAIRE

    Leijh, P C; van Zwet, T L; Furth, R. van

    1980-01-01

    This study shows that the intracellular killing of Streptococcus pyogenes, Streptococcus faecalis, and Streptococcus pneumoniae by human monocytes is stimulated by the extracellular presence of both heat-stable and heat-labile serum factors. A similar kind of stimulation of monocytes has been described in respect of catalase-positive microorganisms. However, killing of these bacteria is negligible in the absence of extracellular serum factors, whereas a large proportion of the ingested catala...

  6. Nucleolin, a shuttle protein promoting infection of human monocytes by Francisella tularensis.

    Directory of Open Access Journals (Sweden)

    Monique Barel

    Full Text Available BACKGROUND: Francisella tularensis is a highly virulent facultative intracellular bacterium, disseminating in vivo mainly within host mononuclear phagocytes. After entry into macrophages, F. tularensis initially resides in a phagosomal compartment, whose maturation is then arrested. Bacteria escape rapidly into the cytoplasm, where they replicate freely. We recently demonstrated that nucleolin, an eukaryotic protein able to traffic from the nucleus to the cell surface, acted as a surface receptor for F. tularensis LVS on human monocyte-like THP-1 cells. METHODOLOGY/PRINCIPAL FINDINGS: Here, we followed the fate of nucleolin once F. tularensis has been endocytosed. We first confirmed by siRNA silencing experiments that expression of nucleolin protein was essential for binding of LVS on human macrophage-type THP-1 cells. We then showed that nucleolin co-localized with intracellular bacteria in the phagosomal compartment. Strikingly, in that compartment, nucleolin also co-localized with LAMP-1, a late endosomal marker. Co-immunoprecipation assays further demonstrated an interaction of nucleolin with LAMP-1. Co-localization of nucleolin with LVS was no longer detectable at 24 h when bacteria were multiplying in the cytoplasm. In contrast, with an iglC mutant of LVS, which remains trapped into the phagosomal compartment, or with inert particles, nucleolin/bacteria co-localization remained almost constant. CONCLUSIONS/SIGNIFICANCE: We herein confirm the importance of nucleolin expression for LVS binding and its specificity as nucleolin is not involved in binding of another intracellular pathogen as L. monocytogenes or an inert particle. Association of nucleolin with F. tularensis during infection continues intracellularly after endocytosis of the bacteria. The present work therefore unravels for the first time the presence of nucleolin in the phagosomal compartment of macrophages.

  7. Activation of human monocytes with proteolytic gliadin fragments

    Czech Academy of Sciences Publication Activity Database

    Jelínková, L.; Tučková, Ludmila; Tlaskalová, Helena

    Elsevier. 02, č. 1 (2002), s. 91. ISSN 1568-9972. [Presentations at the International Congress on Autoimmunity /3./. 01.02.2002-02.02.2002, Geneva] Institutional research plan: CEZ:AV0Z5020903 Keywords : activation * monocytes * proteolytic Subject RIV: EE - Microbiology, Virology

  8. Differential Tropism and Chemokine Receptor Expression of Human Immunodeficiency Virus Type 1 in Neonatal Monocytes, Monocyte-Derived Macrophages, and Placental Macrophages

    OpenAIRE

    Fear, Warwick R.; Kesson, Alison M.; Naif, Hassan; Lynch, Garry W.; Cunningham, Anthony L.

    1998-01-01

    Laboratory-adapted (LA) macrophage-tropic (M-tropic) human immunodeficiency virus type 1 (HIV-1) isolates (e.g., HIV-1Ba-L) and low-passage primary (PR) isolates differed markedly in tropism for syngeneic neonatal monocytes, monocyte-derived macrophages (MDMs), and placental macrophages (PMs). Newly adherent neonatal monocytes and cultured PMs were highly refractory to infection with PR HIV-1 isolates yet were permissive for LA M-tropic isolates. Day 4 MDMs were also permissive for LA M-tropi...

  9. Interaction studies reveal specific recognition of an anti-inflammatory polyphosphorhydrazone dendrimer by human monocytes

    Science.gov (United States)

    Ledall, Jérémy; Fruchon, Séverine; Garzoni, Matteo; Pavan, Giovanni M.; Caminade, Anne-Marie; Turrin, Cédric-Olivier; Blanzat, Muriel; Poupot, Rémy

    2015-10-01

    Dendrimers are nano-materials with perfectly defined structure and size, and multivalency properties that confer substantial advantages for biomedical applications. Previous work has shown that phosphorus-based polyphosphorhydrazone (PPH) dendrimers capped with azabisphosphonate (ABP) end groups have immuno-modulatory and anti-inflammatory properties leading to efficient therapeutic control of inflammatory diseases in animal models. These properties are mainly prompted through activation of monocytes. Here, we disclose new insights into the molecular mechanisms underlying the anti-inflammatory activation of human monocytes by ABP-capped PPH dendrimers. Following an interdisciplinary approach, we have characterized the physicochemical and biological behavior of the lead ABP dendrimer with model and cell membranes, and compared this experimental set of data to predictive computational modelling studies. The behavior of the ABP dendrimer was compared to the one of an isosteric analog dendrimer capped with twelve azabiscarboxylate (ABC) end groups instead of twelve ABP end groups. The ABC dendrimer displayed no biological activity on human monocytes, therefore it was considered as a negative control. In detail, we show that the ABP dendrimer can bind both non-specifically and specifically to the membrane of human monocytes. The specific binding leads to the internalization of the ABP dendrimer by human monocytes. On the contrary, the ABC dendrimer only interacts non-specifically with human monocytes and is not internalized. These data indicate that the bioactive ABP dendrimer is recognized by specific receptor(s) at the surface of human monocytes.Dendrimers are nano-materials with perfectly defined structure and size, and multivalency properties that confer substantial advantages for biomedical applications. Previous work has shown that phosphorus-based polyphosphorhydrazone (PPH) dendrimers capped with azabisphosphonate (ABP) end groups have immuno-modulatory and anti

  10. Modulation of human monocyte/macrophage activity by tocilizumab, abatacept and etanercept: An in vitro study.

    Science.gov (United States)

    Obeng, Joyce Afrakoma; Amoruso, Angela; Camaschella, Gian Luca Ermanno; Sola, Daniele; Brunelleschi, Sandra; Fresu, Luigia Grazia

    2016-06-01

    Tocilizumab, etanercept and abatacept are biological drugs used in the therapy of Rheumatoid Arthritis (RA). Their mechanism of action is well documented but their direct effects on human monocytes/macrophages have not been fully investigated. The objective of this study was to evaluate in vitro the influence of these drugs on monocytes/macrophages from healthy volunteers. Human monocytes were isolated from healthy anonymous volunteers and cultured as such or differentiated to monocyte-derived macrophages (MDMs). The effect of tocilizumab, etanercept and abatacept (at concentrations similar to those in plasma of patients) on superoxide anion production, matrix metalloproteinase-9 (MMP-9) gene expression and activity, Peroxisome Proliferator-Activated Receptor (PPAR)γ expression and cell phenotype was evaluated. Exposure of monocytes/macrophages to tocilizumab, etanercept or abatacept resulted in a significant decrease of the PMA-induced superoxide anion production. Interestingly, the expression of PPARγ was significantly increased only by tocilizumab, while etanercept was the only one able to significantly reduce MMP-9 gene expression and inhibit the LPS-induced MMP-9 activity in monocytes. When etanercept and abatacept were added to the differentiating medium, both significantly reduced the amount of CD206(+)MDM. This study demonstrates that etanercept, abatacept and tocilizumab affect differently human monocytes/macrophages. In particular, the IL-6 antagonist tocilizumab seems to be more effective in inducing an anti-inflammatory phenotype of monocytes/macrophages compared to etanercept and abatacept, also in light of the up-regulation of PPARγ whose anti-inflammatory effects are well recognised. PMID:26997366

  11. A curated compendium of monocyte transcriptome datasets of relevance to human monocyte immunobiology research [version 2; referees: 2 approved

    Directory of Open Access Journals (Sweden)

    Darawan Rinchai

    2016-04-01

    Full Text Available Systems-scale profiling approaches have become widely used in translational research settings. The resulting accumulation of large-scale datasets in public repositories represents a critical opportunity to promote insight and foster knowledge discovery. However, resources that can serve as an interface between biomedical researchers and such vast and heterogeneous dataset collections are needed in order to fulfill this potential. Recently, we have developed an interactive data browsing and visualization web application, the Gene Expression Browser (GXB. This tool can be used to overlay deep molecular phenotyping data with rich contextual information about analytes, samples and studies along with ancillary clinical or immunological profiling data. In this note, we describe a curated compendium of 93 public datasets generated in the context of human monocyte immunological studies, representing a total of 4,516 transcriptome profiles. Datasets were uploaded to an instance of GXB along with study description and sample annotations. Study samples were arranged in different groups. Ranked gene lists were generated based on relevant group comparisons. This resource is publicly available online at http://monocyte.gxbsidra.org/dm3/landing.gsp.

  12. Studies on the metabolism of triphenylphosphate by carboxylesterases and human monocytes

    International Nuclear Information System (INIS)

    Resin workers exposed to triphenylphosphate (TPP), an organophosphate (OP) flame retardant and plasticizer, had a decreased expression of carboxylesterase (CBE) activity in their peripheral blood monocytes. The mechanisms of CBE inhibition by TPP were investigated using purified hog liver CBE and intact human monocytes. TPP inactivated hog liver CBE in a time and dose dependent manner, and this inhibition was partially reversed by alkaline phosphatase (AP). Analysis of [14C]TPP metabolites from the enzymatic reaction by high performance liquid chromatography (HPLC) and gas chromatography/mass spectrometry (GM-C) identified phenol as the hydrolytic metabolite of TPP. Human monocytes cultured with [14C]TPP also released phenol. In addition to phenol, several phenol metabolites, such as catechol, hydroquinone, 2,2 biphenol and 4,4 biphenol were also generated by monocytes. An identical pattern of these metabolites was also formed from monocytes incubated with radiolabelled phenol. This cellular degradation of TPP was inhibited by diisopropylfluorophosphate (DFP), but not observed in neutrophil or lymphocyte cultures. Activation of monocytes with gamma interferon (IFN-g), f-Met-Leu-Phe, and serum treated zymosan (STZ) enhanced the levels of phenolic metabolites and, further, shifted the metabolism of TPP towards the formation of the biphenolic metabolites

  13. The effect of short-chain fatty acids on human monocyte-derived dendritic cells

    DEFF Research Database (Denmark)

    Nastasi, Claudia; Candela, Marco; Bonefeld, Charlotte Menné;

    2015-01-01

    negligible effects, while both butyrate and propionate strongly modulated gene expression in both immature and mature human monocyte-derived DC. An Ingenuity pathway analysis based on the differentially expressed genes suggested that propionate and butyrate modulate leukocyte trafficking, as SCFA strongly......The gut microbiota is essential for human health and plays an important role in the pathogenesis of several diseases. Short-chain fatty acids (SCFA), such as acetate, butyrate and propionate, are end-products of microbial fermentation of macronutrients that distribute systemically via the blood....... The aim of this study was to investigate the transcriptional response of immature and LPS-matured human monocyte-derived DC to SCFA. Our data revealed distinct effects exerted by each individual SCFA on gene expression in human monocyte-derived DC, especially in the mature ones. Acetate only exerted...

  14. Cellular Effects of Insulin in Human THP-1 Monocytes

    OpenAIRE

    Naderi, Jamal

    2015-01-01

    Excess of insulin in the circulating blood relative to the level of glucose (Hyperinsulinemia) may be linked to systemic low-grade inflammation and some chronic disease such as the metabolic syndrome, type 2 diabetes (T2D), and cardiovascular disease (CVD). The function of high levels of insulin in vascular smooth muscle cells (VSMCs), endothelial cells (ECs), and macrophages in relation to CVD has been investigated, but exact role of high insulin levels in monocytes, as an important cel...

  15. Dendritic Cells and Monocytes with Distinct Inflammatory Responses Reside in Lung Mucosa of Healthy Humans.

    Science.gov (United States)

    Baharom, Faezzah; Thomas, Saskia; Rankin, Gregory; Lepzien, Rico; Pourazar, Jamshid; Behndig, Annelie F; Ahlm, Clas; Blomberg, Anders; Smed-Sörensen, Anna

    2016-06-01

    Every breath we take contains potentially harmful pathogens or allergens. Dendritic cells (DCs), monocytes, and macrophages are essential in maintaining a delicate balance of initiating immunity without causing collateral damage to the lungs because of an exaggerated inflammatory response. To document the diversity of lung mononuclear phagocytes at steady-state, we performed bronchoscopies on 20 healthy subjects, sampling the proximal and distal airways (bronchial wash and bronchoalveolar lavage, respectively), as well as mucosal tissue (endobronchial biopsies). In addition to a substantial population of alveolar macrophages, we identified subpopulations of monocytes, myeloid DCs (MDCs), and plasmacytoid DCs in the lung mucosa. Intermediate monocytes and MDCs were highly frequent in the airways compared with peripheral blood. Strikingly, the density of mononuclear phagocytes increased upon descending the airways. Monocytes from blood and airways produced 10-fold more proinflammatory cytokines than MDCs upon ex vivo stimulation. However, airway monocytes were less inflammatory than blood monocytes, suggesting a more tolerant nature. The findings of this study establish how to identify human lung mononuclear phagocytes and how they function in normal conditions, so that dysregulations in patients with respiratory diseases can be detected to elucidate their contribution to immunity or pathogenesis. PMID:27183618

  16. Differential Oxidative Stress Induced by Dengue Virus in Monocytes from Human Neonates, Adult and Elderly Individuals

    Science.gov (United States)

    Valero, Nereida; Mosquera, Jesús; Añez, Germán; Levy, Alegria; Marcucci, Rafael; de Mon, Melchor Alvarez

    2013-01-01

    Changes in immune response during lifespan of man are well known. These changes involve decreased neonatal and elderly immune response. In addition, it has been shown a relationship between immune and oxidative mechanisms, suggesting that altered immune response could be associated to altered oxidative response. Increased expression of nitric oxide (NO) has been documented in dengue and in monocyte cultures infected with different types of dengue virus. However, there is no information about the age-dependent NO oxidative response in humans infected by dengue virus. In this study, monocyte cultures from neonatal, elderly and adult individuals (n = 10 each group) were infected with different dengue virus types (DENV- 1 to 4) and oxidative/antioxidative responses and apoptosis were measured at days 1 and 3 of culture. Increased production of NO, lipid peroxidation and enzymatic and nonenzymatic anti-oxidative responses in dengue infected monocyte cultures were observed. However, neonatal and elderly monocytes had lower values of studied parameters when compared to those in adult-derived cultures. Apoptosis was present in infected monocytes with higher values at day 3 of culture. This reduced oxidant/antioxidant response of neonatal and elderly monocytes could be relevant in the pathogenesis of dengue disease. PMID:24069178

  17. β2-Agonist clenbuterol hinders human monocyte differentiation into dendritic cells.

    Science.gov (United States)

    Giordani, Luciana; Cuzziol, Noemi; Del Pinto, Tamara; Sanchez, Massimo; Maccari, Sonia; Massimi, Alessia; Pietraforte, Donatella; Viora, Marina

    2015-12-10

    Clenbuterol (CLB) is a beta2-adrenergic agonist commonly used in asthma therapy, but is also a non-steroidal anabolic drug often abused in sport doping practices. Here we evaluated the in vitro impact of CLB on the physiology and function of human monocytes and dendritic cells (DCs), instrumental in the development of immune responses. We demonstrate that CLB inhibits the differentiation of monocytes into DCs and this effect is specific and dependent on β2-adrenergic receptor (AR) activation. We found that CLB treatment reduced the percentage of CD1a(+) immature DCs, while increasing the frequency of monocytes retaining CD14 surface expression. Moreover, CLB inhibited tumor necrosis factor-alpha (TNF-alpha) enhanced IL-(interleukin)-10 and IL-6 production. In contrast, CLB did not modulate the phenotypic and functional properties of monocytes and DCs, such as the surface expression of HLA-DR, CD83, CD80 and CD86 molecules, cytokine production, immunostimulatory activity and phagocytic activity. Moreover, we found that CLB did not modulate the activation of NF-kB in DCs. Moreover, we found that the differentiation of monocytes into DCs was associated with a significant decrease of β2-ARs mRNA expression. These results provide new insights on the effect of CLB on monocyte differentiation into DCs. Considering the frequent illegal use of CLB in doping, our work suggests that this drug is potentially harmful to immune responses decreasing the supply of DCs, thus subverting immune surveillance. PMID:26524508

  18. Homocysteine induces production of monocyte chemoattractant protein-1 and interleukin-8 in cultured human whole blood

    Institute of Scientific and Technical Information of China (English)

    Xiao-kun ZENG; Daniel G REMICK; Xian WANG

    2004-01-01

    AIM: To investigate whether increased plasma L-homocysteine (Hcy) level could promote monocyte chemoattractant protein-1 (MCP-1) and interleukin-8 (IL-8) in cultured whole blood. METHODS: Human whole blood or different type of peripheral blood cells from health volunteers were incubated with Hcy and/or the inhibitors. MCP- 1 and IL-8 level were measured by ELISA assay. RESULTS: Hcy 10-1000 μmol/L induced production of MCP-1 and IL-8 in cultured human whole blood (P<0.05). The major cellular source of these chemokines comed from monocytes.Meanwhile,Hcy also promoted the upregulation of MPO level even at the 10 μmol/L in the cultured whole blood.secretion in cultured human whole blood, especially in monocytes via oxidative stress mechanism.

  19. Capacity of human monocytes to phagocytose approved iron oxide MR contrast agents in vitro

    Energy Technology Data Exchange (ETDEWEB)

    Metz, Stephan; Settles, Marcus; Rummeny, Ernst J.; Daldrup-Link, Heike E. [Technical University Munich, Department of Diagnostic Radiology, Munich (Germany); Bonaterra, Gabriel [Ruprecht-Karls-University, Institute of Anatomy and Cell Biology, Heidelberg (Germany); Rudelius, Martina [Technical University Munich, Department of Pathology, Munich (Germany)

    2004-10-01

    To evaluate the capacity of human monocytes to phagocytose various approved iron oxide based magnetic resonance (MR) contrast agents and to optimize in vitro labeling of these cells. Human monocytes were incubated with two superparamagnetic iron oxide particles (SPIO) as well as two ultrasmall SPIO (USPIO) at varying iron oxide concentrations and incubation times. Iron uptake in monocytes was proven by histology, quantified by atomic emission absorption spectrometry and depicted with T2* weighted fast field echo (FFE) MR images at 1.5 T. Additionally, induction of apoptosis in iron oxide labeled monocytes was determined by YO-PRO-1 staining. Cellular iron uptake was significantly (P<0.01) higher after incubation with SPIO compared with USPIO. For SPIO, the iron oxide uptake was significantly (P<0.01) higher after incubation with the ionic Ferucarbotran as compared with the non-ionic Ferumoxides. Efficient cell labeling was achieved after incubation with Ferucarbotran at concentrations {>=}500 {mu}g Fe/ml and incubation times {>=}1 h, resulting in a maximal iron oxide uptake of up to 50 pg Fe/cell without impairment of cell viability. In vitro labeling of human monocytes for MR imaging is most effectively obtained with the approved SPIO Ferucarbotran. Potential subsequent in vivo cell tracking applications comprise, e.g. specific targeting of inflammatory processes. (orig.)

  20. Capacity of human monocytes to phagocytose approved iron oxide MR contrast agents in vitro

    International Nuclear Information System (INIS)

    To evaluate the capacity of human monocytes to phagocytose various approved iron oxide based magnetic resonance (MR) contrast agents and to optimize in vitro labeling of these cells. Human monocytes were incubated with two superparamagnetic iron oxide particles (SPIO) as well as two ultrasmall SPIO (USPIO) at varying iron oxide concentrations and incubation times. Iron uptake in monocytes was proven by histology, quantified by atomic emission absorption spectrometry and depicted with T2* weighted fast field echo (FFE) MR images at 1.5 T. Additionally, induction of apoptosis in iron oxide labeled monocytes was determined by YO-PRO-1 staining. Cellular iron uptake was significantly (P<0.01) higher after incubation with SPIO compared with USPIO. For SPIO, the iron oxide uptake was significantly (P<0.01) higher after incubation with the ionic Ferucarbotran as compared with the non-ionic Ferumoxides. Efficient cell labeling was achieved after incubation with Ferucarbotran at concentrations ≥500 μg Fe/ml and incubation times ≥1 h, resulting in a maximal iron oxide uptake of up to 50 pg Fe/cell without impairment of cell viability. In vitro labeling of human monocytes for MR imaging is most effectively obtained with the approved SPIO Ferucarbotran. Potential subsequent in vivo cell tracking applications comprise, e.g. specific targeting of inflammatory processes. (orig.)

  1. Monoclonal antibodies against human Ia antigens stimulate monocytes to secrete interleukin 1.

    OpenAIRE

    Palacios, R

    1985-01-01

    The monoclonal antibodies (mAb) DA6.147, DA6.164, and HIG.48 against human Ia antigens, but not the W6/32 mAb against human class I major histocompatibility complex antigens or the anti-monocyte OKM1 and 63D3 mAb, stimulated monocytes to secrete interleukin 1 (IL-1). IL-1 was measured by its property of promoting the production of interleukin 2 (IL-2) by phytohemagglutinin-treated LBRM-33 clone 1A5 cells. IL-1 activity induced by anti-Ia antibodies could be detected 24 hr after initiation of ...

  2. Binding of Streptococcus mutans SR protein to human monocytes: production of tumor necrosis factor, interleukin 1, and interleukin 6.

    Science.gov (United States)

    Soell, M; Holveck, F; Schöller, M; Wachsmann, R D; Klein, J P

    1994-05-01

    To examine the possible implication of protein SR, an I/II-related antigen from Streptococcus mutans OMZ 175 (serotype f), in inflammatory reactions, we tested the immunomodulatory effects of protein SR on human monocytes. Using biotinylated protein, we provide evidence that protein SR binds to human monocytes in dose-, time-, and calcium-dependent manners through specific interactions. These results were confirmed by competition experiments using either soluble human monocyte extract or anti-SR immunoglobulin G. Binding occurred through lectin-like interactions between SR and carbohydrate portions of monocyte membrane glycoproteins, since binding could be inhibited by several sugars, especially fucose and N-acetylneuraminic acid (NANA), which were confirmed by ligand blotting to be the primer ligands recognized by SR on human monocyte extracts. The ability of protein SR to stimulate the production of cytokines by human circulating monocytes was then examined. The release of tumor necrosis factor alpha (TNF-alpha), interleukin 1 beta, and interleukin 6 is time and dose dependent and not affected by the addition of polymyxin B. Activation of monocytes resulted from specific binding of SR to NANA and fucose present on cell surface glycoproteins since TNF-alpha release could be inhibited by sialidase and pronase treatment of monocytes and by NANA and fucose. These results confirm that sialic acid and fucose present on cell surface macromolecules and especially glycoproteins are needed for the binding of SR to monocytes and for the release of TNF-alpha. PMID:8168943

  3. HIV-1 induces IL-10 production in human monocytes via a CD4-independent pathway.

    Science.gov (United States)

    Ji, Jiaxiang; Sahu, Gautam K; Braciale, Vivian L; Cloyd, Miles W

    2005-06-01

    In HIV-infected patients, increased levels of IL-10, mainly produced by virally infected monocytes, were reported to be associated with impaired cell-mediated immune responses. In this study, we investigated how HIV-1 induces IL-10 production in human monocytes. We found that CD14(+) monocytes infected by either HIV-1(213) (X4) or HIV-1(BaL) (R5) produced IL-10, IL-6, tumor necrosis factor-alpha (TNF-alpha), and to a lesser extent, IFN-gamma. However, the capacity of HIV-1 to induce these cytokines was not dependent on virus replication since UV-inactivated HIV-1 induced similar levels of these cytokines. In addition, soluble HIV-1 gp160 could induce CD14(+) monocytes to produce IL-10 but at lower levels. Cross-linking CD4 molecules (XLCD4) with anti-CD4 mAbs and goat anti-mouse IgG (GAM) resulted in high levels of IL-6, TNF-alpha and IFN-gamma but no IL-10 production by CD14(+) monocytes. Interestingly, neither anti-CD4 mAbs nor recombinant soluble CD4 (sCD4) receptor could block IL-10 secretion induced by HIV-1(213), HIV-1(BaL) or HIV-1 gp160 in CD14(+) monocytes, whereas anti-CD4 mAb or sCD4 almost completely blocked the secretion of the other cytokines. Furthermore, HIV-1(213) could induce IL-10 mRNA expression in CD14(+) monocytes while XLCD4 by anti-CD4 mAb and GAM failed to do so. As with IL-10 protein levels, HIV-1(213)-induced IL-10 mRNA expression in CD14(+) monocytes could not be inhibited by anti-CD4 mAb or sCD4. Taken together, HIV-1 binding to CD14(+) monocytes can induce CD4-independent IL-10 production at both mRNA and protein levels. This finding suggests that HIV induces the immunosuppressive IL-10 production in monocytes and is not dependent on CD4 molecules and that interference with HIV entry through CD4 molecules may have no impact on counteracting the effects of IL-10 during HIV infection. PMID:15937058

  4. Involvement of Signaling Molecules on Na+/H+ Exchanger-1 Activity in Human Monocytes

    OpenAIRE

    Sarigianni, Maria; Tsapas, Apostolos; Mikhailidis, Dimitri P; Kaloyianni, Martha; Koliakos, George; Paletas, Konstantinos

    2010-01-01

    Background: Sodium/hydrogen exchanger-1 (NHE-1) contributes to maintaining intracellular pH (pHi). We assessed the effect of glucose, insulin, leptin and adrenaline on NHE-1 activity in human monocytes in vitro. These cells play a role in atherogenesis and disturbances in the hormones evaluated are associated with obesity and diabetes. Methods and Results: Monocytes were isolated from 16 healthy obese and 10 lean healthy subjects. NHE-1 activity was estimated by measuring pHi with a fluoresce...

  5. In-Vitro differentiation of mature dendritic cells from human blood monocytes

    OpenAIRE

    Robert Gieseler; Dirk Heise; Afsaneh Soruri; Peter Schwartz; J. Hinrich Peters

    1998-01-01

    Representing the most potent antigen-presenting cells, dendritic cells (DC) can now be generated from human blood monocytes. We recently presented a novel protocol employing GM-CSF, IL-4, and IFN-γ to differentiate monocyte-derived DC in vitro. Here, such cells are characterized in detail. Cells in culture exhibited both dendritic and veiled morphologies, the former being adherent and the latter suspended. Phenotypically, they were CD1a-/dim, CD11a+, CD11b++, CD11c+, CD14dim/-, CD16a-/dim, CD...

  6. Isolation of IL-12p70-competent human monocyte-derived dendritic cells

    DEFF Research Database (Denmark)

    Søndergaard, Jonas Nørskov; Pedersen, Susanne Brix

    Diverse methodologies ranging from experimental immunological studies to immunotherapy involve the application of human monocyte-derived dendritic cells (moDCs). Considerable donor-dependent variations in the moDC production of IL-12p70 affect the outcome of these methodologies. It has been shown...

  7. Cytokine and Eicosanoid Production by Cultured Human Monocytes Exposed to Titanium Particulate Debris

    Science.gov (United States)

    Robinson, Timothy M.; Manley, Paul A.; Sims, Paul A.; Albrecht, Ralph; Darien, Benjamin J.

    1999-10-01

    Phagocytosis of particulate wear debris from arthroplasties by macrophages induces an inflammatory response that has been linked to implant loosening and premature failure of artificial joints. Inflammatory mediators released by phagocytic macrophages such as tumor necrosis factor-a (TNF-[alpha]), interleukin-1[beta] (IL-1[beta]), interleukin-6 (IL-6), and prostaglandin E2 (PGE2) are believed to play a central role in the pathogenesis of aseptic loosening. The objective of this study was to characterize titanium alloy particulates that closely match wear debris found around joint arthroplasties and to study their effects on the biosynthesis of inflammatory mediators by cultured monocytes. Peripheral blood monocytes were isolated from healthy human volunteers. Monocytes were cultured in 96-well plates for 24 h, washed, and exposed to three concentrations of titanium particulates and controls from 18Ð24 h. Supernatants were assayed for TNF-[alpha], IL-1[beta], IL-6, and PGE2 activity. Energy dispersive X-ray spectroscopy (EDX) verified the titanium alloy to be Ti6A14V. Scanning electron microscopy (SEM) analysis showed significant titanium particulate heterogeneity with approximately 95% of the particles titanium particulates utilized for in vitro models of titanium-induced cytokine release by monocytes. Incubation of titanium particulates (in concentrations similar to those found around loosened prosthetic joints) with cultured monocytes significantly increased their production of TNF-[alpha], IL-1[beta], and PGE2.

  8. CD14+ monocytes promote the immunosuppressive effect of human umbilical cord matrix stem cells

    International Nuclear Information System (INIS)

    Here, the effect of CD14+ monocytes on human umbilical cord matrix stem cell (hUC-MSC)-mediated immunosuppression was studied in vitro. hUC-MSCs exerted a potent inhibitory effect on the proliferation and interferon-γ (IFN-γ) secretion capacities of CD4+ and CD8+ T cells in response to anti-CD3/CD28 stimulation. Transwell co-culture system revealed that the suppressive effect was primarily mediated by soluble factors. Addition of prostaglandin synthesis inhibitors (indomethacin or NS-398) almost completely abrogated the immunosuppression activity of hUC-MSCs, identifying prostaglandin E2 (PGE2) as an important soluble mediator. CD14+ monocytes were found to be able to enhance significantly the immunosuppressive effect of hUC-MSCs in a dose-dependent fashion. Moreover, the inflammatory cytokine IL-1β, either exogenously added or produced by CD14+ monocytes in culture, could trigger expression of high levels of PGE2 by hUC-MSCs, whereas inclusion of the IL-1 receptor antagonist (IL-1RA) in the culture down-regulated not only PGE2 expression, but also reversed the promotional effect of CD14+ monocytes and partially restored CD4+ and CD8+ T cell proliferation and IFN-γ secretion. Our data demonstrate an important role of monocytes in the hUC-MSC-induced immunomodulation, which may have important implications in future efforts to explore the clinical potentials of hUC-MSCs.

  9. Effect of glycolipids of Leishmania parasites on human monocyte activity. Inhibition by lipophosphoglycan.

    Science.gov (United States)

    Frankenburg, S; Leibovici, V; Mansbach, N; Turco, S J; Rosen, G

    1990-12-15

    Lipophosphoglycan (LPG) and glycosyl phosphatidylinositol Ag (GPI), are glycolipids present on the membrane of Leishmania parasites. Both glycolipids have been chemically characterized. LPG is a polysaccharide of repeating phosphorylated units linked to a phosphocarbohydrate core that is anchored to the membrane by lysoalkyl phosphatidylinositol (PI). The GPI are smaller glycolipids with a structure resembling the phosphocarbohydrate core of the LPG. They are anchored to the membrane by alkyl acyl PI. Their relative abundance, uniqueness of structure, and cellular location suggest a role in interactions of the parasites with host cells. In the present study we examined the effect of LPG and GPI on the activation of human peripheral blood monocytes. Three parameters were studied: the production of IL-1, chemotactic locomotion, and oxidative burst. We found that whereas neither GPI nor LPG directly affected monocyte activity, preincubation of the monocytes with LPG strongly inhibited further activation: The production of IL-1, after stimulation with LPS, was decreased in a dose-dependent manner. Previous incubation with LPG also inhibited chemotactic locomotion of monocytes and neutrophils in response to diacylglycerol, zymosan-activated serum, FMLP and LTB4. Luminol-dependent chemiluminiscence caused by stimulation of the monocytes with streptococci and histone was also inhibited. After fragmentation of the LPG into phosphoglycan and 1-O-alkylglycerol by phosphatidylinositol-phospholipase C, only the phosphoglycan retained inhibitory activity. No difference in inhibitory activity was found between LPG prepared from Leishmania major or Leishmania donovani promastigotes. These results show that the phosphoglycan of LPG inhibits the immunologic response of normal human monocytes and neutrophils, and suggest that LPG may influence the nature of the inflammatory response surrounding infected cells. PMID:2147940

  10. Interferon-beta induces distinct gene expression response patterns in human monocytes versus T cells.

    Directory of Open Access Journals (Sweden)

    Noa Henig

    Full Text Available BACKGROUND: Monocytes, which are key players in innate immunity, are outnumbered by neutrophils and lymphocytes among peripheral white blood cells. The cytokine interferon-β (IFN-β is widely used as an immunomodulatory drug for multiple sclerosis and its functional pathways in peripheral blood mononuclear cells (PBMCs have been previously described. The aim of the present study was to identify novel, cell-specific IFN-β functions and pathways in tumor necrosis factor (TNF-α-activated monocytes that may have been missed in studies using PBMCs. METHODOLOGY/PRINCIPAL FINDINGS: Whole genome gene expression profiles of human monocytes and T cells were compared following in vitro priming to TNF-α and overnight exposure to IFN-β. Statistical analyses of the gene expression data revealed a cell-type-specific change of 699 transcripts, 667 monocyte-specific transcripts, 21 T cell-specific transcripts and 11 transcripts with either a difference in the response direction or a difference in the magnitude of response. RT-PCR revealed a set of differentially expressed genes (DEGs, exhibiting responses to IFN-β that are modulated by TNF-α in monocytes, such as RIPK2 and CD83, but not in T cells or PBMCs. Known IFN-β promoter response elements, such as ISRE, were enriched in T cell DEGs but not in monocyte DEGs. The overall directionality of the gene expression regulation by IFN-β was different in T cells and monocytes, with up-regulation more prevalent in T cells, and a similar extent of up and down-regulation recorded in monocytes. CONCLUSIONS: By focusing on the response of distinct cell types and by evaluating the combined effects of two cytokines with pro and anti-inflammatory activities, we were able to present two new findings First, new IFN-β response pathways and genes, some of which were monocytes specific; second, a cell-specific modulation of the IFN-β response transcriptome by TNF-α.

  11. Fibronectin and serum amyloid P component stimulate C3b- and C3bi- mediated phagocytosis in cultured human monocytes

    OpenAIRE

    1983-01-01

    Fibronectin (FN) and serum amyloid P component (SAP) markedly enhance phagocytosis mediated by the C3b and C3bi receptors of cultured human monocytes but not of granulocytes. (The C3b and C3bi receptors of granulocytes can be activated by treatment of these phagocytes with PMA.) Activation of monocyte C3 receptors by FN is developmentally regulated: Freshly explanted monocytes respond to FN with a small increase in C3 receptor-mediated phagocytosis while monocytes matured in culture exhibit a...

  12. The Bacillus anthracis cholesterol-dependent cytolysin, Anthrolysin O, kills human neutrophils, monocytes and macrophages

    Directory of Open Access Journals (Sweden)

    Rest Richard F

    2006-06-01

    Full Text Available Abstract Background Bacillus anthracis is an animal and human pathogen whose virulence is characterized by lethal and edema toxin, as well as a poly-glutamic acid capsule. In addition to these well characterized toxins, B. anthracis secretes several proteases and phospholipases, and a newly described toxin of the cholesterol-dependent cytolysin (CDC family, Anthrolysin O (ALO. Results In the present studies we show that recombinant ALO (rALO or native ALO, secreted by viable B. anthracis, is lethal to human primary polymorphonuclear leukocytes (PMNs, monocytes, monocyte-derived macrophages (MDMs, lymphocytes, THP-1 monocytic human cell line and ME-180, Detroit 562, and A549 epithelial cells by trypan blue exclusion or lactate dehydrogenase (LDH release viability assays. ALO cytotoxicity is dose and time dependent and susceptibility to ALO-mediated lysis differs between cell types. In addition, the viability of monocytes and hMDMs was assayed in the presence of vegetative Sterne strains 7702 (ALO+, UT231 (ALO-, and a complemented strain expressing ALO, UT231 (pUTE544, and was dependent upon the expression of ALO. Cytotoxicity of rALO is seen as low as 0.070 nM in the absence of serum. All direct cytotoxic activity is inhibited by the addition of cholesterol or serum concentration as low as 10%. Conclusion The lethality of rALO and native ALO on human monocytes, neutrophils, macrophages and lymphocytes supports the idea that ALO may represent a previously unidentified virulence factor of B. anthracis. The study of other factors produced by B. anthracis, along with the major anthrax toxins, will lead to a better understanding of this bacterium's pathogenesis, as well as provide information for the development of antitoxin vaccines for treating and preventing anthrax.

  13. The inability of human immunodeficiency virus to infect chimpanzee monocytes can be overcome by serial viral passage in vivo.

    OpenAIRE

    Gendelman, H E; Ehrlich, G D; Baca, L M; Conley, S; Ribas, J.; Kalter, D C; Meltzer, M S; Poiesz, B J; Nara, P

    1991-01-01

    Studies of lentivirus infection in ruminants, nonhuman primates, and humans suggest that virus infection of macrophages plays a central role in the disease process. To investigate whether human immunodeficiency virus type 1 (HIV-1) can infect chimpanzee macrophages, we recovered monocytes from peripheral blood mononuclear cells of HIV-1-negative animals and inoculated these and control human monocytes with a panel of four human-passaged monocytotropic virus strains and one chimpanzee-passaged...

  14. Moderate restriction of macrophage-tropic human immunodeficiency virus type 1 by SAMHD1 in monocyte-derived macrophages.

    Directory of Open Access Journals (Sweden)

    Kahoru Taya

    Full Text Available Macrophage-tropic human immunodeficiency virus type 1 (HIV-1 strains are able to grow to high titers in human monocyte-derived macrophages. However, it was recently reported that cellular protein SAMHD1 restricts HIV-1 replication in human cells of the myeloid lineage, including monocyte-derived macrophages. Here we show that degradation of SAMHD1 in monocyte-derived macrophages was associated with moderately enhanced growth of the macrophage-tropic HIV-1 strain. SAMHD1 degradation was induced by treating target macrophages with vesicular stomatitis virus glycoprotein-pseudotyped human immunodeficiency virus type 2 (HIV-2 particles containing viral protein X. For undifferentiated monocytes, HIV-2 particle treatment allowed undifferentiated monocytes to be fully permissive for productive infection by the macrophage-tropic HIV-1 strain. In contrast, untreated monocytes were totally resistant to HIV-1 replication. These results indicated that SAMHD1 moderately restricts even a macrophage-tropic HIV-1 strain in monocyte-derived macrophages, whereas the protein potently restricts HIV-1 replication in undifferentiated monocytes.

  15. Extracellular lipase of Pseudomonas aeruginosa: biochemical characterization and effect on human neutrophil and monocyte function in vitro

    DEFF Research Database (Denmark)

    Jaeger, K E; Kharazmi, A; Høiby, N

    1991-01-01

    concentrations of this lipase preparation were preincubated with human peripheral blood neutrophils and monocytes. The chemotaxis and chemiluminescence of these cells were then determined. It was shown that lipase inhibited the monocyte chemotaxis and chemiluminescence, whereas it had no or very little effect on...

  16. Serine leucocyte proteinase inhibitor-treated monocyte inhibits human CD4(+) lymphocyte proliferation.

    Science.gov (United States)

    Guerrieri, Diego; Tateosian, Nancy L; Maffía, Paulo C; Reiteri, Romina M; Amiano, Nicolás O; Costa, María J; Villalonga, Ximena; Sanchez, Mercedes L; Estein, Silvia M; Garcia, Verónica E; Sallenave, Jean-Michel; Chuluyan, Héctor E

    2011-08-01

    Serine leucocyte proteinase inhibitor (SLPI) is the main serine proteinase inhibitor produced by epithelial cells and has been shown to be a pleiotropic molecule with anti-inflammatory and microbicidal activities. However, the role of SLPI on the adaptive immune response is not well established. Therefore, we evaluated the effect of SLPI on lymphocyte proliferation and cytokine production. Human peripheral blood mononuclear cells (PBMC) were treated with mitogens plus SLPI and proliferation was assessed by [(3) H]thymidine uptake. The SLPI decreased the lymphocyte proliferation induced by interleukin-2 (IL-2) or OKT3 monoclonal antibodies in a dose-dependent manner. Inhibition was not observed when depleting monocytes from the PBMC and it was restored by adding monocytes and SLPI. SLPI-treated monocyte slightly decreased MHC II and increased CD18 expression, and secreted greater amounts of IL-4, IL-6 and IL-10 in the cell culture supernatants. SLPI-treated monocyte culture supernatant inhibited the CD4(+) lymphocyte proliferation but did not affect the proliferation of CD8(+) cells. Moreover, IL-2 increased T-bet expression and the presence of SLPI significantly decreased it. Finally, SLPI-treated monocyte culture supernatant dramatically decreased interferon-γ but increased IL-4, IL-6 and IL-10 in the presence of IL-2-treated T cells. Our results demonstrate that SLPI target monocytes, which in turn inhibit CD4 lymphocyte proliferation and T helper type 1 cytokine secretion. Overall, these results suggest that SLPI is an alarm protein that modulates not only the innate immune response but also the adaptive immune response. PMID:21574992

  17. Serine leucocyte proteinase inhibitor-treated monocyte inhibits human CD4+ lymphocyte proliferation

    Science.gov (United States)

    Guerrieri, Diego; Tateosian, Nancy L; Maffía, Paulo C; Reiteri, Romina M; Amiano, Nicolás O; Costa, María J; Villalonga, Ximena; Sanchez, Mercedes L; Estein, Silvia M; Garcia, Verónica E; Sallenave, Jean-Michel; Chuluyan, Héctor E

    2011-01-01

    Serine leucocyte proteinase inhibitor (SLPI) is the main serine proteinase inhibitor produced by epithelial cells and has been shown to be a pleiotropic molecule with anti-inflammatory and microbicidal activities. However, the role of SLPI on the adaptive immune response is not well established. Therefore, we evaluated the effect of SLPI on lymphocyte proliferation and cytokine production. Human peripheral blood mononuclear cells (PBMC) were treated with mitogens plus SLPI and proliferation was assessed by [3H]thymidine uptake. The SLPI decreased the lymphocyte proliferation induced by interleukin-2 (IL-2) or OKT3 monoclonal antibodies in a dose-dependent manner. Inhibition was not observed when depleting monocytes from the PBMC and it was restored by adding monocytes and SLPI. SLPI-treated monocyte slightly decreased MHC II and increased CD18 expression, and secreted greater amounts of IL-4, IL-6 and IL-10 in the cell culture supernatants. SLPI-treated monocyte culture supernatant inhibited the CD4+ lymphocyte proliferation but did not affect the proliferation of CD8+ cells. Moreover, IL-2 increased T-bet expression and the presence of SLPI significantly decreased it. Finally, SLPI-treated monocyte culture supernatant dramatically decreased interferon-γ but increased IL-4, IL-6 and IL-10 in the presence of IL-2-treated T cells. Our results demonstrate that SLPI target monocytes, which in turn inhibit CD4 lymphocyte proliferation and T helper type 1 cytokine secretion. Overall, these results suggest that SLPI is an alarm protein that modulates not only the innate immune response but also the adaptive immune response. PMID:21574992

  18. Deciphering the transcriptional circuitry of microRNA genes expressed during human monocytic differentiation

    Directory of Open Access Journals (Sweden)

    Schaefer Ulf

    2009-12-01

    Full Text Available Abstract Background Macrophages are immune cells involved in various biological processes including host defence, homeostasis, differentiation, and organogenesis. Disruption of macrophage biology has been linked to increased pathogen infection, inflammation and malignant diseases. Differential gene expression observed in monocytic differentiation is primarily regulated by interacting transcription factors (TFs. Current research suggests that microRNAs (miRNAs degrade and repress translation of mRNA, but also may target genes involved in differentiation. We focus on getting insights into the transcriptional circuitry regulating miRNA genes expressed during monocytic differentiation. Results We computationally analysed the transcriptional circuitry of miRNA genes during monocytic differentiation using in vitro time-course expression data for TFs and miRNAs. A set of TF→miRNA associations was derived from predicted TF binding sites in promoter regions of miRNA genes. Time-lagged expression correlation analysis was utilised to evaluate the TF→miRNA associations. Our analysis identified 12 TFs that potentially play a central role in regulating miRNAs throughout the differentiation process. Six of these 12 TFs (ATF2, E2F3, HOXA4, NFE2L1, SP3, and YY1 have not previously been described to be important for monocytic differentiation. The remaining six TFs are CEBPB, CREB1, ELK1, NFE2L2, RUNX1, and USF2. For several miRNAs (miR-21, miR-155, miR-424, and miR-17-92, we show how their inferred transcriptional regulation impacts monocytic differentiation. Conclusions The study demonstrates that miRNAs and their transcriptional regulatory control are integral molecular mechanisms during differentiation. Furthermore, it is the first study to decipher on a large-scale, how miRNAs are controlled by TFs during human monocytic differentiation. Subsequently, we have identified 12 candidate key controllers of miRNAs during this differentiation process.

  19. Monocytes increase human cardiac myofibroblast-mediated extracellular matrix remodeling through TGF-β1.

    Science.gov (United States)

    Mewhort, Holly E M; Lipon, Brodie D; Svystonyuk, Daniyil A; Teng, Guoqi; Guzzardi, David G; Silva, Claudia; Yong, V Wee; Fedak, Paul W M

    2016-03-15

    Following myocardial infarction (MI), cardiac myofibroblasts remodel the extracellular matrix (ECM), preventing mechanical complications. However, prolonged myofibroblast activity leads to dysregulation of the ECM, maladaptive remodeling, fibrosis, and heart failure (HF). Chronic inflammation is believed to drive persistent myofibroblast activity; however, the mechanisms are unclear. We assessed the influence of peripheral blood monocytes on human cardiac myofibroblast activity in a three-dimensional (3D) ECM microenvironment. Human cardiac myofibroblasts isolated from surgical biopsies of the right atrium and left ventricle were seeded into 3D collagen matrices. Peripheral blood monocytes were isolated from healthy human donors and cocultured with myofibroblasts. Monocytes increased myofibroblast activity measured by collagen gel contraction (baseline: 57.6 ± 5.9% vs. coculture: 65.2 ± 7.1% contraction; P < 0.01) and increased local ECM remodeling quantified by confocal microscopy. Under coculture conditions that allow indirect cellular interaction via paracrine factors but prevent direct cell-cell contact, monocytes had minimal effects on myofibroblast activity (17.9 ± 11.1% vs. 6.4 ± 7.0% increase, respectively; P < 0.01). When cells were cultured under direct contact conditions, multiplex analysis of the coculture media revealed an increase in the paracrine factors TGF-β1 and matrix metalloproteinase 9 compared with baseline (122.9 ± 10.1 pg/ml and 3,496.0 ± 190.4 pg/ml, respectively, vs. 21.5 ± 16.3 pg/ml and 183.3 ± 43.9 pg/ml; P < 0.001). TGF-β blockade abolished the monocyte-induced increase in cardiac myofibroblast activity. These data suggest that direct cell-cell interaction between monocytes and cardiac myofibroblasts stimulates TGF-β-mediated myofibroblast activity and increases remodeling of local matrix. Peripheral blood monocyte interaction with human cardiac myofibroblasts stimulates myofibroblast activity through release of TGF-β1

  20. Human monocytes and macrophages undergo M1-type inflammatory polarization in response to high levels of glucose.

    Science.gov (United States)

    Torres-Castro, Israel; Arroyo-Camarena, Úrsula D; Martínez-Reyes, Camilo P; Gómez-Arauz, Angélica Y; Dueñas-Andrade, Yareth; Hernández-Ruiz, Joselín; Béjar, Yadira L; Zaga-Clavellina, Verónica; Morales-Montor, Jorge; Terrazas, Luis I; Kzhyshkowska, Julia; Escobedo, Galileo

    2016-08-01

    Emerging data suggest that elevated glucose may promote inflammatory activation of monocytic lineage cells with the ability to injure vascular endothelial tissue of diabetic patients, however evidence in primary human monocytes and macrophages is still insufficient. We investigated the effect of high glucose concentration on the inflammatory capacity of human macrophages in vitro and examined whether similar responses were detectable in circulating monocytes from prediabetic patients. Primary monocytes were isolated from healthy blood donors and differentiated into macrophages. Differentiated macrophages were exposed to normal levels of glucose (NG), high glucose (HG) or high mannitol as osmotic pressure control (OP) for three days. Using PCR, ELISA and flow cytometry, we found that HG macrophages showed overexpression of CD11c and inducible nitric oxide synthase as well as down-regulation of arginase-1 and interleukin (IL)-10 with respect to NG and OP macrophages. Consistent with in vitro results, circulating monocytes from hyperglycemic patients exhibited higher levels of CD11c and lower expression of CD206 than monocytes from normoglycemic controls. In subjects with hyperglycemia, elevation in CD11c(+) monocytes was associated with increased obesity, insulin resistance, and triglyceridemia as well as low serum IL-10. Our data suggest that human monocytes and macrophages undergo M1-like inflammatory polarization when exposed to high levels of glucose on in vitro culture conditions and in patients with hyperglycemia. These results demonstrate that excess glucose has direct effects on macrophage activation though the molecular mechanisms mediating such a response remain to be elucidated. PMID:27269375

  1. The Bioenergetic Health Index is a sensitive measure of oxidative stress in human monocytes.

    Science.gov (United States)

    Chacko, Balu K; Zhi, Degui; Darley-Usmar, Victor M; Mitchell, Tanecia

    2016-08-01

    Metabolic and bioenergetic dysfunction are associated with oxidative stress and thought to be a common underlying mechanism of chronic diseases such as atherosclerosis, diabetes, and neurodegeneration. Recent findings support an emerging concept that circulating leukocytes and platelets can act as sensors or biomarkers of mitochondrial function in patients subjected to metabolic diseases. It is proposed that systemic stress-induced alterations in leukocyte bioenergetics are the consequence of several factors including reactive oxygen species. This suggests that oxidative stress mediated changes in leukocyte mitochondrial function could be used as an indicator of bioenergetic health in individuals. To test this concept, we investigated the effect of the redox cycling agent, 2,3 dimethoxynaphthoquinone (DMNQ) on the bioenergetic profiles of monocytes isolated from healthy human subjects using the extracellular flux analyzer. In addition, we tested the hypothesis that the bioenergetic health index (BHI), a single value that represents the bioenergetic health of individuals, is dynamically sensitive to oxidative stress in human monocytes. DMNQ decreased monocyte ATP-linked respiration, maximal respiration, and reserve capacity and caused an increase in proton leak and non-mitochondrial respiration compared to monocytes not treated with DMNQ. The BHI was a more sensitive indicator of the DMNQ-dependent changes in bioenergetics than any individual parameter. These data suggest that monocytes are susceptible to oxidative stress mediated by DMNQ and this can be accurately assessed by the BHI. Taken together, our findings suggest that the BHI has the potential to act as a functional biomarker of the impact of systemic oxidative stress in patients with metabolic disorders. PMID:26748041

  2. The effect of short-chain fatty acids on human monocyte-derived dendritic cells.

    Science.gov (United States)

    Nastasi, Claudia; Candela, Marco; Bonefeld, Charlotte Menné; Geisler, Carsten; Hansen, Morten; Krejsgaard, Thorbjørn; Biagi, Elena; Andersen, Mads Hald; Brigidi, Patrizia; Ødum, Niels; Litman, Thomas; Woetmann, Anders

    2015-01-01

    The gut microbiota is essential for human health and plays an important role in the pathogenesis of several diseases. Short-chain fatty acids (SCFA), such as acetate, butyrate and propionate, are end-products of microbial fermentation of macronutrients that distribute systemically via the blood. The aim of this study was to investigate the transcriptional response of immature and LPS-matured human monocyte-derived DC to SCFA. Our data revealed distinct effects exerted by each individual SCFA on gene expression in human monocyte-derived DC, especially in the mature ones. Acetate only exerted negligible effects, while both butyrate and propionate strongly modulated gene expression in both immature and mature human monocyte-derived DC. An Ingenuity pathway analysis based on the differentially expressed genes suggested that propionate and butyrate modulate leukocyte trafficking, as SCFA strongly reduced the release of several pro-inflammatory chemokines including CCL3, CCL4, CCL5, CXCL9, CXCL10, and CXCL11. Additionally, butyrate and propionate inhibited the expression of lipopolysaccharide (LPS)-induced cytokines such as IL-6 and IL-12p40 showing a strong anti-inflammatory effect. This work illustrates that bacterial metabolites far from the site of their production can differentially modulate the inflammatory response and generally provides new insights into host-microbiome interactions. PMID:26541096

  3. Effect of Dexamethasone on Expression of Glucocorticoid Receptor in Human Monocyte Cell Line THP-1

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    The effect of dexamethasone with differentconcentrations and different stimulating periods on the expression of glucocorticoid receptors (GRα, GRβ) protein was investigated in human monocyte cell line THP-1. The cultured human monocyte line THP-1 cells were stimulated by dexamethasone with different concentrations and different periods. The expression of GRα and GRβ protein was detected by Western blotting. The results showed that the expression of GRα and GRβ was detected in the THP-1 cells. The quantity of GRα expression was reduced by dexamethasone under the same concentration with the prolongation of the stimulating periods. The quantity of GRβ expression was increased by dexamethasone treatment in a time- and dose-dependent manner. It was concluded that dexamethasone stimulation time-dependently reduced the GRα expression in THP-1 cells. Dexamethasone stimulation time- and dose-dependently increased the GRβ expression in THP1 cells. The expression of GRα and GRβ was regulated by glucocorticoid.

  4. Gram-positive pathogenic bacteria induce a common early response in human monocytes

    OpenAIRE

    Ghai Rohit; Tchatalbachev Svetlin; Hossain Hamid; Chakraborty Trinad

    2010-01-01

    Abstract Background We infected freshly isolated human peripheral monocytes with live bacteria of three clinically important gram-positive bacterial species, Staphylococcus aureus, Streptococcus pneumoniae and Listeria monocytogenes and studied the ensuing early transcriptional response using expression microarrays. Thus the observed response was unbiased by signals originating from other helper and effector cells of the host and was not limited to induction by solitary bacterial constituents...

  5. Monocytes of individual human subjects display heterogeneous bacterial uptake and antilisterial activity.

    OpenAIRE

    Zerlauth, G; Chehadeh, H E; Maier, E; Schaff, Z.; Eibl, M M; Mannhalter, J W

    1996-01-01

    Peripheral blood monocytes (Mo) of normal human donors simultaneously exhibit two subsets differing in their functional activity towards the facultative intracellular bacterium Listeria monocytogenes. One subset (on average, 25% of total Mo) was characteristically able to ingest a large number of L. monocytogenes bacteria and permitted intracellular growth of these bacteria. The other Mo subpopulation (on average, 75% of total Mo) was far less active in phagocytosing L. monocytogenes and rest...

  6. CD14{sup +} monocytes promote the immunosuppressive effect of human umbilical cord matrix stem cells

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Ding, E-mail: qqhewd@gmail.com [The State Key Laboratory of Experimental Hematology, Institute of Hematology and Hospital of Blood Diseases, Chinese Academy of Medical Sciences and Peking Union of Medical College, 288 Nanjing Road, Tianjin 300020 (China); TEDA Life and Technology Research Center, Institute of Hematology, Chinese Academy of Medical Sciences, TEDA, Tianjin (China); Chen, Ke, E-mail: chenke_59@hotmail.com [The State Key Laboratory of Experimental Hematology, Institute of Hematology and Hospital of Blood Diseases, Chinese Academy of Medical Sciences and Peking Union of Medical College, 288 Nanjing Road, Tianjin 300020 (China); TEDA Life and Technology Research Center, Institute of Hematology, Chinese Academy of Medical Sciences, TEDA, Tianjin (China); Du, Wei Ting, E-mail: duwtpumc@yahoo.com.cn [The State Key Laboratory of Experimental Hematology, Institute of Hematology and Hospital of Blood Diseases, Chinese Academy of Medical Sciences and Peking Union of Medical College, 288 Nanjing Road, Tianjin 300020 (China); Han, Zhi-Bo, E-mail: zhibohan@hotmail.com [The State Key Laboratory of Experimental Hematology, Institute of Hematology and Hospital of Blood Diseases, Chinese Academy of Medical Sciences and Peking Union of Medical College, 288 Nanjing Road, Tianjin 300020 (China); TEDA Life and Technology Research Center, Institute of Hematology, Chinese Academy of Medical Sciences, TEDA, Tianjin (China); Ren, He, E-mail: knifesharp2000@hotmail.com [National Engineering Research Center of Cell Products, AmCellGene Co. Ltd, TEDA, Tianjin (China); Chi, Ying, E-mail: caizhuying@hotmail.com [The State Key Laboratory of Experimental Hematology, Institute of Hematology and Hospital of Blood Diseases, Chinese Academy of Medical Sciences and Peking Union of Medical College, 288 Nanjing Road, Tianjin 300020 (China); TEDA Life and Technology Research Center, Institute of Hematology, Chinese Academy of Medical Sciences, TEDA, Tianjin (China); and others

    2010-09-10

    Here, the effect of CD14{sup +} monocytes on human umbilical cord matrix stem cell (hUC-MSC)-mediated immunosuppression was studied in vitro. hUC-MSCs exerted a potent inhibitory effect on the proliferation and interferon-{gamma} (IFN-{gamma}) secretion capacities of CD4{sup +} and CD8{sup +} T cells in response to anti-CD3/CD28 stimulation. Transwell co-culture system revealed that the suppressive effect was primarily mediated by soluble factors. Addition of prostaglandin synthesis inhibitors (indomethacin or NS-398) almost completely abrogated the immunosuppression activity of hUC-MSCs, identifying prostaglandin E{sub 2} (PGE{sub 2}) as an important soluble mediator. CD14{sup +} monocytes were found to be able to enhance significantly the immunosuppressive effect of hUC-MSCs in a dose-dependent fashion. Moreover, the inflammatory cytokine IL-1{beta}, either exogenously added or produced by CD14{sup +} monocytes in culture, could trigger expression of high levels of PGE{sub 2} by hUC-MSCs, whereas inclusion of the IL-1 receptor antagonist (IL-1RA) in the culture down-regulated not only PGE{sub 2} expression, but also reversed the promotional effect of CD14{sup +} monocytes and partially restored CD4{sup +} and CD8{sup +} T cell proliferation and IFN-{gamma} secretion. Our data demonstrate an important role of monocytes in the hUC-MSC-induced immunomodulation, which may have important implications in future efforts to explore the clinical potentials of hUC-MSCs.

  7. Synthesis of pro-inflammatory cytokines and adhesion molecules expression by the irradiated human monocyte/macrophage

    International Nuclear Information System (INIS)

    As lesions induced by ionizing radiations are essentially noticed in organs the functional and structural organisation of which depend on the highly proliferative stem cell pool, the author reports an in-vivo investigation of the effect of a gamma irradiation on the expression and secretion of pro-inflammatory cytokines par human monocytes/macrophages. In order to study the role of the cell environment in the radiation-induced inflammation, the author studied whether a co-stimulation of monocytes/macrophages by gamma irradiation, or the exposure of co-cultures of monocytes/macrophages and lymphocytes, could modulate the regulation of inflammatory cytokines. The author also studied the modulation of the expression of adhesion molecules mainly expressed by the monocyte/macrophage, and the membrane density of the CD14 receptor after irradiation of monocytes/macrophages during 24 hours, and of totally differentiated macrophages after seven days of culture

  8. Homocysteine induces production of monocyte chemoattractant protein-1 and interleukin-8 in cultured human whole blood

    Institute of Scientific and Technical Information of China (English)

    Xiao-kunZENG; DanielGREMICK; XianWANG

    2004-01-01

    AIM: To investigate whether increased plasma L-homocysteine (Hcy) level could promote monocyte chemoattract antprotein-1 (MCP-1) and interleukin-8 (IL-8) in cultured whole blood. METHODS: Human whole blood or differenttype of peripheral blood cells from health volunteers were incubated with Hcy and/or the inhibitors. MCP-1 and IL-8 level were measured by ELISA assay. RESULTS: Hcy 10-1000 μmol/L induced production of MCP-1and IL-8 in cultured human whole blood (P<0.05). The major cellular source of these chemokines comed from monocytes. Meanwhile,Hcy also promoted the upregulation of MPO level even at the 10 μmol/L in the cultured whole blood.The intracellular ROS, particular the OH radicals, play extremely important role in the Hcy-induced MCP-1 and IL-8 production. CONCLUSION: Increased Hcy level in plasma (hyperhomocysteinemia) induced MCP-1 and IL-8secretion in cultured human whole blood, especially in monocytes via oxidative stress mechanism,

  9. Human monocytes undergo functional re-programming during differentiation to dendritic cell mediated by human extravillous trophoblasts.

    Science.gov (United States)

    Zhao, Lei; Shao, Qianqian; Zhang, Yun; Zhang, Lin; He, Ying; Wang, Lijie; Kong, Beihua; Qu, Xun

    2016-01-01

    Maternal immune adaptation is required for a successful pregnancy to avoid rejection of the fetal-placental unit. Dendritic cells within the decidual microenvironment lock in a tolerogenic profile. However, how these tolerogenic DCs are induced and the underlying mechanisms are largely unknown. In this study, we show that human extravillous trophoblasts redirect the monocyte-to-DC transition and induce regulatory dendritic cells. DCs differentiated from blood monocytes in the presence of human extravillous trophoblast cell line HTR-8/SVneo displayed a DC-SIGN(+)CD14(+)CD1a(-) phenotype, similar with decidual DCs. HTR8-conditioned DCs were unable to develop a fully mature phenotype in response to LPS, and altered the cytokine secretory profile significantly. Functionally, conditioned DCs poorly induced the proliferation and activation of allogeneic T cells, whereas promoted CD4(+)CD25(+)Foxp3(+) Treg cells generation. Furthermore, the supernatant from DC and HTR-8/SVneo coculture system contained significant high amount of M-CSF and MCP-1. Using neutralizing antibodies, we discussed the role of M-CSF and MCP-1 during monocyte-to-DCs differentiation mediated by extravillous trophoblasts. Our data indicate that human extravillous trophoblasts play an important role in modulating the monocyte-to-DC differentiation through M-CSF and MCP-1, which facilitate the establishment of a tolerogenic microenvironment at the maternal-fetal interface. PMID:26857012

  10. Tetranectin: a novel secretory protein from human monocytes

    DEFF Research Database (Denmark)

    Nielsen, H; Clemmensen, I; Kharazmi, A

    1993-01-01

    stimulated by preincubation with purified tetranectin, whereas chemotactic and chemiluminescence responses to fMLP and C5a were unchanged. Neutrophil functions were not affected. It is concluded that tetranectin is secreted from human mononuclear phagocytes upon weak stimulation, and that the secreted...

  11. Cerebral blood flow in humans following resuscitation from cardiac arrest

    International Nuclear Information System (INIS)

    Cerebral blood flow was measured by xenon-133 washout in 13 patients 6-46 hours after being resuscitated from cardiac arrest. Patients regaining consciousness had relatively normal cerebral blood flow before regaining consciousness, but all patients who died without regaining consciousness had increased cerebral blood flow that appeared within 24 hours after resuscitation (except in one patient in whom the first measurement was delayed until 28 hours after resuscitation, by which time cerebral blood flow was increased). The cause of the delayed-onset increase in cerebral blood flow is not known, but the increase may have adverse effects on brain function and may indicate the onset of irreversible brain damage

  12. Difference of cell cycle arrests induced by lidamycin in human breast cancer cells.

    Science.gov (United States)

    Liu, Xia; He, Hongwei; Feng, Yun; Zhang, Min; Ren, Kaihuan; Shao, Rongguang

    2006-02-01

    Lidamycin (LDM) is a member of the enediyne antibiotic family. It is undergoing phase I clinical trials in China as a potential chemotherapeutic agent. In the present study, we investigated the mechanism by which LDM induced cell cycle arrest in human breast cancer cells. The results showed that LDM induced G1 arrest in p53 wild-type MCF-7 cells at low concentrations, and caused both G1 and G2/M arrests at higher concentrations. In contrast, LDM induced only G2/M arrest in p53-mutant MCF-7/DOX cells. Western blotting analysis indicated that LDM-induced G1 and G2/M arrests in MCF-7 cells were associated with an increase of p53 and p21, and a decrease of phosphorylated retinoblastoma tumor suppressor protein, cyclin-dependent kinase (Cdk), Cdc2 and cyclin B1 protein levels. However, LDM-induced G2/M arrest in MCF-7/DOX cells was correlated with the reduction of cyclin B1 expression. Further study indicated that the downregulation of cyclin B1 by LDM in MCF-7 cells was associated with decreasing cyclin B1 mRNA levels and promoting protein degradation, whereas it was only due to inducing cyclin B1 protein degradation in MCF-7/DOX cells. In addition, activation of checkpoint kinases Chk1 or Chk2 maybe contributed to LDM-induced cell cycle arrest. Taken together, we provide the first evidence that LDM induces different cell cycle arrests in human breast cancer cells, which are dependent on drug concentration and p53 status. These findings are helpful in understanding the molecular anti-cancer mechanisms of LDM and support its clinical trials. PMID:16428935

  13. Human TLR8 is activated upon recognition of Borrelia burgdorferi RNA in the phagosome of human monocytes

    Science.gov (United States)

    Cervantes, Jorge L.; La Vake, Carson J.; Weinerman, Bennett; Luu, Stephanie; O'Connell, Caitlin; Verardi, Paulo H.; Salazar, Juan C.

    2013-01-01

    Phagocytosed Borrelia burgdorferi (Bb), the Lyme disease spirochete, induces a robust and complex innate immune response in human monocytes, in which TLR8 cooperates with TLR2 in the induction of NF-κB-mediated cytokine production, whereas TLR8 is solely responsible for transcription of IFN-β through IRF7. We now establish the role of Bb RNA in TLR8-mediated induction of IFN-β. First, using TLR2-transfected HEK.293 cells, which were unable to phagocytose intact Bb, we observed TLR2 activation by lipoprotein-rich borrelial lysates and TLR2 synthetic ligands but not in response to live spirochetes. Purified Bb RNA, but not borrelial DNA, triggered TLR8 activation. Neither of these 2 ligands induced activation of TLR7. Using purified human monocytes we then show that phagocytosed live Bb, as well as equivalent amounts of borrelial RNA delivered into the phagosome by polyethylenimine (PEI), induces transcription of IFN-β and secretion of TNF-α. The cytokine response to purified Bb RNA was markedly impaired in human monocytes naturally deficient in IRAK-4 and in cells with knockdown TLR8 expression by small interfering RNA. Using confocal microscopy we provide evidence that TLR8 colocalizes with internalized Bb RNA in both early (EEA1) and late endosomes (LAMP1). Live bacterial RNA staining indicates that spirochetal RNA does not transfer from the phagosome into the cytosol. Using fluorescent dextran particles we show that phagosomal integrity in Bb-infected monocytes is not affected. We demonstrate, for the first time, that Bb RNA is a TLR8 ligand in human monocytes and that transcription of IFN-β in response to the spirochete is induced from within the phagosomal vacuole through the TLR8-MyD88 pathway. PMID:23906644

  14. Deciphering the transcriptional circuitry of microRNA genes expressed during human monocytic differentiation

    KAUST Repository

    Schmeier, Sebastian

    2009-12-10

    Background: Macrophages are immune cells involved in various biological processes including host defence, homeostasis, differentiation, and organogenesis. Disruption of macrophage biology has been linked to increased pathogen infection, inflammation and malignant diseases. Differential gene expression observed in monocytic differentiation is primarily regulated by interacting transcription factors (TFs). Current research suggests that microRNAs (miRNAs) degrade and repress translation of mRNA, but also may target genes involved in differentiation. We focus on getting insights into the transcriptional circuitry regulating miRNA genes expressed during monocytic differentiation. Results: We computationally analysed the transcriptional circuitry of miRNA genes during monocytic differentiation using in vitro time-course expression data for TFs and miRNAs. A set of TF?miRNA associations was derived from predicted TF binding sites in promoter regions of miRNA genes. Time-lagged expression correlation analysis was utilised to evaluate the TF?miRNA associations. Our analysis identified 12 TFs that potentially play a central role in regulating miRNAs throughout the differentiation process. Six of these 12 TFs (ATF2, E2F3, HOXA4, NFE2L1, SP3, and YY1) have not previously been described to be important for monocytic differentiation. The remaining six TFs are CEBPB, CREB1, ELK1, NFE2L2, RUNX1, and USF2. For several miRNAs (miR-21, miR-155, miR-424, and miR-17-92), we show how their inferred transcriptional regulation impacts monocytic differentiation. Conclusions: The study demonstrates that miRNAs and their transcriptional regulatory control are integral molecular mechanisms during differentiation. Furthermore, it is the first study to decipher on a large-scale, how miRNAs are controlled by TFs during human monocytic differentiation. Subsequently, we have identified 12 candidate key controllers of miRNAs during this differentiation process. 2009 Schmeier et al; licensee Bio

  15. Leishmania major surface protease Gp63 interferes with the function of human monocytes and neutrophils in vitro

    DEFF Research Database (Denmark)

    Sørensen, A L; Hey, A S; Kharazmi, A

    1994-01-01

    In the present study the effect of Leishmania major surface protease Gp63 on the chemotaxis and oxidative burst response of human peripheral blood monocytes and neutrophils was investigated. It was shown that prior incubation of cells with Gp63 inhibited chemotaxis of neutrophils but not monocytes...... by heat inactivation of the protease at 70 degrees C for 15 min. Neither neutrophil nor monocyte chemiluminescence was inhibited by Gp63 when cells were stimulated with PMA. Our data suggest that the major surface protease Gp63 might play an important role in the initial stages of Leishmania...

  16. Identification of genes responsive to solar simulated UV radiation in human monocyte-derived dendritic cells.

    Directory of Open Access Journals (Sweden)

    Hortensia de la Fuente

    Full Text Available Ultraviolet (UV irradiation has profound effects on the skin and the systemic immune system. Several effects of UV radiation on Dendritic cells (DCs functions have been described. However, gene expression changes induced by UV radiation in DCs have not been addressed before. In this report, we irradiated human monocyte-derived DCs with solar-simulated UVA/UVB and analyzed regulated genes on human whole genome arrays. Results were validated by RT-PCR and further analyzed by Gene Set Enrichment Analysis (GSEA. Solar-simulated UV radiation up-regulated expression of genes involved in cellular stress and inflammation, and down-regulated genes involved in chemotaxis, vesicular transport and RNA processing. Twenty four genes were selected for comparison by RT-PCR with similarly treated human primary keratinocytes and human melanocytes. Several genes involved in the regulation of the immune response were differentially regulated in UVA/UVB irradiated human monocyte-derived DCs, such as protein tyrosine phosphatase, receptor type E (PTPRE, thrombospondin-1 (THBS1, inducible costimulator ligand (ICOSL, galectins, Src-like adapter protein (SLA, IL-10 and CCR7. These results indicate that UV-exposure triggers the regulation of a complex gene repertoire involved in human-DC-mediated immune responses.

  17. Gram-positive pathogenic bacteria induce a common early response in human monocytes

    Directory of Open Access Journals (Sweden)

    Ghai Rohit

    2010-11-01

    Full Text Available Abstract Background We infected freshly isolated human peripheral monocytes with live bacteria of three clinically important gram-positive bacterial species, Staphylococcus aureus, Streptococcus pneumoniae and Listeria monocytogenes and studied the ensuing early transcriptional response using expression microarrays. Thus the observed response was unbiased by signals originating from other helper and effector cells of the host and was not limited to induction by solitary bacterial constituents. Results Activation of monocytes was demonstrated by the upregulation of chemokine rather than interleukin genes except for the prominent expression of interleukin 23, marking it as the early lead cytokine. This activation was accompanied by cytoskeleton rearrangement signals and a general anti-oxidative stress and anti-apoptotic reaction. Remarkably, the expression profiles also provide evidence that monocytes participate in the regulation of angiogenesis and endothelial function in response to these pathogens. Conclusion Regardless of the invasion properties and survival mechanisms of the pathogens used, we found that the early response comprised of a consistent and common response. The common response was hallmarked by the upregulation of interleukin 23, a rather unexpected finding regarding Listeria infection, as this cytokine has been linked primarily to the control of extracellular bacterial dissemination.

  18. Prostaglandin E2 and thromboxane B2 release from human monocytes treated with bacterial lipopolysaccharide

    International Nuclear Information System (INIS)

    We investigated the capacity of counterflow-isolated human monocytes to independently synthesize thromboxane B2 (TxB2) and prostaglandin E2 (PGE2) when stimulated with bacterial lipopolysaccharide (LPS). Independent metabolism was confirmed by establishing different specific activities (dpm/ng) of TxB2 and PGE2 released from LPS-treated cells. For metabolites released during the initial 2-hr treatment period, the specific activity of PGE2 was approximately threefold higher than that of TxB2 regardless of labeling with [3H]arachidonic acid (AA) or [14C]AA. Cells that were pulse-labeled for 2 hr with [3H]AA demonstrated a decreasing PGE2 specific activity over 24 hr, whereas the TxB2 specific activity remained unchanged. In contrast, cells continuously exposed to [14C]AA demonstrated an increasing TxB2 specific activity that approached the level of PGE2 by 24 hr. These results suggest the presence of at least 2 cyclooxygenase metabolic compartments in counterflow-isolated monocytes. Although freshly isolated monocytes have been reported to contain variable numbers of adherent platelets, additional experiments demonstrated that counterflow-isolated platelets are not capable of releasing elevated levels of TxB2 or PGE2 when treated with LPS. It is proposed from these findings that at least two subsets of monocytes exist in peripheral blood that can be distinguished on the basis of independent conversion of AA to TxB2 and PGE2

  19. Circulating Blood Monocyte Subclasses and Lipid-Laden Adipose Tissue Macrophages in Human Obesity.

    Directory of Open Access Journals (Sweden)

    Tal Pecht

    Full Text Available Visceral adipose tissue foam cells are increased in human obesity, and were implicated in adipose dysfunction and increased cardio-metabolic risk. In the circulation, non-classical monocytes (NCM are elevated in obesity and associate with atherosclerosis and type 2 diabetes. We hypothesized that circulating NCM correlate and/or are functionally linked to visceral adipose tissue foam cells in obesity, potentially providing an approach to estimate visceral adipose tissue status in the non-surgical obese patient.We preformed ex-vivo functional studies utilizing sorted monocyte subclasses from healthy donors. Moreover, we assessed circulating blood monocyte subclasses and visceral fat adipose tissue macrophage (ATM lipid content by flow-cytometry in paired blood and omental-fat samples collected from patients (n = 65 undergoing elective abdominal surgery.Ex-vivo, NCM and NCM-derived macrophages exhibited lower lipid accumulation capacity compared to classical or intermediate monocytes/-derived macrophages. Moreover, of the three subclasses, NCM exhibited the lowest migration towards adipose tissue conditioned-media. In a cohort of n = 65, increased %NCM associated with higher BMI (r = 0.250,p<0.05 and ATM lipid content (r = 0.303,p<0.05. Among patients with BMI≥25Kg/m2, linear regression models adjusted for age, sex or BMI revealed that NCM independently associate with ATM lipid content, particularly in men.Collectively, although circulating blood NCM are unlikely direct functional precursor cells for adipose tissue foam cells, their increased percentage in the circulation may clinically reflect higher lipid content in visceral ATMs.

  20. Receptor-mediated uptake of remnant lipoproteins by cholesterol-loaded human monocyte-macrophages

    International Nuclear Information System (INIS)

    Normal human monocyte-macrophages were cholesterol-loaded, and the rates of uptake and degradation of several lipoproteins were measured and compared to rates in control cells. Receptor activities for 125I-rabbit beta-very low density lipoproteins (beta-VLDL), 125I-human low density lipoprotein, and 125I-human chylomicrons were down-regulated in cholesterol-loaded cells; however, the rate of uptake and degradation of 125I-human chylomicron remnants was unchanged from control cells. Cholesterol-loaded alveolar macrophages from a Watanabe heritable hyperlipidemic rabbit, which lack low density lipoprotein receptors, showed receptor down-regulation for 125I-beta-VLDL but not for 125I-human chylomicron remnants. In addition to chylomicron remnants, apo-E-phospholipid complexes competed for 125I-chylomicron remnant uptake, but apo-A-I-phospholipid complexes did not. Chylomicron remnants and beta-VLDL were equally effective in competing for 125I-beta-VLDL and 125I-chylomicron remnant uptake in cholesterol-loaded macrophages. The authors conclude: 1) specific lipoprotein receptor activity persists in cholesterol-loaded cells; 2) this receptor activity recognizes lipo-proteins (at least in part) by their apo-E content; and 3) cholesteryl ester accumulation can occur in monocyte-macrophages incubated with chylomicron remnants

  1. Cerium dioxide nanoparticles do not modulate the lipopolysaccharide-induced inflammatory response in human monocytes

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    Hussain S

    2012-03-01

    Full Text Available Salik Hussain1,*, Faris Al-Nsour1,*, Annette B Rice1, Jamie Marshburn1, Zhaoxia Ji2, Jeffery I Zink2, Brenda Yingling1, Nigel J Walker3, Stavros Garantziotis11Clinical Research Unit, National Institute of Environmental Health Sciences/National Institute of Health, Research Triangle Park, NC, 2UC Center for Environmental Implications of Nanotechnology University of California, Los Angeles, CA, 3Division of National Toxicology Program, National Institute of Environmental Health Sciences/National Institute of Health, Research Triangle Park, NC, USA*Both are principal authorsBackground: Cerium dioxide (CeO2 nanoparticles have potential therapeutic applications and are widely used for industrial purposes. However, the effects of these nanoparticles on primary human cells are largely unknown. The ability of nanoparticles to exacerbate pre-existing inflammatory disorders is not well documented for engineered nanoparticles, and is certainly lacking for CeO2 nanoparticles. We investigated the inflammation-modulating effects of CeO2 nanoparticles at noncytotoxic concentrations in human peripheral blood monocytes.Methods: CD14+ cells were isolated from peripheral blood samples of human volunteers. Cells were exposed to either 0.5 or 1 µg/mL of CeO2 nanoparticles over a period of 24 or 48 hours with or without lipopolysaccharide (10 ng/mL prestimulation. Modulation of the inflammatory response was studied by measuring secreted tumor necrosis factor-alpha, interleukin-1beta, macrophage chemotactic protein-1, interferon-gamma, and interferon gamma-induced protein 10.Results: CeO2 nanoparticle suspensions were thoroughly characterized using dynamic light scattering analysis (194 nm hydrodynamic diameter, zeta potential analysis (-14 mV, and transmission electron microscopy (irregular-shaped particles. Transmission electron microscopy of CD14+ cells exposed to CeO2 nanoparticles revealed that these nanoparticles were efficiently internalized by monocytes and

  2. Effects of PM10 in human peripheral blood monocytes and J774 macrophages

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    Donaldson K

    2004-12-01

    Full Text Available Abstract The effects of PM10, one of the components of particulate air pollution, was investigated using human monocytes and a mouse macrophage cell line (J774. The study aimed to investigate the role of these nanoparticles on the release of the pro-inflammatory cytokine TNF-α and IL-1α gene expression. We also investigated the role of intracellular calcium signalling events and oxidative stress in control of these cytokines and the effect of the particles on the functioning of the cell cytoskeleton. We showed that there was an increase in intracellular calcium concentration in J774 cells on treatment with PM10 particles which could be significantly reduced with concomitant treatment with the calcium antagonists verapamil, the intracellular calcium chelator BAPTA-AM but not with the antioxidant nacystelyn or the calmodulin inhibitor W-7. In human monocytes, PM10 stimulated an increase in intracellular calcium which was reduced by verapamil, BAPTA-AM and nacystelyn. TNF-α release was increased with particle treatment in human monocytes and reduced by only verapamil and BAPTA-AM. IL-1α gene expression was increased with particle treatment and reduced by all of the inhibitors. There was increased F-actin staining in J774 cells after treatment with PM10 particles, which was significantly reduced to control levels with all the antagonists tested. The present study has shown that PM10 particles may exert their pro-inflammatory effects by modulating intracellular calcium signalling in macrophages leading to expression of pro-inflammatory cytokines. Impaired motility and phagocytic ability as shown by changes in the F-actin cytoskeleton is likely to play a key role in particle clearance from the lung.

  3. Response of the Nrf2 protection system in human monocytic cells after ionising irradiation

    International Nuclear Information System (INIS)

    In response to reactive oxygen species (ROS) or electrophiles, the transcription factor nuclear factor-erythroid 2 (NF-E2)-related factor 2 (Nrf2) rapidly trans-locates into the nucleus and induces the expression of various antioxidant genes, such as heme oxygenase-1 (HO-1). Low linear energy transfer (LET) ionising radiations such as X-rays generate ROS, which cause biological damage. However, little is known about whether the Nrf2 system in human monocytic cells is activated by low LET ionising irradiation. Therefore, in this study, the response of the Nrf2 system to X-irradiation in human monocytic THP1 cells was investigated. Following exposure of THP1 cells to X-rays (1-5 Gy), intracellular ROS levels were measured using 2',7'-dichlorodihydro-fluorescein diacetate, Nrf2 localisation was determined using immunofluorescence staining and HO-1 mRNA and protein expression were examined. Although ROS were generated by irradiation in a dose-dependent manner, they disappeared immediately after irradiation. Nrf2 translocation into the nucleus was observed 6 h after 5 Gy X irradiation but was not detected following 1-2 Gy irradiation or in non-irradiated controls. HO-1 expression was significantly higher in 5 Gy-irradiated cells after 24 h than in non-irradiated controls. These results indicate that high-dose irradiation (5 Gy) activates Nrf2 and that the Nrf2 protection system may function from 24 h after irradiation in human monocytic cells. (authors)

  4. Role of superoxide anion radicals in ethanol metabolism by blood monocyte-derived human macrophages

    OpenAIRE

    1989-01-01

    The effects of a number of additives on the rate of conversion of ethanol (1 mg/ml; 21.7 mM) to acetate by monolayers of blood monocyte- derived human macrophages were investigated. The additives studied were superoxide dismutase (SOD; 1,500 U/ml), catalase (1,500 U/ml), tetrahydrofurane (20 mM), and PMA (20 nM), either singly or in various combinations. SOD, catalase, SOD plus catalase, tetrahydrofurane, and tetrahydrofurane plus SOD inhibited ethanol oxidation by 49.2, 12.1, 52.9, 60.4, and...

  5. Transcriptional profiling of human monocytes identifies the inhibitory receptor CD300a as regulator of transendothelial migration.

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    Sharang Ghavampour

    Full Text Available Local inflammatory responses are characterized by the recruitment of circulating leukocytes from the blood to sites of inflammation, a process requiring the directed migration of leukocytes across the vessel wall and hence a penetration of the endothelial lining. To identify underlying signalling events and novel factors involved in these processes we screened for genes differentially expressed in human monocytes following their adhesion to and passage through an endothelial monolayer. Functional annotation clustering of the genes identified revealed an overrepresentation of those associated with inflammation/immune response, in particular early monocyte to macrophage differentiation. Among the gene products so far not implicated in monocyte transendothelial migration was the inhibitory immune receptor CD300a. CD300a mRNA and protein levels were upregulated following transmigration and engagement of the receptor by anti-CD300a antibodies markedly reduced monocyte transendothelial migration. In contrast, siRNA mediated downregulation of CD300a in human monocytes increased their rate of migration. CD300a colocalized and cosedimented with actin filaments and, when activated, caused F-actin cytoskeleton alterations. Thus, monocyte transendothelial migration is accompanied by an elevation of CD300a which serves an inhibitory function possibly required for termination of the actual transmigration.

  6. Phenotypic and functional characteristics of dendritic cells derived from human peripheral blood monocytes

    Institute of Scientific and Technical Information of China (English)

    TANG Ling-ling; ZHANG Zhe; ZHENG Jie-sheng; SHENG Ji-fang; LIU Ke-zhou

    2005-01-01

    Objective: This study is aimed at developing a simple and easy way to generate dendritic cells (DCs) from human peripheral blood monocytes (PBMCs) in vitro. Methods: PBMCs were isolated directly from white blood cell rather than whole blood and purified by patching methods (collecting the attached cell and removing the suspension cell). DCs were then generated by culturing PBMCs for six days with 30 ng/ml recombinant human granulocyte-macrophage stimulating factor (rhGM-CSF) and 20 ng/ml recombinant human interleukin-4 (rhIL-4) in vitro. On the sixth day, TNF-alpha (TNFα) 30 ng/ml was added into some DC cultures, which were then incubated for two additional days. The morphology was monitored by light microscopy and transmission electronic microscopy, and the phenotypes were determined by flow cytometry. Autologous mixed leukocyte reactions (MLR) were used to characterize DC function after TNFα or lipopolysaccharide (LPS) stimulations for 24 h. Results: After six days of culture, the monocytes developed significant dendritic morphology and a portion of cells expressed CD 1 a, CD80 and CD86, features of DCs. TNFα treatment induced DCs maturation and up-regulation of CD80, CD86 and CD83. Autologous MLR demonstrated that these DCs possess potent T-cell stimulatory capacity. Conclusion: This study developed a simple and easy way to generate DCs from PBMCs exposed to rhGM-CSF and rhIL-4. The DCs produced by this method acquired morphologic and antigenic characteristics of DCs.

  7. Human glioblastoma-associated microglia/monocytes express a distinct RNA profile compared to human control and murine samples.

    Science.gov (United States)

    Szulzewsky, Frank; Arora, Sonali; de Witte, Lot; Ulas, Thomas; Markovic, Darko; Schultze, Joachim L; Holland, Eric C; Synowitz, Michael; Wolf, Susanne A; Kettenmann, Helmut

    2016-08-01

    Glioblastoma (GBM) is the most aggressive brain tumor in adults. It is strongly infiltrated by microglia and peripheral monocytes that support tumor growth. In the present study we used RNA sequencing to compare the expression profile of CD11b(+) human glioblastoma-associated microglia/monocytes (hGAMs) to CD11b(+) microglia isolated from non-tumor samples. Hierarchical clustering and principal component analysis showed a clear separation of the two sample groups and we identified 334 significantly regulated genes in hGAMs. In comparison to human control microglia hGAMs upregulated genes associated with mitotic cell cycle, cell migration, cell adhesion, and extracellular matrix organization. We validated the expression of several genes associated with extracellular matrix organization in samples of human control microglia, hGAMs, and the hGAMs-depleted fraction via qPCR. The comparison to murine GAMs (mGAMs) showed that both cell populations share a significant fraction of upregulated transcripts compared with their respective controls. These genes were mostly related to mitotic cell cycle. However, in contrast to murine cells, human GAMs did not upregulate genes associated to immune activation. Comparison of human and murine GAMs expression data to several data sets of in vitro-activated human macrophages and murine microglia showed that, in contrast to mGAMs, hGAMs share a smaller overlap to these data sets in general and in particular to cells activated by proinflammatory stimulation with LPS + INFγ or TNFα. Our findings provide new insights into the biology of human glioblastoma-associated microglia/monocytes and give detailed information about the validity of murine experimental models. GLIA 2016 GLIA 2016;64:1416-1436. PMID:27312099

  8. Nordihydroguaiaretic Acid Attenuates the Oxidative Stress-Induced Decrease of CD33 Expression in Human Monocytes

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    Silvia Guzmán-Beltrán

    2013-01-01

    Full Text Available Nordihydroguaiaretic acid (NDGA is a natural lignan with recognized antioxidant and beneficial properties that is isolated from Larrea tridentata. In this study, we evaluated the effect of NDGA on the downregulation of oxidant stress-induced CD33 in human monocytes (MNs. Oxidative stress was induced by iodoacetate (IAA or hydrogen peroxide (H2O2 and was evaluated using reactive oxygen species (ROS production, and cell viability. NDGA attenuates toxicity, ROS production and the oxidative stress-induced decrease of CD33 expression secondary to IAA or H2O2 in human MNs. It was also shown that NDGA (20 μM attenuates cell death in the THP-1 cell line that is caused by treatment with either IAA or H2O2. These results suggest that NDGA has a protective effect on CD33 expression, which is associated with its antioxidant activity in human MNs.

  9. Differential regulation of human T cell responsiveness by mucosal versus blood monocytes.

    Science.gov (United States)

    Qiao, L; Braunstein, J; Golling, M; Schürmann, G; Autschbach, F; Möller, P; Meuer, S

    1996-04-01

    Human intestinal T lymphocytes are constantly exposed to a large number of foreign antigens without developing a systemic immune response. One crucial mechanisms leading to this intestinal hyporesponsiveness is based on impaired signal transduction through the T cell receptor/CD3 complex in lamina propria T lymphocytes (LP-T). In this study, we addressed the question whether a lack of co-stimulatory/progression signals might also contribute to LP-T hyporesponsiveness. To this end, isolated human monocyte populations from the intestinal lamina propria were obtained and their phenotypes as well as their capacity to promote T cell activation studied. Here, we demonstrate that lamina propria macrophages (LP-MO), in contrast to peripheral blood monocytes (PB-MO), do not support proliferation of either LP-T or PB-T. This may be due to the low expression of ligands (CD54, CD58, CD80) for the T cell accessory receptors CD11/18, CD2 and CD28/CTLA-4 on mucosal macrophages. Thus, down-regulation of both recognition/competence and co-stimulatory/progression signals contribute to intestinal hypo- or unresponsiveness. PMID:8625989

  10. Differentiation and function of human monocyte-derived dendritic cells under the influence of leflunomide

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    Stojić-Vukanić Z.

    2011-01-01

    Full Text Available Leflunomide is an immunosuppressive drug effective in experimental models of transplantation and autoimmune diseases and in the treatment of active rheumatoid arthritis (RA. Having in mind that it has been shown that some other immunosuppressive drugs (glucocorticoids, mycophenolate mofetil, sirolimus etc. impair dendritic cell (DC phenotype and function, we investigated the effect of A77 1726, an active metabolite of leflunomide, on the differentiation and function of human monocyte-derived dendritic cells (MDDC in vitro. Immature MDDC were generated by cultivating monocytes in medium supplemented with GM-CSF and IL-4. To induce maturation, immature MDDC were cultured for 2 additional days with LPS. A77 1726 (100 μM was added at the beginning of cultivation. Flow cytometric analysis showed that MDDC differentiated in the presence of A77 1726 exhibited an altered phenotype, with a down-regulated surface expression of CD80, CD86, CD54 and CD40 molecules. Furthermore, the continuous presence of A77 1726 during differentiation and maturation prevented successful maturation, judging by the decreased expression of maturation marker CD83, costimulatory and adhesive molecules on A77 1726-treated mature MDDC. In addition, A77 1726-pretreated MDDC exhibited a poor stimulatory capacity of the allogeneic T cells and a low production of IL-10 and IL-18. These data suggest that leflunomide impairs the differentiation, maturation and function of human MDDC in vitro, which is an additional mechanism of its immunosuppressive effect.

  11. Development of human connective tissue mast cells from purified blood monocytes.

    Science.gov (United States)

    Czarnetzki, B M; Figdor, C G; Kolde, G; Vroom, T; Aalberse, R; de Vries, J E

    1984-01-01

    Highly purified subfractions of human peripheral blood monocytes, when cultured in the presence of 30% L cell supernatant and 30% horse serum, assumed all the characteristics that define human connective tissue mast cells. After three weeks of culture, 75% of the cells developed metachromasia and granular chloroacetate esterase staining, and their intracellular histamine levels increased from 0.0 to 50.5 ng/10(6) cells. On electron microscopy, the cells developed intracytoplasmic granules with all the features typical for mature and immature mast cells. Cultured cells bound 55 pg 125I-IgE/10(6) cells, while labelling was negligible with cells prior to culture and with heat-denatured 125I-IgE. Fluorescent staining with anti-IgE increased slightly as well, while staining with monoclonal anti-monocyte and anti-HLA-Dr markers decreased. Purified lymphocytes did not assume mast cell characteristics, and lymphokines did not induce or enhance in vitro mast cell development or IgE binding. The data therefore further support the concept that connective tissue mast cells arise from the monocytoid lineage. Images Figure 1 PMID:6698581

  12. Monoclonal antibody to a subset of human monocytes found only in the peripheral blood and inflammatory tissues

    Energy Technology Data Exchange (ETDEWEB)

    Zwadlo, G.; Schlegel, R.; Sorg, C.

    1986-07-15

    A monoclonal antibody is described that was generated by immunizing mice with cultured human blood monocytes. The antibody (27E10) belongs to the IgG1 subclass and detects a surface antigen at M/sub r/ 17,000 that is found on 20% of peripheral blood monocytes. The antigen is increasingly expressed upon culture of monocytes, reaching a maximum between days 2 and 3. Stimulation of monocytes with interferon-..gamma.. (IFN-..gamma..), 12-O-tetradecanoyl-phorbol-13-acetate (TPA), and lipopolysaccharide (LPS) Ylalanine (fMLP) increased the 27E10 antigen density. The amount of 27E10-positive cells is not or is only weakly affected. The antigen is absent from platelets, lymphotyces, and all tested human cell lines, yet it cross-reacts with 15% of freshly isolated granulocytes. By using the indirect immunoperoxidase technique, the antibody is found to be negative on cryostat sections of normal human tissue (skin, lung, and colon) and positive on only a few monocyte-like cells in liver and on part of the cells of the splenic red pulp. In inflammatory tissue, however, the antibody is positive on monocytes/macrophages and sometimes on endothelial cells and epidermal cells, depending on the stage and type of inflammation, e.g., BCG ranulomas are negative, whereas psoriasis vulgaris, atopic dermatitis, erythrodermia, pressure urticaria, and periodontitis contain positively staining cells. In contact eczemas at different times after elicitation (6 hr, 24 hr, and 72 hr), the 27E10 antigen is seen first after 24 hr on a few infiltrating monocytes/macrophages, which increase in numbers after 72 hr.

  13. FUCOIDIN INHIBITS OXIDIZED LOW DENSITY LIPOPROTEIN FROM INDUCING HUMAN PERIPHERAL BLOOD MONOCYTE EXPRESSION OF PROINFLAMMATORY CYTOKINES mRNA

    Institute of Scientific and Technical Information of China (English)

    雷新军; 马爱群; 任冰稳; 耿涛; 张葳; 白玲

    2003-01-01

    Objective To study the significance of scavenger receptor class A(SR-A)in mediating human peripheral blood monocyte to uptake oxidized low density lipoprotein(OxLDL) and promoting the atherosclerotic immuno-pathological lesion in the local blood vessel. Methods With the Digoxenin-labeled Oligonucleotide-probes In situ Hybridization, this research investigated the effects of OxLDL on the mRNA expression of proinflammatory cytokines including MCP-1, bFGF, PDGF and IL-10 in the human peripheral blood monocyte and whether fucoidin, a peculiarly inhibitory ligand for SR-A, would influence this process. Results Monocyte was significantly increased the mRNA expression of MCP-1, bFGF, PDGF and IL-10 in a dose-dependent manner after incubating with OxLDL (10,15,20,25,30·mg·L-1, respectively)for 24 hours(P<0.001). Fucoidin(50,100,150,200,250·mg·mL-1, respectively)completely inhibited OxLDL(20·mg·L-1)from inducing monocyte the mRNA expression of above proinflammatory cytokines(P<0.001). Conclusion OxLDL can stimulate human peripheral blood monocyte to give expression to proinflammatory cytokines mRNA in a dose-dependent manner, while a peculiarly inhibitory ligand for SR-A-fucoidin has an obviously opposed role.

  14. Reversible and irreversible loss of Fc receptor function of human monocytes as a consequence of interaction with immunoglobulin G.

    OpenAIRE

    Kurlander, R J

    1980-01-01

    The effects of IgG in different configurations on the Fc receptor function of human monocytes were studied. Receptor function was assessed by quantitating immune adherence and/or ingestion of human erythrocytes coated with IgG anti-D antibody. Monomeric IgGl in solution inhibited the Fc receptor function of monocytes, but this function was restored completely after washing. In contrast, monomeric IgG that was adsorbed nonspecifically to a plastic surface inhibited the Fc receptor function of ...

  15. Human peripheral blood monocytes display surface antigens recognized by monoclonal antinuclear antibodies

    International Nuclear Information System (INIS)

    The authors used monoclonal anti-nuclear autoantibodies and indirect immunofluorescence to examine normal human peripheral blood mononuclear leukocytes for the presence of cell surface nuclear antigens. Only one monoclonal anti-histone antibody (MH-2) was found to bind to freshly isolated PBL, staining approximately 10% of large cells. However, after cells were placed into culture for 16-24 h, a high percentage (up to 60%) of large-sized cells were recognized by an anti-DNA (BWD-1) and several different antihistone monoclonal antibodies (BWH-1, MH-1, and MH-2). These antibodies recognize separate antigenic determinants on chromatin and histones extracted from chromatin. The histone antigen-positive cells were viable, and the monoclonal antibodies could be shown to be binding to the cell surface and not to the nucleus. Using monoclonal antibodies specific for monocytes and T cells, and complement-mediated cytotoxicity, the cells bearing histone antigens were shown to be primarily monocytes. The appearance of histone and DNA antigen-positive cells was nearly completely inhibited by the addition of low concentrations of cycloheximide at initiation of the cultures. In contrast, little effect on the percentage of positive cells was detected if cells were exposed to high doses of gamma irradiation before culture. These data further support the existence of cell surface nuclear antigens on selected cell subsets, which may provide insight into the immunopathogenesis of systemic lupus erythematosus and related autoimmune diseases

  16. Human Monocyte-Derived Osteoclasts Are Targeted by Staphylococcal Pore-Forming Toxins and Superantigens

    Science.gov (United States)

    Flammier, Sacha; Rasigade, Jean-Philippe; Badiou, Cédric; Henry, Thomas; Vandenesch, François; Laurent, Frédéric; Trouillet-Assant, Sophie

    2016-01-01

    Staphylococcus aureus is the leading cause of bone and joint infections (BJIs). Staphylococcal pathogenesis involves numerous virulence factors including secreted toxins such as pore-forming toxins (PFTs) and superantigens. The role of these toxins on BJI outcome is largely unknown. In particular, few studies have examined how osteoclasts, the bone-resorbing cells, respond to exposure to staphylococcal PFTs and superantigens. We investigated the direct impact of recombinant staphylococcal toxins on human primary mature monocyte-derived osteoclasts, in terms of cytotoxicity and cell activation with cell death and bone resorption assays, using macrophages of the corresponding donors as a reference. Monocyte-derived osteoclasts displayed similar toxin susceptibility profiles compared to macrophages. Specifically, we demonstrated that the Panton-Valentine leukocidin, known as one of the most powerful PFT which lyses myeloid cells after binding to the C5a receptor, was able to induce the death of osteoclasts. The archetypal superantigen TSST-1 was not cytotoxic but enhanced the bone resorption activity of osteoclasts, suggesting a novel mechanism by which superantigen-producing S. aureus can accelerate the destruction of bone tissue during BJI. Altogether, our data indicate that the diverse clinical presentations of BJIs could be related, at least partly, to the toxin profiles of S. aureus isolates involved in these severe infections. PMID:26934588

  17. Costunolide causes mitotic arrest and enhances radiosensitivity in human hepatocellular carcinoma cells

    Directory of Open Access Journals (Sweden)

    Chen Chih-Jen

    2011-05-01

    Full Text Available Abstract Purpose This work aimed to investigate the effect of costunolide, a sesquiterpene lactone isolated from Michelia compressa, on cell cycle distribution and radiosensitivity of human hepatocellular carcinoma (HCC cells. Methods The assessment used in this study included: cell viability assay, cell cycle analysis by DNA histogram, expression of phosphorylated histone H3 (Ser 10 by flow cytometer, mitotic index by Liu's stain and morphological observation, mitotic spindle alignment by immunofluorescence of alpha-tubulin, expression of cell cycle-related proteins by Western blotting, and radiation survival by clonogenic assay. Results Our results show that costunolide reduced the viability of HA22T/VGH cells. It caused a rapid G2/M arrest at 4 hours shown by DNA histogram. The increase in phosphorylated histone H3 (Ser 10-positive cells and mitotic index indicates costunolide-treated cells are arrested at mitosis, not G2, phase. Immunofluorescence of alpha-tubulin for spindle formation further demonstrated these cells are halted at metaphase. Costunolide up-regulated the expression of phosphorylated Chk2 (Thr 68, phosphorylated Cdc25c (Ser 216, phosphorylated Cdk1 (Tyr 15 and cyclin B1 in HA22T/VGH cells. At optimal condition causing mitotic arrest, costunolide sensitized HA22T/VGH HCC cells to ionizing radiation with sensitizer enhancement ratio up to 1.9. Conclusions Costunolide could reduce the viability and arrest cell cycling at mitosis in hepatoma cells. Logical exploration of this mitosis-arresting activity for cancer therapeutics shows costunolide enhanced the killing effect of radiotherapy against human HCC cells.

  18. Costunolide causes mitotic arrest and enhances radiosensitivity in human hepatocellular carcinoma cells

    International Nuclear Information System (INIS)

    This work aimed to investigate the effect of costunolide, a sesquiterpene lactone isolated from Michelia compressa, on cell cycle distribution and radiosensitivity of human hepatocellular carcinoma (HCC) cells. The assessment used in this study included: cell viability assay, cell cycle analysis by DNA histogram, expression of phosphorylated histone H3 (Ser 10) by flow cytometer, mitotic index by Liu's stain and morphological observation, mitotic spindle alignment by immunofluorescence of alpha-tubulin, expression of cell cycle-related proteins by Western blotting, and radiation survival by clonogenic assay. Our results show that costunolide reduced the viability of HA22T/VGH cells. It caused a rapid G2/M arrest at 4 hours shown by DNA histogram. The increase in phosphorylated histone H3 (Ser 10)-positive cells and mitotic index indicates costunolide-treated cells are arrested at mitosis, not G2, phase. Immunofluorescence of alpha-tubulin for spindle formation further demonstrated these cells are halted at metaphase. Costunolide up-regulated the expression of phosphorylated Chk2 (Thr 68), phosphorylated Cdc25c (Ser 216), phosphorylated Cdk1 (Tyr 15) and cyclin B1 in HA22T/VGH cells. At optimal condition causing mitotic arrest, costunolide sensitized HA22T/VGH HCC cells to ionizing radiation with sensitizer enhancement ratio up to 1.9. Costunolide could reduce the viability and arrest cell cycling at mitosis in hepatoma cells. Logical exploration of this mitosis-arresting activity for cancer therapeutics shows costunolide enhanced the killing effect of radiotherapy against human HCC cells

  19. Oxygen-Loaded Nanodroplets Effectively Abrogate Hypoxia Dysregulating Effects on Secretion of MMP-9 and TIMP-1 by Human Monocytes

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    Giulia Rossana Gulino

    2015-01-01

    Full Text Available Monocytes play a key role in the inflammatory stage of the healing process. To allow monocyte migration to injured tissues, the balances between secreted matrix metalloproteinases (MMPs and their inhibitors (TIMPs must be finely modulated. However, a reduction of blood supply and local oxygen tension can modify the phenotype of immune cells. Intriguingly, hypoxia might be targeted by new effective oxygenating devices such as 2H,3H-decafluoropentane- (DFP- based oxygen-loaded nanodroplets (OLNs. Here, hypoxia effects on gelatinase/TIMP release from human peripheral monocytes were investigated, and the therapeutic potential of dextran-shelled OLNs was evaluated. Normoxic monocytes constitutively released ~500 ng/mL MMP-9, ~1.3 ng/mL TIMP-1, and ~0.6 ng/mL TIMP-2 proteins. MMP-2 was not detected. After 24 hours, hypoxia significantly altered MMP-9/TIMP-1 balance by reducing MMP-9 and increasing TIMP-1, without affecting TIMP-2 secretion. Interestingly OLNs, not displaying toxicity to human monocytes after cell internalization, effectively counteracted hypoxia, restoring a normoxia-like MMP-9/TIMP-1 ratio. The action of OLNs was specifically dependent on time-sustained oxygen diffusion up to 24 h from their DFP-based core. Therefore, OLNs appear as innovative, nonconventional, cost-effective, and nontoxic therapeutic tools, to be potentially employed to restore the physiological invasive phenotype of immune cells in hypoxia-associated inflammation.

  20. Macrophage Inflammatory Protein-1alpha mediates Matrix Metalloproteinase-9 enhancement in human adherent monocytes fed with malarial pigment

    Institute of Scientific and Technical Information of China (English)

    Giuliana Giribaldi; Elena Valente; Amina Khadjavi; Manuela Polimeni; Mauro Prato

    2011-01-01

    Objective:To investigate the role of macrophage inflammatory protein-1alpha (MIP-1alpha) in the detrimental enhancement of matrix metalloproteinase-9 (MMP-9)expression, release and activity induced by phagocytosis of malarial pigment (haemozoin,HZ) in human monocytes. Methods: Human adherent monocytes were unfed/fed with nativeHZ for 2 h. After 24 hours, MIP-1alpha production was evaluated by ELISA in cell supernatants. Alternatively,HZ-unfed/fed monocytes were treated in presence/absence of anti-humanMIP-1alpha blocking antibodies or recombinant humanMIP-1alpha for15 h (RNA studies) or 24 h (protein studies); therefore,MMP-9mRNA expression was evaluated in cell lysates by Real TimeRT-PCR, whereas proMMP-9and activeMMP-9protein release were measured in cell supernatants by Western blotting and gelatin zymography.Results: Phagocytosis ofHZ by human monocytes increased production ofMIP-1alpha, mRNA expression ofMMP-9and protein release of proMMP-9 and activeMMP-9. All theHZ-enhancing effects onMMP-9 were abrogated by anti-humanMIP-1alpha blocking antibodies and mimicked by recombinant humanMIP-1alpha.Conclusions:The present work suggests a role for MIP-1alpha in theHZ-dependent enhancement ofMMP-9 expression, release and activity observed in human monocytes, highlighting new detrimental effects ofHZ-triggered proinflammatory response by phagocytic cells in falciparum malaria.

  1. Activated human mast cells induce LOX-1-specific scavenger receptor expression in human monocyte-derived macrophages.

    Directory of Open Access Journals (Sweden)

    Mervi Alanne-Kinnunen

    Full Text Available Activated mast cells in atherosclerotic lesions degranulate and release bioactive compounds capable of regulating atherogenesis. Here we examined the ability of activated human primary mast cells to regulate the expression of the major scavenger receptors in cultured human primary monocyte-derived macrophages (HMDMs.Components released by immunologically activated human primary mast cells induced a transient expression of lectin-like oxidized LDL receptor (LOX-1 mRNA in HMDMs, while the expression of two other scavenger receptors, MSR1 and CD36, remained unaffected. The LOX-1-inducing secretory components were identified as histamine, tumor necrosis factor alpha (TNF-α, and transforming growth factor beta (TGF-β1, which exhibited a synergistic effect on LOX-1 mRNA expression. Histamine induced a transient expression of LOX-1 protein. Mast cell -induced increase in LOX-1 expression was not associated with increased uptake of oxidized LDL by the macrophages.Mast cell-derived histamine, TNF-α, and TGF-β1 act in concert to induce a transient increase in LOX-1 expression in human primary monocyte-derived macrophages. The LOX-1-inducing activity potentially endows mast cells a hitherto unrecognized role in the regulation of innate immune reactions in atherogenesis.

  2. Clinical significance of monocyte heterogeneity

    OpenAIRE

    Stansfield, Brian K.; Ingram, David A.

    2015-01-01

    Monocytes are primitive hematopoietic cells that primarily arise from the bone marrow, circulate in the peripheral blood and give rise to differentiated macrophages. Over the past two decades, considerable attention to monocyte diversity and macrophage polarization has provided contextual clues into the role of myelomonocytic derivatives in human disease. Until recently, human monocytes were subdivided based on expression of the surface marker CD16. “Classical” monocytes express surface marke...

  3. Antibody-dependent cell-mediated cytotoxicity (ADCC) toward human O+ red cells coated with anti-D antibody: comparison between lymphocyte and monocyte ADCC activity.

    OpenAIRE

    Sunada,Mitsutoshi; Suzuki, Shinya; Ota, Zensuke

    1985-01-01

    We investigated the antibody dependent cell-mediated cytotoxicity (ADCC) of lymphocytes and monocytes toward human O+ red cells coated with anti-D antibody using a 51Cr release assay. Lysis of sensitized red cells by lymphocytes occurred rapidly, but monocyte-mediated lysis occurred slowly. This difference might be due to postphagocytic 51Cr release by monocytes. ADCC of lymphocytes increased in proportion to the effector cell number, but large amounts of antibodies were required. In contrast...

  4. Development of a new protocol for 2-day generation of mature dendritic cells from human monocytes

    Directory of Open Access Journals (Sweden)

    Obermaier Bianca

    2003-01-01

    Full Text Available We developed a new 2-day protocol for the generation of dendritic cells (DCs from human monocytes in vitro. First, we demonstrated that 24 hours of culture with GM-CSF and IL-4 are sufficient to generate immature DCs capable of antigen uptake. We then compared two different strategies for DC maturation: proinflammatory mediators were either added together with GM-CSF and IL-4 from the beginning of cell culture or added after 24 hours of differentiation with GM-CSF and IL-4. After 48 hours of total culture period, expression of activation markers was more pronounced in cells generated by the 2-step differentiation and activation method. Our new protocol for 2-day DC differentiation reduces labor, cost and time and also reliably renders high numbers of mature and viable DCs.

  5. The interplay between the master transcription factor PU.1 and miR-424 regulates human monocyte/macrophage differentiation

    OpenAIRE

    Rosa, A.; Ballarino, M.; Sorrentino, A; Sthandier, O.; De Angelis, F. G.; Marchioni, M; Masella, B; Guarini, A.; Fatica, A.; Peschle, C; Bozzoni, I

    2007-01-01

    We describe a pathway by which the master transcription factor PU.1 regulates human monocyte/macrophage differentiation. This includes miR-424 and the transcriptional factor NFI-A. We show that PU.1 and these two components are interlinked in a finely tuned temporal and regulatory circuitry: PU.1 activates the transcription of miR-424, and this up-regulation is involved in stimulating monocyte differentiation through miR-424-dependent translational repression of NFI-A. In turn, the decrease i...

  6. The immune response during the luteal phase of the ovarian cycle : increasing sensitivity of human monocytes to endotoxin

    NARCIS (Netherlands)

    Bouman, Annechien; Moes, H; Heineman, MJ; de Leij, LFMH; Faas, MM

    2001-01-01

    Objective: To test the hypothesis that during the luteal phase of the human ovarian cycle, as compared with the follicular phase, the percentage of cytokines producing peripheral monocytes after in vitro stimulation with endotoxin is increased. Design: Prospective study. Setting: Academic research i

  7. 17 beta-estradiol and progesterone do not influence the production of cytokines from lipopolysaccharide-stimulated monocytes in humans

    NARCIS (Netherlands)

    Schipper, M; Heineman, MJ; Faas, M; Bouman, A.

    2004-01-01

    Objective: To test whether 17beta-estradiol or progesterone influence the cytokine productive capacity of lipopolysaccharide (LPS)-stimulated monocytes in humans. Design: Prospective study. Setting: Academic research institution. Patient(s): Seven women in the luteal phase of a normal ovarian cycle,

  8. Cell surface expression and function of the macromolecular C1 complex on the surface of human monocytes

    Directory of Open Access Journals (Sweden)

    Kinga K Hosszu

    2012-03-01

    Full Text Available The synthesis of the subunits of the C1 complex (C1q, C1s, C1r, and its regulator C1 inhibitor (C1-Inh by human monocytes has been previously established. However, surface expression of these molecules by monocytes has not been shown. Using flow cytometry and antigen-capture ELISA, we show here for the first time that, in addition to C1q, PB monocytes and the monocyte-derived U937 cells express C1s and C1r, as well as Factor B and C1-Inh on their surface. C1s and C1r immunoprecipitated with C1q, suggesting that at least some of the C1q on these cells is part of the C1 complex. Furthermore, the C1 complex on U937 cells was able to trigger complement activation via the classical pathway. The presence of C1-Inh may ensure that an unwarranted autoactivation of the C1 complex does not take place. Since C1-Inh closely monitors the activation of the C1 complex in a sterile or infectious inflammatory environment, further elucidation of the role of C1 complex is crucial to dissect its function in monocyte, DC and T cell activities, and its implications in host defense and tolerance.

  9. Measles virus infection enhances IL-1 beta but reduces tumor necrosis factor-alpha expression in human monocytes.

    Science.gov (United States)

    Leopardi, R; Vainionpää, R; Hurme, M; Siljander, P; Salmi, A A

    1992-10-01

    Monocytes may play a role in the immunologic abnormalities caused by measles. The effect of measles virus (MV) infection on peripheral blood monocyte functions is poorly known. We report that MV-infected PBM have an altered pattern of IL-1 beta and TNF-alpha production in response to stimulation with LPS and PMA in vitro. MV-infected peripheral blood monocytes produced higher amounts of IL-1 beta, whereas the production of TNF-alpha was reduced. The same effect was observed in the human monocytic cell line THP-1, which was used for RNA analysis. An increased steady-state level of IL-1 beta mRNA was observed in MV-infected cells, and the level of TNF-alpha mRNA was reduced. However, both IL-1 beta and TNF-alpha had about 50% increased transcription rate. Analysis of the mRNA stability after transcriptional block by actinomycin D showed that the TNF-alpha mRNA had a reduced half-life in MV-infected cells (about 30 vs 80 min in uninfected cells), whereas IL-1 beta mRNA stability was similar in uninfected and MV-infected cells. These results indicate that MV infection disturbs the immunoregulatory network by interfering with the monocyte functions. PMID:1527385

  10. Flagella from five Cronobacter species induce pro-inflammatory cytokines in macrophage derivatives from human monocytes.

    Directory of Open Access Journals (Sweden)

    Ariadnna Cruz-Córdova

    Full Text Available Cronobacter spp. are opportunistic pathogens linked to lie-threatening infections in neonates and contaminated powdered infant formula that has been epidemiologically associated with these cases. Clinical symptoms of Cronobacter include necrotizing enterocolitis, bacteremia, and meningitis. Flagella from C. sakazakii are involved in biofilm formation and its adhesion to epithelial cells. We investigated the role of flagella from C. sakazakii ST1 and ST4, C. malonaticus, C. muytjensii, C. turicensis and C. dublinensis during the activation of cytokines (IL-8, TNF-α, and IL-10 in macrophage derivatives from human monocytes, which has not been extensively studied. The production and identity of flagella from the five Cronobacter species were visualized and recognized with anti-flagella antibodies by immunogold labeling through transmission electron microscopy. Purified flagella were dissociated into monomers in 12% SDS-PAGE Coomassie blue-stained gels showing a band of ∼28 kDa and, in addition, mass spectrometry revealed the presence of several peptides that correspond to flagellin. Flagella (100 ng induced the release of IL-8 (3314-6025 pg/ml, TNF-α (39-359 pg/ml, and IL-10 (2-96 pg/ml, in macrophage isolates from human monocytes and similar results were obtained when flagella were dissociated into monomers. Inhibition assays using three dilutions of anti-flagella antibodies (1∶10, 1∶100, and 1∶200 suppressed the secretion of IL-8, TNF-α, and IL-10 between 95-100% using 100 ng of protein. A transfection assay using 293-hTLR5 cells showed IL-8 release of 197 pg/ml and suppression in the secretion of IL-8 when anti-hTLR5-IgA antibodies were used at different concentrations. These observations suggest that flagella and flagellin are involved in an inflammatory response dependent on TLR5 recognition, which could contribute to the pathogenesis of the bacteria.

  11. Surface expression of Mo3e antigen by activated human monocytes and U-937 cells

    Energy Technology Data Exchange (ETDEWEB)

    Todd R.F. III; Bury, M.J.; Liu, D.Y.

    1986-03-05

    The surface expression of a protease-sensitive antigen, Mo3e, by activated human monocytes and U-937 cells is a plasma membrane feature of the activated state. Mo3e, which is an 80 kD protein on Western blot analysis, may represent the surface receptor for migration inhibitory factor (MIF), as evidenced by inhibition of MIF responsiveness produced by anti-Mo3e monoclonal antibody. Mo3e is barely detectable (by surface immunofluorescence) on freshly isolated monocytes but becomes expressed in high antigen density during 18-24 hrs culture in medium containing E. coli lipopolysaccharide (> 1 ng/ml), 4..beta..-phorbol 12-myristate 13-acetate (PMA) (5-10 nM), or muramyl dipeptide (0.1-1 ..mu..M). In U-937 cells, Mo3e surface expression is detectable after 24 hrs exposure to PMA and other pharmacological activators of protein kinase C: 4..beta..-phorbol 12, 13 dibutyrate, 4..beta..-phorbol 12, 13 didecanoate, mezerein, or Sn-1,2-dioctanoylglycerol. The biologically-inactivate phorbol compounds, 4..cap alpha..-phorbol 12, 13 didecanoate and 4/sub ..beta../-phorbol do not stimulate Mo3e expression. The calcium ionophore, ionomycin, has a synergistic effect on Mo3e expression stimulated by PMA; conversely, calcium antagonists block PMA-induced Mo3e expression. These results suggest the involvement of protein kinase C activation and intracellular calcium mobilization in the stimulated expression of Mo3e by activated human mononuclear phagocytes.

  12. Human neutrophil Fc receptor-mediated adhesion under flow: a hollow fibre model of intravascular arrest.

    Science.gov (United States)

    D'Arrigo, C; Candal-Couto, J J; Greer, M; Veale, D J; Woof, J M

    1995-04-01

    Human polymorphonuclear cells (PMN) were found to adhere to a novel model of blood vessel wall-associated IgG. The internal surfaces of cellulose acetate hollow fibres, of comparable internal diameter to small blood vessels, were coated with normal serum human IgG, heat-aggregated IgG (HAIgG), laminin or fibrinogen. Under conditions of flow mimicking those in a small vessel, PMN were found to adhere markedly only to immunoglobulin-coated fibres. Arrest on HAIgG was inhibited by excess soluble IgG but not by bovine serum albumin (BSA), demonstrating that the adhesion was IgG-specific and presumably mediated by Fc gamma R on the PMN surface. Pre-adsorption of serum components onto HAIgG-coated fibres enhanced PMN arrest, due most probably to fixation of complement components by immobilized HAIgG, resulting in additional potential to entrap PMN via complement receptors such as CR3. Treatment of PMN with the regulatory neuropeptide substance P also enhanced adhesion to HAIgG-coated fibres and caused increased surface expression of Fc gamma RI, Fc gamma RII and Fc gamma RIII. A mouse cell line derived from L cells, hR4C6, stably transfected with human Fc gamma RII, was found to adhere under flow to HAIgG-coated fibres, whilst untransfected parent L cells did not. This adhesion was similarly inhibited by excess soluble IgG, confirming the capability of Fc gamma R to mediate cell arrest. The study strongly suggests that Fc gamma R may play an important role in intravascular PMN arrest and we speculate that in inflammatory diseases PMN may adhere via Fc gamma R to immobilized immunoglobulin on the vascular endothelium, with subsequent degranulation and tissue damage. PMID:7535210

  13. Modulation of neutrophil and monocyte function by recombinant human granulocyte macrophage colony-stimulating factor in patients with lymphoma

    DEFF Research Database (Denmark)

    Kharazmi, A; Nielsen, H; Hovgaard, D;

    1991-01-01

    Granulocyte macrophage colony-stimulating factor (GM-CSF) has been shown to inhibit the chemotaxis and enhance the oxidative burst response of human neutrophils in vitro. The present study describes the effect of recombinant GM-CSF on the neutrophil and monocyte function in patients with lymphoma...... undergoing GM-CSF treatment. Patients with either Hodgkin's or non-Hodgkin's lymphoma were treated with various dosages (2-16 micrograms kg-1 body weight per day for 5 days) of rhGM-CSF by intravenous or subcutaneous route. Prior to and on day 5 of rhGM-CSF treatment, neutrophil and monocyte chemotaxis...... by up to 43-fold. rhGM-CSF treatment did not affect degranulation of the neutrophils as measured by release of vitamin B12 binding protein. Degree of modulation of neutrophil and monocyte function by rhGM-CSF was independent of rhGM-CSF dosages administered. These data suggest that phagocytic defence...

  14. Effect of a standardized liver and spleen fraction of peptides on the differentiation of human monocyte-derived macrophages.

    Science.gov (United States)

    Spessotto, P; Bulla, R; Mittenzwei, H; Dri, P

    1994-06-01

    The effect of Factor AF2 (AF2), a standardized fraction of peptides with a molecular weight of < 10,000 Dalton obtained from livers and spleens of newborn lambs, on the differentiation of human monocyte-derived macrophages was studied, in view of the central role played by these cells in inflammation and tumor cytotoxicity. The results show that the drug 1. increases the cell density of cultures, 2. favours the morphologic differentiation of monocytes into macrophages, and 3. increases the macrophages phagocytic capacity. The first two effects are observed when monocytes are cultured in 1% serum but not in 10% serum while the enhancement of phagocytic activity is detected at both serum concentrations. PMID:8053979

  15. RRR-α-tocopheryl succinate inhibits human gastric cancer SGC-7901 cell growth by inducing apoptosis and DNA synthesis arrest

    OpenAIRE

    Wu, Kun; ZHAO Yan; Liu, Bai-He; Li, Yao; Liu, Fang; Guo, Jian; Yu, Wei-Ping

    2002-01-01

    AIM: To investigate the effects of growth inhibition of human gastric cancer SGC-7901 cell with RRR-α-tocopheryl succinate (VES), a derivative of natural Vitamin E, via inducing apoptosis and DNA synthesis arrest.

  16. Oligonol, a lychee fruit-derived low-molecular form of polyphenol mixture, suppresses inflammatory cytokine production from human monocytes.

    Science.gov (United States)

    Lee, Naeun; Shin, Min Sun; Kang, Youna; Park, Kieyoung; Maeda, Takahiro; Nishioka, Hiroshi; Fujii, Hajime; Kang, Insoo

    2016-06-01

    Monocytes produce high levels of inflammatory cytokines including IL-6 and TNF-α that are involved in autoimmunity, inflammatory diseases, cardiovascular disease and obesity. Therapies targeting IL-6 and TNF-α have been utilized in treating chronic inflammatory diseases. Oligonol is a lychee fruit-derived low-molecular form of polyphenol mixture, typically catechin-type monomers and oligomers of proanthocyanidins, which are produced by an oligomerization process. Although previous studies reported anti-inflammatory properties of Oligonol, it is unknown whether and how Oligonol suppresses IL-6 and TNF-α production in human monocytes. The results of our study demonstrate that Oligonol (25μg/ml) decreases the production of IL-6 and TNF-α from human primary monocytes as measured by flow cytometry and ELISA. Such an anti-cytokine effect was likely mediated by the suppression of NF-κB activation without inducing cell death. Our findings raise the possibility of exploring the benefits of Oligonol in controlling inflammatory conditions, especially those associated with monocytes, in humans. PMID:27079270

  17. MIF inhibition reverts the gene expression profile of human melanoma cell line-induced MDSCs to normal monocytes

    Directory of Open Access Journals (Sweden)

    Sabine Waigel

    2016-03-01

    Full Text Available Myeloid-derived suppressor cells (MDSCs are potently immunosuppressive innate immune cells that accumulate in advanced cancer patients and actively inhibit anti-tumor T lymphocyte responses [1]. Increased numbers of circulating MDSCs directly correlate with melanoma patient morbidity and reduced anti-tumor immune responses [2,3]. Previous studies have revealed that monocyte-derived macrophage migration inhibitory factor (MIF is necessary for the immune suppressive function of MDSCs in mouse models of melanoma [4,5]. To investigate whether MIF participates in human melanoma-induced MDSC differentiation and/or suppressive function, we have established an in vitro MDSC induction model using primary, normal human monocytes co-cultured with human melanoma cell lines in the presence or absence of the MIF antagonist—4-IPP [4,6–9]. To identify potential mechanistic effectors, we have performed transcriptome analyses on cultured monocytes and on melanoma-induced MDSCs obtained from either untreated or 4-IPP-treated A375:monocyte co-cultures. Here, we present a detailed protocol, which can facilitate easy reproduction of the microarray results (NCBI GEO accession number GSE73333 published by Yaddanapudi et al. (2015 in Cancer Immunology Research [10].

  18. MIF inhibition reverts the gene expression profile of human melanoma cell line-induced MDSCs to normal monocytes.

    Science.gov (United States)

    Waigel, Sabine; Rendon, Beatriz E; Lamont, Gwyneth; Richie, Jamaal; Mitchell, Robert A; Yaddanapudi, Kavitha

    2016-03-01

    Myeloid-derived suppressor cells (MDSCs) are potently immunosuppressive innate immune cells that accumulate in advanced cancer patients and actively inhibit anti-tumor T lymphocyte responses [1]. Increased numbers of circulating MDSCs directly correlate with melanoma patient morbidity and reduced anti-tumor immune responses [2], [3]. Previous studies have revealed that monocyte-derived macrophage migration inhibitory factor (MIF) is necessary for the immune suppressive function of MDSCs in mouse models of melanoma [4], [5]. To investigate whether MIF participates in human melanoma-induced MDSC differentiation and/or suppressive function, we have established an in vitro MDSC induction model using primary, normal human monocytes co-cultured with human melanoma cell lines in the presence or absence of the MIF antagonist-4-IPP [4], [6], [7], [8], [9]. To identify potential mechanistic effectors, we have performed transcriptome analyses on cultured monocytes and on melanoma-induced MDSCs obtained from either untreated or 4-IPP-treated A375:monocyte co-cultures. Here, we present a detailed protocol, which can facilitate easy reproduction of the microarray results (NCBI GEO accession number GSE73333) published by Yaddanapudi et al. (2015) in Cancer Immunology Research [10]. PMID:26981417

  19. Prolyl oligopeptidase inhibition-induced growth arrest of human gastric cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Suzuki, Kanayo [Laboratory of Cell Biology, Osaka University of Pharmaceutical Sciences, 4-20-1 Nasahara, Takatsuki, Osaka 569-1094 (Japan); Sakaguchi, Minoru, E-mail: sakaguti@gly.oups.ac.jp [Laboratory of Cell Biology, Osaka University of Pharmaceutical Sciences, 4-20-1 Nasahara, Takatsuki, Osaka 569-1094 (Japan); Tanaka, Satoshi [Laboratory of Cell Biology, Osaka University of Pharmaceutical Sciences, 4-20-1 Nasahara, Takatsuki, Osaka 569-1094 (Japan); Yoshimoto, Tadashi [Department of Life Science, Setsunan University, 17-8 Ikeda-Nakamachi, Neyagawa, Osaka 572-8508 (Japan); Takaoka, Masanori [Laboratory of Cell Biology, Osaka University of Pharmaceutical Sciences, 4-20-1 Nasahara, Takatsuki, Osaka 569-1094 (Japan)

    2014-01-03

    Highlights: •We examined the effects of prolyl oligopeptidase (POP) inhibition on p53 null gastric cancer cell growth. •POP inhibition-induced cell growth suppression was associated with an increase in a quiescent G{sub 0} state. •POP might regulate the exit from and/or reentry into the cell cycle. -- Abstract: Prolyl oligopeptidase (POP) is a serine endopeptidase that hydrolyzes post-proline peptide bonds in peptides that are <30 amino acids in length. We recently reported that POP inhibition suppressed the growth of human neuroblastoma cells. The growth suppression was associated with pronounced G{sub 0}/G{sub 1} cell cycle arrest and increased levels of the CDK inhibitor p27{sup kip1} and the tumor suppressor p53. In this study, we investigated the mechanism of POP inhibition-induced cell growth arrest using a human gastric cancer cell line, KATO III cells, which had a p53 gene deletion. POP specific inhibitors, 3-((4-[2-(E)-styrylphenoxy]butanoyl)-L-4-hydroxyprolyl)-thiazolidine (SUAM-14746) and benzyloxycarbonyl-thioprolyl-thioprolinal, or RNAi-mediated POP knockdown inhibited the growth of KATO III cells irrespective of their p53 status. SUAM-14746-induced growth inhibition was associated with G{sub 0}/G{sub 1} cell cycle phase arrest and increased levels of p27{sup kip1} in the nuclei and the pRb2/p130 protein expression. Moreover, SUAM-14746-mediated cell cycle arrest of KATO III cells was associated with an increase in the quiescent G{sub 0} state, defined by low level staining for the proliferation marker, Ki-67. These results indicate that POP may be a positive regulator of cell cycle progression by regulating the exit from and/or reentry into the cell cycle by KATO III cells.

  20. p38 mitogen-activated protein kinase mediates IL-8 induction by the ribotoxin deoxynivalenol in human monocytes

    International Nuclear Information System (INIS)

    The effects of the ribotoxic trichothecene deoxynivalenol (DON) on mitogen-activated protein kinase (MAPK)-mediated IL-8 expression were investigated in cloned human monocytes and peripheral blood mononuclear cells (PBMC). DON (250 to 1000 ng/ml) induced both IL-8 mRNA and IL-8 heteronuclear RNA (hnRNA), an indicator of IL-8 transcription, in the human U937 monocytic cell line in a concentration-dependent manner. Expression of IL-8 hnRNA, mRNA and protein correlated with p38 phosphorylation and was completely abrogated by the p38 MAPK inhibitor SB203580. DON at 500 ng/ml similarly induced p38-dependent IL-8 protein and mRNA expression in PBMC cultures from healthy volunteers. Significantly increased IL-6 and IL-1β intracellular protein and mRNA expression was also observed in PBMC treated with DON (500 ng/ml) which were also partially p38-dependent. Flow cytometry of PBMC revealed that DON-induced p38 phosphorylation varied among individuals relative to both threshold toxin concentrations (25-100 ng/ml) and relative increases in percentages of phospho-p38+ cells. DON-induced p38 activation occurred exclusively in the CD14+ monocyte population. DON was devoid of agonist activity for human Toll-like receptors 2, 3, 4, 5, 7, 8 and 9. However, two other ribotoxins, emetine and anisomycin, induced p38 phosphorylation in PBMC similarly to DON. Taken together, these data suggest that (1) p38 activation was required for induction of IL-8 and proinflammatory gene expression in the monocyte and (2) DON induced p38 activation in human monocytes via the ribotoxic stress response

  1. TRIGGERING OF A CYTOLYTIC FACTOR WITH TNF-LIKE ACTIVITY FROM HUMAN MONOCYTES

    Science.gov (United States)

    Monocytes have a nonoxidative mechanism of tumor cytolysis operative in CGD and in normal monocytes under anaerobic conditions. The absence of competitive inhibition provides negative evidence for a contact-independent mechanism of cytolysis, confirmed by the demonstration of a s...

  2. Salidroside induces cell-cycle arrest and apoptosis in human breast cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Hu, Xiaolan, E-mail: huxiaolan1998@yahoo.com.cn [Department of Pathology and Pathophysiology, Zhejiang University School of Medicine, Hangzhou (China); Zhang, Xianqi [The 2nd Affiliated Hospital of Zhejiang University School of Medicine, Hangzhou (China); Qiu, Shuifeng [Department of Pathology and Pathophysiology, Zhejiang University School of Medicine, Hangzhou (China); Yu, Daihua; Lin, Shuxin [Fourth Military Medical University, Xi' an (China)

    2010-07-16

    Research highlights: {yields} Salidroside inhibits the growth of human breast cancer cells. {yields} Salidroside induces cell-cycle arrest of human breast cancer cells. {yields} Salidroside induces apoptosis of human breast cancer cell lines. -- Abstract: Recently, salidroside (p-hydroxyphenethyl-{beta}-D-glucoside) has been identified as one of the most potent compounds isolated from plants of the Rhodiola genus used widely in traditional Chinese medicine, but pharmacokinetic data on the compound are unavailable. We were the first to report the cytotoxic effects of salidroside on cancer cell lines derived from different tissues, and we found that human breast cancer MDA-MB-231 cells (estrogen receptor negative) were sensitive to the inhibitory action of low-concentration salidroside. To further investigate the cytotoxic effects of salidroside on breast cancer cells and reveal possible ER-related differences in response to salidroside, we used MDA-MB-231 cells and MCF-7 cells (estrogen receptor-positive) as models to study possible molecular mechanisms; we evaluated the effects of salidroside on cell growth characteristics, such as proliferation, cell cycle duration, and apoptosis, and on the expression of apoptosis-related molecules. Our results demonstrated for the first time that salidroside induces cell-cycle arrest and apoptosis in human breast cancer cells and may be a promising candidate for breast cancer treatment.

  3. Isolation of Human Monocytes by Double Gradient Centrifugation and Their Differentiation to Macrophages in Teflon-coated Cell Culture Bags

    OpenAIRE

    Menck, Kerstin; Behme, Daniel; Pantke, Mathias; Reiling, Norbert; Binder, Claudia; Pukrop, Tobias; Klemm, Florian

    2014-01-01

    Human macrophages are involved in a plethora of pathologic processes ranging from infectious diseases to cancer. Thus they pose a valuable tool to understand the underlying mechanisms of these diseases. We therefore present a straightforward protocol for the isolation of human monocytes from buffy coats, followed by a differentiation procedure which results in high macrophage yields. The technique relies mostly on commonly available lab equipment and thus provides a cost and time effective wa...

  4. Human Podoplanin-positive Monocytes and Platelets Enhance Lymphangiogenesis Through the Activation of the Podoplanin/CLEC-2 Axis

    OpenAIRE

    Hur, Jin; Jang, Jae Hee; Oh, Il-Young; Choi, Jae-Il; Yun, Ji-Yeon; Kim, Joonoh; Choi, Young-Eun; Ko, Seung-Bum; Kang, Jin-A; Kang, Jeehoon; Lee, Sang Eun; LEE, Hwan; Park, Young-Bae; Kim, Hyo-Soo

    2014-01-01

    Emerging studies suggested that murine podoplanin-positive monocytes (PPMs) are involved in lymphangiogenesis. The goal of this study was to demonstrate the therapeutic lymphangiogenesis of human PPMs by the interaction with platelets. Aggregation culture of human peripheral blood mononuclear cells (PBMCs) resulted in cellular aggregates termed hematospheres. During 5-day culture, PPMs expanded exponentially and expressed several lymphatic endothelial cell–specific markers including vascular ...

  5. Suppression of blood monocyte and neutrophil chemotaxis in acute human malaria

    DEFF Research Database (Denmark)

    Nielsen, H; Kharazmi, A; Theander, T G

    1986-01-01

    tested monocyte chemotactic responsiveness in 19 patients with acute primary attack malaria. In addition, the neutrophil chemotaxis was measured in 12 patients. Before the initiation of antimalarial treatment a significant depression of monocyte chemotaxis was observed in approximately half of the...... suppressed. The monocyte chemotaxis was followed in 14 of the patients, during treatment and after complete recovery. After 3 days of treatment the response had improved in most of the patients, and after 7 days all patients had a normal monocyte chemotaxis, which remained normal after one month. No...... significant differences between P. falciparum and P. vivax/ovale malaria was observed with respect to blood monocyte chemotactic responsiveness. Neutrophil chemotaxis in patients with P. falciparum infections was similarly suppressed before treatment (54% of controls), was still defective after 3 days of...

  6. The interplay between the master transcription factor PU.1 and miR-424 regulates human monocyte/macrophage differentiation.

    Science.gov (United States)

    Rosa, A; Ballarino, M; Sorrentino, A; Sthandier, O; De Angelis, F G; Marchioni, M; Masella, B; Guarini, A; Fatica, A; Peschle, C; Bozzoni, I

    2007-12-11

    We describe a pathway by which the master transcription factor PU.1 regulates human monocyte/macrophage differentiation. This includes miR-424 and the transcriptional factor NFI-A. We show that PU.1 and these two components are interlinked in a finely tuned temporal and regulatory circuitry: PU.1 activates the transcription of miR-424, and this up-regulation is involved in stimulating monocyte differentiation through miR-424-dependent translational repression of NFI-A. In turn, the decrease in NFI-A levels is important for the activation of differentiation-specific genes such as M-CSFr. In line with these data, both RNAi against NFI-A and ectopic expression of miR-424 in precursor cells enhance monocytic differentiation, whereas the ectopic expression of NFI-A has an opposite effect. The interplay among these three components was demonstrated in myeloid cell lines as well as in human CD34+ differentiation. These data point to the important role of miR-424 and NFI-A in controlling the monocyte/macrophage differentiation program. PMID:18056638

  7. Estradiol coupling to human monocyte nitric oxide release is dependent on intracellular calcium transients: evidence for an estrogen surface receptor.

    Science.gov (United States)

    Stefano, G B; Prevot, V; Beauvillain, J C; Fimiani, C; Welters, I; Cadet, P; Breton, C; Pestel, J; Salzet, M; Bilfinger, T V

    1999-10-01

    We tested the hypothesis that estrogen acutely stimulates constitutive NO synthase (cNOS) activity in human peripheral monocytes by acting on an estrogen surface receptor. NO release was measured in real time with an amperometric probe. 17beta-estradiol exposure to monocytes stimulated NO release within seconds in a concentration-dependent manner, whereas 17alpha-estradiol had no effect. 17beta-estradiol conjugated to BSA (E2-BSA) also stimulated NO release, suggesting mediation by a membrane surface receptor. Tamoxifen, an estrogen receptor inhibitor, antagonized the action of both 17beta-estradiol and E2-BSA, whereas ICI 182,780, a selective inhibitor of the nuclear estrogen receptor, had no effect. We further showed, using a dual emission microfluorometry in a calcium-free medium, that the 17beta-estradiol-stimulated release of monocyte NO was dependent on the initial stimulation of intracellular calcium transients in a tamoxifen-sensitive process. Leeching out the intracellular calcium stores abolished the effect of 17beta-estradiol on NO release. RT-PCR analysis of RNA obtained from the cells revealed a strong estrogen receptor-alpha amplification signal and a weak beta signal. Taken together, a physiological dose of estrogen acutely stimulates NO release from human monocytes via the activation of an estrogen surface receptor that is coupled to increases in intracellular calcium. PMID:10490972

  8. Apoptosis and cell-cycle arrest in human and murine tumor cells are initiated by isoprenoids.

    Science.gov (United States)

    Mo, H; Elson, C E

    1999-04-01

    Diverse classes of phytochemicals initiate biological responses that effectively lower cancer risk. One class of phytochemicals, broadly defined as pure and mixed isoprenoids, encompasses an estimated 22,000 individual components. A representative mixed isoprenoid, gamma-tocotrienol, suppresses the growth of murine B16(F10) melanoma cells, and with greater potency, the growth of human breast adenocarcinoma (MCF-7) and human leukemic (HL-60) cells. beta-Ionone, a pure isoprenoid, suppresses the growth of B16 cells and with greater potency, the growth of MCF-7, HL-60 and human colon adenocarcinoma (Caco-2) cells. Results obtained with diverse cell lines differing in ras and p53 status showed that the isoprenoid-mediated suppression of growth is independent of mutated ras and p53 functions. beta-Ionone suppressed the growth of human colon fibroblasts (CCD-18Co) but only when present at three-fold the concentration required to suppress the growth of Caco-2 cells. The isoprenoids initiated apoptosis and, concomitantly arrested cells in the G1 phase of the cell cycle. Both suppress 3-hydroxy-3-methylglutaryl CoA reductase activity. beta-Ionone and lovastatin interfered with the posttranslational processing of lamin B, an activity essential to assembly of daughter nuclei. This interference, we postulate, renders neosynthesized DNA available to the endonuclease activities leading to apoptotic cell death. Lovastatin-imposed mevalonate starvation suppressed the glycosylation and translocation of growth factor receptors to the cell surface. As a consequence, cells were arrested in the G1 phase of the cell cycle. This rationale may apply to the isoprenoid-mediated G1-phase arrest of tumor cells. The additive and potentially synergistic actions of these isoprenoids in the suppression of tumor cell proliferation and initiation of apoptosis coupled with the mass action of the diverse isoprenoid constituents of plant products may explain, in part, the impact of fruit, vegetable

  9. Different effects of phorbol ester derivates on human immunodeficiency virus 1 replication in lymphocytic and monocytic human cells

    International Nuclear Information System (INIS)

    The mode of action of the phorbol 12,13-dibutyrate (PDBu) and phorbol 12-myristate 13-acetate (PMA) on the human immunodeficiency virus 1 (HIV-l) replication in human lymphocytes and monocytes was studied. PDBu and PMA appear to have similar effects on the regulation of HIV-l replication in acutely infected cells. Here we show a significantly increased replication of HIV-l induced by PDBu and PMA in Molt-4 and Jurkat cells, but a reduced replication in THP-l and U-937 cells. Moreover, quantitatively different activity of the two derivatives in relation to HIV-l replication was observed. PDBu proved to be a stronger stimulator or suppressor of HIV-l replication as compared to PMA. Although the precise mechanism of the activation of HIV-l replication by phorbol ester derivatives is not clear, it can be assumed that the hydrophilicity of PDBu may cause its stronger effect. (authors)

  10. Differential regulation of the human immunodeficiency virus type 2 enhancer in monocytes at various stages of differentiation.

    Science.gov (United States)

    Hilfinger, J M; Clark, N; Smith, M; Robinson, K; Markovitz, D M

    1993-07-01

    We have demonstrated that stimulation of the human immunodeficiency virus type 2 (HIV-2) enhancer in T cells is dependent upon at least four cis-acting elements, including two purine-rich binding sites, PuB1 and PuB2, which are capable of binding members of the ets family of proto-oncogenes, the pets (peri-ets) site, which lies just upstream of the PuB2 site, and a single kappa B site (D. M. Markovitz, M. Smith, J. M. Hilfinger, M. C. Hannibal, B. Petryniak, and G. J. Nabel, J. Virol. 66:5479-5484, 1992). In this study, we examined the regulation of the HIV-2 enhancer in cells of monocytic lineage. We found that in immature monocytic cell lines, the HIV-2 enhancer is markedly induced by phorbol esters and that all four cis-acting elements are required for activation. In mature monocytic cells, constitutive activity is high, with only modest stimulation following phorbol ester treatment. Mutation of any of the four cis-acting elements resulted in greatly reduced basal expression in mature monocytes. This is in contrast to HIV-1, in which developmentally controlled expression of the enhancer in monocytes is mediated largely through the kappa B sites alone [G. E. Griffin, K. Leung, T. M. Folks, S. Kunkel, and G. J. Nabel, Nature (London) 339:70-73, 1989]. Further, we demonstrated that although both Elf-1, an ets family member with significant similarity to the drosophila developmental regulatory protein E74, and Pu.1, a monocyte- and B-cell-specific member of the ets family, bind the purine-rich enhancer region, Elf-1 is the protein which binds predominantly in vivo. A nuclear factor(s) which binds the pets site, an element which has been described only in HIV-2, was detected in extracts of all of the monocytic cells tested. These findings indicate that the mechanism by which cellular factors regulate HIV-2 enhancer function in monocytic cells differs significantly from that of HIV-1 and may offer a partial explanation for the differences in the biological and

  11. The transcriptome of Legionella pneumophila-infected human monocyte-derived macrophages.

    Directory of Open Access Journals (Sweden)

    Christopher T D Price

    Full Text Available Legionella pneumophila is an intracellular bacterial pathogen that invades and replicates within alveolar macrophages through injection of ∼ 300 effector proteins by its Dot/Icm type IV translocation apparatus. The bona fide F-box protein, AnkB, is a nutritional virulence effector that triggers macrophages to generate a surplus of amino acids, which is essential for intravacuolar proliferation. Therefore, the ankB mutant represents a novel genetic tool to determine the transcriptional response of human monocyte-derived macrophages (hMDMs to actively replicating L. pneumophila.Here, we utilized total human gene microarrays to determine the global transcriptional response of hMDMs to infection by wild type or the ankB mutant of L. pneumophila. The transcriptomes of hMDMs infected with either actively proliferating wild type or non-replicative ankB mutant bacteria were remarkably similar. The transcriptome of infected hMDMs was predominated by up-regulation of inflammatory pathways (IL-10 anti-inflammatory, interferon signaling and amphoterin signaling, anti-apoptosis, and down-regulation of protein synthesis pathways. In addition, L. pneumophila modulated diverse metabolic pathways, particularly those associated with bio-active lipid metabolism, and SLC amino acid transporters expression.Taken together, the hMDM transcriptional response to L. pneumophila is independent of intra-vacuolar replication of the bacteria and primarily involves modulation of the immune response and metabolic as well as nutritional pathways.

  12. Piperlongumine Suppresses Proliferation of Human Oral Squamous Cell Carcinoma through Cell Cycle Arrest, Apoptosis and Senescence.

    Science.gov (United States)

    Chen, San-Yuan; Liu, Geng-Hung; Chao, Wen-Ying; Shi, Chung-Sheng; Lin, Ching-Yen; Lim, Yun-Ping; Lu, Chieh-Hsiang; Lai, Peng-Yeh; Chen, Hau-Ren; Lee, Ying-Ray

    2016-01-01

    Oral squamous cell carcinoma (OSCC), an aggressive cancer originating in the oral cavity, is one of the leading causes of cancer deaths in males worldwide. This study investigated the antitumor activity and mechanisms of piperlongumine (PL), a natural compound isolated from Piper longum L., in human OSCC cells. The effects of PL on cell proliferation, the cell cycle, apoptosis, senescence and reactive oxygen species (ROS) levels in human OSCC cells were investigated. PL effectively inhibited cell growth, caused cell cycle arrest and induced apoptosis and senescence in OSCC cells. Moreover, PL-mediated anti-human OSCC behavior was inhibited by an ROS scavenger N-acetyl-l-cysteine (NAC) treatment, suggesting that regulation of ROS was involved in the mechanism of the anticancer activity of PL. These findings suggest that PL suppresses tumor growth by regulating the cell cycle and inducing apoptosis and senescence and is a potential chemotherapy agent for human OSCC cells. PMID:27120594

  13. Piperlongumine Suppresses Proliferation of Human Oral Squamous Cell Carcinoma through Cell Cycle Arrest, Apoptosis and Senescence

    Directory of Open Access Journals (Sweden)

    San-Yuan Chen

    2016-04-01

    Full Text Available Oral squamous cell carcinoma (OSCC, an aggressive cancer originating in the oral cavity, is one of the leading causes of cancer deaths in males worldwide. This study investigated the antitumor activity and mechanisms of piperlongumine (PL, a natural compound isolated from Piper longum L., in human OSCC cells. The effects of PL on cell proliferation, the cell cycle, apoptosis, senescence and reactive oxygen species (ROS levels in human OSCC cells were investigated. PL effectively inhibited cell growth, caused cell cycle arrest and induced apoptosis and senescence in OSCC cells. Moreover, PL-mediated anti-human OSCC behavior was inhibited by an ROS scavenger N-acetyl-l-cysteine (NAC treatment, suggesting that regulation of ROS was involved in the mechanism of the anticancer activity of PL. These findings suggest that PL suppresses tumor growth by regulating the cell cycle and inducing apoptosis and senescence and is a potential chemotherapy agent for human OSCC cells.

  14. Isolation of Human Monocytes by Double Gradient Centrifugation and Their Differentiation to Macrophages in Teflon-coated Cell Culture Bags

    Science.gov (United States)

    Menck, Kerstin; Behme, Daniel; Pantke, Mathias; Reiling, Norbert; Binder, Claudia; Pukrop, Tobias; Klemm, Florian

    2014-01-01

    Human macrophages are involved in a plethora of pathologic processes ranging from infectious diseases to cancer. Thus they pose a valuable tool to understand the underlying mechanisms of these diseases. We therefore present a straightforward protocol for the isolation of human monocytes from buffy coats, followed by a differentiation procedure which results in high macrophage yields. The technique relies mostly on commonly available lab equipment and thus provides a cost and time effective way to obtain large quantities of human macrophages. Briefly, buffy coats from healthy blood donors are subjected to a double density gradient centrifugation to harvest monocytes from the peripheral blood. These monocytes are then cultured in fluorinated ethylene propylene (FEP) Teflon-coated cell culture bags in the presence of macrophage colony-stimulating factor (M-CSF). The differentiated macrophages can be easily harvested and used for subsequent studies and functional assays. Important methods for quality control and validation of the isolation and differentiation steps will be highlighted within the protocol. In summary, the protocol described here enables scientists to routinely and reproducibly isolate human macrophages without the need for cost intensive tools. Furthermore, disease models can be studied in a syngeneic human system circumventing the use of murine macrophages. PMID:25226391

  15. DNA Damage, Cell Cycle Arrest, and Apoptosis Induction Caused by Lead in Human Leukemia Cells

    OpenAIRE

    Yedjou, Clement G.; Tchounwou, Hervey M.; Tchounwou, Paul B.

    2015-01-01

    In recent years, the industrial use of lead has been significantly reduced from paints and ceramic products, caulking, and pipe solder. Despite this progress, lead exposure continues to be a significant public health concern. The main goal of this research was to determine the in vitro mechanisms of lead nitrate [Pb(NO3)2] to induce DNA damage, apoptosis, and cell cycle arrest in human leukemia (HL-60) cells. To reach our goal, HL-60 cells were treated with different concentrations of Pb(NO3)...

  16. Phenotypic and functional characterization of macrophages with therapeutic potential generated from human cirrhotic monocytes in a cohort study

    Science.gov (United States)

    Moore, Joanna K.; Mackinnon, Alison C.; Wojtacha, Dvina; Pope, Caroline; Fraser, Alasdair R.; Burgoyne, Paul; Bailey, Laura; Pass, Chloe; Atkinson, Anne; Mcgowan, Neil W.A.; Manson, Lynn; Turner, Mark L.; Campbell, John D.M.; Forbes, Stuart J.

    2015-01-01

    Background aims Macrophages have complex roles in the liver. The aim of this study was to compare profiles of human monocyte-derived macrophages between controls and cirrhotic patients, to determine whether chronic inflammation affects precursor number or the phenotype, with the eventual aim to develop a cell therapy for cirrhosis. Methods Infusion of human macrophages in a murine liver fibrosis model demonstrated a decrease in markers of liver injury (alanine transaminase, bilirubin, aspartate transaminase) and fibrosis (transforming growth factor-β, α-smooth muscle actin, phosphatidylserine receptor) and an increase in markers of liver regeneration (matrix metalloproteinases [MMP]-9, MMP-12 and TNF-related weak inducer of apoptosis). CD14+ monocytes were then isolated from controls. Monocytes were matured into macrophages for 7 days using a Good Manufacturing Practice–compatible technique. Results There was no significant difference between the mean number of CD14+ monocytes isolated from cirrhotic patients (n = 9) and controls (n = 10); 2.8 ± SEM 0.54 × 108 and 2.5 ± 0.56 × 108, respectively. The mean yield of mature macrophages cultured was also not significantly different between cirrhotic patients and controls (0.9 × 108 ± 0.38 × 108, with more than 90% viability and 0.65 × 108 ± 0.16 × 108, respectively. Maturation to macrophages resulted in up-regulation of a number of genes (MMP-9, CCL2, interleukin [IL]-10 and TNF-related weak inducer of apoptosis). A cytokine and chemokine polymerase chain reaction array, comparing the control and cirrhotic macrophages, revealed no statistically significant differences. Conclusions Macrophages can be differentiated from cirrhotic patients' apheresis-derived CD14 monocytes and develop the same pro-resolution phenotype as control macrophages, indicating their suitability for clinical therapy. PMID:26342993

  17. Human monocytes stimulation by particles of hydroxyapatite, silicon carbide and diamond: in vitro studies of new prosthesis coatings.

    Science.gov (United States)

    Nordsletten, L; Høgåsen, A K; Konttinen, Y T; Santavirta, S; Aspenberg, P; Aasen, A O

    1996-08-01

    Aseptic loosening due to wear and debris formation constitutes the major problem in longevity of joint replacements. Diamond coated onto the prosthesis surface may reduce wear, owing to its excellent tribological properties. A thin diamond coating may be brittle, and we plan eventually to reinforce it with silicon carbide whiskers (SiC). In the present study we compared particles of diamond, SiC and hydroxyapatite (HA) in serum-free cultures of human monocytes. All particles were found to be phagocytozed, and monocyte morphology changed except after the ingestion of diamond. Interleukin-1 beta production was increased on average 30-fold and 38-fold in cultures exposed to HA and SiC, respectively, compared to control and diamond cultures (n = 6). Addition of the phagocytosis inhibitor cytochalasin B inhibited the morphological changes of the monocytes and reduced interleukin-1 beta production. In some experiments particles of polymethylmethacrylate were also included, and the interleukin-1 beta stimulation was in the same range as after HA and SiC stimulation. The results show that diamond particles in serum-free monocyte culture are inert, while SiC and HA have a stimulatory effect comparable to polymethylmethacrylate. With its excellent tribological and biocompatible properties, future studies with diamond coating are warranted. PMID:8853123

  18. Amelioration of Glucolipotoxicity-Induced Endoplasmic Reticulum Stress by a “Chemical Chaperone” in Human THP-1 Monocytes

    Directory of Open Access Journals (Sweden)

    Raji Lenin

    2012-01-01

    Full Text Available Chronic ER stress is emerging as a trigger that imbalances a number of systemic and arterial-wall factors and promote atherosclerosis. Macrophage apoptosis within advanced atherosclerotic lesions is also known to increase the risk of atherothrombotic disease. We hypothesize that glucolipotoxicity might mediate monocyte activation and apoptosis through ER stress. Therefore, the aims of this study are (a to investigate whether glucolipotoxicity could impose ER stress and apoptosis in THP-1 human monocytes and (b to investigate whether 4-Phenyl butyric acid (PBA, a chemical chaperone could resist the glucolipotoxicity-induced ER stress and apoptosis. Cells subjected to either glucolipotoxicity or tunicamycin exhibited increased ROS generation, gene and protein (PERK, GRP-78, IRE1α, and CHOP expression of ER stress markers. In addition, these cells showed increased TRPC-6 channel expression and apoptosis as revealed by DNA damage and increased caspase-3 activity. While glucolipotoxicity/tunicamycin increased oxidative stress, ER stress, mRNA expression of TRPC-6, and programmed the THP-1 monocytes towards apoptosis, all these molecular perturbations were resisted by PBA. Since ER stress is one of the underlying causes of monocyte dysfunction in diabetes and atherosclerosis, our study emphasize that chemical chaperones such as PBA could alleviate ER stress and have potential to become novel therapeutics.

  19. Osthole inhibits proliferation of human breast cancer cells by inducing cell cycle arrest and apoptosis

    Institute of Scientific and Technical Information of China (English)

    Lintao Wang; Yanyan Peng; Kaikai Shi; Haixiao Wang; Jianlei Lu; Yanli Li; Changyan Ma

    2015-01-01

    Recent studies have revealed that osthole,an active constituent isolated from the fruit of Cnidium monnieri (L.) Cusson,a traditional Chinese medicine,possesses anticancer activity.However,its effect on breast cancer cells so far has not been elucidated clearly.In the present study,we evaluated the effects of osthole on the proliferation,cell cycle and apoptosis of human breast cancer cells MDA-MB 435.We demonstrated that osthole is effective in inhibiting the proliferation of MDA-MB 435 cells,The mitochondrion-mediated apoptotic pathway was involved in apoptosis induced by osthole,as indicated by activation of caspase-9 and caspase-3 followed by PARP degradation.The mechanism underlying its effect on the induction of G1 phase arrest was due to the up-regulation of p53 and p21 and down-regulation of Cdk2 and cyclin D1 expression.Were observed taken together,these findings suggest that the anticancer efficacy of osthole is mediated via induction of cell cycle arrest and apoptosis in human breast cancer cells and osthole may be a potential chemotherapeutic agent against human breast cancer.

  20. Carrageenan Induces Cell Cycle Arrest in Human Intestinal Epithelial Cells in Vitro1–3

    Science.gov (United States)

    Bhattacharyya, Sumit; Borthakur, Alip; Dudeja, Pradeep K.; Tobacman, Joanne K.

    2016-01-01

    Multiple studies in animal models have shown that the commonly used food additive carrageenan (CGN) induces inflammation and intestinal neoplasia. We performed the first studies to determine the effects of CGN exposure on human intestinal epithelial cells (IEC) in tissue culture and tested the effect of very low concentrations (1–10 mg/L) of undegraded, high-molecular weight CGN. These concentrations of CGN are less than the anticipated exposure of the human colon to CGN from the average Western diet. In the human colonic epithelial cell line NCM460 and in primary human colonic epithelial cells that were exposed to CGN for 1–8 d, we found increased cell death, reduced cell proliferation, and cell cycle arrest compared with unexposed control cells. After 6–8 d of CGN exposure, the percentage of cells reentering G0–G1 significantly decreased and the percentages of cells in S and G2-M phases significantly increased. Increases in activated p53, p21, and p15 followed CGN exposure, consistent with CGN-induced cell cycle arrest. Additional data, including DNA ladder, poly ADP ribose polymerase Western blot, nuclear DNA staining, and activities of caspases 3 and 7, indicated no evidence of increased apoptosis following CGN exposure and were consistent with CGN-induced necrotic cell death. These data document for the first time, to our knowledge, marked adverse effects of low concentrations of CGN on survival of normal human IEC and suggest that CGN exposure may have a role in development of human intestinal pathology. PMID:18287351

  1. The Immuno-Regulatory Effects of Schisandra chinensis and Its Constituents on Human Monocytic Leukemia Cells

    Directory of Open Access Journals (Sweden)

    Mei-Hsien Lee

    2011-06-01

    Full Text Available Many diseases occur when the immune system is weakened. Intracellular signals activate immuno-responsive cells to produce cytokines that modulate the immune response. Schisandra chinensis has been used traditionally to treat general fatigue, neurasthenia, and spontaneous sweating. In the present study, the effect of constituents of S. chinensis on cytokine release by human monocytic leukemia cells (THP-1 was tested using microparticle-based flow cytometric analysis. Two major lignans, schizandrin (Sch and gomisin A (Gom A, were identified and shown to induce interleukin (IL-8, macrophage inflammatory protein-1β (MIP-1β, and granulocyte-macrophage-colony stimulating factor (GM-CSF release by THP-1 cells. By reverse transcription polymerase chain reaction (RT-PCR or quantitative real-time PCR, there was a dose-dependent increase of IL-8, MIP-1β and GM-CSF mRNA levels. Thus, Sch and Gom A from S. chinensis enhance cytokine release by THP-1 cells and this effect occurs through mRNA upregulation. Upregulation of MIP-1β and GM-CSF in particular may have clinical applications. Therefore, S. chinensis may be therapeutically beneficial by promoting humoral and cell-mediated immune responses.

  2. Is the uptake of pefloxacin in human blood monocytes a simple diffusion process?

    Science.gov (United States)

    Memin, E; Panteix, G; Revol, A

    1996-11-01

    Effective therapy of infections caused by facultative intracellular micro-organisms, which persist after ingestion by mononuclear phagocytic cells, requires antibiotics which inactivate these intraphagocytic bacteria. A previous study demonstrated that intracellular concentrations of normethylpefloxacin, Ci, were the result of simple diffusion and an active efflux involving organic anion transporters. In our work, we studied pefloxacin transport in human monocytes. A kinetic approach was followed to establish the processes involved. Extracellular concentrations, Ce, corresponding to in-vivo pharmacological levels of the drug were used. Uptake of the antibiotic was assessed principally by HPLC. Pefloxacin was accumulated by the cells (Ci/Ce = 3). The uptake was rapid, non saturable, reversible and temperature dependent. At 4 degrees C, the Ci/Ce was equal to 1. At high temperatures, an active and initial process appeared that was not visible at lower temperatures and disappeared in presence of energetic metabolism inhibitor: At 42 degrees C, NaCN and 2.4 DNP (1 mM) reduced the initial transport but they had no significant effect on the subsequent curve. PMID:8961048

  3. Human monocyte-derived dendritic cells turn into foamy dendritic cells with IL-17A.

    Science.gov (United States)

    Salvatore, Giulia; Bernoud-Hubac, Nathalie; Bissay, Nathalie; Debard, Cyrille; Daira, Patricia; Meugnier, Emmanuelle; Proamer, Fabienne; Hanau, Daniel; Vidal, Hubert; Aricò, Maurizio; Delprat, Christine; Mahtouk, Karène

    2015-06-01

    Interleukin 17A (IL-17A) is a proinflammatory cytokine involved in the pathogenesis of chronic inflammatory diseases. In the field of immunometabolism, we have studied the impact of IL-17A on the lipid metabolism of human in vitro-generated monocyte-derived dendritic cells (DCs). Microarrays and lipidomic analysis revealed an intense remodeling of lipid metabolism induced by IL-17A in DCs. IL-17A increased 2-12 times the amounts of phospholipids, cholesterol, triglycerides, and cholesteryl esters in DCs. Palmitic (16:0), stearic (18:0), and oleic (18:ln-9c) acid were the main fatty acid chains present in DCs. They were strongly increased in response to IL-17A while their relative proportion remained unchanged. Capture of extracellular lipids was the major mechanism of lipid droplet accumulation, visualized by electron microscopy and Oil Red O staining. Besides this foamy phenotype, IL-17A induced a mixed macrophage-DC phenotype and expression of the nuclear receptor NR1H3/liver X receptor-α, previously identified in the context of atherosclerosis as the master regulator of cholesterol homeostasis in macrophages. These IL-17A-treated DCs were as competent as untreated DCs to stimulate allogeneic naive T-cell proliferation. Following this first characterization of lipid-rich DCs, we propose to call these IL-17A-dependent cells "foamy DCs" and discuss the possible existence of foamy DCs in atherosclerosis, a metabolic and inflammatory disorder involving IL-17A. PMID:25833686

  4. The Salmonella virulence plasmid spv genes are required for cytopathology in human monocyte-derived macrophages.

    Science.gov (United States)

    Libby, S J; Lesnick, M; Hasegawa, P; Weidenhammer, E; Guiney, D G

    2000-02-01

    The pathogenesis of serious systemic Salmonella infections is characterized by survival and proliferation of bacteria inside macrophages. Infection of human monocyte-derived macrophages in vitro with S. typhimurium or S. dublin produces cytopathology characterized by detachment of cells that contain large numbers of proliferating bacteria. This cytopathology is dependent on the expression of the bacterial spv genes, a virulence locus previously shown to markedly enhance the ability of Salmonella to produce systemic disease. After 24 h of infection, macrophage cultures contain two populations of bacteria: (i) proliferating organisms present in a detached cell fraction; and (ii) a static bacterial population in macrophages remaining attached to the culture well. Mutations in either the essential transcriptional activator SpvR or the key SpvB protein markedly reduce the cytopathic effect of Salmonella infection. The spv-dependent cytopathology in macrophages exhibits characteristics of apoptosis, with release of nucleosomes into the cytoplasm, nuclear condensation and DNA fragmentation. The current findings suggest that the mechanism of the spv effect is through induction of increased cytopathology in host macrophages. PMID:11207562

  5. Induction of proliferation and monocytic differentiation of human CD34+ cells by CD137 ligand signaling.

    Science.gov (United States)

    Jiang, Dongsheng; Yue, Pei Shan Eunice; Drenkard, Daniela; Schwarz, Herbert

    2008-09-01

    CD137 is a member of the tumor necrosis factor receptor family and is involved in the regulation of activation, proliferation, differentiation, and cell death of leukocytes. Bidirectional signaling exists for the CD137 receptor/ligand system, as CD137 ligand, which is expressed as a transmembrane protein, can also transduce signals into the cells on which it is expressed. In this study, we have identified expression of CD137 in human bone marrow and expression of CD137 ligand on a subset of CD34+ cells. Cross-linking of CD137 ligand on CD34+ cells by CD137 ligand agonists induces activation, prolongation of survival, proliferation, and colony formation. CD137 ligand agonists induce differentiation of early hematopoietic progenitor cells to colony-forming units-granulocyte/macrophage and subsequently to monocytes and macrophages but not to dendritic cells. These data uncover a novel function of CD137 and CD137 ligand by showing their participation in the growth and differentiation of hematopoietic progenitor cells. PMID:18566330

  6. TLR2 ligands induce NF-κB activation from endosomal compartments of human monocytes.

    Directory of Open Access Journals (Sweden)

    Karim J Brandt

    Full Text Available Localization of Toll-like receptors (TLR in subcellular organelles is a major strategy to regulate innate immune responses. While TLR4, a cell-surface receptor, signals from both the plasma membrane and endosomal compartments, less is known about the functional role of endosomal trafficking upon TLR2 signaling. Here we show that the bacterial TLR2 ligands Pam3CSK4 and LTA activate NF-κB-dependent signaling from endosomal compartments in human monocytes and in a NF-κB sensitive reporter cell line, despite the expression of TLR2 at the cell surface. Further analyses indicate that TLR2-induced NF-κB activation is controlled by a clathrin/dynamin-dependent endocytosis mechanism, in which CD14 serves as an important upstream regulator. These findings establish that internalization of cell-surface TLR2 into endosomal compartments is required for NF-κB activation. These observations further demonstrate the need of endocytosis in the activation and regulation of TLR2-dependent signaling pathways.

  7. Lobohedleolide induces interleukin-8 production in LPS-stimulated human monocytic cell line THP-1

    Directory of Open Access Journals (Sweden)

    T Oda

    2011-09-01

    Full Text Available Summary: Lobohedleolide (1 has been isolated from a soft coral, Sarcophyton sp., as one of three responsible substances for observed inhibitory activity against TNF-a production in lipopolysaccharide (LPS-stimulated murine macrophage-like RAW 264.7 cells. During the examination of other inflammatory cytokines, we found that only 1 induced the production of interleukin (IL-8 in LPS-stimulated human monocytic THP-1 cells, while the other two compounds did not affect the production of IL-8. Although 1 showed an inhibitory effect on nuclear factor-kappaB (NF-kB activation, this compound induced IL-8 promoter activity, which led to the induction of IL-8 production in LPS-stimulated THP-1 cells. Industrial Relevance: Marine organisms are a rich resource of biologically active secondary metabolites. Two marine natural products and two derivatives of marine sponge compounds have been utilized for medical treatment, and marked numbers of marine natural products and their derivatives are now in clinical and pre-clinical trials. Therefore, marine natural products are attractive sources of medicines and their lead compounds, and elucidation of new bioactivity and the mechanism of action of marine natural products are also a very important study for drug discovery. This report describes a new bioactivity of a diterpene previously obtained from an Indonesian soft corral.

  8. iPLA2β: front and center in human monocyte chemotaxis to MCP-1

    OpenAIRE

    Mishra, Ravi S.; Carnevale, Kevin A.; Cathcart, Martha K.

    2008-01-01

    Monocyte chemoattractant protein-1 (MCP-1) directs migration of blood monocytes to inflamed tissues. Despite the central role of chemotaxis in immune responses, the regulation of chemotaxis by signal transduction pathways and their in vivo significance remain to be thoroughly deciphered. In this study, we examined the intracellular location and functions of two recently identified regulators of chemotaxis, Ca2+-independent phospholipase (iPLA2β) and cytosolic phospholipase (cPLA2α), and subst...

  9. Tenascin expression patterns and cells of monocyte lineage: relationship in human gliomas.

    Science.gov (United States)

    Kulla, A; Liigant, A; Piirsoo, A; Rippin, G; Asser, T

    2000-01-01

    Stromal extracellular matrix (ECM) components are thought to play an important role in regulating invasion of human gliomas. Macrophages and microglial cells may heavily influence the integrity of the extracellular compartment of gliomas, and the affected ECM may play a key role in regulating migratory activity of both tumor cells and macrophages/microglia. The aim of this investigation was to study immunohistochemically the expression patterns of four ECM components: fibronectin, laminin, collagen IV, and tenascin (TN) in human gliomas, with special attention to TN. Our main goal was to study the possible correlation between TN expression and macrophagic/microglial infiltration in gliomas. Altogether, 90 gliomas were studied. Tumors included 46 glioblastomas, 19 anaplastic gliomas, 22 low grade gliomas, and 3 pilocytic astrocytomas. Vascular TN prevailed in perinecrotic areas of glioblastomas, whereas interstitial TN was more often expressed distant from necrosis and in the ECM of anaplastic and low grade gliomas. Double staining with CD68 and anti-TN antibodies showed that macrophagic/microglial density was significantly higher in TN-positive areas of most of the glioblastomas and anaplastic gliomas, whereas microglial percentage from total number of CD68-positive cells was in most of the cases significantly higher in TN-negative areas. In addition, we saw a morphologically spatial correlation between higher densities of macrophagic/microglial infiltration and TN expression in perinecrotic areas in glioblastomas. Attachment of macrophages to TN-positive basement membrane zones of newly formed stromal blood vessels was evident. On the basis of our results, we conclude that TN may play a crucial role in regulating trafficking of cells of monocyte lineage in human gliomas. PMID:10658911

  10. Responsiveness of human monocyte-derived dendritic cells to thimerosal and mercury derivatives

    International Nuclear Information System (INIS)

    Several cases of skin sensitization have been reported following the application of thimerosal, which is composed of ethyl mercury and thiosalicylic acid (TSA). However, few in vitro studies have been carried out on human dendritic cells (DCs) which play an essential role in the initiation of allergic contact dermatitis. The aim of the present study was to identify the effect of thimerosal and other mercury compounds on human DCs. To address this purpose, DCs derived from monocytes (mono-DCs) were used. Data show that thimerosal and mercury derivatives induced DC activation, as monitored by CD86 and HLA-DR overexpression associated with the secretion of tumor necrosis factor α and interleukin 8, similarly to lipopolysaccharide and the sensitizers, 1-chloro-2,4-dinitrobenzene (DNCB) and nickel sulfate, which were used as positive controls. In contrast, TSA, the non-mercury part of thimerosal, as well as dichloronitrobenzene, a DNCB negative control, and the irritant, sodium dodecyl sulfate, had no effect. Moreover, oxidative stress, monitored by ROS induction and depolarization of the mitochondrial membrane potential, was induced by thimerosal and mercury compounds, as well as DNCB, in comparison with hydrogen peroxide, used as a positive control. The role of thiol oxidation in the initiation of mono-DC activation was confirmed by a pre-treatment with N-acetyl-L-cysteine which strongly decreased chemical-induced CD86 overexpression. These data are in agreement with several clinical observations of the high relevance of thimerosal in patch-test reactions and prove that human mono-DCs are useful in vitro tools for determining the allergenic potency of chemicals.

  11. Methods for defining distinct bioenergetic profiles in platelets, lymphocytes, monocytes, and neutrophils, and the oxidative burst from human blood

    OpenAIRE

    Chacko, Balu K; Kramer, Philip A.; Ravi, Saranya; Johnson, Michelle S.; Hardy, Robert W.; Ballinger, Scott W.; Darley-Usmar, Victor M.

    2013-01-01

    Peripheral blood mononuclear cells and platelets have long been recognized as having the potential to act as sensitive markers for mitochondrial dysfunction in a broad range of pathological conditions. However, the bioenergetic function of these cells has not been examined from the same donors, yet this is important for the selection of cell types for translational studies. Here, we demonstrate the measurement of cellular bioenergetics in isolated human monocytes, lymphocytes, and platelets, ...

  12. MicroRNA-155 modulates the interleukin-1 signaling pathway in activated human monocyte-derived dendritic cells

    OpenAIRE

    Ceppi, Maurizio; Pereira, Patricia M.; Dunand-Sauthier, Isabelle; Barras, Emmanuèle; Reith, Walter; Santos, Manuel A.; Pierre, Philippe

    2009-01-01

    In response to inflammatory stimulation, dendritic cells (DCs) have a remarkable pattern of differentiation (maturation) that exhibits specific mechanisms to control immunity. Here, we show that in response to Lipopolysaccharides (LPS), several microRNAs (miRNAs) are regulated in human monocyte-derived dendritic cells. Among these miRNAs, miR-155 is highly up-regulated during maturation. Using LNA silencing combined to microarray technology, we have identified the Toll-like receptor/interleuk...

  13. Growth inhibitory effect of 4-phenyl butyric acid on human gastric cancer cells is associated with cell cycle arrest

    Institute of Scientific and Technical Information of China (English)

    Long-Zhu Li; Hong-Xia Deng; Wen-Zhu Lou; Xue-Yan Sun; Meng-Wan Song; Jing Tao; Bing-Xiu Xiao; Jun-Ming Guo

    2012-01-01

    AIM: To investigate the growth effects of 4-phenyl butyric acid (PBA) on human gastric carcinoma cells and their mechanisms. METHODS: Moderately-differentiated human gastric carcinoma SGC-7901 and lowly-differentiated MGC-803 cells were treated with 5, 10, 20, 40, and 60 μmol/L PBA for 1-4 d. Cell proliferation was detected using the MTT colorimetric assay. Cell cycle distributions were examined using flow cytometry. RESULTS: The proliferation of gastric carcinoma cells was inhibited by PBA in a dose- and time-dependent fashion. Flow cytometry showed that SGC-7901 cells treated with low concentrations of PBA were arrested at the G0/G1 phase, whereas cells treated with high concentrations of PBA were arrested at the G2/M phase. Although MGC-803 cells treated with low concentrations of PBA were also arrested at the G0/G1 phase, cells treated with high concentrations of PBA were arrested at the S phase. CONCLUSION: The growth inhibitory effect of PBA on gastric cancer cells is associated with alteration of the cell cycle. For moderately-differentiated gastric cancer cells, the cell cycle was arrested at the G0/G1 and G2/M phases. For lowly-differentiated gastric cancer cells, the cell cycle was arrested at the G0/G1 and S phases.

  14. Organochlorine insecticides induce NADPH oxidase-dependent reactive oxygen species in human monocytic cells via phospholipase A2/arachidonic acid.

    Science.gov (United States)

    Mangum, Lee C; Borazjani, Abdolsamad; Stokes, John V; Matthews, Anberitha T; Lee, Jung Hwa; Chambers, Janice E; Ross, Matthew K

    2015-04-20

    Bioaccumulative organohalogen chemicals, such as organochlorine (OC) insecticides, have been increasingly associated with disease etiology; however, the mechanistic link between chemical exposure and diseases, such as atherosclerosis, cancer, and diabetes, is complex and poorly defined. Systemic oxidative stress stemming from OC exposure might play a vital role in the development of these pathologies. Monocytes are important surveillance cells of the innate immune system that respond to extracellular signals possessing danger-associated molecular patterns by synthesizing oxyradicals, such as superoxide, for the purpose of combating infectious pathogens. We hypothesized that OC chemicals can be toxic to monocytes because of an inappropriate elevation in superoxide-derived reactive oxygen species (ROS) capable of causing cellular oxidative damage. Reactive oxyradicals are generated in monocytes in large part by NADPH oxidase (Nox). The present study was conducted to examine the ability of two chlorinated cyclodiene compounds, trans-nonachlor and dieldrin, as well as p,p'-DDE, a chlorinated alicyclic metabolite of DDT, to stimulate Nox activity in a human monocytic cell line and to elucidate the mechanisms for this activation. Human THP-1 monocytes treated with either trans-nonachlor or dieldrin (0.1-10 μM in the culture medium) exhibited elevated levels of intracellular ROS, as evidenced by complementary methods, including flow cytometry analysis using the probe DCFH-DA and hydroethidine-based fluorometric and UPLC-MS assays. In addition, the induced reactive oxygen flux caused by trans-nonachlor was also observed in two other cell lines, murine J774 macrophages and human HL-60 cells. The central role of Nox in OC-mediated oxidative stress was demonstrated by the attenuated superoxide production in OC-exposed monocytes treated with the Nox inhibitors diphenyleneiodonium and VAS-2870. Moreover, monocytes challenged with OCs exhibited increased phospho-p47(phox

  15. Yersinia enterocolitica serotype O:3 alters the expression of serologic HLA-B27 epitopes on human monocytes.

    OpenAIRE

    Wuorela, M; Jalkanen, S; Kirveskari, J; Laitio, P; Granfors, K

    1997-01-01

    The expression of serologic HLA-B27 epitopes on leukocytes of patients with reactive arthritis or ankylosing spondylitis has been shown to be modified in the course of the disease. The purpose of this work was to study whether phagocytosis of arthritis-triggering microbes in vitro alters the expression of HLA-B27 molecules on human antigen-presenting cells and to characterize the underlying mechanisms. Human monocytes and HLA-B27- or HLA-A2-transfected human U-937 cells were exposed to Yersin...

  16. DNA Damage, Cell Cycle Arrest, and Apoptosis Induction Caused by Lead in Human Leukemia Cells.

    Science.gov (United States)

    Yedjou, Clement G; Tchounwou, Hervey M; Tchounwou, Paul B

    2016-01-01

    In recent years, the industrial use of lead has been significantly reduced from paints and ceramic products, caulking, and pipe solder. Despite this progress, lead exposure continues to be a significant public health concern. The main goal of this research was to determine the in vitro mechanisms of lead nitrate [Pb(NO₃)₂] to induce DNA damage, apoptosis, and cell cycle arrest in human leukemia (HL-60) cells. To reach our goal, HL-60 cells were treated with different concentrations of Pb(NO₃)₂ for 24 h. Live cells and necrotic death cells were measured by the propidium idiode (PI) assay using the cellometer vision. Cell apoptosis was measured by the flow cytometry and DNA laddering. Cell cycle analysis was evaluated by the flow cytometry. The result of the PI demonstrated a significant (p rupture by Pb(NO₃)₂ compared to the control. Data generated from the comet assay indicated a concentration-dependent increase in DNA damage, showing a significant increase (p < 0.05) in comet tail-length and percentages of DNA cleavage. Data generated from the flow cytometry assessment indicated that Pb(NO₃)₂ exposure significantly (p < 0.05) increased the proportion of caspase-3 positive cells (apoptotic cells) compared to the control. The flow cytometry assessment also indicated Pb(NO₃)₂ exposure caused cell cycle arrest at the G₀/G₁ checkpoint. The result of DNA laddering assay showed presence of DNA smear in the agarose gel with little presence of DNA fragments in the treated cells compared to the control. In summary, Pb(NO₃)₂ inhibits HL-60 cells proliferation by not only inducing DNA damage and cell cycle arrest at the G₀/G₁ checkpoint but also triggering the apoptosis through caspase-3 activation and nucleosomal DNA fragmentation accompanied by secondary necrosis. We believe that our study provides a new insight into the mechanisms of Pb(NO₃)₂ exposure and its associated adverse health effects. PMID:26703663

  17. Higher infection of dengue virus serotype 2 in human monocytes of patients with G6PD deficiency.

    Directory of Open Access Journals (Sweden)

    Yuan-Chang Chao

    Full Text Available The prevalence of glucose-6-phosphate dehydrogenase (G6PD deficiency is high in Asia. An ex vivo study was conducted to elucidate the association of G6PD deficiency and dengue virus (DENV infection when many Asian countries are hyper-endemic. Human monocytes from peripheral mononuclear cells collected from 12 G6PD-deficient patients and 24 age-matched controls were infected with one of two DENV serotype 2 (DENV-2 strains-the New Guinea C strain (from a case of dengue fever or the 16681 strain (from a case of dengue hemorrhagic fever with a multiplicity of infection of 0.1. The infectivity of DENV-2 in human monocytes was analyzed by flow cytometry. Experimental results indicated that the monocytes of G6PD-deficient patients exhibited a greater levels of infection with DENV-2 New Guinea C strain than did those in healthy controls [mean+/-SD:33.6%+/-3.5 (27.2% approximately 39.2% vs 20.3%+/-6.2 (8.0% approximately 30.4%, P<0.01]. Similar observations were made of infection with the DENV-2 16681 strain [40.9%+/-3.9 (35.1% approximately 48.9% vs 27.4%+/-7.1 (12.3% approximately 37.1%, P<0.01]. To our knowledge, this study demonstrates for the first time higher infection of human monocytes in G6PD patients with the dengue virus, which may be important in increasing epidemiological transmission and perhaps with the potential to develop more severe cases pathogenically.

  18. Proteomic analysis of arginine methylation sites in human cells reveals dynamic regulation during transcriptional arrest

    DEFF Research Database (Denmark)

    Sylvestersen, Kathrine B; Horn, Heiko; Jungmichel, Stephanie;

    2014-01-01

    contain regulated functions on their own. Collectively, we present a site-specific MMA dataset in human cells and demonstrate for the first time that MMA is a dynamic post-translational modification regulated during transcriptional arrest by a hitherto uncharacterized arginine demethylase....... mono-methylation (MMA) sites. We thereby identify 1,027 site-specific MMA sites on 494 human proteins, discovering numerous novel mono-methylation targets and confirming the majority of currently known MMA substrates. Nuclear RNA-binding proteins involved in RNA processing, RNA localization......, transcription, and chromatin remodeling are predominantly found modified with MMA. Despite this, MMA sites prominently are located outside RNA-binding domains as compared to the proteome-wide distribution of arginine residues. Quantification of arginine methylation in cells treated with Actinomycin D uncovers...

  19. Clinical significance of monocyte heterogeneity.

    Science.gov (United States)

    Stansfield, Brian K; Ingram, David A

    2015-01-01

    Monocytes are primitive hematopoietic cells that primarily arise from the bone marrow, circulate in the peripheral blood and give rise to differentiated macrophages. Over the past two decades, considerable attention to monocyte diversity and macrophage polarization has provided contextual clues into the role of myelomonocytic derivatives in human disease. Until recently, human monocytes were subdivided based on expression of the surface marker CD16. "Classical" monocytes express surface markers denoted as CD14(++)CD16(-) and account for greater than 70% of total monocyte count, while "non-classical" monocytes express the CD16 antigen with low CD14 expression (CD14(+)CD16(++)). However, recognition of an intermediate population identified as CD14(++)CD16(+) supports the new paradigm that monocytes are a true heterogeneous population and careful identification of specific subpopulations is necessary for understanding monocyte function in human disease. Comparative studies of monocytes in mice have yielded more dichotomous results based on expression of the Ly6C antigen. In this review, we will discuss the use of monocyte subpopulations as biomarkers of human disease and summarize correlative studies in mice that may yield significant insight into the contribution of each subset to disease pathogenesis. PMID:25852821

  20. Mechanisms involved in ceramide-induced cell cycle arrest in human hepatocarcinoma cells

    Institute of Scientific and Technical Information of China (English)

    Jing Wang; Xiao-Wen Lv; Jie-Ping Shi; Xiao-Song Hu

    2007-01-01

    AIM:To investigate the effect of ceramide on the cell cycle in human hepatocarcinoma Bel7402 cells.Possible molecular mechanisms were explored.METHODS:[3-(4,5)-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide(MTT)assay,plasmid transfection,reporter assay,FACS and Western blotting analyses were employed to investigate the effect and the related molecular mechanisms of C2-ceramide on the cell cycle of Bel7402 cells.RESULTS:C2-ceramide was found to inhibit the growth of Bel7402 cells by inducing cell cycle arrest.During the process,the expression of p21 protein increased,while that of cyclinD1,phospho-ERK1/2 and c-myc decreased.Furthermore,the level of CDK7 was downregulated,while the transcriptional activity of PPARγ was upregulated.Addition of GW9662,which is a PPARγ specific antagonist,could reserve the modulation action on CDK7.CONCLUSION:Our results support the hypothesis that cell cycle arrest induced by C2-ceramide may be mediated via accumulation of p21 and reduction of cyclinD1 and CDK7,at least partly,through PPARγ activation.The ERK signaling pathway was involved in this process.

  1. Berberine induces p53-dependent cell cycle arrest and apoptosis of human osteosarcoma cells by inflicting DNA damage

    Energy Technology Data Exchange (ETDEWEB)

    Liu Zhaojian; Liu Qiao; Xu Bing; Wu Jingjing [Key Laboratory of Experimental Teratology of Ministry of Education and Institute of Molecular Medicine and Genetics, Shandong University School of Medicine, Jinan, Shandong 250012 (China); Guo Chun; Zhu Faliang [Institute of Immunology, Shandong University School of Medicine, Jinan, Shandong 250012 (China); Yang Qiaozi [Department of Genetics, Rutgers University, Piscataway, NJ 08854 (United States); Gao Guimin [Key Laboratory of Experimental Teratology of Ministry of Education and Institute of Molecular Medicine and Genetics, Shandong University School of Medicine, Jinan, Shandong 250012 (China); Gong Yaoqin [Key Laboratory of Experimental Teratology of Ministry of Education and Institute of Molecular Medicine and Genetics, Shandong University School of Medicine, Jinan, Shandong 250012 (China)], E-mail: yxg8@sdu.edu.cn; Shao Changshun [Key Laboratory of Experimental Teratology of Ministry of Education and Institute of Molecular Medicine and Genetics, Shandong University School of Medicine, Jinan, Shandong 250012 (China); Department of Genetics, Rutgers University, Piscataway, NJ 08854 (United States)], E-mail: shao@biology.rutgers.edu

    2009-03-09

    Alkaloid berberine is widely used for the treatment of diarrhea and other diseases. Many laboratory studies showed that it exhibits anti-proliferative activity against a wide spectrum of cancer cells in culture. In this report we studied the mechanisms underlying the inhibitory effects of berberine on human osteosarcoma cells and on normal osteoblasts. The inhibition was largely attributed to cell cycle arrest at G1 and G2/M, and to a less extent, to apoptosis. The G1 arrest was dependent on p53, as G1 arrest was abolished in p53-deficient osteosarcoma cells. The induction of G1 arrest and apoptosis was accompanied by a p53-dependent up-regulation of p21 and pro-apoptotic genes. However, the G2/M arrest could be induced by berberine regardless of the status of p53. Interestingly, DNA double-strand breaks, as measured by the phosphorylation of H2AX, were remarkably accumulated in berberine-treated cells in a dose-dependent manner. Thus, one major mechanism by which berberine exerts its growth-inhibitory effect is to inflict genomic lesions on cells, which in turn trigger the activation of p53 and the p53-dependent cellular responses including cell cycle arrest and apoptosis.

  2. Heat shock protein and heat shock factor 1 expression and localization in vaccinia virus infected human monocyte derived macrophages

    Directory of Open Access Journals (Sweden)

    Dziedzic Jakub

    2005-10-01

    Full Text Available Abstract Background Viruses remain one of the inducers of the stress response in the infected cells. Heat shock response induced by vaccinia virus (VV infection was studied in vitro in human blood monocyte derived macrophages (MDMs as blood cells usually constitute the primary site of the infection. Methods Human blood monocytes were cultured for 12 – 14 days. The transcripts of heat shock factor 1 (HSF1, heat shock protein 70 (HSP70, heat shock protein 90 (HSP90 and two viral genes (E3L and F17R were assayed by reverse transcriptase-polymerase chain reaction (RT-PCR, and the corresponding proteins measured by Western blot. Heat shock factor 1 DNA binding activities were estimated by electrophoretic mobility shift assay (EMSA and its subcellular localization analyzed by immunocytofluorescence. Results It appeared that infection with vaccinia virus leads to activation of the heat shock factor 1. Activation of HSF1 causes increased synthesis of an inducible form of the HSP70 both at the mRNA and the protein level. Although HSP90 mRNA was enhanced in vaccinia virus infected cells, the HSP90 protein content remained unchanged. At the time of maximum vaccinia virus gene expression, an inhibitory effect of the infection on the heat shock protein and the heat shock factor 1 was most pronounced. Moreover, at the early phase of the infection translocation of HSP70 and HSP90 from the cytoplasm to the nucleus of the infected cells was observed. Conclusion Preferential nuclear accumulation of HSP70, the major stress-inducible chaperone protein, suggests that VV employs this particular mechanism of cytoprotection to protect the infected cell rather than to help viral replication. The results taken together with our previuos data on monocytes or MDMs infected with VV or S. aureus strongly argue that VV employs multiple cellular antiapoptotic/cytoprotective mechanisms to prolong viability and proinflammatory activity of the cells of monocytic

  3. STAT3 activation in monocytes accelerates liver cancer progression

    International Nuclear Information System (INIS)

    Signal transducer and activator of transcription 3 (STAT3) is an important transcription factor ubiquitously expressed in different cell types. STAT3 plays an essential role in cell survival, proliferation, and differentiation. Aberrantly hyper-activated STAT3 signaling in cancer cells and in the tumor microenvironment has been detected in a wide variety of human cancers and is considered an important factor for cancer initiation, development, and progression. However, the role of STAT3 activation in monocytes in the development of HCC has not been well understood. Immunohistochemical analysis of phosphorylated STAT3 was performed on tissue microarray from HCC patients. Using a co-culture system in vivo, HCC cell growth was determined by the MTT assay. In vivo experiments were conducted with mice given diethylinitrosamine (DEN), which induces HCC was used to investigate the role of STAT3 expression in monocytes on tumor growth. Real-time PCR was used to determine the expression of cell proliferation and cell arrest associated genes in the tumor and nontumor tissue from liver. Phosphorylated STAT3 was found in human hepatocellular carcinoma tissue samples and was expressed in tumor cells and also in monocytes. Phosphorylated STAT3 expression in monocyte was significantly correlated to advanced clinical stage of HCC and a poor prognosis. Using a co-culture system in vivo, monocytes promoted HCC cell growth via the IL-6/STAT3 signaling pathway. The STAT3 inhibitor, NSC 74859, significantly suppressed tumor growth in vivo in mice with diethylinitrosamine (DEN)-induced HCC. In this animal model, blockade of STAT3 with NSC 74859 induced tumor cell apoptosis, while inhibiting both tumor cells and monocytes proliferation. Furthermore, NSC 74859 treatment suppressed cancer associated inflammation in DEN-induce HCC. Our data suggest constitutively activated STAT3 monocytes promote liver tumorigenesis in clinical patients and animal experiments. Thus, STAT3 in tumor

  4. STAT3 activation in monocytes accelerates liver cancer progression

    Directory of Open Access Journals (Sweden)

    Wu Wen-Yong

    2011-12-01

    Full Text Available Abstract Background Signal transducer and activator of transcription 3 (STAT3 is an important transcription factor ubiquitously expressed in different cell types. STAT3 plays an essential role in cell survival, proliferation, and differentiation. Aberrantly hyper-activated STAT3 signaling in cancer cells and in the tumor microenvironment has been detected in a wide variety of human cancers and is considered an important factor for cancer initiation, development, and progression. However, the role of STAT3 activation in monocytes in the development of HCC has not been well understood. Methods Immunohistochemical analysis of phosphorylated STAT3 was performed on tissue microarray from HCC patients. Using a co-culture system in vivo, HCC cell growth was determined by the MTT assay. In vivo experiments were conducted with mice given diethylinitrosamine (DEN, which induces HCC was used to investigate the role of STAT3 expression in monocytes on tumor growth. Real-time PCR was used to determine the expression of cell proliferation and cell arrest associated genes in the tumor and nontumor tissue from liver. Results Phosphorylated STAT3 was found in human hepatocellular carcinoma tissue samples and was expressed in tumor cells and also in monocytes. Phosphorylated STAT3 expression in monocyte was significantly correlated to advanced clinical stage of HCC and a poor prognosis. Using a co-culture system in vivo, monocytes promoted HCC cell growth via the IL-6/STAT3 signaling pathway. The STAT3 inhibitor, NSC 74859, significantly suppressed tumor growth in vivo in mice with diethylinitrosamine (DEN-induced HCC. In this animal model, blockade of STAT3 with NSC 74859 induced tumor cell apoptosis, while inhibiting both tumor cells and monocytes proliferation. Furthermore, NSC 74859 treatment suppressed cancer associated inflammation in DEN-induce HCC. Conclusion Our data suggest constitutively activated STAT3 monocytes promote liver tumorigenesis in clinical

  5. Human cytomegalovirus inhibits maturation and impairs function of monocyte-derived dendritic cells.

    Science.gov (United States)

    Moutaftsi, Magdalena; Mehl, Anja M; Borysiewicz, Leszek K; Tabi, Zsuzsanna

    2002-04-15

    Dendritic cells (DCs) play a pivotal role in the generation of virus-specific cytotoxic T-cell responses, but some viruses can render DCs inefficient in stimulating T cells. We studied whether infection of DCs with human cytomegalovirus (HCMV) results in a suppression of DC function which may assist HCMV in establishing persistence. The effect of HCMV infection on the phenotype and function of monocyte-derived DCs and on their ability to mature following infection with an endothelial cell-adapted clinical HCMV isolate were studied. HCMV infection induced no maturation of DCs; instead, it efficiently down-regulated the expression of surface major histocompatibility complex (MHC) class I, CD40, and CD80 molecules. Slight down-regulation of MHC class II and CD86 molecules was also observed. Lipopolysaccharide (LPS)-induced maturation of infected DCs was strongly inhibited, as indicated by lower levels of surface expression of MHC class I, class II, costimulatory, and CD83 molecules. The down-regulation or inhibition of these surface markers occurred only in HCMV antigen-positive DCs. DCs produced no interleukin 12 (IL-12) and only low levels of tumor necrosis factor alpha (TNF-alpha) upon HCMV infection. Furthermore, cytokine production upon stimulation with LPS or CD40L was significantly impaired. Inhibition of cytokine production did not depend on viral gene expression as UV-irradiated HCMV resulted in the same effect. Proliferation and cytotoxicity of T cells specific to a recall antigen presented by DCs were also reduced when DCs were HCMV infected. This study shows that HCMV inhibits DC function, revealing a powerful viral strategy to delay or prevent the generation of virus-specific cytotoxic T cells. PMID:11929782

  6. Role of ATM in bystander signaling between human monocytes and lung adenocarcinoma cells.

    Science.gov (United States)

    Ghosh, Somnath; Ghosh, Anu; Krishna, Malini

    2015-12-01

    The response of a cell or tissue to ionizing radiation is mediated by direct damage to cellular components and indirect damage mediated by radiolysis of water. Radiation affects both irradiated cells and the surrounding cells and tissues. The radiation-induced bystander effect is defined by the presence of biological effects in cells that were not themselves in the field of irradiation. To establish the contribution of the bystander effect in the survival of the neighboring cells, lung carcinoma A549 cells were exposed to gamma-irradiation, 2Gy. The medium from the irradiated cells was transferred to non-irradiated A549 cells. Irradiated A549 cells as well as non-irradiated A549 cells cultured in the presence of medium from irradiated cells showed decrease in survival and increase in γ-H2AX and p-ATM foci, indicating a bystander effect. Bystander signaling was also observed between different cell types. Phorbol-12-myristate-13-acetate (PMA)-stimulated and gamma-irradiated U937 (human monocyte) cells induced a bystander response in non-irradiated A549 (lung carcinoma) cells as shown by decreased survival and increased γ-H2AX and p-ATM foci. Non-stimulated and/or irradiated U937 cells did not induce such effects in non-irradiated A549 cells. Since ATM protein was activated in irradiated cells as well as bystander cells, it was of interest to understand its role in bystander effect. Suppression of ATM with siRNA in A549 cells completely inhibited bystander effect in bystander A549 cells. On the other hand suppression of ATM with siRNA in PMA stimulated U937 cells caused only a partial inhibition of bystander effect in bystander A549 cells. These results indicate that apart from ATM, some additional factor may be involved in bystander effect between different cell types. PMID:26653982

  7. What are the roles of macrophages and monocytes in human pregnancy?

    Science.gov (United States)

    Tang, Mao-Xing; Hu, Xiao-Hui; Liu, Zhao-Zhao; Kwak-Kim, Joanne; Liao, Ai-Hua

    2015-11-01

    During pregnancy, the maternal immune system is challenged by the semi-allogeneic fetus, which leads to systemic and local immunity. Systemic immunity, including enhanced innate immunity with increased activation of monocytes, is induced by various placental factors. Maternal immune adaptations are most evident at the feto-maternal interface, where macrophages are enriched and communicate with various decidual leukocytes. These cells are not only contributing to the protection of the growing fetus from microorganisms, but also aiding placental development by promoting trophoblast invasion and spiral artery remodeling, and the parturition process. Thus, monocytes and macrophages concurrently play important roles throughout the trimesters. Dysregulation of these cells may thus lead to pregnancy complications, such as pre-eclampsia and preterm labor. In this review, monocytes and macrophage subsets and their roles in normal and pathological pregnancies are reviewed. PMID:26340023

  8. Expression of interleukin 2 receptors and binding of interleukin 2 by gamma interferon-induced human leukemic and normal monocytic cells

    OpenAIRE

    1985-01-01

    Gamma interferon induced surface expression of interleukin 2 (IL-2) receptors on normal human monocytes and the monocytoid cell lines U937 and HL60. These receptors were detected by anti-IL-2 receptor monoclonal antibodies, and U937 IL-2 receptors were indistinguishable from T lymphocyte IL-2 receptors by immunoprecipitation. Also, U937 IL- 2 receptors bound biologically active IL-2. These results suggest a role for monocyte IL-2 receptors in T cell/monocyte interaction during an immune respo...

  9. Comparison of Ultrastructural Cytotoxic Effects of Carbon and Carbon/Iron Particulates on Human Monocyte-Derived Macrophages

    OpenAIRE

    Long, John F.; Waldman, W James; Kristovich, Robert; Williams, Marshall; Knight, Deborah; Dutta, Prabir K

    2004-01-01

    In this study, we tested the hypothesis that the presence of iron in carbon particulates enhances ultrastructural perturbation in human monocyte-derived macrophages (MDMs) after phagocytosis. We used 1-μm synthetic carbon-based particulates, designed to simulate environmental particulates of mass median aerodynamic diameter ≤ 2.5 μm (PM2.5). Cultures of human MDMs or T-lymphocytes (as a nonphagocytic control) were exposed to carbon or carbon/iron particulates for various time periods and exam...

  10. OSCAR Is a Receptor for Surfactant Protein D That Activates TNF-α Release from Human CCR2+ Inflammatory Monocytes

    DEFF Research Database (Denmark)

    Barrow, Alexander D; Palarasah, Yaseelan; Bugatti, Mattia;

    2015-01-01

    collagenous region of recombinant SP-D and captured native SP-D from human bronchoalveolar lavage. OSCAR localized in an intracellular compartment of alveolar macrophages together with SP-D. Moreover, we found OSCAR on the surface of interstitial lung and blood CCR2(+) inflammatory monocytes, which secreted...... TNF-α when exposed to SP-D in an OSCAR-dependent fashion. OSCAR and SP-D did not exclusively colocalize in lung, as they were also highly expressed in atherosclerotic plaques of human aorta, supporting a role for this interaction in atherosclerosis. Our results identify the OSCAR:SP-D interaction as a...

  11. ApoE Production in Human Monocytes and Its Regulation by Inflammatory Cytokines

    OpenAIRE

    Braesch-Andersen, Sten; Paulie, Staffan; Smedman, Christian; Mia, Sohel; Kumagai-Braesch, Makiko

    2013-01-01

    The apoE production by tissue macrophages is crucial for the prevention of atherosclerosis and the aim of this study was to further elucidate how this apolipoprotein is regulated by cytokines present during inflammation. Here we studied apoE production in peripheral blood mononuclear cells (PBMC) and analysis was made with a newly developed apoE ELISpot assay. In PBMC, apoE secretion was restricted to monocytes with classical (CD14++CD16−) and intermediate (CD14+CD16+) monocytes being the mai...

  12. Genistein inhibits human TNF-α-induced porcine endothelial cell adhesiveness for human monocytes and natural killer cells

    Institute of Scientific and Technical Information of China (English)

    2002-01-01

    Cellular immune response is a major barrier to xenotransplantation. Human tumor necrosis factor-α (hTNF-α) possesses cross-species activity and directly amplifies the immune rejection via the upregulation of adhesion molecules on porcine endothelium. We investigated the role of protein tyrosine phosphorylation in the induction of expression of E-sclectin and vascular cell adhesion molecule-1 (VCAM-1), and the augmentation of adhesion of human peripheral blood monocytes (PBMo) and natural killer cells (PBNK), after rhTNF-α-stimulation of porcine aortic endothelial cells (PAEC) in vitro, rhTNF-α-increased adhesiveness of PAEC for both PBMo and PBNK was dose-dependently reduced by pretreatment of PAEC with the selective protein tyrosine kinase (PTK) inhibitor genistein. The inhibitory effect occurred at the early time of PAEC activation triggered by rhTNF-α, and was completely reversible. PTK activity assay indicated that genistein also suppressed rhTNF-α stimulated activation of protein tyrosine kinases (PTKs) in PAEC in a dose-dependent manner. Flow cytometric analysis showed that genistein inhibited the upregulation of E-selectin and VCAM-1 by rhTNF-α. These results suggest that PTKs may regulate the expression of E-selectin and VCAM-1 on PAEC and the adherence of PBMo and PBNK induced by rhTNF-α. Moreover, dietary genistein, used as an adhesion antagonist, may contribute to managing the cell-mediated rejection in the clinical application.

  13. Macrophage-derived human resistin is induced in multiple helminth infections and promotes inflammatory monocytes and increased parasite burden.

    Science.gov (United States)

    Jang, Jessica C; Chen, Gang; Wang, Spencer H; Barnes, Mark A; Chung, Josiah I; Camberis, Mali; Le Gros, Graham; Cooper, Philip J; Steel, Cathy; Nutman, Thomas B; Lazar, Mitchell A; Nair, Meera G

    2015-01-01

    Parasitic helminth infections can be associated with lifelong morbidity such as immune-mediated organ failure. A better understanding of the host immune response to helminths could provide new avenues to promote parasite clearance and/or alleviate infection-associated morbidity. Murine resistin-like molecules (RELM) exhibit pleiotropic functions following helminth infection including modulating the host immune response; however, the relevance of human RELM proteins in helminth infection is unknown. To examine the function of human resistin (hResistin), we utilized transgenic mice expressing the human resistin gene (hRetnTg+). Following infection with the helminth Nippostrongylus brasiliensis (Nb), hResistin expression was significantly upregulated in infected tissue. Compared to control hRetnTg- mice, hRetnTg+ mice suffered from exacerbated Nb-induced inflammation characterized by weight loss and increased infiltration of inflammatory monocytes in the lung, along with elevated Nb egg burdens and delayed parasite expulsion. Genome-wide transcriptional profiling of the infected tissue revealed that hResistin promoted expression of proinflammatory cytokines and genes downstream of toll-like receptor signaling. Moreover, hResistin preferentially bound lung monocytes, and exogenous treatment of mice with recombinant hResistin promoted monocyte recruitment and proinflammatory cytokine expression. In human studies, increased serum resistin was associated with higher parasite load in individuals infected with soil-transmitted helminths or filarial nematode Wuchereria bancrofti, and was positively correlated with proinflammatory cytokines. Together, these studies identify human resistin as a detrimental factor induced by multiple helminth infections, where it promotes proinflammatory cytokines and impedes parasite clearance. Targeting the resistin/proinflammatory cytokine immune axis may provide new diagnostic or treatment strategies for helminth infection and associated

  14. Regulation of interleukin-1 beta production by glucocorticoids in human monocytes: the mechanism of action depends on the activation signal.

    Science.gov (United States)

    Hurme, M; Siljander, P; Anttila, H

    1991-11-14

    Glucocorticoids are known to downregulate interleukin-1 beta production in monocytic cells by two different mechanims: direct inhibition of the gene transcription and destabilization of the preformed interleukin-1 beta mRNA. Now we have examined the effect of the nature of the monocyte activating signal on these two inhibitory mechanims. When human monocytes were preincubated with dexamethasone for 1 hour and then stimulated either with bacterial lipopolysaccharide or phorbol myristate, it was found that dexamethasone inhibited the lipopolysaccharide-induced interleukin-1 beta protein production, but the phorbol myristate-induced production was increased 3-10 fold. This difference was also seen at the mRNA level. When dexamethasone was added to the cultures 3 hours after the stimulators, it clearly decreased the interleukin-1 beta mRNA levels regardless of the stimulator used (although the effect was clearly weaker on the PMA-induced mRNA). Thus these data suggest that the phorbol myristate-induced signal (prolonged protein kinase C activation?) cannot be inhibited by prior incubation with dexamethasone and it also protects the induced mRNA for the degradative action of dexamethasone. PMID:1953785

  15. Palmitate-induced inflammatory pathways in human adipose microvascular endothelial cells promote monocyte adhesion and impair insulin transcytosis.

    Science.gov (United States)

    Pillon, Nicolas J; Azizi, Paymon M; Li, Yujin E; Liu, Jun; Wang, Changsen; Chan, Kenny L; Hopperton, Kathryn E; Bazinet, Richard P; Heit, Bryan; Bilan, Philip J; Lee, Warren L; Klip, Amira

    2015-07-01

    Obesity is associated with inflammation and immune cell recruitment to adipose tissue, muscle and intima of atherosclerotic blood vessels. Obesity and hyperlipidemia are also associated with tissue insulin resistance and can compromise insulin delivery to muscle. The muscle/fat microvascular endothelium mediates insulin delivery and facilitates monocyte transmigration, yet its contribution to the consequences of hyperlipidemia is poorly understood. Using primary endothelial cells from human adipose tissue microvasculature (HAMEC), we investigated the effects of physiological levels of fatty acids on endothelial inflammation and function. Expression of cytokines and adhesion molecules was measured by RT-qPCR. Signaling pathways were evaluated by pharmacological manipulation and immunoblotting. Surface expression of adhesion molecules was determined by immunohistochemistry. THP1 monocyte interaction with HAMEC was measured by cell adhesion and migration across transwells. Insulin transcytosis was measured by total internal reflection fluorescence microscopy. Palmitate, but not palmitoleate, elevated the expression of IL-6, IL-8, TLR2 (Toll-like receptor 2), and intercellular adhesion molecule 1 (ICAM-1). HAMEC had markedly low fatty acid uptake and oxidation, and CD36 inhibition did not reverse the palmitate-induced expression of adhesion molecules, suggesting that inflammation did not arise from palmitate uptake/metabolism. Instead, inhibition of TLR4 to NF-κB signaling blunted palmitate-induced ICAM-1 expression. Importantly, palmitate-induced surface expression of ICAM-1 promoted monocyte binding and transmigration. Conversely, palmitate reduced insulin transcytosis, an effect reversed by TLR4 inhibition. In summary, palmitate activates inflammatory pathways in primary microvascular endothelial cells, impairing insulin transport and increasing monocyte transmigration. This behavior may contribute in vivo to reduced tissue insulin action and enhanced tissue

  16. Successful isolation of infectious and high titer human monocyte-derived HIV-1 from two subjects with discontinued therapy.

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    Tong Wang

    Full Text Available BACKGROUND: HIV-1 DNA in blood monocytes is considered a viral source of various HIV-1 infected tissue macrophages, which is also known as "Trojan horse" hypothesis. However, whether these DNA can produce virions has been an open question for years, due to the inability of isolating high titer and infectious HIV-1 directly from monocytes. RESULTS: In this study, we demonstrated successful isolation of two strains of M-HIV-1 (1690 M and 1175 M from two out of four study subjects, together with their in vivo controls, HIV-1 isolated from CD4+ T-cells (T-HIV-1, 1690 T and 1175 T. All M- and T- HIV-1 isolates were detected CCR5-tropic. Both M- HIV-1 exhibited higher levels of replication in monocyte-derived macrophages (MDM than the two T- HIV-1. Consistent with our previous reports on the subject 1175 with late infection, compartmentalized env C2-V3-C3 sequences were identified between 1175 M and 1175 T. In contrast, 1690 M and 1690 T, which were isolated from subject 1690 with relatively earlier infection, showed homogenous env C2-V3-C3 sequences. However, multiple reverse transcriptase (RT inhibitor resistance-associated variations were detected in the Gag-Pol region of 1690 M, but not of 1690 T. By further measuring HIV DNA intracellular copy numbers post-MDM infection, 1690 M was found to have significantly higher DNA synthesis efficiency than 1690 T in macrophages, indicating a higher RT activity, which was confirmed by AZT inhibitory assays. CONCLUSIONS: These results suggested that the M- and T- HIV-1 are compartmentalized in the two study subjects, respectively. Therefore, we demonstrated that under in vitro conditions, HIV-1 infected human monocytes can productively release live viruses while differentiating into macrophages.

  17. Complement component C3 fixes selectively to the major outer membrane protein (MOMP) of Legionella pneumophila and mediates phagocytosis of liposome-MOMP complexes by human monocytes

    OpenAIRE

    1990-01-01

    Legionella pneumophila is a facultative intracellular bacterial pathogen that parasitizes human monocytes and alveolar macrophages. Previous studies from this laboratory have shown that monocyte complement receptors CR1 and CR3 and complement component C3 in serum mediate L. pneumophila phagocytosis. In this study, we have explored C3 fixation to L. pneumophila. We developed a whole-cell enzyme-linked immunosorbent assay (ELISA) to measure C3 fixation to the bacterial surface. By this assay, ...

  18. Genome-wide promoter analysis of histone modifications in human monocyte-derived antigen presenting cells

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    Peterson Hedi

    2010-11-01

    Full Text Available Abstract Background Monocyte-derived macrophages and dendritic cells (DCs are important in inflammatory processes and are often used for immunotherapeutic approaches. Blood monocytes can be differentiated into macrophages and DCs, which is accompanied with transcriptional changes in many genes, including chemokines and cell surface markers. Results To study the chromatin modifications associated with this differentiation, we performed a genome wide analysis of histone H3 trimethylation on lysine 4 (H3K4me3 and 27 (H3K27me3 as well as acetylation of H3 lysines (AcH3 in promoter regions. We report that both H3K4me3 and AcH3 marks significantly correlate with transcriptionally active genes whereas H3K27me3 mark is associated with inactive gene promoters. During differentiation, the H3K4me3 levels decreased on monocyte-specific CD14, CCR2 and CX3CR1 but increased on DC-specific TM7SF4/DC-STAMP, TREM2 and CD209/DC-SIGN genes. Genes associated with phagocytosis and antigen presentation were marked by H3K4me3 modifications. We also report that H3K4me3 levels on clustered chemokine and surface marker genes often correlate with transcriptional activity. Conclusion Our results provide a basis for further functional correlations between gene expression and histone modifications in monocyte-derived macrophages and DCs.

  19. Induction of oxygen free radical generation in human monocytes by lipoprotein(a)

    DEFF Research Database (Denmark)

    Riis Hansen, P; Kharazmi, A; Jauhiainen, M;

    1994-01-01

    The mechanism behind the association of elevated plasma lipoprotein(a) [Lp(a)] levels with atherosclerotic disease is unknown. In the present study, Lp(a) induced generation of oxygen free radicals by monocytes from selected healthy individuals in vitro. This observation may provide a link between...

  20. HIGH GLUCOSE INDUCES TOLL-LIKE RECEPTOR EXPRESSION IN HUMAN MONOCYTES: MECHANISM OF ACTIVATION

    Science.gov (United States)

    Objective: Hyperglycemia induced inflammation is central in diabetes complications and monocytes are important in orchestrating these effects. Toll-like receptors (TLRs) play a key role in innate immune responses as well as inflammation. However, there is a paucity of data examining the expression a...

  1. Fibronectin Binding Is Required for Acquisition of Mesenchymal/Endothelial Differentiation Potential in Human Circulating Monocytes

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    Noriyuki Seta

    2012-01-01

    Full Text Available We previously reported monocyte-derived multipotential cells (MOMCs, which include progenitors capable of differentiating into a variety of mesenchymal cells and endothelial cells. In vitro generation of MOMCs from circulating CD14+ monocytes requires their binding to extracellular matrix (ECM protein and exposure to soluble factor(s derived from circulating CD14- cells. Here, we investigated the molecular factors involved in MOMC generation by examining the binding of monocytes to ECM proteins. We found that MOMCs were obtained on the fibronectin, but not on type I collagen, laminin, or poly-L-lysine. MOMC generation was followed by changes in the expression profiles of transcription factors and was completely inhibited by either anti-α5 integrin antibody or a synthetic peptide that competed with the RGD domain for the β1-integrin binding site. These results indicate that acquisition of the multidifferentiation potential by circulating monocytes depends on their binding to the RGD domain of fibronectin via cell-surface α5β1 integrin.

  2. DNA Damage, Cell Cycle Arrest, and Apoptosis Induction Caused by Lead in Human Leukemia Cells

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    Clement G. Yedjou

    2015-12-01

    Full Text Available In recent years, the industrial use of lead has been significantly reduced from paints and ceramic products, caulking, and pipe solder. Despite this progress, lead exposure continues to be a significant public health concern. The main goal of this research was to determine the in vitro mechanisms of lead nitrate [Pb(NO32] to induce DNA damage, apoptosis, and cell cycle arrest in human leukemia (HL-60 cells. To reach our goal, HL-60 cells were treated with different concentrations of Pb(NO32 for 24 h. Live cells and necrotic death cells were measured by the propidium idiode (PI assay using the cellometer vision. Cell apoptosis was measured by the flow cytometry and DNA laddering. Cell cycle analysis was evaluated by the flow cytometry. The result of the PI demonstrated a significant (p < 0.05 increase of necrotic cell death in Pb(NO32-treated cells, indicative of membrane rupture by Pb(NO32 compared to the control. Data generated from the comet assay indicated a concentration-dependent increase in DNA damage, showing a significant increase (p < 0.05 in comet tail-length and percentages of DNA cleavage. Data generated from the flow cytometry assessment indicated that Pb(NO32 exposure significantly (p < 0.05 increased the proportion of caspase-3 positive cells (apoptotic cells compared to the control. The flow cytometry assessment also indicated Pb(NO32 exposure caused cell cycle arrest at the G0/G1 checkpoint. The result of DNA laddering assay showed presence of DNA smear in the agarose gel with little presence of DNA fragments in the treated cells compared to the control. In summary, Pb(NO32 inhibits HL-60 cells proliferation by not only inducing DNA damage and cell cycle arrest at the G0/G1 checkpoint but also triggering the apoptosis through caspase-3 activation and nucleosomal DNA fragmentation accompanied by secondary necrosis. We believe that our study provides a new insight into the mechanisms of Pb(NO32 exposure and its associated adverse

  3. Differences in PGE2 production between primary human monocytes and differentiated macrophages: role of IL-1β and TRIF/IRF3.

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    Yukinori Endo

    Full Text Available Prostaglandin E2 (PGE2 is induced in vivo by bacterial products including TLR agonists. To determine whether PGE2 is induced directly or via IL-1β, human monocytes and macrophages were cultured with LPS or with Pam3CSK4 in presence of caspase-1 inhibitor, ZVAD, or IL-1R antagonist, Kineret. TLR agonists induced PGE2 in macrophages exclusively via IL-1β-independent mechanisms. In contrast, ZVAD and Kineret reduced PGE2 production in LPS-treated (but not in Pam3CSK4-treated monocytes, by 30-60%. Recombinant human IL-1β augmented COX-2 and mPGES-1 mRNA and PGE2 production in LPS-pretreated monocytes but not in un-primed or Pam3CSK4-primed monocytes. This difference was explained by the finding that LPS but not Pam3CSK4 induced phosphorylation of IRF3 in monocytes suggesting activation of the TRIF signaling pathway. Knocking down TRIF, TRAM, or IRF3 genes by siRNA inhibited IL-1β-induced COX-2 and mPGES-1 mRNA. Blocking of TLR4 endocytosis during LPS priming prevented the increase in PGE2 production by exogenous IL-1β. Our data showed that TLR2 agonists induce PGE2 in monocytes independently from IL-1β. In the case of TLR4, IL-1β augments PGE2 production in LPS-primed monocytes (but not in macrophages through a mechanism that requires TLR4 internalization and activation of the TRIF/IRF3 pathway. These findings suggest a key role for blood monocytes in the rapid onset of fever in animals and humans exposed to bacterial products and some novel adjuvants.

  4. Lipid-associated membrane proteins of Mycoplasma penetrans induce production of proinflammatory cytokines in human monocytic cells

    Institute of Scientific and Technical Information of China (English)

    YAN HUA ZENG; YI MOU WU; MIN JUN YU; LI ZHI TAN; ZHONG LIANG DENG; XIAO XING YOU

    2006-01-01

    The aim of this study is to explore potential pathogenicity of Mycoplasma penetrans, and to investigate whether M. penetrans lipid-associated membrane proteins (LAMPs) could induce human monocytic cell line (THP-1) to produce some proinflammatory cytokines in vitro, including interleukin-1β (IL1β), tumor necrosis factor alpha (TNF-α), and IL-8. THP-1 was stimulated with different concentrations of M. penetrans LAMPs and at different time to analyze the production of human IL-1β, TNF-α and IL-8.The protein levels of human IL-1β, TNF-α and IL-8 were measured by enzyme-linked immunoadsorbent assay (ELISA) and the mRNA levels of these proinflammatory cytokines were detected by reverse transcriptase-PCR (RT-PCR). It was demonstrated in the present study that the production of IL-1β, TNF-αand IL-8 increased in dose- and time-dependent manner after stimulation with M. penetrans LAMPs in THP-1 cells. M.penetrans LAMPs also induced the expression of IL-1β, TNF-α and IL-8 mRNA. The production of IL-1β, TNF-α and IL-8 and the expression of mRNA were down-regulated by pyrrolidine dithiocarbamate (PDTC). This study demonstrated that M. penetrans LAMPs can induce the production of proinflammatory cytokines in human monocytic cells in vitro, thus suggesting that it may be an important etiological factor.

  5. Unlike PPARγ, PPARα or PPARβ/δ activation does not promote human monocyte differentiation toward alternative macrophages

    International Nuclear Information System (INIS)

    Macrophages adapt their response to micro-environmental signals. While Th1 cytokines promote pro-inflammatory M1 macrophages, Th2 cytokines promote an 'alternative' anti-inflammatory M2 macrophage phenotype. Peroxisome proliferator-activated receptors (PPARs) are ligand-activated transcription factors expressed in macrophages where they control the inflammatory response. It has been shown that PPARγ promotes the differentiation of monocytes into anti-inflammatory M2 macrophages in humans and mice, while a role for PPARβ/δ in this process has been reported only in mice and no data are available for PPARα. Here, we show that in contrast to PPARγ, expression of PPARα and PPARβ/δ overall does not correlate with the expression of M2 markers in human atherosclerotic lesions, whereas a positive correlation with genes of lipid metabolism exists. Moreover, unlike PPARγ, PPARα or PPARβ/δ activation does not influence human monocyte differentiation into M2 macrophages in vitro. Thus, PPARα and PPARβ/δ do not appear to modulate the alternative differentiation of human macrophages.

  6. Berberine induces cell cycle arrest and apoptosis in human gastric carcinoma SNU-5 cell line

    Institute of Scientific and Technical Information of China (English)

    Jing-Pin Lin; Jai-Sing Yang; Jau-Hong Lee; Wen-Tsong Hsieh; Jing-Gung Chung

    2006-01-01

    AIM: To investigate the relationship between the inhibited growth (cytotoxic activity) of berberine and apoptotic pathway with its molecular mechanism of action.METHODS: The in vitro cytotoxic techniques were complemented by cell cycle analysis and determination of sub-G1 for apoptosis in human gastric carcinoma SNU-5 cells. Percentage of viable cells, cell cycle, and sub-G1 group (apoptosis) were examined and determined by the flow cytometric methods. The associated proteins for cell cycle arrest and apoptosis were examined by Western blotting.RESULTS: For SNU-5 cell line, the IC (50) was found to be 48 μmol/L of berberine. In SNU-5 cells treated with 25-200 μmol/L berberine, G2/M cell cycle arrest was observed which was associated with a marked increment of the expression of p53, Wee1 and CDk1 proteins and decreased cyclin B. A concentration-dependent decrease of cells in G0/G1 phase and an increase in G2/M phase were detected. In addition, apoptosis detected as sub-G0 cell population in cell cycle measurement was proved in 25-200 μmol/L berberine-treated cells by monitoring the apoptotic pathway. Apoptosis was identified by sub-G0 cell population, and upregulation of Bax, downregulation of Bcl-2, release of Ca2+, decreased the mitochondrial membrane potential and then led to the release of mitochondrial cytochrome C into the cytoplasm and caused the activation of caspase-3, and finally led to the occurrence of apoptosis.CONCLUSION: Berberine induces p53 expression and leads to the decrease of the mitochondrial membrane potential, Cytochrome C release and activation of caspase-3 for the induction of apoptosis.

  7. Cordycepin Induces S Phase Arrest and Apoptosis in Human Gallbladder Cancer Cells

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    Xu-An Wang

    2014-07-01

    Full Text Available Gallbladder cancer is the most common malignant tumor of the biliary tract, and this condition has a rather dismal prognosis, with an extremely low five-year survival rate. To improve the outcome of unresectable and recurrent gallbladder cancer, it is necessary to develop new effective treatments and drugs. The purpose of the present study was to evaluate the effects of cordycepin on human gallbladder cells and uncover the molecular mechanisms responsible for these effects. The Cell Counting Kit-8 (CCK-8 and colony formation assays revealed that cordycepin affected the viability and proliferation of human gallbladder cancer cells in a dose- and time-dependent manner. Flow cytometric analysis showed that cordycepin induced S phase arrest in human gallbladder cancer cell lines(NOZ and GBC-SD cells. Cordycepin-induced apoptosis was observed using an Annexin V/propidium iodide (PI double-staining assay, and the mitochondrial membrane potential (ΔΨm decreased in a dose-dependent manner. Additionally, western blot analysis revealed the upregulation of cleaved-caspase-3, cleaved-caspase-9, cleaved-PARP and Bax and the downregulation of Bcl-2, cyclin A and Cdk-2 in cordycepin-treated cells. Moreover, cordycepin inhibited tumor growth in nude mice bearing NOZ tumors. Our results indicate that this drug may represent an effective treatment for gallbladder carcinoma.

  8. The Pelargonium sidoides Extract EPs 7630 Drives the Innate Immune Defense by Activating Selected MAP Kinase Pathways in Human Monocytes.

    Science.gov (United States)

    Witte, Katrin; Koch, Egon; Volk, Hans-Dieter; Wolk, Kerstin; Sabat, Robert

    2015-01-01

    Pelargonium sidoides is a medical herb and respective extracts are used very frequently for the treatment of respiratory tract infections. However, the effects of Pelargonium sidoides and a special extract prepared from its roots (EPs 7630) on human immune cells are not fully understood. Here we demonstrate that EPs 7630 induced a rapid and dose-dependent production of TNF-α, IL-6, and IL-10 by human blood immune cells. This EPs 7630-induced cytokine profile was more pro-inflammatory in comparison with the profile induced by viral or bacterial infection-mimicking agents. The search for EPs 7630 target cells revealed that T-cells did not respond to EPs 7630 stimulation by production of TNF-α, IL-6, or IL-10. Furthermore, pretreatment of T-cells with EPs 7630 did not modulate their TNF-α, IL-6, and IL-10 secretion during subsequent activation. In contrast to lymphocytes, monocytes showed clear intracellular TNF-α staining after EPs 7630 treatment. Accordingly, EPs 7630 predominantly provoked activation of MAP kinases and inhibition of p38 strongly reduced the monocyte TNF-α production. The pretreatment of blood immune cells with EPs 7630 lowered their secretion of TNF-α and IL-10 and caused an IL-6 dominant response during second stimulation with viral or bacterial infection-mimicking agents. In summary, we demonstrate that EPs 7630 activates human monocytes, induces MAP kinase-dependent pro-inflammatory cytokines in these cells, and specifically modulates their production capacity of mediators known to lead to an increase of acute phase protein production in the liver, neutrophil generation in the bone marrow, and the generation of adaptive Th17 and Th22 cells. PMID:26406906

  9. Effect of Bacterial Lipopolysaccharide Contamination on Gutta Percha- versus Resilon-Induced Human Monocyte Cell Line Toxicity.

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    Jamshid Hadjati

    2015-04-01

    Full Text Available Cytotoxic effects of obturation materials were tested in presence and absence of endotoxin on human monocytes in vitro.Human monocytes from THP-1 cell line were cultured. Three millimeters from the tip of each Resilon and gutta percha points were cut and directly placed at the bottom of the culture wells. Cultured cells were exposed to gutta percha (groups G1 and G2 and Resilon (R1 and R2. Ten μg/ml bacterial lipopolysaccharide (LPS was added to the culture wells in groups G1 and R1. Positive control included the bacterial LPS without the root canal filling material and the negative control contained the cells in culture medium only. Viability of cells was tested in all groups after 24, 48, and 72 hours using the methylthiazolyldiphenyl-tetrazolium bromide (MTT assay for at least 3 times to obtain reproducible results. Optical density values were read and the data were analyzed using three-way ANOVA and post hoc statistical test.The results showed that cells in G2 had the lowest rate of viability at 24 hours, but the lowest rate of viable cells was recorded in G1 at 48 and 72 hours. The effect of LPS treatment was not statistically significant. Resilon groups showed cell viability values higher than those of gutta percha groups, although statistically non-significant (P=0.105. Cell viability values were lower in gutta percha than Resilon groups when LPS-treated and LPS-untreated groups were compared independently at each time point.It could be concluded that none of the tested root canal filling materials had toxic effects on cultured human monocyte cells whether in presence or absence of LPS contamination.

  10. Generation of Monocyte-Derived Insulin-Producing Cells from Non-Human Primates According to an Optimized Protocol for the Generation ofPCMO-Derived Insulin-Producing Cells

    OpenAIRE

    Walter, Jessica; Harder, Ole; Faendrich, Fred; Schulze, Maren

    2014-01-01

    Ob­jec­ti­ve: The vision of potential autologous cell therapy for the cure of diabetes encourages ongoing research. According to a previously published protocol for the generation of insulin-producing cells from human monocytes, we analyzed whether the addition of growth factors could increase insulin production. This protocol was then transferred to a non-human primate model by using either blood- or spleen-derived monocytes. Methods: Human monocytes were treated to dedifferentiate into prog...

  11. Human circulating monocytes internalize 125I-insulin in a similar fashion to rat hepatocytes: relevance to receptor regulation in target and nontarget tissues

    International Nuclear Information System (INIS)

    Circulating monocytes bind 125I-insulin in a specific fashion and have been used to analyze the ambient receptor status in humans. When freshly isolated circulating monocytes are incubated with 125I-insulin and examined by electron microscopic autoradiography, approximately 18% of the labeled material is internalized after 15 minutes at 37 degrees C. By 2 hours at 37 degrees C, approximately one half of the 125I-insulin is internalized. Internalization occurs also at 15 degrees C but at a slower rate. Furthermore, the monocytes bind and internalize 125I-insulin in a manner that mirrors that of major target tissues, such as rat hepatocytes. These data suggest that the insulin receptor of the circulating monocyte might be regulated by adsorptive endocytosis in a manner analogous to that of target tissue, such as the liver

  12. Induction of DNA repair synthesis in human monocytes/B-lymphocytes compared with T-lymphocytes after exposure to N-acetoxy-N-acetylaminofluorene and dimethylsulfate in vitro

    DEFF Research Database (Denmark)

    Knudsen, Lisbeth E.; Ryder, L P; Wassermann, K

    1992-01-01

    We have explored the induction of DNA repair synthesis in monocyte/B- and T-lymphocyte enriched cell fractions from 12 different human mononuclear blood cell populations. Unscheduled DNA synthesis was measured in monocyte/B- and T-cells after exposure to the DNA-damaging agents dimethylsulfate (DMS......) and N-acetoxy-N-acetylaminofluorene in vitro. Also, the binding of DMS to DNA was measured. An increased DNA repair synthesis was measured in monocyte/B-lymphocytes after induction of the two different types of DNA lesions, whereas no induction of unscheduled DNA synthesis was observed in T-lymphocytes....... A significantly higher DMS-DNA binding was also observed in monocyte/B-lymphocytes when compared with T-lymphocytes. Specific characterization of mononuclear blood cell populations used in biomonitoring of DNA adducts and repair is recommended....

  13. New method to differentiate human peripheral blood monocytes into insulin producing cells: Human hematosphere culture.

    Science.gov (United States)

    Hur, Jin; Yang, Ji Min; Choi, Jae-Il; Yun, Ji-Yeon; Jang, Jae Hee; Kim, Joonoh; Kim, Ju-Young; Oh, Il-Young; Yoon, Chang-Hwan; Cho, Hyun-Jai; Park, Young-Bae; Kim, Hyo-Soo

    2012-02-24

    Strategy to differentiate stem cells into insulin producing cells (IPCs) in vitro has been a promising one to get cell source of β-cell replacement therapy for diabetes. It has been suggested that islets and neurons share features and nestin-positive cells could differentiate into IPCs. We have recently developed a three-dimensional culture system using human peripheral blood cells named as blood-born hematosphere (BBHS). Here we showed that most of BBHS were composed of nestin-positive cells. Under the four-stage differentiation protocol for IPCs, we plated nestin-positive BBHS onto fibronectin-coated dish. These cells form islet-like clusters and most of them expressed insulin. Pancreatic specific genes were turned on, such as transcription factors (Pdx-1, Ngn3 and Nkx6.1), genes related to endocrine function (Glut-2 and PC2) or β cell function (Kir6.2, SUR1). Furthermore islet differentiation was confirmed by dithizone (DTZ) staining to detect zinc ion which binds insulin protein within the cells. Finally, IPCs derived from BBHS showed capability to secrete insulin in response to glucose stimulation. Taken together, our novel protocol successfully induced islet-like human insulin producing cells out of BBHS. This strategy of ex vivo expansion of IPCs using BBHS provides an autologous therapeutic cell source for the treatment of diabetes. PMID:22310720

  14. Immature dendritic cells generated from cryopreserved human monocytes show impaired ability to respond to LPS and to induce allogeneic lymphocyte proliferation.

    Directory of Open Access Journals (Sweden)

    Guilherme Ferreira Silveira

    Full Text Available Dendritic cells play a key role in the immune system, in the sensing of foreign antigens and triggering of an adaptive immune response. Cryopreservation of human monocytes was investigated to understand its effect on differentiation into immature monocyte-derived dendritic cells (imdDCs, the response to inflammatory stimuli and the ability to induce allogeneic lymphocyte proliferation. Cryopreserved (crp-monocytes were able to differentiate into imdDCs, albeit to a lesser extent than freshly (frh-obtained monocytes. Furthermore, crp-imdDCs had lower rates of maturation and cytokine/chemokine secretion in response to LPS than frh-imdDCs. Lower expression of Toll-like receptor 4 (at 24 and 48 h and higher susceptibility to apoptosis in crp-imdDCs than in fresh cells would account for the impaired maturation and cytokine/chemokine secretion observed. A mixed leukocyte reaction showed that lymphocyte proliferation was lower with crp-imdDCs than with frh-imdDCs. These findings suggested that the source of monocytes used to generate human imdDCs could influence the accuracy of results observed in studies of the immune response to pathogens, lymphocyte activation, vaccination and antigen sensing. It is not always possible to work with freshly isolated monocytes but the possible effects of freezing/thawing on the biology and responsiveness of imdDCs should be taken into account.

  15. Palmitate and insulin synergistically induce IL-6 expression in human monocytes

    OpenAIRE

    Lumpkin Charles K; Thrailkill Kathryn M; Ou Yang; Cockrell Gael E; Bunn Robert C; Fowlkes John L

    2010-01-01

    Abstract Background Insulin resistance is associated with a proinflammatory state that promotes the development of complications such as type 2 diabetes mellitus (T2DM) and atherosclerosis. The metabolic stimuli that initiate and propagate proinflammatory cytokine production and the cellular origin of proinflammatory cytokines in insulin resistance have not been fully elucidated. Circulating proinflammatory monocytes show signs of enhanced inflammation in obese, insulin resistant subjects and...

  16. Gene expression pattern in human monocytes as a surrogate marker for systemic inflammatory response syndrome (SIRS).

    OpenAIRE

    Wiegand, G.; Selleng, K.; Gründling, M.; Jack, R S

    1999-01-01

    BACKGROUND: Systemic inflammatory response syndrome (SIRS) is a mild inflammatory episode which, in a minority of patients, may deteriorate into septic shock. In the mouse, injection of bacteria or bacterial endotoxin induces systemic inflammation through the activation of blood monocytes, which leads to lethal shock. A number of intervention strategies have been shown to prevent progression to shock in mouse model systems. However, recent clinical trials of a number of these therapeutic stra...

  17. Impact of Human Granulocyte and Monocyte Isolation Procedures on Functional Studies

    OpenAIRE

    Zhou, Lu; Somasundaram, Rajesh; Nederhof, Rosa F.; Dijkstra, Gerard; Faber, Klaas Nico; Peppelenbosch, Maikel P.; Fuhler, Gwenny M.

    2012-01-01

    One of the first lines of defense against infection is the activation of the innate immune system. It is becoming clear that autoimmune diseases, such as rheumatoid arthritis and Crohn's disease, may be caused by disturbed innate immunity, and relating granulocyte and monocyte functions to the patient genotype has become an important part of contemporary research. Although it is essential to move this field forward, a systematic study comparing the efficacy and suitability for functional stud...

  18. Piceatannol and Its Metabolite, Isorhapontigenin, Induce SIRT1 Expression in THP-1 Human Monocytic Cell Line

    OpenAIRE

    Shinpei Kawakami; Yosuke Kinoshita; Hiroko Maruki-Uchida; Koji Yanae; Masahiko Sai; Tatsuhiko Ito

    2014-01-01

    Piceatannol is a phytochemical that is present in large amounts in passion fruit (Passiflora edulis) seeds, and is an analog of resveratrol. Recently, the absorption and metabolism of piceatannol were investigated in rats, and isorhapontigenin, O-methyl piceatannol, was detected as a piceatannol metabolite in rat plasma. To elucidate the function of piceatannol and its metabolites, we investigated the expression of sirtuin 1 (SIRT1) in THP-1 monocytic cells after treatment with piceatannol an...

  19. Human Adipose Derived Stem Cells Induced Cell Apoptosis and S Phase Arrest in Bladder Tumor

    Directory of Open Access Journals (Sweden)

    Xi Yu

    2015-01-01

    Full Text Available The aim of this study was to determine the effect of human adipose derived stem cells (ADSCs on the viability and apoptosis of human bladder cancer cells. EJ and T24 cells were cocultured with ADSCs or cultured with conditioned medium of ADSCs (ADSC-CM, respectively. The cell counting and colony formation assay showed ADSCs inhibited the proliferation of EJ and T24 cells. Cell viability assessment revealed that the secretions of ADSCs, in the form of conditioned medium, were able to decrease cancer cell viability. Wound-healing assay suggested ADSC-CM suppressed migration of T24 and EJ cells. Moreover, the results of the flow cytometry indicated that ADSC-CM was capable of inducing apoptosis of T24 cells and inducing S phase cell cycle arrest. Western blot revealed ADSC-CM increased the expression of cleaved caspase-3 and cleaved PARP, indicating that ADSC-CM induced apoptosis in a caspase-dependent way. PTEN/PI3K/Akt pathway and Bcl-2 family proteins were involved in the mechanism of this reaction. Our study indicated that ADSCs may provide a promising and practicable manner for bladder tumor therapy.

  20. From human monocytes to genome-wide binding sites--a protocol for small amounts of blood: monocyte isolation/ChIP-protocol/library amplification/genome wide computational data analysis.

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    Sebastian Weiterer

    Full Text Available Chromatin immunoprecipitation in combination with a genome-wide analysis via high-throughput sequencing is the state of the art method to gain genome-wide representation of histone modification or transcription factor binding profiles. However, chromatin immunoprecipitation analysis in the context of human experimental samples is limited, especially in the case of blood cells. The typically extremely low yields of precipitated DNA are usually not compatible with library amplification for next generation sequencing. We developed a highly reproducible protocol to present a guideline from the first step of isolating monocytes from a blood sample to analyse the distribution of histone modifications in a genome-wide manner.The protocol describes the whole work flow from isolating monocytes from human blood samples followed by a high-sensitivity and small-scale chromatin immunoprecipitation assay with guidance for generating libraries compatible with next generation sequencing from small amounts of immunoprecipitated DNA.

  1. RNA of Enterococcus faecalis Strain EC-12 Is a Major Component Inducing Interleukin-12 Production from Human Monocytic Cells.

    Directory of Open Access Journals (Sweden)

    Ryoichiro Nishibayashi

    Full Text Available Interleukin-12 (IL-12 is an important cytokine for the immunomodulatory effects of lactic acid bacteria (LAB. Using murine immune cells, we previously reported that the RNA of Enterococcus faecalis EC-12, a LAB strain exerting probiotic-like beneficial effects, is the major IL-12-inducing immunogenic component. However, it was recently revealed that bacterial RNA can be a ligand for Toll-like receptor (TLR 13, which is only expressed in mice. Because TLR13 is not expressed in humans, the immuno-stimulatory and -modulatory effects of LAB RNA in human cells should be augmented excluding TLR13 contribution. In experiment 1 of this study, the role of LAB RNA in IL-12 induction in human immune cells was studied using three LAB strains, E.faecalis EC-12, Lactobacillus gasseri JCM5344, and Bifidobacterium breve JCM1192. RNase A treatment of heat-killed LAB significantly decreased the IL-12 production of human peripheral blood mononuclear cells on stimulation, while RNase III treatment revealed virtually no effects. Further, IL-12 production against heat-killed E. faecalis EC-12 was abolished by depleting monocytes. These results demonstrated that single stranded RNA (ssRNA of LAB is a strong inducer of IL-12 production from human monocytes. In experiment 2, major receptor for ssRNA of E. faecalis EC-12 was identified using THP-1 cells, a human monocytic cell line. The type of RNA molecules of E. faecalis EC-12 responsible for IL-12 induction was also identified. IL-12 production induced by the total RNA of E. faecalis EC-12 was significantly reduced by the treatment of siRNA for TLR8 but not for TLR7. Furthermore, both 23S and 16S rRNA, but not mRNA, of E. faecalis EC-12 markedly induced IL-12 production from THP-1 cells. These results suggested that the recognition of ssRNA of E. faecalis EC-12 was mediated by TLR8 and that rRNA was the RNA molecule that exhibited IL-12-inducing ability in human cells.

  2. HSV-1-induced chemokine expression via IFI16-dependent and IFI16-independent pathways in human monocyte-derived macrophages

    DEFF Research Database (Denmark)

    Søby, Stine; Laursen, Rune R; Østergaard, Lars Jørgen; Melchjorsen, Jesper

    2012-01-01

    -mediated chemokine responses are not dependent on TLR2 and TLR9 in human macrophages. Here, we investigated the role of the recently identified innate IFN-inducible DNA receptor IFI16 during HSV-1 infection in human macrophages. METHODS: Peripheral blood mononuclear cells were purified from buffy coats and...... monocytes were differentiated to macrophages. Macrophages infected with HSV-1 were analyzed using siRNA-mediated knock-down of IFI16 by real-time PCR, ELISA, and Western blotting. RESULTS: We determined that both CXCL10 and CCL3 are induced independent of HSV-1 replication. IFI16 mediates CCL3 m......RNA accumulation during early HSV-1 infection. In contrast, CXCL10 was induced independently of IFI16. CONCLUSIONS: Our data provide the first evidence of HSV-1-induced innate immune responses via IFI16 in human primary macrophages. In addition, the data suggest that at least one additional unidentified receptor...

  3. Ubiquinol decreases monocytic expression and DNA methylation of the pro-inflammatory chemokine ligand 2 gene in humans

    Directory of Open Access Journals (Sweden)

    Fischer Alexandra

    2012-10-01

    Full Text Available Abstract Background Coenzyme Q10 is an essential cofactor in the respiratory chain and serves in its reduced form, ubiquinol, as a potent antioxidant. Studies in vitro and in vivo provide evidence that ubiquinol reduces inflammatory processes via gene expression. Here we investigate the putative link between expression and DNA methylation of ubiquinol sensitive genes in monocytes obtained from human volunteers supplemented with 150 mg/ day ubiquinol for 14 days. Findings Ubiquinol decreases the expression of the pro-inflammatory chemokine (C-X-C motif ligand 2 gene (CXCL2 more than 10-fold. Bisulfite-/ MALDI-TOF-based analysis of regulatory regions of the CXCL2 gene identified six adjacent CpG islands which showed a 3.4-fold decrease of methylation status after ubiquinol supplementation. This effect seems to be rather gene specific, because ubiquinol reduced the expression of two other pro-inflammatory genes (PMAIP1, MMD without changing the methylation pattern of the respective gene. Conclusion In conclusion, ubiquinol decreases monocytic expression and DNA methylation of the pro-inflammatory CXCL2 gene in humans. Current Controlled Trials ISRCTN26780329.

  4. Comparison of two different strategies for human monocyte subsets gating within the large-scale prospective CARE FOR HOMe Study.

    Science.gov (United States)

    Zawada, Adam M; Fell, Lisa H; Untersteller, Kathrin; Seiler, Sarah; Rogacev, Kyrill S; Fliser, Danilo; Ziegler-Heitbrock, Loems; Heine, Gunnar H

    2015-08-01

    Monocytes are heterogeneous cells consisting of (at least) three subsets: classical, intermediate, and nonclassical monocytes. Correct enumeration of cell counts necessitates well-defined gating strategies, which are essentially based upon CD14 and CD16 expression. For the delineation of intermediate from nonclassical monocytes, a "rectangular gating (RG) strategy" and a "trapezoid gating (TG) strategy" have been proposed. We compared the two gating strategies in a well-defined clinical cohort of patients with chronic kidney disease (CKD). Within the ongoing CARE FOR HOMe study, monocyte subsets were reanalyzed in 416 CKD patients, who were followed 3.6 ± 1.6 years for the occurrence of a cardiovascular event. Gating was performed by either RG or TG. We analyzed the expression of surface markers, and compared the predictive role of cell counts of monocyte subsets, as defined by RG and TG, respectively. With both gating strategies, higher intermediate monocyte counts predicted the cardiovascular endpoint in Kaplan-Meier analyses (P strategy toward quartiles of intermediate monocytes counts with TG strategy. In expression analysis, those monocytes which are defined as intermediate monocytes by the RG strategy and as nonclassical monocytes by the TG strategy share characteristics of both subsets. In conclusion, intermediate monocytes were independent predictors of cardiovascular outcome irrespective of the applied gating strategy. Future studies should aim to identify markers that allow for an unequivocal definition of intermediate monocytes, which may further improve their power to predict cardiovascular events. PMID:26062127

  5. Effects of allitridi on cell cycle arrest of human gastric cancer cells

    Institute of Scientific and Technical Information of China (English)

    Min-Wen Ha; Rui Ma; Li-Ping Shun; Yue-Hua Gong; Yuan Yuan

    2005-01-01

    AIM: To determine the effect of allitridi on cell cycle of human gastric cancer (HGC) cell lines MGC803 and SGC7901 and its possible mechanism.METHODS: Trypan blue dye exclusion was used to evaluate the proliferation, inhibition of cells and damages of these cells were detected with electron microscope.Flow cytometry and cell mitotic index were used to analyze the change of cell cycle, immunohistochemistry, and RT-PCR was used to examine expression of the p21WAF1 gene.RESULTS: MGC803 cell growth was inhibited by allitridi with 24 h IC50 being 6.4 μg/mL. SGC7901 cell growth was also inhibited by allitridi with 24 h IC50 being 7.3 μg/mL.After being treated with allitridi at the concentration of 12 μg/mL for 24 h, cells were found to have direct cytotoxic effects, including broken cellular membrane, swollen and vesiculated mitochondria and rough endoplasmic reticula,and mass lipid droplet. When cells were treated with allitridi at the concentration of 3, 6, and 9 μg/mL for 24 h, the percentage of G0/G1 phase cells was decreased and that of G2/M phase cells was significantly increased (P = 0.002)compared with those in the group. When cells were treated with allitridi at the concentration of 6 μg/mL, cell mitotic index was much higher (P = 0.003) than that of control group, indicating that allitridi could cause gastric cancer cell arrest in M phase. Besides, the expression levels of p21WAF1 gene of MGC803 cells and p21WAF1 gene of SGC7901 cells were remarkably upregulated after treatment.CONCLUSION: Allitridi can cause gastric cancer cell arrest in M phase, and this may be one of the mechanisms for inhibiting cell proliferation. Effect of allitridi on cells in M phas e may be associated with the upregulation of p21WAF1 genes. This study provides experimental data for clinical use of allitridi in the treatment of gastric carcinoma.

  6. The rs1143679 (R77H) lupus associated variant of ITGAM (CD11b) impairs complement receptor 3 mediated functions in human monocytes

    OpenAIRE

    Rhodes, B.; Furnrohr, B. G.; Roberts, A.L.; Tzircotis, G.; Schett, G; Spector, T. D.; Vyse, T J

    2012-01-01

    Objectives The rs1143679 variant of ITGAM, encoding the R77H variant of CD11b (part of complement receptor 3; CR3), is among the strongest genetic susceptibility effects in human systemic lupus erythematosus (SLE). The authors aimed to demonstrate R77H function in ex-vivo human cells. Methods Monocytes/monocyte-derived macrophages from healthy volunteers homozygous for either wild type (WT) or 77H CD11b were studied. The genotype-specific expression of CD11b, and CD11b activation using confor...

  7. The immunosuppressive domain of the transmembrane envelope protein gp41 of HIV-1 binds to human monocytes and B cells.

    Science.gov (United States)

    Mühle, Michael; Kroniger, Tobias; Hoffmann, Kerstin; Denner, Joachim

    2016-06-01

    The induction of the acquired immunodeficiency syndrome by the human immunodeficiency virus-1 (HIV-1) is a complex process which is not yet understood in full detail. Still open is the question whether the highly conserved so-called immunosuppressive (Isu) domain in the transmembrane envelope (TM) protein gp41 of HIV-1 is actively participating in immunopathogenesis. Inactivated virus particles, recombinant gp41 and peptides corresponding to the Isu domain have been reported to inhibit lymphocyte proliferation, as well as to alter cytokine release and gene expression. Here we demonstrate, using fluorescence-activated cell sorting and competition experiments, that homopolymers of the Isu peptide of HIV-1 are binding specifically to human peripheral blood mononuclear cells, mainly to monocytes and B cells. These data suggest that a putative receptor might be involved in the immunomodulatory effects observed previously. PMID:26754765

  8. Taxifolin enhances andrographolide-induced mitotic arrest and apoptosis in human prostate cancer cells via spindle assembly checkpoint activation.

    Directory of Open Access Journals (Sweden)

    Zhong Rong Zhang

    Full Text Available Andrographolide (Andro suppresses proliferation and triggers apoptosis in many types of cancer cells. Taxifolin (Taxi has been proposed to prevent cancer development similar to other dietary flavonoids. In the present study, the cytotoxic and apoptotic effects of the addition of Andro alone and Andro and Taxi together on human prostate carcinoma DU145 cells were assessed. Andro inhibited prostate cancer cell proliferation by mitotic arrest and activation of the intrinsic apoptotic pathway. Although the effect of Taxi alone on DU145 cell proliferation was not significant, the combined use of Taxi with Andro significantly potentiated the anti-proliferative effect of increased mitotic arrest and apoptosis by enhancing the cleavage of poly(ADP-ribose polymerase, and caspases-7 and -9. Andro together with Taxi enhanced microtubule polymerization in vitro, and they induced the formation of twisted and elongated spindles in the cancer cells, thus leading to mitotic arrest. In addition, we showed that depletion of MAD2, a component in the spindle assembly checkpoint (SAC, alleviated the mitotic block induced by the two compounds, suggesting that they trigger mitotic arrest by SAC activation. This study suggests that the anti-cancer activity of Andro can be significantly enhanced in combination with Taxi by disrupting microtubule dynamics and activating the SAC.

  9. Human monocyte-derived cells with individual hepatocyte characteristics: a novel tool for personalized in vitro studies.

    Science.gov (United States)

    Benesic, Andreas; Rahm, Nora L; Ernst, Samuel; Gerbes, Alexander L

    2012-06-01

    Gender, ethnicity and individual differences in hepatic metabolism have major impact on individual drug response, adverse events and attrition rate during drug development. Therefore, there is an urgent need for reliable test systems based on human cells. Yet, the use of primary human hepatocytes (PHHs) is restricted by limited availability, invasive preparation and short-term stability in culture. All other cellular approaches proposed so far have major disadvantages. We investigated whether peripheral human monocytes after cultivation according to our novel protocol (monocyte-derived hepatocyte-like cells (MH cells)) can serve as an in vitro model for hepatocyte metabolism. Enzyme activities, synthesis parameters (coagulation factor VII and urea) and cytochrome (CY) P450 activities and induction were investigated. Furthermore, MH cells were compared with PHH from the same donor. Using our protocol, we could generate cells that exhibit hepatocyte-like properties: These cells show 71±9% of specific ALT activity, 41±3% of CYP3A4 activity and 65±13% of factor VII secretion when compared with PHHs. Consequently, CYP-mediated acetaminophen toxicity and drug interactions could be shown. Moreover, the investigated parameters were stable in culture over at least 4 weeks. Furthermore, MH cells retain gender-specific and donor-specific CYP activities and toxicity profiles, respectively. MH cells show quantitative and qualitative approximation to human hepatocytes concerning CYP-metabolism and toxicity. Our data support individual prediction of toxicity and CYP metabolism. MH cells are a novel tool to investigate long-term hepatic toxicity, metabolism and drug interactions. PMID:22469698

  10. Cell cycle arrest biomarkers in human lung cancer cells after treatment with selenium in culture.

    Science.gov (United States)

    Swede, Helen; Dong, Yan; Reid, Mary; Marshall, James; Ip, Clement

    2003-11-01

    In the planning of future intervention trials using chemopreventive agents against lung cancer, it is critical to evaluate the effect on biomarkers implicated specifically in lung carcinogenesis. With the use of the H520 and H522 human lung cancer cell lines, the present study showed that treatment with selenium (in the form of methylseleninic acid) inhibited cell growth, arrested cell cycle progression at G(1), and induced apoptosis as a late event. Because H520 cells were more sensitive to selenium than H522 cells (IC(50) of MSA was 2.5 or 10 micro M for H520 or H522 cells, respectively, at 24 h), a panel of nine cell cycle regulatory proteins known to be involved in G(1)-->S transition was assessed by Western analysis using whole cell lysate from H520 cells. These nine proteins (DP1, cdc25A, cyclin A, cyclin B(1), cyclin D(1), cdk1, cdk5, p21(WAF1), and GADD153) have been reported previously by our laboratory to be modulated by MSA in human breast and prostate cancer cells. Our data showed that only four (DP1, cdc25A, p21(WAF1), and GADD153) of nine biomarkers produced the expected changes after treatment of lung cancer cells with MSA. This finding raises the possibility that the molecular targets sensitive to selenium modulation may be tissue specific. Thus, the selection of selenium biomarkers for evaluation in an intervention trial must be based on empirical data derived from the cancer cell type of interest. PMID:14652289

  11. Mutation of Growth Arrest Specific 8 Reveals a Role in Motile Cilia Function and Human Disease.

    Science.gov (United States)

    Lewis, Wesley R; Malarkey, Erik B; Tritschler, Douglas; Bower, Raqual; Pasek, Raymond C; Porath, Jonathan D; Birket, Susan E; Saunier, Sophie; Antignac, Corinne; Knowles, Michael R; Leigh, Margaret W; Zariwala, Maimoona A; Challa, Anil K; Kesterson, Robert A; Rowe, Steven M; Drummond, Iain A; Parant, John M; Hildebrandt, Friedhelm; Porter, Mary E; Yoder, Bradley K; Berbari, Nicolas F

    2016-07-01

    Ciliopathies are genetic disorders arising from dysfunction of microtubule-based cellular appendages called cilia. Different cilia types possess distinct stereotypic microtubule doublet arrangements with non-motile or 'primary' cilia having a 9+0 and motile cilia have a 9+2 array of microtubule doublets. Primary cilia are critical sensory and signaling centers needed for normal mammalian development. Defects in their structure/function result in a spectrum of clinical and developmental pathologies including abnormal neural tube and limb patterning. Altered patterning phenotypes in the limb and neural tube are due to perturbations in the hedgehog (Hh) signaling pathway. Motile cilia are important in fluid movement and defects in motility result in chronic respiratory infections, altered left-right asymmetry, and infertility. These features are the hallmarks of Primary Ciliary Dyskinesia (PCD, OMIM 244400). While mutations in several genes are associated with PCD in patients and animal models, the genetic lesion in many cases is unknown. We assessed the in vivo functions of Growth Arrest Specific 8 (GAS8). GAS8 shares strong sequence similarity with the Chlamydomonas Nexin-Dynein Regulatory Complex (NDRC) protein 4 (DRC4) where it is needed for proper flagella motility. In mammalian cells, the GAS8 protein localizes not only to the microtubule axoneme of motile cilia, but also to the base of non-motile cilia. Gas8 was recently implicated in the Hh signaling pathway as a regulator of Smoothened trafficking into the cilium. Here, we generate the first mouse with a Gas8 mutation and show that it causes severe PCD phenotypes; however, there were no overt Hh pathway phenotypes. In addition, we identified two human patients with missense variants in Gas8. Rescue experiments in Chlamydomonas revealed a subtle defect in swim velocity compared to controls. Further experiments using CRISPR/Cas9 homology driven repair (HDR) to generate one of these human missense variants in

  12. Human circulating monocytes can express receptor activator of nuclear factor-kappaB ligand and differentiate into functional osteoclasts without exogenous stimulation.

    Science.gov (United States)

    Seta, Noriyuki; Okazaki, Yuka; Kuwana, Masataka

    2008-07-01

    Osteoclast formation from mononuclear precursors is believed to require accessory cells expressing receptor activator of nuclear factor-kappaB ligand (RANKL). We recently identified a human cell population originated from circulating CD14(+) monocytes, called monocyte-derived multipotential cells (MOMCs), which can differentiate into several distinct mesenchymal cells, neuron and endothelial cells. This study was undertaken to examine whether MOMCs can differentiate into functional osteoclasts. MOMCs prepared from peripheral blood of healthy volunteers cultured on fibronectin for 7 days at high density (8 x 10(5) cells cm(-2)), but not at regular density (2 x 10(4) cells cm(-2)), resulted in the appearance of tartrate-resistant acid phosphatase-positive giant multi-nucleated cells forming actin ring without exogenous osteoclastogenic factors. A subset of these cells showed bone resorption capacity on dentine slices and expression of genes for cathepsin K and calcitonin receptor, characteristic of functional osteoclasts. Such osteoclast differentiation was not observed in high-density culture of circulating monocytes, macrophages or dendritic cells, or the high-density culture of MOMCs on type I collagen. Among cells of the monocyte lineage, untreated MOMCs exclusively showed gene and protein expression of RANKL. When osteoprotegerin/IgG1 Fc chimera was added to high-density MOMC cultures, osteoclast formation was completely inhibited by neutralizing the endogenous RANKL. These results indicate that human MOMCs derived from circulating monocytes can express RANKL and differentiate into functional osteoclasts without RANKL-expressing accessory cells. PMID:18301383

  13. Outer membrane vesicles from Brucella abortus promote bacterial internalization by human monocytes and modulate their innate immune response.

    Directory of Open Access Journals (Sweden)

    Cora N Pollak

    Full Text Available Outer membrane vesicles (OMVs released by some gram-negative bacteria have been shown to exert immunomodulatory effects that favor the establishment of the infection. The aim of the present study was to assess the interaction of OMVs from Brucella abortus with human epithelial cells (HeLa and monocytes (THP-1, and the potential immunomodulatory effects they may exert. Using confocal microscopy and flow cytometry, FITC-labeled OMVs were shown to be internalized by both cell types. Internalization was shown to be partially mediated by clathrin-mediated endocytosis. Pretreatment of THP-1 cells with Brucella OMVs inhibited some cytokine responses (TNF-α and IL-8 to E. coli LPS, Pam3Cys or flagellin (TLR4, TLR2 and TLR5 agonists, respectively. Similarly, pretreatment with Brucella OMVs inhibited the cytokine response of THP-1 cells to B. abortus infection. Treatment of THP-1 cells with OMVs during IFN-γ stimulation reduced significantly the inducing effect of this cytokine on MHC-II expression. OMVs induced a dose-dependent increase of ICAM-1 expression on THP-1 cells and an increased adhesion of these cells to human endothelial cells. The addition of OMVs to THP-1 cultures before the incubation with live B. abortus resulted in increased numbers of adhered and internalized bacteria as compared to cells not treated with OMVs. Overall, these results suggest that OMVs from B. abortus exert cellular effects that promote the internalization of these bacteria by human monocytes, but also downregulate the innate immune response of these cells to Brucella infection. These effects may favor the persistence of Brucella within host cells.

  14. The gene region UL128-UL131A of human cytomegalovirus (HCMV) is essential for monocyte infection and block of migration - Characterisation of the infection of primary human monocytes

    OpenAIRE

    Straschewski, Sarah

    2011-01-01

    Monocytes are targets of HCMV infection and vehicles for viral dissemination in vivo. By using two TB40E-derived BAC clones expressing the full-length (BAC4) or a truncated form of pUL128 (BAC1), the role of UL128 for infection and motility of monocytes has been investigated. Like TB40E, BAC4 was able to infect and to express IE gene products in monocytes. BAC4-infected monocytes exhibited normal cytoskeleton architecture but a complete inability to migrate in response to chemokines as a cons...

  15. Vitamin A induces inhibitory histone methylation modifications and down-regulates trained immunity in human monocytes

    DEFF Research Database (Denmark)

    Arts, Rob J W; Blok, Bastiaan A; van Crevel, Reinout;

    2015-01-01

    assessed. ATRA inhibited cytokine responses upon restimulation of monocytes, and this effect was exerted through increased expression of SUV39H2, a histone methyltransferase that induces the inhibitory mark H3K9me3. H3K9me3 at promoter sites of several cytokines was up-regulated by ATRA, and inhibition of...... SUV39H2 restored cytokine production. In addition to H3K9me3, the stimulatory histone mark H3K4me3 was down-regulated by ATRA at several promoter locations of cytokine genes. Therefore, we can conclude that ATRA inhibits cytokine production in models of direct stimulation or BCG-induced trained...

  16. Glucocorticoid mediated regulation of inflammation in human monocytes is associated with depressive mood and obesity.

    Science.gov (United States)

    Cheng, Tiefu; Dimitrov, Stoyan; Pruitt, Christopher; Hong, Suzi

    2016-04-01

    Dysregulation of the hypothalamic-pituitary-adrenal (HPA) axis is observed in various conditions, including depression and obesity, which are also often related. Glucocorticoid (GC) resistance and desensitization of peripheral GC receptors (GRs) are often the case in HPA dysregulation seen in depression, and GC plays a critical role in regulation of inflammation. Given the growing evidence that inflammation is a central feature of some depression cases and obesity, we aimed to investigate the immune-regulatory role of GC-GR in relation to depressive mood and obesity in 35 healthy men and women. Depressive mood and level of obesity were assessed, using Beck Depression Inventory (BDI-Ia) and body mass index (BMI), respectively. We measured plasma cortisol levels via enzyme-linked immunosorbent assay and lipopolysaccharide-stimulated intracellular tumor necrosis factor (TNF) production by monocytes, using flow cytometry. Cortisol sensitivity was determined by the difference in monocytic TNF production between the conditions of 1 and 0μM cortisol incubation ("cortisol-mediated inflammation regulation, CoMIR"). GR vs. mineralocorticoid receptor (MR) antagonism for CoMIR was examined by using mifepristone and spironolactone. A series of multiple regression analyses were performed to investigate independent contribution of depressive mood vs. obesity after controlling for age, gender, systolic blood pressure (SBP), and plasma cortisol in predicting CoMIR. CoMIR was explained by somatic subcomponents of depressive mood (BDI-S: β=-0.499, p=0.001), or BMI (β=-0.466, pinitial indication that greater obesity and somatic depressive symptoms were associated with smaller efficacy of the blockers, which warrants further investigation. Our findings, although in a preclinical sample, signify the shared pathophysiology of immune dysregulation in depression and obesity and warrant further mechanistic investigation. PMID:26829709

  17. Phosphorylation of STAT-1 serine 727 is prolonged in HLA-B27-expressing human monocytic cells.

    Directory of Open Access Journals (Sweden)

    Marja Ruuska

    Full Text Available A tissue antigen, HLA-B27, is strongly associated with a group of rheumatic diseases called spondyloarthritides. Despite the intensive research, the exact role of HLA-B27 in the pathogenesis of these diseases is still unclear. Here we studied whether HLA-B27 modulates the phosphorylation of signal transducer and activator of transcription 1 (STAT-1 serine 727 residue and the localization of STAT-1 in Salmonella-infected human monocytic cells. In addition, we studied the role of signaling molecule double-stranded RNA activated protein kinase (PKR in these modulatory effects. U937 human monocytic cell transfectants stably expressing wild type HLA-B27 or mutated HLA-B27 heavy chains with amino acid substitutions in the B pocket were prepared. The PMA-differentiated cells were infected with S. enteritidis. Western blotting was used to detect the phosphorylation of STAT-1, and to visualize the localization of STAT-1 in the cells confocal microscopy was used. Specific inhibitors were employed to study the role of PKR in STAT-1 phosphorylation. We discovered that the phosphorylation of STAT-1 serine 727 is prolonged in cells expressing misfolding forms of HLA-B27 after S. enteritidis infection, whereas in mock cells and in cells expressing mutated, non-misfolding HLA-B27 the phosphorylation of serine 727 is transient. Interestingly, STAT-1 serine 727 phosphorylation is partly dependent on PKR. In addition, more STAT-1 is localized in the nucleus of HLA-B27-expressing cells, even before an external trigger, when compared to mock cells. In conclusion, our results show that the phosphorylation of STAT-1 serine 727 residue is prolonged in HLA-B27-expressing monocyte-macrophage U937 cells after bacterial infection. This is of interest since the phosphorylation of serine 727 on STAT-1 is suggested to contribute to macrophage activation and promote inflammatory responses. Therefore, our results provide a mechanism which explains how the expression of an HLA

  18. Phagocytic function of monocyte-derived macrophages is not affected by human immunodeficiency virus type 1 infection.

    Science.gov (United States)

    Nottet, H S; de Graaf, L; de Vos, N M; Bakker, L J; van Strijp, J A; Visser, M R; Verhoef, J

    1993-07-01

    The immunopathogenesis of human immunodeficiency virus (HIV) infection is characterized by the failure to control opportunistic infections. Here, the direct effect of HIV on macrophage phagocytic function was studied. HIV-1-infected monocyte-derived macrophages expressed as many Fc gamma and complement receptors as did control macrophages. The function of these receptors was not affected by HIV-1 infection since binding and internalization of opsonized Escherichia coli and Staphylococcus aureus were not impaired. Production of reactive oxygen species induced by stimulation of the HIV-1-infected macrophages with opsonized E. coli, zymosan, or PMA was intact. HIV-1-infected macrophages killed opsonized E. coli and Candida albicans as effectively as did control macrophages. These results, therefore, do not support the hypothesis that HIV-1 infection of macrophages causes phagocytic dysfunction and suggest that HIV-induced abnormalities outside the mononuclear phagocyte system may lead to the inability to control opportunistic pathogens. PMID:8390549

  19. Expression of adhesion molecules, monocyte interactions and oxidative stress in human endothelial cells exposed to wood smoke and diesel exhaust particulate matter

    DEFF Research Database (Denmark)

    Forchhammer, Lykke Ali; Loft, Steffen; Roursgaard, Martin; Cao, Yi; Riddervold, Ingunn Skogstad; Sigsgaard, Torben; Møller, Peter

    2012-01-01

    Toxicological effects of wood smoke particles are less investigated than traffic-related combustion particles. We investigated the effect of wood smoke particles, generated by smouldering combustion conditions, on human umbilical endothelial cells (HUVECs) co-cultured with or without monocytic THP...... the wood smoke particles caused increased level of DNA strand breaks and oxidised guanines at concentrations with low cytotoxicity. In conclusion, our results indicate that the adherence of monocytes on endothelial cells in wood smoke particle exposed cultures depend on activation of both cell types....

  20. Differences in Rate and Variability of Intracellular Growth of a Panel of Mycobacterium tuberculosis Clinical Isolates within a Human Monocyte Model

    OpenAIRE

    Li, Qing; Whalen, Christopher C.; Albert, Jeffrey M.; Larkin, Rhonda; Zukowski, Lynn; Cave, M. Donald; Silver, Richard F.

    2002-01-01

    Significant differences were observed in the capacities of Mycobacterium tuberculosis clinical isolates to grow within human monocytes. Genotyping indicated that the four most rapidly growing isolates were members of the Beijing strain family. M. tuberculosis strain H37Rv provided more reproducible infection than the clinical isolates or M. tuberculosis Erdman.

  1. Urokinase receptor mRNA level and gene transcription are strongly and rapidly increased by phorbol myristate acetate in human monocyte-like U937 cells

    DEFF Research Database (Denmark)

    Lund, L R; Rønne, E; Roldan, A L;

    1991-01-01

    We have studied the effect of the tumor promotor phorbol myristate acetate (PMA) on the level of mRNA for the receptor for urokinase-type plasminogen activator (u-PAR) in the human monocyte-like cell line U937. PMA causes an early increase in the u-PAR mRNA level which reaches a maximal 50-fold e...

  2. DEAD-box proteins, like Leishmania eIF4A, modulate interleukin (IL)-12, IL-10 and tumour necrosis factor-alpha production by human monocytes.

    Science.gov (United States)

    Barhoumi, M; Meddeb-Garnaoui, A; Tanner, N K; Banroques, J; Kaabi, B; Guizani, I

    2013-01-01

    Previously we showed that His-tagged, recombinant, Leishmania infantum eukaryotic initiation factor (LeIF) was both an RNA-dependent ATPase and an ATP-dependent RNA helicase in vitro, as described for other members of the DEAD-box helicase family. In addition, we showed that LeIF induces the production of IL-12, IL-10, and TNF-α by human monocytes. This study aims to characterize the cytokine-inducing activity in human monocytes of several proteins belonging to the DEAD-box family from mammals and yeast. All tested proteins contained the 11 conserved motifs (Q, I, Ia, GG Ib, II, III, IV, QxxR, V and VI) characteristic of DEAD-box proteins, but they have different biological functions and different percentages of identities with LeIF. We show that these mammalian or yeast recombinant proteins also are able to induce IL-12, IL-10 and TNF-α secretion by monocytes of healthy human subjects. This cytokine-inducing activity is proteinase K sensitive and polymyxin B resistant. Our results show that the induction of cytokines in human monocytes is not unique to the protein LeIF of Leishmania, and it suggests that the activity of certain DEAD-box proteins can be exploited as adjuvant and/or to direct immune responses towards a Th1 profile in vaccination or immunotherapy protocols. PMID:23363368

  3. Epicubenol and Ferruginol induce DC from human monocytes and differentiate IL-10-producing regulatory T cells in vitro

    International Nuclear Information System (INIS)

    Epicubenol and 19-hydroxyferruginol (Ferruginol) are sesquiterpenes isolated from the black heartwood of Cryptomeria japonica. Dendritic cells (DC) are specialized antigen-presenting cells that monitor the antigenic environment and activate naive T cells. The role of DC is not only to sense danger but also to tolerize the immune system to antigens encountered in the absence of maturation/inflammatory stimuli. In this study, we attempted to investigate the effects of Epicubenol and Ferruginol on the phenotypic and functional maturation of human monocytes-derived DC in vitro. Human monocytes were cultured with GM-CSF and IL-4 for 6 days under standard conditions, followed by another 2 days with Epicubenol or Ferruginol. The expression levels of CD1a, CD83, and HLA-DR as expressed by mean fluorescence intensity (MFI) on Epicubenol-primed DC or Ferruginol-primed DC were enhanced. Allogeneic Epicubenol-primed DC or Ferruginol-primed DC co-cultured with naive T cells at 1:5 ratio, secreted IL-10 and TGF-β, but little IL-4. Moreover, T cells that develop in co-culture of Epicubenol-primed DC or Ferruginol-primed DC and naive T cells at 1:5 ratio suppressed the proliferation of autologous T cells at Treg cells: Ttarget cells and this suppression of proliferation was inhibited by anti-IL-10 mAb. The expression of FoxP3 mRNA on T cells that develop in co-culture of Epicubenol-primed DC or Ferruginol-primed DC and naive T cells was lower. From these results, Epicubenol and Ferruginol may induce IL-10-producing Treg 1 cells from naive T cells by modulating DC function. It seems that Epicubenol and Ferruginol appear to be a target for tolerance after transplantation and in autoimmune diseases

  4. Granulocyte-Macrophage Colony-Stimulating Factor- and Tumor Necrosis Factor Alpha-Mediated Matrix Metalloproteinase Production by Human Osteoblasts and Monocytes after Infection with Brucella abortus ▿

    OpenAIRE

    Scian, Romina; Barrionuevo, Paula; Giambartolomei, Guillermo H.; Fossati, Carlos A.; Baldi, Pablo C.; Delpino, M. Victoria

    2010-01-01

    Osteoarticular complications are common in human brucellosis, but the pathogenic mechanisms involved are largely unknown. Since matrix metalloproteinases (MMPs) are involved in joint and bone damage in inflammatory and infectious diseases, we investigated the production of MMPs by human osteoblasts and monocytes, either upon Brucella abortus infection or upon reciprocal stimulation with factors produced by each infected cell type. B. abortus infection of the normal human osteoblastic cell lin...

  5. Deoxyelephantopin from Elephantopus scaber L. induces cell-cycle arrest and apoptosis in the human nasopharyngeal cancer CNE cells

    International Nuclear Information System (INIS)

    Highlights: → Deoxyelephantopin (ESD) inhibited cell proliferation in the human nasopharyngeal cancer CNE cells. → ESD induced cell cycle arrest in S and G2/M phases via modulation of cell cycle regulatory proteins. → ESD triggered apoptosis by dysfunction of mitochondria and induction of both intrinsic and extrinsic apoptotic signaling pathways. → ESD also triggered Akt, ERK, and JNK signaling pathways. -- Abstract: Deoxyelephantopin (ESD), a naturally occurring sesquiterpene lactone present in the Chinese medicinal herb, Elephantopus scaber L. exerted anticancer effects on various cultured cancer cells. However, the cellular mechanisms by which it controls the development of the cancer cells are unavailable, particularly the human nasopharyngeal cancer CNE cells. In this study, we found that ESD inhibited the CNE cell proliferation. Cell cycle arrest in S and G2/M phases was also found. Western blotting analysis showed that modulation of cell cycle regulatory proteins was responsible for the ESD-induced cell cycle arrest. Besides, ESD also triggered apoptosis in CNE cells. Dysfunction in mitochondria was found to be associated with the ESD-induced apoptosis as evidenced by the loss of mitochondrial membrane potential (ΔΨm), the translocation of cytochrome c, and the regulation of Bcl-2 family proteins. Despite the Western blotting analysis showed that both intrinsic and extrinsic apoptotic pathways (cleavage of caspases-3, -7, -8, -9, and -10) were triggered in the ESD-induced apoptosis, additional analysis also showed that the induction of apoptosis could be achieved by the caspase-independent manner. Besides, Akt, ERK and JNK pathways were found to involve in ESD-induced cell death. Overall, our findings provided the first evidence that ESD induced cell cycle arrest, and apoptosis in CNE cells. ESD could be a potential chemotherapeutic agent in the treatment of nasopharyngeal cancer (NPC).

  6. HIV-1 infection of in vitro cultured human monocytes: early events and influence of anti HIV-1 antibodies

    DEFF Research Database (Denmark)

    Arendrup, M; Olofsson, S; Nielsen, Jens Ole; Hansen, J E

    To characterize the role of the humoral immune response on HIV-1 infection of monocytes and macrophages (M phi s) we examined the susceptibility of in vitro cultured monocyte/M phi s to various HIV-1 isolates and the influence of heterologous and particularly autologous anti HIV-1 sera on this...... infection. Depending on the period of in vitro cultivation and the virus isolate used different patterns of susceptibility were detected. One week old monocyte/M phi s were highly susceptible to HIV-1 infection, in contrast to monocyte/M phi s cultured 4 weeks. The infection by virus isolated immediately...

  7. Dharmendra antigen but not integral M. leprae is an efficient inducer of immunostimulant cytokine production by human monocytes, and M. leprae lipids inhibit the cytokine production.

    Science.gov (United States)

    Nakamura, C; Fukutomi, Y; Kashiwabara, Y; Oomoto, Y; Kojima, M; Hayashi, H; Onozaki, K

    1997-03-01

    Killed integral Mycobacterium leprae, Mitsuda antigen, and chloroform-treated M. leprae, Dharmendra antigen (Dh-Ag), have been used for the classification of leprosy patients based on cell-mediated immunity. Heat-killed M. leprae also were used as a component of the Convit vaccine. Human blood monocytes were stimulated with M. leprae or Dh-Ag and their cytokine-inducing ability was compared. Monocytes were cultured in the presence of fresh human serum because of the efficiency of cytokine induction and the phagocytosis of M. leprae have been shown to be optimal in the presence of fresh serum. M. leprae and Dh-Ag were equally phagocytosed by monocytes. Dh-Ag was more potent than M. leprae in the induction of immunostimulatory/proinflammatory cytokines, interleukin-1 (IL-1), IL-6 and tumor necrosis factor (TNF). In contrast, a comparable level of IL-1ra, an immunosuppressive cytokine, was induced by M. leprae and Dh-Ag. The lipids extracted from M. leprae induced none of these cytokines by monocytes. Nevertheless, when monocytes were pretreated with the lipids followed by stimulation with Dh-Ag, productions of IL-1, IL-6 and TNF were all inhibited in a dose-dependent manner. However, the lipids did not inhibit the cytokine production induced by other stimuli including BCG and lipopolysaccharide. Moreover the lipids did not affect the production of IL-1ra. These results suggest that the lipids from M. leprae are responsible for the poor cytokine-inducing ability of M. leprae, thus favoring their infection. These results also suggest that Dh-Ag rather than integral M. leprae may be useful as a vaccine candidate because Dh-Ag is able to induce a large amount of cytokines from monocytes. PMID:9207755

  8. Monocyte-induced recovery of inflammation-associated hepatocellular dysfunction in a biochip-based human liver model.

    Science.gov (United States)

    Gröger, Marko; Rennert, Knut; Giszas, Benjamin; Weiß, Elisabeth; Dinger, Julia; Funke, Harald; Kiehntopf, Michael; Peters, Frank T; Lupp, Amelie; Bauer, Michael; Claus, Ralf A; Huber, Otmar; Mosig, Alexander S

    2016-01-01

    Liver dysfunction is an early event in sepsis-related multi-organ failure. We here report the establishment and characterization of a microfluidically supported in vitro organoid model of the human liver sinusoid. The liver organoid is composed of vascular and hepatocyte cell layers integrating non-parenchymal cells closely reflecting tissue architecture and enables physiological cross-communication in a bio-inspired fashion. Inflammation-associated liver dysfunction was mimicked by stimulation with various agonists of toll-like receptors. TLR-stimulation induced the release of pro- and anti-inflammatory cytokines and diminished expression of endothelial VE-cadherin, hepatic MRP-2 transporter and apolipoprotein B (ApoB), resulting in an inflammation-related endothelial barrier disruption and hepatocellular dysfunction in the liver organoid. However, interaction of the liver organoid with human monocytes attenuated inflammation-related cell responses and restored MRP-2 transporter activity, ApoB expression and albumin/urea production. The cellular events observed in the liver organoid closely resembled pathophysiological responses in the well-established sepsis model of peritoneal contamination and infection (PCI) in mice and clinical observations in human sepsis. We therefore conclude that this human liver organoid model is a valuable tool to investigate sepsis-related liver dysfunction and subsequent immune cell-related tissue repair/remodeling processes. PMID:26902749

  9. Alcohol and cannabinoids differentially affect HIV infection and function of human monocyte-derived dendritic cells (MDDC

    Directory of Open Access Journals (Sweden)

    Marisela eAgudelo

    2015-12-01

    Full Text Available During human immunodeficiency virus (HIV infection, alcohol has been known to induce inflammation while cannabinoids have been shown to have an anti-inflammatory role. For instance cannabinoids have been shown to reduce susceptibility to HIV-1 infection and attenuate HIV replication in macrophages. Recently, we demonstrated that alcohol induces cannabinoid receptors and regulates cytokine production by monocyte-derived dendritic cells (MDDC. However, the ability of alcohol and cannabinoids to alter MDDC function during HIV infection has not been clearly elucidated yet. In order to study the potential impact of alcohol and cannabinoids on differentiated MDDC infected with HIV, monocytes were cultured for 7 days with GM-CSF and IL-4, differentiated MDDC were infected with HIV-1Ba-L and treated with EtOH (0.1 and 0.2%, THC (5 and 10 uM, or JWH-015 (5 and 10 uM for 4-7 days. HIV infection of MDDC was confirmed by p24 and Long Terminal Repeats (LTR estimation. MDDC endocytosis assay and cytokine array profiles were measured to investigate the effects of HIV and substances of abuse on MDDC function. Our results show the HIV+EtOH treated MDDC had the highest levels of p24 production and expression when compared with the HIV positive controls and the cannabinoid treated cells. Although both cannabinoids, THC and JWH-015 had lower levels of p24 production and expression, the HIV+JWH-015 treated MDDC had the lowest levels of p24 when compared to the HIV+THC treated cells. In addition, MDDC endocytic function and cytokine production were also differentially altered after alcohol and cannabinoid treatments. Our results show a differential effect of alcohol and cannabinoids, which may provide insights into the divergent inflammatory role of alcohol and cannabinoids to modulate MDDC function in the context of HIV infection.

  10. Spleen Tyrosine Kinase (Syk) Mediates IL-1β Induction by Primary Human Monocytes during Antibody-enhanced Dengue Virus Infection.

    Science.gov (United States)

    Callaway, Justin B; Smith, Scott A; McKinnon, Karen P; de Silva, Aravinda M; Crowe, James E; Ting, Jenny P-Y

    2015-07-10

    Approximately 500,000 people are hospitalized with severe dengue illness annually. Antibody-dependent enhancement (ADE) of dengue virus (DENV) infection is believed to contribute to the pathogenic cytokine storm described in severe dengue patients, but the precise signaling pathways contributing to elevated cytokine production are not elucidated. IL-1β is a potent inflammatory cytokine that is frequently elevated during severe dengue, and the unique dual regulation of IL-1β provides an informative model to study ADE-induced cytokines. This work utilizes patient-derived anti-DENV mAbs and primary human monocytes to study ADE-induced IL-1β and other cytokines. ADE of DENV serotype 2 (DENV-2) elevates mature IL-1β secretion by monocytes independent of DENV replication by 4 h postinoculation (hpi). Prior to this, DENV immune complexes activate spleen tyrosine kinase (Syk) within 1 hpi. Syk induces elevated IL1B, TNF, and IL6 mRNA by 2 hpi. Syk mediates elevated IL-1β secretion by activating ERK1/2, and both Syk and ERK1/2 inhibitors ablated ADE-induced IL-1β secretion. Maturation of pro-IL-1β during ADE requires caspase-1 and NLRP3, but caspase-1 is suboptimally increased by ADE and can be significantly enhanced by a typical inflammasome agonist, ATP. Importantly, this inflammatory Syk-ERK signaling axis requires DENV immune complexes, because DENV-2 in the presence of serotype-matched anti-DENV-2 mAb, but not anti-DENV-1 mAb, activates Syk, ERK, and IL-1β secretion. This study provides evidence that DENV-2 immune complexes activate Syk to mediate elevated expression of inflammatory cytokines. Syk and ERK may serve as new therapeutic targets for interfering with ADE-induced cytokine expression during severe dengue. PMID:26032420

  11. Erythropoietin receptor is expressed on human peripheral blood T and B lymphocytes and monocytes and is modulated by recombinant human erythropoietin treatment.

    Science.gov (United States)

    Lisowska, Katarzyna A; Debska-Slizień, Alicja; Bryl, Ewa; Rutkowski, Bolesław; Witkowski, Jacek M

    2010-08-01

    Erythropoietin receptor (EPO-R) appears on the cell surface in the early stages of erythropoiesis. It has also been found on endothelial cells and polymorphonuclear leukocytes, suggesting erythropoietin (EPO) role beyond erythropoiesis itself. Earlier reports have shown that treatment with recombinant human erythropoietin (rhEPO) in chronic renal failure (CRF) patients improves interleukin-2 production and restores the T lymphocyte function. We decided to investigate possible expression of EPO-R on circulating peripheral blood lymphocytes and monocytes of CRF patients in order to assess the possibility of rhEPO direct action on these cells. Flow cytometry was used for detection and quantification of EPO-R, and reverse transcription polymerase chain reaction for detection of the EPO receptor mRNA. Our results show for the first time the existence of EPO-R on cell surface of human T and B lymphocytes and monocytes as well as at the transcriptional activity of the EPO-R gene in these cells, both in healthy and CRF individuals. We have also found significant differences between the numbers of EPO-R molecules on T and B lymphocytes of CRF patients not treated and treated with rhEPO and healthy control. Discovery of EPO-R expression on human lymphocytes suggests that EPO is probably able to directly modulate some signaling pathways important for these cells. PMID:20528849

  12. Activation of human monocytes by streptococcal rhamnose glucose polymers is mediated by CD14 antigen, and mannan binding protein inhibits TNF-alpha release.

    Science.gov (United States)

    Soell, M; Lett, E; Holveck, F; Schöller, M; Wachsmann, D; Klein, J P

    1995-01-15

    The present work was initiated to define mechanisms that account for the binding on human monocytes of streptococcal cell wall polysaccharides formed by rhamnose glucose polymers (RGPs), and subsequent stimulatory activities. We show here that RGPs bind to and stimulate human monocytes to produce TNF-alpha in a dose-dependent manner. To detect cell surface RGPs binding proteins, intact monocytes were biotinylated before lysis with Nonidet P-40 and solubilized proteins were incubated with RGPs Affi-Prep beads. One major membrane protein of 55 kDa was specifically detected and identified as CD14 because it reacted with anti-CD14 mAbs. Furthermore, anti-CD14 mAbs were able to perform a dose-dependent inhibition of RGPs binding, and suppressed TNF-alpha release from RGPs-stimulated monocytes. Moreover, we demonstrated that RGPs also bind to CD11b; however, this binding is not implicated in synthesis of TNF-alpha. Interestingly, RGPs binding to monocytes was enhanced by human normal serum (HNS) whereas HNS inhibits the TNF-alpha-stimulating activity of RGPs. Western blotting analysis of HNS proteins purified on RGPs Affi-prep beads revealed three specific bands of 75, 55, and 32 kDa reactive with anti-C3 Abs, anti-CD14 mAbs (TUK4), and anti-human mannan binding protein (hMBP)-derived peptide IgG, respectively. These results suggest that C3, soluble CD14, and hMBP form complexes that are probably active in enhancing the binding of RGPs to monocytes. Additional studies have shown that hMBP that recognizes RGPs prevents, unlike the LPS binding protein, TNF-alpha release by inhibiting the binding of RGPs to CD14 Ag. By incubating cells with a constant amount of RGPs-hMBP complexes in the presence or absence of increasing concentrations of C1q, we also demonstrated that C1q receptor mediates the binding and probably the uptake of RGPs-hMBP complexes by human monocytes. PMID:7529289

  13. Redox Stimulation of Human THP-1 Monocytes in Response to Cold Physical Plasma

    Directory of Open Access Journals (Sweden)

    Sander Bekeschus

    2016-01-01

    Full Text Available In plasma medicine, cold physical plasma delivers a delicate mixture of reactive components to cells and tissues. Recent studies suggested a beneficial role of cold plasma in wound healing. Yet, the biological processes related to the redox modulation via plasma are not fully understood. We here used the monocytic cell line THP-1 as a model to test their response to cold plasma in vitro. Intriguingly, short term plasma treatment stimulated cell growth. Longer exposure only modestly compromised cell viability but apparently supported the growth of cells that were enlarged in size and that showed enhanced metabolic activity. A significantly increased mitochondrial content in plasma treated cells supported this notion. On THP-1 cell proteome level, we identified an increase of protein translation with key regulatory proteins being involved in redox regulation (hypoxia inducible factor 2α, differentiation (retinoic acid signaling and interferon inducible factors, and cell growth (Yin Yang 1. Regulation of inflammation is a key element in many chronic diseases, and we found a significantly increased expression of the anti-inflammatory heme oxygenase 1 (HMOX1 and of the neutrophil attractant chemokine interleukin-8 (IL-8. Together, these results foster the view that cold physical plasma modulates the redox balance and inflammatory processes in wound related cells.

  14. Piceatannol and Its Metabolite, Isorhapontigenin, Induce SIRT1 Expression in THP-1 Human Monocytic Cell Line

    Directory of Open Access Journals (Sweden)

    Shinpei Kawakami

    2014-10-01

    Full Text Available Piceatannol is a phytochemical that is present in large amounts in passion fruit (Passiflora edulis seeds, and is an analog of resveratrol. Recently, the absorption and metabolism of piceatannol were investigated in rats, and isorhapontigenin, O-methyl piceatannol, was detected as a piceatannol metabolite in rat plasma. To elucidate the function of piceatannol and its metabolites, we investigated the expression of sirtuin 1 (SIRT1 in THP-1 monocytic cells after treatment with piceatannol and its metabolites, and compared their effects with those of resveratrol and its metabolites. Piceatannol and resveratrol upregulated the expression levels of SIRT1 mRNA and SIRT1 protein. An extract of passion fruit seeds, which contained high levels of piceatannol, also upregulated SIRT1 mRNA expression. As for the metabolites, isorhapontigenin upregulated SIRT1 mRNA expression, whereas resveratrol glucuronides and sulfate did not affect SIRT1 expression. These findings indicate that after intake of piceatannol, not only piceatannol itself, but also its metabolite, isorhapontigenin, contributed to the upregulation of SIRT1 expression.

  15. Piceatannol and its metabolite, isorhapontigenin, induce SIRT1 expression in THP-1 human monocytic cell line.

    Science.gov (United States)

    Kawakami, Shinpei; Kinoshita, Yosuke; Maruki-Uchida, Hiroko; Yanae, Koji; Sai, Masahiko; Ito, Tatsuhiko

    2014-11-01

    Piceatannol is a phytochemical that is present in large amounts in passion fruit (Passiflora edulis) seeds, and is an analog of resveratrol. Recently, the absorption and metabolism of piceatannol were investigated in rats, and isorhapontigenin, O-methyl piceatannol, was detected as a piceatannol metabolite in rat plasma. To elucidate the function of piceatannol and its metabolites, we investigated the expression of sirtuin 1 (SIRT1) in THP-1 monocytic cells after treatment with piceatannol and its metabolites, and compared their effects with those of resveratrol and its metabolites. Piceatannol and resveratrol upregulated the expression levels of SIRT1 mRNA and SIRT1 protein. An extract of passion fruit seeds, which contained high levels of piceatannol, also upregulated SIRT1 mRNA expression. As for the metabolites, isorhapontigenin upregulated SIRT1 mRNA expression, whereas resveratrol glucuronides and sulfate did not affect SIRT1 expression. These findings indicate that after intake of piceatannol, not only piceatannol itself, but also its metabolite, isorhapontigenin, contributed to the upregulation of SIRT1 expression. PMID:25360511

  16. Human papillomavirus 16E6 and NFX1-123 potentiate notch signaling and differentiation without activating cellular arrest

    International Nuclear Information System (INIS)

    High-risk human papillomavirus (HR HPV) oncoproteins bind host cell proteins to dysregulate and uncouple apoptosis, senescence, differentiation, and growth. These pathways are important for both the viral life cycle and cancer development. HR HPV16 E6 (16E6) interacts with the cellular protein NFX1-123, and they collaboratively increase the growth and differentiation master regulator, Notch1. In 16E6 expressing keratinocytes (16E6 HFKs), the Notch canonical pathway genes Hes1 and Hes5 were increased with overexpression of NFX1-123, and their expression was directly linked to the activation or blockade of the Notch1 receptor. Keratinocyte differentiation genes Keratin 1 and Keratin 10 were also increased, but in contrast their upregulation was only indirectly associated with Notch1 receptor stimulation and was fully unlinked to growth arrest, increased p21Waf1/CIP1, or decreased proliferative factor Ki67. This leads to a model of 16E6, NFX1-123, and Notch1 differently regulating canonical and differentiation pathways and entirely uncoupling cellular arrest from increased differentiation. - Highlights: • 16E6 and NFX1-123 increased the Notch canonical pathway through Notch1. • 16E6 and NFX1-123 increased the differentiation pathway indirectly through Notch1. • 16E6 and NFX1-123 increased differentiation gene expression without growth arrest. • Increased NFX1-123 with 16E6 may create an ideal cellular phenotype for HPV

  17. Human papillomavirus 16E6 and NFX1-123 potentiate notch signaling and differentiation without activating cellular arrest

    Energy Technology Data Exchange (ETDEWEB)

    Vliet-Gregg, Portia A.; Hamilton, Jennifer R. [Center for Global Infectious Disease Research, Seattle Children' s Research Institute, 1900 Ninth Ave., Seattle, WA 98101 (United States); Katzenellenbogen, Rachel A., E-mail: rkatzen@uw.edu [Center for Global Infectious Disease Research, Seattle Children' s Research Institute, 1900 Ninth Ave., Seattle, WA 98101 (United States); Department of Pediatrics, Division of Adolescent Medicine, University of Washington, Seattle WA (United States)

    2015-04-15

    High-risk human papillomavirus (HR HPV) oncoproteins bind host cell proteins to dysregulate and uncouple apoptosis, senescence, differentiation, and growth. These pathways are important for both the viral life cycle and cancer development. HR HPV16 E6 (16E6) interacts with the cellular protein NFX1-123, and they collaboratively increase the growth and differentiation master regulator, Notch1. In 16E6 expressing keratinocytes (16E6 HFKs), the Notch canonical pathway genes Hes1 and Hes5 were increased with overexpression of NFX1-123, and their expression was directly linked to the activation or blockade of the Notch1 receptor. Keratinocyte differentiation genes Keratin 1 and Keratin 10 were also increased, but in contrast their upregulation was only indirectly associated with Notch1 receptor stimulation and was fully unlinked to growth arrest, increased p21{sup Waf1/CIP1}, or decreased proliferative factor Ki67. This leads to a model of 16E6, NFX1-123, and Notch1 differently regulating canonical and differentiation pathways and entirely uncoupling cellular arrest from increased differentiation. - Highlights: • 16E6 and NFX1-123 increased the Notch canonical pathway through Notch1. • 16E6 and NFX1-123 increased the differentiation pathway indirectly through Notch1. • 16E6 and NFX1-123 increased differentiation gene expression without growth arrest. • Increased NFX1-123 with 16E6 may create an ideal cellular phenotype for HPV.

  18. Sensitivity to sodium arsenite in human melanoma cells depends upon susceptibility to arsenite-induced mitotic arrest

    International Nuclear Information System (INIS)

    Arsenic induces clinical remission in patients with acute promyelocytic leukemia and has potential for treatment of other cancers. The current study examines factors influencing sensitivity to arsenic using human malignant melanoma cell lines. A375 and SK-Mel-2 cells were sensitive to clinically achievable concentrations of arsenite, whereas SK-Mel-3 and SK-Mel-28 cells required supratherapeutic levels for toxicity. Inhibition of glutathione synthesis, glutathione S-transferase (GST) activity, and multidrug resistance protein (MRP) transporter function attenuated arsenite resistance, consistent with studies suggesting that arsenite is extruded from the cell as a glutathione conjugate by MRP-1. However, MRP-1 was not overexpressed in resistant lines and GST-π was only slightly elevated. ICP-MS analysis indicated that arsenite-resistant SK-Mel-28 cells did not accumulate less arsenic than arsenite-sensitive A375 cells, suggesting that resistance was not attributable to reduced arsenic accumulation but rather to intrinsic properties of resistant cell lines. The mode of arsenite-induced cell death was apoptosis. Arsenite-induced apoptosis is associated with cell cycle alterations. Cell cycle analysis revealed arsenite-sensitive cells arrested in mitosis whereas arsenite-resistant cells did not, suggesting that induction of mitotic arrest occurs at lower intracellular arsenic concentrations. Higher intracellular arsenic levels induced cell cycle arrest in the S-phase and G2-phase in SK-Mel-3 and SK-Mel-28 cells, respectively. The lack of arsenite-induced mitotic arrest in resistant cell lines was associated with a weakened spindle checkpoint resulting from reduced expression of spindle checkpoint protein BUBR1. These data suggest that arsenite has potential for treatment of solid tumors but a functional spindle checkpoint is a prerequisite for a positive response to its clinical application

  19. Lidamycin induces marked G2 cell cycle arrest in human colon carcinoma HT-29 cells through activation of p38 MAPK pathway.

    Science.gov (United States)

    Liu, Xia; Bian, Chunjing; Ren, Kaihuan; Jin, Haixia; Li, Baowei; Shao, Rong-Guang

    2007-03-01

    Lidamycin (LDM), a member of the enediyne antibiotic family, is presently undergoing phase I clinical trials in P.R. China. In this study, we investigated the mechanisms of LDM-induced cell cycle arrest in order to support its use in clinical cancer therapy. Using human colon carcinoma HT-29 cells, we observed that LDM induced G2 cell cycle arrest in a time- and dose-dependent manner. LDM-induced G2 arrest was associated with increasing phosphorylation of Chk1, Chk2, Cdc25C, Cdc2 and expression of Cdc2 and cyclin B1. In addition, cytoplasmic localization of cyclin B1 was also involved in LDM-induced G2 arrest. Moreover, we found that p38 MAPK pathway contributed to LDM-induced G2 arrest. Inhibition of p38 MAPK by its inhibitor SB203580 not only attenuated LDM-induced G2 arrest but also potentiated LDM-induced apoptosis, which was accompanied by decreasing phosphorylation of Cdc2 and increasing expression of FasL and phosphorylation of JNK. Finally, we demonstrated that cells at G1 phase were more sensitive to LDM. Together, our findings suggest that p38 MAPK signaling pathway is involved in LDM-induced G2 arrest, at least partly, and a combination of LDM with p38 MAPK inhibitor may represent a new strategy for human colon cancer therapy. PMID:17273739

  20. In vitro Effects of Selected Saponins on the Production and Release of Lysozyme Activity of Human Monocytic and Epithelial Cell Lines

    OpenAIRE

    Helal, Racha; Melzig, Matthias F.

    2011-01-01

    Lysozyme is one of the most important factors of innate immunity and a unique enzybiotic in that it exerts not only antibacterial activity, but also antiviral, anti-inflammatory, anticancer and immunomodulatory activities. The purpose of the present study was to investigate whether in vitro exposure to saponins can affect the release and production of lysozyme activity in human monocytic cells THP-1, and in human epithelial cells HT-29. Lysozyme activity levels in cell culture fluids were mea...

  1. The interleukin 1beta-converting enzyme, caspase 1, is activated during Shigella flexneri-induced apoptosis in human monocyte-derived macrophages.

    OpenAIRE

    Hilbi, H.; Chen, Y; Thirumalai, K; Zychlinsky, A

    1997-01-01

    Shigella, the etiological agent of bacillary dysentery, rapidly kills human monocyte-derived macrophages in vitro. Wild-type Shigella flexneri, but not a nonvirulent derivative, induced human macrophage apoptosis as determined by morphology and terminal deoxynucleotidyltransferase-mediated dUTP-biotin nick end labeling (TUNEL). Shigella-mediated macrophage cell death was blocked by the peptide inhibitors of caspases, acetyl-Tyr-Val-Ala-Asp-aldehyde (acetyl-YVAD-CHO) and acetyl-Tyr-Val-Ala-Asp...

  2. HIV-1 infection of in vitro cultured human monocytes: early events and influence of anti HIV-1 antibodies

    DEFF Research Database (Denmark)

    Arendrup, M; Olofsson, S; Nielsen, Jens Ole;

    1994-01-01

    To characterize the role of the humoral immune response on HIV-1 infection of monocytes and macrophages (M phi s) we examined the susceptibility of in vitro cultured monocyte/M phi s to various HIV-1 isolates and the influence of heterologous and particularly autologous anti HIV-1 sera on this...... infection. Depending on the period of in vitro cultivation and the virus isolate used different patterns of susceptibility were detected. One week old monocyte/M phi s were highly susceptible to HIV-1 infection, in contrast to monocyte/M phi s cultured 4 weeks. The infection by virus isolated immediately...... CD4 and that post binding events may be common to the infection of lymphocytes. Anti HIV-1 sera showed neutralizing activity against heterologous and even autologous escape virus. This finding, together with the observation that monocytes and M phi s are infected in vivo, suggests that protection...

  3. Candida albicans Targets a Lipid Raft/Dectin-1 Platform to Enter Human Monocytes and Induce Antigen Specific T Cell Responses.

    Directory of Open Access Journals (Sweden)

    Valeria de Turris

    Full Text Available Several pathogens have been described to enter host cells via cholesterol-enriched membrane lipid raft microdomains. We found that disruption of lipid rafts by the cholesterol-extracting agent methyl-β-cyclodextrin or by the cholesterol-binding antifungal drug Amphotericin B strongly impairs the uptake of the fungal pathogen Candida albicans by human monocytes, suggesting a role of raft microdomains in the phagocytosis of the fungus. Time lapse confocal imaging indicated that Dectin-1, the C-type lectin receptor that recognizes Candida albicans cell wall-associated β-glucan, is recruited to lipid rafts upon Candida albicans uptake by monocytes, supporting the notion that lipid rafts act as an entry platform. Interestingly disruption of lipid raft integrity and interference with fungus uptake do not alter cytokine production by monocytes in response to Candida albicans but drastically dampen fungus specific T cell response. In conclusion, these data suggest that monocyte lipid rafts play a crucial role in the innate and adaptive immune responses to Candida albicans in humans and highlight a new and unexpected immunomodulatory function of the antifungal drug Amphotericin B.

  4. Monocyte and macrophage differentiation: circulation inflammatory monocyte as biomarker for inflammatory diseases

    OpenAIRE

    Yang, Jiyeon; Zhang, Lixiao; Yu, Caijia; Yang, Xiao-Feng; Wang, Hong

    2014-01-01

    Monocytes express various receptors, which monitor and sense environmental changes. Monocytes are highly plastic and heterogeneous, and change their functional phenotype in response to environmental stimulation. Evidence from murine and human studies has suggested that monocytosis can be an indicator of various inflammatory diseases. Monocytes can differentiate into inflammatory or anti-inflammatory subsets. Upon tissue damage or infection, monocytes are rapidly recruited to the tissue, where...

  5. Inhibition of HIV-1 replication in human monocyte-derived macrophages by parasite Trypanosoma cruzi.

    Directory of Open Access Journals (Sweden)

    Guadalupe Andreani

    Full Text Available BACKGROUND: Cells of monocyte/macrophage lineage are one of the major targets of HIV-1 infection and serve as reservoirs for viral persistence in vivo. These cells are also the target of the protozoa Trypanosoma cruzi, the causative agent of Chagas disease, being one of the most important endemic protozoonoses in Latin America. It has been demonstrated in vitro that co-infection with other pathogens can modulate HIV replication. However, no studies at cellular level have suggested an interaction between T. cruzi and HIV-1 to date. METHODOLOGY/PRINCIPAL FINDINGS: By using a fully replicative wild-type virus, our study showed that T. cruzi inhibits HIV-1 antigen production by nearly 100% (p99% being stronger than HIV-T. cruzi (approximately 90% for BaL and approximately 85% for VSV-G infection. In MDM with established HIV-1 infection, T. cruzi significantly inhibited luciferate activity (p<0.01. By quantifying R-U5 and U5-gag transcripts by real time PCR, our study showed the expression of both transcripts significantly diminished in the presence of trypomastigotes (p<0.05. Thus, T. cruzi inhibits viral post-integration steps, early post-entry steps and entry into MDM. Trypomastigotes also caused a approximately 60-70% decrease of surface CCR5 expression on MDM. Multiplication of T. cruzi inside the MDM does not seem to be required for inhibiting HIV-1 replication since soluble factors secreted by trypomastigotes have shown similar effects. Moreover, the major parasite antigen cruzipain, which is secreted by the trypomastigote form, was able to inhibit viral production in MDM over 90% (p<0.01. CONCLUSIONS/SIGNIFICANCE: Our study showed that T. cruzi inhibits HIV-1 replication at several replication stages in macrophages, a major cell target for both pathogens.

  6. Cortical brain microdialysis and temperature monitoring during hypothermic circulatory arrest in humans

    OpenAIRE

    Mendelowitsch, A; Mergner, G; Shuaib, A.; Sekhar, L

    1998-01-01

    OBJECTIVES—Critical vascular surgery of the brain or the heart occasionally requires total cessation of the circulatory system. Profound hypothermia is used to protect the brain from ischaemic injury. This study explores the use of microdialysis to measure metabolic indices of ischaemia: glutamate, lactate, and pH, and cerebral temperature during profound hypothermia and circulatory arrest.
METHODS—Effluent from a microdialysis catheter placed in the cerebral cortex of three...

  7. Phagocytosis of haemozoin (malarial pigment enhances metalloproteinase-9 activity in human adherent monocytes: Role of IL-1beta and 15-HETE

    Directory of Open Access Journals (Sweden)

    Giribaldi Giuliana

    2008-08-01

    Full Text Available Abstract Background It has been shown previously that human monocytes fed with haemozoin (HZ or trophozoite-parasitized RBCs displayed increased matrix metalloproteinase-9 (MMP-9 enzyme activity and protein/mRNA expression and increased TNF production, and showed higher matrix invasion ability. The present study utilized the same experimental model to analyse the effect of phagocytosis of: HZ, delipidized HZ, beta-haematin (lipid-free synthetic HZ and trophozoites on production of IL-1beta and MMP-9 activity and expression. The second aim was to find out which component of HZ was responsible for the effects. Methods Native HZ freshly isolated from Plasmodium falciparum (Palo Alto strain, Mycoplasma-free, delipidized HZ, beta-haematin (lipid-free synthetic HZ, trophozoites and control meals such as opsonized non-parasitized RBCs and inert latex particles, were fed to human monocytes. The production of IL-1beta by differently fed monocytes, in presence or absence of specific MMP-9 inhibitor or anti-hIL-1beta antibodies, was quantified in supernatants by ELISA. Expression of IL-1beta was analysed by quantitative real-time RT-PCR. MMP-9 activity and protein expression were quantified by gelatin zymography and Western blotting. Results Monocytes fed with HZ or trophozoite-parasitized RBCs generated increased amounts of IL-1beta and enhanced enzyme activity (in cell supernatants and protein/mRNA expression (in cell lysates of monocyte MMP-9. The latter appears to be causally related to enhanced IL-1beta production, as enhancement of both expression and enzyme activity were abrogated by anti-hIL-1beta Abs. Upregulation of IL-1beta and MMP-9 were absent in monocytes fed with beta-haematin or delipidized HZ, indicating a role for HZ-attached or HZ-generated lipid components. 15-HETE (15(S,R-hydroxy-6,8,11,13-eicosatetraenoic acid a potent lipoperoxidation derivative generated by HZ from arachidonic acid via haem-catalysis was identified as one mediator

  8. Regulation of tumor necrosis factor gene expression by ionizing radiation in human myeloid leukemia cells and peripheral blood monocytes

    International Nuclear Information System (INIS)

    Previous studies have demonstrated that ionizing radiation induces the expression of certain cytokines, such as TNF alpha/cachectin. However, there is presently no available information regarding the molecular mechanisms responsible for the regulation of cytokine gene expression by ionizing radiation. In this report, we describe the regulation of the TNF gene by ionizing radiation in human myeloid leukemia cells. The increase in TNF transcripts by x rays was both time- and dose-dependent as determined by Northern blot analysis. Similar findings were obtained in human peripheral blood monocytes. Transcriptional run-on analyses have demonstrated that ionizing radiation stimulates the rate of TNF gene transcription. Furthermore, induction of TNF mRNA was increased in the absence of protein synthesis. In contrast, ionizing radiation had little effect on the half-life of TNF transcripts. These findings indicate that the increase in TNF mRNA observed after irradiation is regulated by transcriptional mechanisms and suggest that production of this cytokine by myeloid cells may play a role in the pathophysiologic effects of ionizing radiation

  9. Antiproliferative effect of rapamycin on human T-cell leukemia cell line Jurkat by cell cycle arrest and telomerase inhibition

    Institute of Scientific and Technical Information of China (English)

    Yan-min ZHAO; Qian ZHOU; Yun XU; Xiao-yu LAI; He HUANG

    2008-01-01

    Aim:To examine the ability of rapamycin to suppress growth and regulate telomerase activity in the human T-cell leukemia cell line Jurkat. Methods:Cell proliferation was assessed after exposure to rapamycin by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. Cell cycle progression and apoptosis were determined by flow cytometry. The proteins important for cell cycle progres-sion and Akt/mammalian target of rapamycin signaling cascade were assessed by Western blotting. Telomerase activity was quantified by telomeric repeat amplication protocol assay. The human telomerase reverse transcriptase (hTERT) mRNA levels were determined by semi-quantitative RT-PCR. Results:Rapamycin inhibited the proliferation of Jurkat, induced G1 phase arrest, unregulated the pro-tein level of p21 as well as p27, and downregulated cyclinD3, phospho-p70s6k, and phospho-s6, but had no effect on apoptosis. Treatment with rapamycin reduced telomerase activity, and reduced hTERT mRNA and protein expression. Conclusion:Rapamycin displayed a potent antileukemic effect in the human T-cell leukemia cell line by inhibition of cell proliferation through G1 cell cycle arrest and also through the suppression of telomerase activity, suggesting that rapamycin may have potential clinical implications in the treatment of some leukemias.

  10. Ganoderma lucidum polysaccharides can induce human monocytic leukemia cells into dendritic cells with immuno-stimulatory function

    Directory of Open Access Journals (Sweden)

    Lau Yu

    2008-07-01

    Full Text Available Abstract Background Previous studies demonstrated Ganoderma lucidum polysaccharides (GL-PS, a form of bioactive β-glucan can stimulate the maturation of monocyte-derived dendritic cells (DC. The question of how leukemic cells especially in monocytic lineage respond to GL-PS stimuli remains unclear. Results In this study, we used in vitro culture model with leukemic monocytic cell-lines THP-1 and U937 as monocytic effectors cells for proliferation responses and DCs induction. We treated the THP-1 and U937 cells with purified GL-PS (100 μg/mL or GL-PS with GM-CSF/IL-4. GL-PS alone induced proliferative response on both THP-1 and U937 cells but only THP-1 transformed into typical DC morphology when stimulated with GL-PS plus GM-CSF/IL-4. The transformed THP-1 DCs had significant increase expression of HLA-DR, CD40, CD80 and CD86 though not as high as the extent of normal monocyte-derived DCs. They had similar antigen-uptake ability as the normal monocyte-derived DCs positive control. However, their potency in inducing allogeneic T cell proliferation was also less than that of normal monocyte-derived DCs. Conclusion Our findings suggested that GL-PS could induce selected monocytic leukemic cell differentiation into DCs with immuno-stimulatory function. The possible clinical impact of using this commonly used medicinal mushroom in patients with monocytic leukemia (AML-M4 and M5 deserved further investigation.

  11. Hellebrigenin induces cell cycle arrest and apoptosis in human hepatocellular carcinoma HepG2 cells through inhibition of Akt.

    Science.gov (United States)

    Deng, Li-Juan; Hu, Li-Ping; Peng, Qun-Long; Yang, Xiao-Lin; Bai, Liang-Liang; Yiu, Anita; Li, Yong; Tian, Hai-Yan; Ye, Wen-Cai; Zhang, Dong-Mei

    2014-08-01

    Hellebrigenin, one of bufadienolides belonging to cardioactive steroids, was found in skin secretions of toads and plants of Helleborus and Kalanchoe genera. In searching for natural constituents with anti-hepatoma activities, we found that hellebrigenin, isolated from traditional Chinese medicine Venenum Bufonis, potently reduced the viability and colony formation of human hepatocellular carcinoma cells HepG2, and went on to explore the underlying molecular mechanisms. Our results demonstrated that hellebrigenin triggered DNA damage through DNA double-stranded breaks and subsequently induced cell cycle G2/M arrest associated with up-regulation of p-ATM (Ser(1981)), p-Chk2 (Tyr(68)), p-CDK1 (Tyr(15)) and Cyclin B1, and down-regulation of p-CDC25C (Ser(216)). It was also found that hellebrigenin induced mitochondrial apoptosis, characterized by Bax translocation to mitochondria, disruption of mitochondrial membrane potential, release of cytochrome c into cytosol and sequential activation of caspases and PARP. In addition, Akt expression and phosphorylation were inhibited by hellebrigenin, whereas Akt silencing with siRNA significantly blocked cell cycle arrest but enhanced apoptosis induced by hellebrigenin. Activation of Akt by human insulin-like growth factor I (hIGF-I) could obviously attenuate hellebrigenin-induced cell death. In summary, our study is the first to report the efficacy of hellebrigenin against HepG2 and elucidated its molecular mechanisms including DNA damage, mitochondria collapse, cell cycle arrest and apoptosis, which will contribute to the development of hellebrigenin into a chemotherapeutic agent in the treatment of liver cancer. PMID:24954031

  12. Subcellular proteomic analysis of host-pathogen interactions using human monocytes exposed to Yersinia pestis and Yersinia pseudotuberculosis

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, C G; Gonzales, A D; Choi, M W; Chromy, B A; Fitch, J P; McCutchen-Maloney, S L

    2004-05-20

    Yersinia pestis, the etiological agent of plague, is of concern to human health both from an infectious disease and a civilian biodefense perspective. While Y. pestis and Y. pseudotuberculosis share more than 90% DNA homology, they have significantly different clinical manifestations. Plague is often fatal if untreated, yet Y. pseudotuberculosis causes severe intestinal distress and is rarely fatal. A better understanding of host response to these closely related pathogens may help explain the different mechanisms of virulence and pathogenesis that result in such different clinical outcomes. The aim of this study was to characterize host protein expression changes in human monocyte-like U937 cells after exposure to Y. pestis and Y. pseudotuberculosis. In order to gain global proteomic coverage of host response, proteins from cytoplasmic, nuclear and membrane fractions of host cells were studied by 2-dimensional differential gel electrophoresis (2-D DIGE) and relative protein expression differences were quantitated. Differentially expressed proteins, with at least 1.5 fold expression changes and p values of 0.01 or less, were identified by MALDI-MS or LC/MS/MS. With these criteria, differential expression was detected in 16 human proteins after Y. pestis exposure and 13 human proteins after Y. pseudotuberculosis exposure, of which only two of the differentially expressed proteins identified were shared between the two exposures. Proteins identified in this study are reported to be involved in a wide spectrum of cellular functions and host defense mechanisms including apoptosis, cytoskeletal rearrangement, protein synthesis and degradation, DNA replication and transcription, metabolism, protein folding, and cell signaling. Notably, the differential expression patterns observed can distinguish the two pathogen exposures from each other and from unexposed host cells. The functions of the differentially expressed proteins identified provide insight on the different

  13. Fas expression correlates with human germ cell degeneration in meiotic and post-meiotic arrest of spermatogenesis.

    Science.gov (United States)

    Francavilla, Sandro; D'Abrizio, Piera; Cordeschi, Giuliana; Pelliccione, Fiore; Necozione, Stefano; Ulisse, Salvatore; Properzi, Giuliana; Francavilla, Felice

    2002-03-01

    Degeneration of human male germ cells was analysed by means of light (LM) and transmission electron (TEM) microscopy. The frequency of degenerating cells was correlated with that of Fas-expressing germ cells in human testes with normal spermatogenesis (n = 10), complete early maturation arrest (EMA) (n = 10) or incomplete late maturation arrest (LMA; n = 10) of spermatogenesis. LM analysis of testis sections with normal spermatogenesis indicated that degenerating germ cells were localized in the adluminal compartment of the seminiferous epithelium. TEM showed that apoptotic cells were mostly primary spermatocytes and, to a lesser extent, round or early elongating spermatids. Apoptotic germ cells appeared to be eliminated either in the seminiferous lumen or by Sertoli cell phagocytosis. An increased number of degenerating cells was observed in testes with LMA as compared with normal testes and testes with EMA of spermatogenesis (P < 0.001, Wilcoxon's rank sum test). Comparison of these results with those obtained from immunohistochemistry experiments demonstrated a tight correlation between the number of apoptotic cells and the number of Fas-expressing germ cells (P = 0.001, Spearman's rank = 0.69). These findings suggest that altered meiotic and post-meiotic germ cell maturation might be associated with an up-regulation of Fas gene expression capable of triggering apoptotic elimination of defective germ cells. PMID:11870228

  14. Grifola frondosa Glycoprotein GFG-3a Arrests S phase, Alters Proteome, and Induces Apoptosis in Human Gastric Cancer Cells.

    Science.gov (United States)

    Cui, Fengjie; Zan, Xinyi; Li, Yunhong; Sun, Wenjing; Yang, Yan; Ping, Lifeng

    2016-01-01

    GFG-3a is a novel glycoprotein previously purified from the fermented mycelia of Grifola frondosa with novel sugar compositions and protein sequencing. The present study aims to investigate its effects on the cell cycle, differential proteins expression, and apoptosis of human gastric cancer SGC-7901 cells. Our findings revealed that GFG-3a induced the cell apoptosis and arrested cell cycle at S phase. GFG-3a treatment resulted in the differential expression of 21 proteins in SGC-7901 cells by upregulating 10 proteins including RBBP4 associated with cell cycle arrest and downregulating 11 proteins including RUVBL1, NPM, HSP90AB1, and GRP78 involved in apoptosis and stress response. qRT-PCR and Western blot analysis also suggested that GFG-3a could increase the expressions of Caspase-8/-3, p53, Bax, and Bad while decrease the expressions of Bcl2, Bcl-xl, PI3K, and Akt1. These results indicated that the stress response, p53-dependent mitochondrial-mediated, Caspase-8/-3-dependent, and PI3k/Akt pathways were involved in the GFG-3a-induced apoptosis process in SGC-7901 cells. These findings might provide a basis to prevent or treat human gastric cancer with GFG-3a and understand the tumor-inhibitory molecular mechanisms of mushroom glycoproteins. PMID:27040446

  15. Asparanin A induces G(2)/M cell cycle arrest and apoptosis in human hepatocellular carcinoma HepG2 cells.

    Science.gov (United States)

    Liu, Wei; Huang, Xue-Feng; Qi, Qi; Dai, Qin-Sheng; Yang, Li; Nie, Fei-Fei; Lu, Na; Gong, Dan-Dan; Kong, Ling-Yi; Guo, Qing-Long

    2009-04-17

    We recently established that asparanin A, a steroidal saponin extracted from Asparagus officinalis L., is an active cytotoxic component. The molecular mechanisms by which asparanin A exerts its cytotoxic activity are currently unknown. In this study, we show that asparanin A induces G(2)/M phase arrest and apoptosis in human hepatocellular carcinoma HepG2 cells. Following treatment of HepG2 cells with asparanin A, cell cycle-related proteins such as cyclin A, Cdk1 and Cdk4 were down-regulated, while p21(WAF1/Cip1) and p-Cdk1 (Thr14/Tyr15) were up-regulated. Additionally, we observed poly (ADP-ribose) polymerase (PARP) cleavage and activation of caspase-3, caspase-8 and caspase-9. The expression ratio of Bax/Bcl-2 was increased in the treated cells, where Bax was also up-regulated. We also found that the expression of p53, a modulator of p21(WAF1/Cip1) and Bax, was not affected in asparanin A-treated cells. Collectively, our findings demonstrate that asparanin A induces cell cycle arrest and triggers apoptosis via a p53-independent manner in HepG2 cells. These data indicate that asparanin A shows promise as a preventive and/or therapeutic agent against human hepatoma. PMID:19254688

  16. Exposed hydrophobic residues in human immunodeficiency virus type 1 Vpr helix-1 are important for cell cycle arrest and cell death.

    Directory of Open Access Journals (Sweden)

    R Anthony Barnitz

    Full Text Available The human immunodeficiency virus type 1 (HIV-1 accessory protein viral protein R (Vpr is a major determinant for virus-induced G2/M cell cycle arrest and cytopathicity. Vpr is thought to perform these functions through the interaction with partner proteins. The NMR structure of Vpr revealed solvent exposed hydrophobic amino acids along helices 1 and 3 of Vpr, which could be putative protein binding domains. We previously showed that the hydrophobic patch along helix-3 was important for G2/M blockade and cytopathicity. Mutations of the exposed hydrophobic residues along helix-1 were found to reduce Vpr-induced cell cycle arrest and cell death as well. The levels of toxicity during virion delivery of Vpr correlated with G2/M arrest. Thus, the exposed hydrophobic amino acids in the amino-terminal helix-1 are important for the cell cycle arrest and cytopathicity functions of Vpr.

  17. Integrating genome-wide genetic variations and monocyte expression data reveals trans-regulated gene modules in humans.

    Directory of Open Access Journals (Sweden)

    Maxime Rotival

    2011-12-01

    Full Text Available One major expectation from the transcriptome in humans is to characterize the biological basis of associations identified by genome-wide association studies. So far, few cis expression quantitative trait loci (eQTLs have been reliably related to disease susceptibility. Trans-regulating mechanisms may play a more prominent role in disease susceptibility. We analyzed 12,808 genes detected in at least 5% of circulating monocyte samples from a population-based sample of 1,490 European unrelated subjects. We applied a method of extraction of expression patterns-independent component analysis-to identify sets of co-regulated genes. These patterns were then related to 675,350 SNPs to identify major trans-acting regulators. We detected three genomic regions significantly associated with co-regulated gene modules. Association of these loci with multiple expression traits was replicated in Cardiogenics, an independent study in which expression profiles of monocytes were available in 758 subjects. The locus 12q13 (lead SNP rs11171739, previously identified as a type 1 diabetes locus, was associated with a pattern including two cis eQTLs, RPS26 and SUOX, and 5 trans eQTLs, one of which (MADCAM1 is a potential candidate for mediating T1D susceptibility. The locus 12q24 (lead SNP rs653178, which has demonstrated extensive disease pleiotropy, including type 1 diabetes, hypertension, and celiac disease, was associated to a pattern strongly correlating to blood pressure level. The strongest trans eQTL in this pattern was CRIP1, a known marker of cellular proliferation in cancer. The locus 12q15 (lead SNP rs11177644 was associated with a pattern driven by two cis eQTLs, LYZ and YEATS4, and including 34 trans eQTLs, several of them tumor-related genes. This study shows that a method exploiting the structure of co-expressions among genes can help identify genomic regions involved in trans regulation of sets of genes and can provide clues for understanding the

  18. Scavenger receptor-mediated recognition of maleyl bovine plasma albumin and the demaleylated protein in human monocyte macrophages

    International Nuclear Information System (INIS)

    Maleyl bovine plasma albumin competed on an equimolar basis with malondialdehyde low density lipoprotein (LDL) in suppressing the lysosomal hydrolysis of 125I-labeled malondialdehyde LDL mediated by the scavenger receptor of human monocyte macrophages. Maleyl bovine plasma albumin, in which 94% of the amino groups were modified, exhibited an anodic mobility in agarose electrophoresis 1.7 times that of the native protein. Incubation of maleyl bovine plasma albumin at pH 3.5 regenerated the free amino groups and restored the protein to the same electrophoretic mobility as native albumin. Although ligands recognized by the scavenger receptor typically are anionic, the authors propose that addition of new negative charge achieved by maleylation, rather than directly forming the receptor binding site(s), induces conformational changes in albumin as a prerequisite to expression of the recognition domain(s). They conclude that the primary sequence of albumin, rather than addition of new negative charge, provides the recognition determinant(s) essential for interaction of maleyl bovine plasma albumin with the scavenger receptor

  19. Lymph node hemophagocytosis in rickettsial diseases: a pathogenetic role for CD8 T lymphocytes in human monocytic ehrlichiosis (HME?

    Directory of Open Access Journals (Sweden)

    Dumler J Stephen

    2006-07-01

    Full Text Available Abstract Background Human monocytic ehrlichiosis (HME and Rocky Mountain spotted fever (RMSF are caused by Ehrlichia chaffeensis and Rickettsia rickettsii, respectively. The pathogenesis of RMSF relates to rickettsia-mediated vascular injury, but it is unclear in HME. Methods To study histopathologic responses in the lymphatic system for correlates of immune injury, lymph nodes from patients with HME (n = 6 and RMSF (n = 5 were examined. H&E-stained lymph node tissues were examined for five histopathologic features, including hemophagocytosis, cellularity, necrosis, and vascular congestion and edema. The relative proportions of CD68 macrophages, CD8 and CD4 T lymphocytes, and CD20 B lymphocytes were evaluated by immunohistochemical staining. Results Hemophagocytosis was similar in HME and RMSF, and was greater than in control cases (p = .015. Cellularity in HME was not different from controls, whereas RMSF lymph nodes were markedly less cellular (p E. chaffeensis-infected mononuclear phagocytes were infrequent compared to R. rickettsii-infected endothelial cells. More CD8 cells in lymph nodes were observed with HME (p Conclusion Hemophagocytosis, CD8 T cell expansion, and the paucity of infected cells in HME, suggest that E. chaffeensis infection leads to macrophage activation and immune-mediated injury.

  20. Annexin 2: a novel human immunodeficiency virus type 1 Gag binding protein involved in replication in monocyte-derived macrophages.

    Science.gov (United States)

    Ryzhova, Elena V; Vos, Robin M; Albright, Andrew V; Harrist, Alexia V; Harvey, Thomas; González-Scarano, Francisco

    2006-03-01

    Human immunodeficiency virus (HIV) replication in the major natural target cells, CD4+ T lymphocytes and macrophages, is parallel in many aspects of the virus life cycle. However, it differs as to viral assembly and budding, which take place on plasma membranes in T cells and on endosomal membranes in macrophages. It has been postulated that cell type-specific host factors may aid in directing viral assembly to distinct destinations. In this study we defined annexin 2 (Anx2) as a novel HIV Gag binding partner in macrophages. Anx2-Gag binding was confined to productively infected macrophages and was not detected in quiescently infected monocyte-derived macrophages (MDM) in which an HIV replication block was mapped to the late stages of the viral life cycle (A. V. Albright, R. M. Vos, and F. Gonzalez-Scarano, Virology 325:328-339, 2004). We demonstrate that the Anx2-Gag interaction likely occurs at the limiting membranes of late endosomes/multivesicular bodies and that Anx2 depletion is associated with a significant decline in the infectivity of released virions; this coincided with incomplete Gag processing and inefficient incorporation of CD63. Cumulatively, our data suggest that Anx2 is essential for the proper assembly of HIV in MDM. PMID:16501079

  1. Oxygen-Loaded Nanodroplets Effectively Abrogate Hypoxia Dysregulating Effects on Secretion of MMP-9 and TIMP-1 by Human Monocytes

    OpenAIRE

    2015-01-01

    Monocytes play a key role in the inflammatory stage of the healing process. To allow monocyte migration to injured tissues, the balances between secreted matrix metalloproteinases (MMPs) and their inhibitors (TIMPs) must be finely modulated. However, a reduction of blood supply and local oxygen tension can modify the phenotype of immune cells. Intriguingly, hypoxia might be targeted by new effective oxygenating devices such as 2H,3H-decafluoropentane- (DFP-) based oxygen-loaded nanodroplets (...

  2. Daily Arrests

    Data.gov (United States)

    Montgomery County of Maryland — This dataset provides the public with arrest information from the Montgomery County Central Processing Unit (CPU) systems. The data presented is derived from every...

  3. Human immunodeficiency virus infection of T cells and monocytes proceeds via receptor-mediated endocytosis

    OpenAIRE

    1988-01-01

    The rates of internalization and uncoating of 32P-labelled human immunodeficiency virus (HIV) in the human T lymphoid cell line CEM are consonant with a receptor-mediated endocytosis mechanism of entry. This interpretation was affirmed by electron microscopic observation of virions within endosomes. Virus binding and infectivity were inhibited to the same extent by pretreatment with OKT4A antibody, therefore, the CD4 receptor-dependent pathway of internalization appears to be the infectious r...

  4. Establishment of long-term monocyte suspension cultures from normal human peripheral blood

    OpenAIRE

    1982-01-01

    The long-term suspension growth of normal, immature myeloid cells from fresh human cord blood was recently reported and required cells separated on supplemented discontinuous Percoll gradients, growth in media containing hydrocortisone and vitamins D3, and gentle, continuous agitation (13). When normal adult bone marrow (six donors) or blood from Epstein-Barr virus (EBV)-seropositive donors (nine donors) was used as a source of fresh human leukocytes, only short-term proliferation of myeloid ...

  5. Monocyte/macrophage trafficking in acquired immunodeficiency syndrome encephalitis: Lessons from human and nonhuman primate studies

    OpenAIRE

    Fischer-Smith, Tracy; Bell, Christie; Croul, Sidney; Lewis, Mark; Rappaport, Jay

    2008-01-01

    Here the authors discuss evidence in human and animal models supporting two opposing views regarding the pathogenesis of human immunodeficiency virus (HIV) in the central nervous system (CNS): (1) HIV infection in the CNS is a compartmentalized infection, with the virus-infected macrophages entering the CNS early, infecting resident microglia and astrocytes, and achieving a state of latency with evolution toward a fulminant CNS infection late in the course of disease; or alternatively, (2) ev...

  6. Genetic Requirements for Salmonella-Induced Cytopathology in Human Monocyte-Derived Macrophages

    OpenAIRE

    Browne, Sara H; Lesnick, Marc L.; Guiney, Donald G.

    2002-01-01

    Infection of human macrophages with Salmonella enterica serovar Typhimurium or Salmonella enterica serovar Dublin produces delayed cytotoxicity characterized by cell detachment and associated apoptosis. Using a site-specific mutant in the SpvB active site, we verify that the ADP-ribosylation activity of SpvB is required for delayed cytotoxicity in human macrophages infected with Salmonella. SipB and the type III protein secretion system (TTSS) encoded by Salmonella pathogenicity island 1 (SPI...

  7. Evaluation of suppressive and pro-resolving effects of EPA and DHA in human primary monocytes and T-helper cells[S

    OpenAIRE

    Jaudszus, Anke; Gruen, Michael; Watzl, Bernhard; Ness, Christina; Roth, Alexander; Lochner, Alfred; Barz, Dagmar; Gabriel, Holger; Rothe, Michael; Jahreis, Gerhard

    2013-01-01

    Despite their beneficial anti-inflammatory properties, eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) may increase the infection risk at high doses, likely by generating an immune-depressed state. To assess the contribution of different immune cell populations to the immunomodulatory fatty acid effect, we comparatively investigated several aspects of inflammation in human T-helper (Th) cells and monocytes. Both fatty acids, but DHA to a lesser extent compared with EPA, selectively...

  8. Circulating CD14brightCD16+ 'intermediate' monocytes exhibit enhanced parasite pattern recognition in human helminth infection.

    Directory of Open Access Journals (Sweden)

    Joseph D Turner

    2014-04-01

    Full Text Available Circulating monocyte sub-sets have recently emerged as mediators of divergent immune functions during infectious disease but their role in helminth infection has not been investigated. In this study we evaluated whether 'classical' (CD14brightCD16-, 'intermediate' (CD14brightCD16+, and 'non-classical' (CD14dimCD16+ monocyte sub-sets from peripheral blood mononuclear cells varied in both abundance and ability to bind antigenic material amongst individuals living in a region of Northern Senegal which is co-endemic for Schistosoma mansoni and S. haematobium. Monocyte recognition of excretory/secretory (E/S products released by skin-invasive cercariae, or eggs, of S. mansoni was assessed by flow cytometry and compared between S. mansoni mono-infected, S. mansoni and S. haematobium co-infected, and uninfected participants. Each of the three monocyte sub-sets in the different infection groups bound schistosome E/S material. However, 'intermediate' CD14brightCD16+ monocytes had a significantly enhanced ability to bind cercarial and egg E/S. Moreover, this elevation of ligand binding was particularly evident in co-infected participants. This is the first demonstration of modulated parasite pattern recognition in CD14brightCD16+ intermediate monocytes during helminth infection, which may have functional consequences for the ability of infected individuals to respond immunologically to infection.

  9. Nonclassical Patrolling Monocyte Function in the Vasculature

    OpenAIRE

    Thomas, Graham; Tacke, Robert; Hedrick, Catherine C.; Hanna, Richard N.

    2015-01-01

    Nonclassical patrolling monocytes are characterized by their unique ability to actively patrol the vascular endothelium under homeostatic and inflammatory conditions. Patrolling monocyte subsets (CX3CR1highLy6C− in mouse, and CX3CR1highCD14dimCD16+ in humans) are distinct from the classical monocyte subsets (CCR2highLy6C+ in mouse, and CCR2highCD14+CD16− in humans) and exhibit unique functions in the vasculature and inflammatory disease. Patrolling monocytes function in a number of disease se...

  10. Knockdown of human deubiquitinase PSMD14 induces cell cycle arrest and senescence

    Energy Technology Data Exchange (ETDEWEB)

    Byrne, Ann; McLaren, Rajashree P.; Mason, Paul; Chai, Lilly; Dufault, Michael R.; Huang, Yinyin; Liang, Beirong; Gans, Joseph D.; Zhang, Mindy; Carter, Kara; Gladysheva, Tatiana B.; Teicher, Beverly A.; Biemann, Hans-Peter N.; Booker, Michael; Goldberg, Mark A.; Klinger, Katherine W.; Lillie, James [Genzyme Corporation, 49 New York Avenue, Framingham, MA 01701 (United States); Madden, Stephen L., E-mail: steve.madden@genzyme.com [Genzyme Corporation, 49 New York Avenue, Framingham, MA 01701 (United States); Jiang, Yide, E-mail: yide.jiang@genzyme.com [Genzyme Corporation, 49 New York Avenue, Framingham, MA 01701 (United States)

    2010-01-15

    The PSMD14 (POH1, also known as Rpn11/MPR1/S13/CepP1) protein within the 19S complex (19S cap; PA700) is responsible for substrate deubiquitination during proteasomal degradation. The role of PSMD14 in cell proliferation and senescence was explored using siRNA knockdown in carcinoma cell lines. Our results reveal that down-regulation of PSMD14 by siRNA transfection had a considerable impact on cell viability causing cell arrest in the G0-G1 phase, ultimately leading to senescence. The molecular events associated with decreased cell proliferation, cell cycle arrest and senescence include down-regulation of cyclin B1-CDK1-CDC25C, down-regulation of cyclin D1 and up-regulation of p21{sup /Cip} and p27{sup /Kip1}. Most notably, phosphorylation of the retinoblastoma protein was markedly reduced in PSMD14 knockdown cells. A comparative study with PSMB5, a subunit of the 20S proteasome, revealed that PSMB5 and PSMD14 have different effects on cell cycle, senescence and associated molecular events. These data support the view that the 19S and 20S subunits of the proteasome have distinct biological functions and imply that targeting 19S and 20S would have distinct molecular consequences on tumor cells.

  11. Monocyte dysfunction in Sydenham's chorea patients.

    Science.gov (United States)

    Torres, Karen C; Dutra, Walderez O; de Rezende, Vitor Bortolo; Cardoso, Francisco; Gollob, Kenneth J; Teixeira, Antonio L

    2010-04-01

    Until now, there are no conclusive data about the mechanisms involved in motor symptoms of Sydenham's chorea (SC). Taking into account the autoreactive antibody-mediated hypothesis of SC pathogenesis, the SC may be associated with uncontrolled immune mechanisms. Besides the antibody hypothesis, the innate immune system has been underappreciated. Hence, we evaluated the activation state of monocytes, cells that are precursors of macrophages, to characterize the inflammation profile of patients. We assessed the surface molecules CD80, CD86, and human leukocyte antigen DR expression in patients with SC by flow cytometry analysis. Our results showed a decreased CD14(+) (monocyte) frequency, with concomitant increased CD14(-) frequency inside monocyte population. Although monocyte population showed a decreased human leukocyte antigen DR and CD86 frequencies, the CD14(-) population showed an increased frequency of CD80(+) monocyte from SC compared with controls. These data suggest that monocytes showed a reduced costimulatory potential in SC. PMID:20080141

  12. Overexpression of cyclin L2 induces apoptosis and cell-cycle arrest in human lung cancer cells

    Institute of Scientific and Technical Information of China (English)

    LI Hong-li; WANG Tong-shan; LI Xiao-yu; LI Nan; HUANG Ding-zhi; CHEN Qi; BA Yi

    2007-01-01

    Background Uncontrolled cell division is one of the hallmarks of tumor growth. Researches have been focused on numerous molecules involved in this process. Cyclins are critical regulatory proteins of cell cycle progression and/or transcription. The present study aimed to investigate the anti-proliferative effect of cyclin L2, and to define its growth regulatory mechanisms using human lung adenocarcinoma cell line A549.Methods Human cyclin L2 was transfected into human lung adenocarcinoma cells (A549 cell), and was expressed in a mammalian expression vector pcDNA3.1. The effects and mechanisms of the cyclin L2 in cell growth, cell cycle analysis and apoptosis were studied by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT), flow cytometry or Western blot, respectively.Results Overexpression of cyclin L2 inhibited the growth of A549 cells. Cell cycle analysis in cells transfected with pCCNL2 revealed an increment in proportion in G0/G1 phase ((68.07 ± 4.2)%) in contrast to (60.39 ± 2.82)% of the cells transfected with mock vector. Apoptosis occurred in (7.25 ± 0.98)% cells transfected with pCCNL2, as compared with (1.25 ± 0.21)% of the mock vector control group. Cyclin L2-induced-G0/G1 arrest and apoptosis involved upregulation of caspase-3 and downregulation of Bcl-2 and survivin.Conclusion The results indicate that overexpression of cyclin L2 protein may promote efficient growth inhibition of human lung adenocarcinoma cells by inducing G0/G1 cell cycle arrest and apoptosis.

  13. Effector functions of a monoclonal aglycosylated mouse IgG2a: binding and activation of complement component C1 and interaction with human monocyte Fc receptor.

    Science.gov (United States)

    Leatherbarrow, R J; Rademacher, T W; Dwek, R A; Woof, J M; Clark, A; Burton, D R; Richardson, N; Feinstein, A

    1985-04-01

    Aglycosylated monoclonal anti-DNP mouse IgG2a produced in the presence of tunicamycin was compared with the native monoclonal IgG2a with respect to its ability to interact with the first component of complement, C1, and to compete with human IgG for binding to human monocyte Fc receptors. The aglycosylated IgG2a was found to bind subcomponent C1q with an equivalent capacity to the native IgG2a, but the dissociation constant was found to be increased three-fold. When activation of C1 by the glycosylated and aglycosylated IgG2a was compared, the rate of C1 activation by the aglycosylated IgG2a was reduced approximately three-fold. In contrast aglycosylation was accompanied by a large decrease (greater than or equal to 50-fold) in the apparent binding constant of monomeric IgG2a to human monocytes. The data suggest that the aglycosylated IgG2a has a structure which differs in the CH2 domain from the native IgG2a, and that the heterogeneous N-linked oligosaccharides of this monoclonal IgG2a which occur at a conserved position in the CH2 domain play a role in maintaining the integrity of its monocyte-binding site. This lack of monocyte binding may result either from a localized conformational change occurring in a single CH2 domain or from an alteration in the CH2-CH2 cross-domain architecture which is normally structured by a pair of opposing and interacting oligosaccharides. The minimal changes in C1q binding and C1 activation suggest that the oligosaccharides are, at most, indirectly involved in these events. PMID:4033665

  14. Honokiol, a chemopreventive agent against skin cancer, induces cell cycle arrest and apoptosis in human epidermoid A431 cells.

    Science.gov (United States)

    Chilampalli, Chandeshwari; Guillermo, Ruth; Kaushik, Radhey S; Young, Alan; Chandrasekher, Gudiseva; Fahmy, Hesham; Dwivedi, Chandradhar

    2011-11-01

    Honokiol is a plant lignan isolated from bark and seed cones of Magnolia officinalis. Recent studies from our laboratory indicated that honokiol pretreatment decreased ultraviolet B-induced skin cancer development in SKH-1 mice. The aim of the present investigation was to study the effects of honokiol on human epidermoid squamous carcinoma A431 cells and to elucidate possible mechanisms involved in preventing skin cancer. A431 cells were pretreated with different concentrations of honokiol for a specific time period and investigated for effects on apoptosis and cell cycle analysis. Treatment with honokiol significantly decreased cell viability and cell proliferation in a concentration- and time-dependent manner. Honokiol pretreatment at 50 μmol/L concentration induced G0/G1 cell cycle arrest significantly (P Cdk4 and Cdk6 proteins and up-regulated the expression of Cdk's inhibitor proteins p21 and p27. Pretreatment of A431 cells with honokiol leads to induction of apoptosis and DNA fragmentation. These findings indicate that honokiol provides its effects in squamous carcinoma cells by inducing cell cycle arrest at G0/G1 phase and apoptosis. PMID:21908486

  15. Induction of apoptosis and antiproliferative activity of naringenin in human epidermoid carcinoma cell through ROS generation and cell cycle arrest.

    Directory of Open Access Journals (Sweden)

    Md Sultan Ahamad

    Full Text Available A natural predominant flavanone naringenin, especially abundant in citrus fruits, has a wide range of pharmacological activities. The search for antiproliferative agents that reduce skin carcinoma is a task of great importance. The objective of this study was to analyze the anti-proliferative and apoptotic mechanism of naringenin using MTT assay, DNA fragmentation, nuclear condensation, change in mitochondrial membrane potential, cell cycle kinetics and caspase-3 as biomarkers and to investigate the ability to induce reactive oxygen species (ROS initiating apoptotic cascade in human epidermoid carcinoma A431 cells. Results showed that naringenin exposure significantly reduced the cell viability of A431 cells (p<0.01 with a concomitant increase in nuclear condensation and DNA fragmentation in a dose dependent manner. The intracellular ROS generation assay showed statistically significant (p<0.001 dose-related increment in ROS production for naringenin. It also caused naringenin-mediated epidermoid carcinoma apoptosis by inducing mitochondrial depolarization. Cell cycle study showed that naringenin induced cell cycle arrest in G0/G1 phase of cell cycle and caspase-3 analysis revealed a dose dependent increment in caspase-3 activity which led to cell apoptosis. This study confirms the efficacy of naringenin that lead to cell death in epidermoid carcinoma cells via inducing ROS generation, mitochondrial depolarization, nuclear condensation, DNA fragmentation, cell cycle arrest in G0/G1 phase and caspase-3 activation.

  16. Interaction of E-cadherin and PTEN regulates morphogenesis and growth arrest in human mammary epithelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Fournier, Marcia V.; Fata, Jimmie E.; Martin, Katherine J.; Yaswen, Paul; Bissell, Mina J.

    2009-06-03

    PTEN is a dual function phosphatase with tumor suppressor function compromised in a wide spectrum of cancers. Because tissue polarity and architecture are crucial modulators of normal and malignant behavior, we postulated that PTEN may play a role in maintenance of tissue integrity. We used two non-malignant human mammary epithelial cell lines (HMECs) that form polarized, growth-arrested structures (acini) when cultured in 3-dimensional laminin-rich extracellular matrix gels (3D lrECM). As acini begin to form, PTEN accumulates in both the cytoplasm, and at cell-cell contacts where it colocalizes with E-cadherin/{beta}-catenin complex. Reduction of PTEN levels by shRNA in lrECM prevents formation of organized breast acini and disrupts growth arrest. Importantly, disruption of acinar polarity and cell-cell contact by E-cadherin function-blocking antibodies reduces endogenous PTEN protein levels and inhibits its accumulation at cell-cell contacts. Conversely, in SKBR3 breast cancer cells lacking endogenous E-cadherin expression, exogenous introduction of E-cadherin gene causes induction of PTEN expression and its accumulation at sites of cell interactions. These studies provide evidence that E-cadherin regulates both the PTEN protein levels and its recruitment to cell-cell junctions in 3D lrECM indicating a dynamic reciprocity between architectural integrity and the levels and localization of PTEN. This interaction thus appears to be a critical integrator of proliferative and morphogenetic signaling in breast epithelial cells.

  17. Human Monocytes Promote Th1 and Th17 Responses to Streptococcus pneumoniae ▿

    OpenAIRE

    Olliver, Marie; Hiew, Jeffni; Mellroth, Peter; Henriques-Normark, Birgitta; Bergman, Peter

    2011-01-01

    Streptococcus pneumoniae is a leading cause of bacterial pneumonia, meningitis, and sepsis in children. Human immunity to pneumococcal infections has been assumed to depend on anticapsular antibodies. However, recent findings from murine models suggest that alternative mechanisms, dependent on T helper cells, are also involved. Although the immunological events in which T helper cells contribute to acquired immunity have been studied in mice, little is known about how these responses are gene...

  18. Omega-3 Polyunsaturated Fatty Acids Trigger Cell Cycle Arrest and Induce Apoptosis in Human Neuroblastoma LA-N-1 Cells

    Directory of Open Access Journals (Sweden)

    Wai Wing So

    2015-08-01

    Full Text Available Omega-3 (n-3 fatty acids are dietary long-chain fatty acids with an array of health benefits. Previous research has demonstrated the growth-inhibitory effect of n-3 fatty acids on different cancer cell lines in vitro, yet their anti-tumor effects and underlying action mechanisms on human neuroblastoma LA-N-1 cells have not yet been reported. In this study, we showed that docosahexaenoic acid (DHA and eicosapentaenoic acid (EPA exhibited time- and concentration-dependent anti-proliferative effect on the human neuroblastoma LA-N-1 cells, but had minimal cytotoxicity on the normal or non-tumorigenic cells, as measured by MTT reduction assay. Mechanistic studies indicated that DHA and EPA triggered G0/G1 cell cycle arrest in LA-N-1 cells, as detected by flow cytometry, which was accompanied by a decrease in the expression of CDK2 and cyclin E proteins. Moreover, DHA and EPA could also induce apoptosis in LA-N-1 cells as revealed by an increase in DNA fragmentation, phosphatidylserine externalization and mitochondrial membrane depolarization. Up-regulation of Bax, activated caspase-3 and caspase-9 proteins, and down-regulation of Bcl-XL protein, might account for the occurrence of apoptotic events. Collectively, our results suggest that the growth-inhibitory effect of DHA and EPA on LA-N-1 cells might be mediated, at least in part, via triggering of cell cycle arrest and apoptosis. Therefore, DHA and EPA are potential anti-cancer agents which might be used for the adjuvant therapy or combination therapy with the conventional anti-cancer drugs for the treatment of some forms of human neuroblastoma with minimal toxicity.

  19. Radical intermediate generation and cell cycle arrest by an aqueous extract of Thunbergia Laurifolia Linn. In human breast cancer cells.

    Science.gov (United States)

    Jetawattana, Suwimol; Boonsirichai, Kanokporn; Charoen, Savapong; Martin, Sean M

    2015-01-01

    Thunbergia Laurifolia Linn. (TL) is one of the most familiar plants in Thai traditional medicine that is used to treat various conditions, including cancer. However, the antitumor activity of TL or its constituents has never been reported at the molecular level to support the folklore claim. The present study was designed to investigate the antitumor effect of an aqueous extract of TL in human breast cancer cells and the possible mechanism(s) of action. An aqueous crude extract was prepared from dried leaves of TL. Folin-Ciocalteu colorimetric assays were used to determine the total phenolic content. Antiproliferative and cell cycle effects were evaluated in human breast adenocarcinoma MCF-7 cells by MTT reduction assay, cell growth inhibition, clonogenic cell survival, and flow cytometric analysis. Free radical generation by the extracts was detected using electron paramagnetic resonance spectroscopy. The exposure of human breast adenocarcinoma MCF-7 cells to a TL aqueous extract resulted in decreases in cell growth, clonogenic cell survival, and cell viability in a concentration-dependent manner with an IC50 value of 843 μg/ml. Treatments with extract for 24 h at 250 μg/ml or higher induced cell cycle arrest as indicated by a significant increase of cell population in the G1 phase and a significant decrease in the S phase of the cell cycle. The capability of the aqueous extract to generate radical intermediates was observed at both high pH and near-neutral pH conditions. The findings suggest the antitumor bioactivities of TL against selected breast cancer cells may be due to induction of a G1 cell cycle arrest. Cytotoxicity and cell cycle perturbation that are associated with a high concentration of the extract could be in part explained by the total phenolic contents in the extract and the capacity to generate radical intermediates to modulate cellular proliferative signals. PMID:26028099

  20. Mycobacterium tuberculosis-infected human monocytes down-regulate microglial MMP-2 secretion in CNS tuberculosis via TNFα, NFκB, p38 and caspase 8 dependent pathways

    Directory of Open Access Journals (Sweden)

    Elkington Paul T

    2011-05-01

    Full Text Available Abstract Tuberculosis (TB of the central nervous system (CNS is a deadly disease characterized by extensive tissue destruction, driven by molecules such as Matrix Metalloproteinase-2 (MMP-2 which targets CNS-specific substrates. In a simplified cellular model of CNS TB, we demonstrated that conditioned medium from Mycobacterium tuberculosis-infected primary human monocytes (CoMTb, but not direct infection, unexpectedly down-regulates constitutive microglial MMP-2 gene expression and secretion by 72.8% at 24 hours, sustained up to 96 hours (P M.tb-infected monocyte-dependent networks paradoxically involves the pro-inflammatory mediators TNF-α, p38 MAP kinase and NFκB in addition to a novel caspase 8-dependent pathway.

  1. Human peripheral blood monocytes display surface antigens recognized by monoclonal antinuclear antibodies.

    OpenAIRE

    Holers, V.M.; Kotzin, B L

    1985-01-01

    We used monoclonal anti-nuclear autoantibodies and indirect immunofluorescence to examine normal human peripheral blood mononuclear leukocytes for the presence of cell surface nuclear antigens. Only one monoclonal anti-histone antibody (MH-2) was found to bind to freshly isolated PBL, staining approximately 10% of large cells. However, after cells were placed into culture for 16-24 h, a high percentage (up to 60%) of large-sized cells were recognized by an anti-DNA (BWD-1) and several differe...

  2. Epigenome-wide effects of vitamin D and their impact on the transcriptome of human monocytes involve CTCF.

    Science.gov (United States)

    Seuter, Sabine; Neme, Antonio; Carlberg, Carsten

    2016-05-19

    The physiological functions of vitamin D are mediated by its metabolite 1α,25-dihydroxyvitamin D3 (1,25(OH)2D3) activating the transcription factor vitamin D receptor (VDR). In THP-1 human monocytes we demonstrated epigenome-wide effects of 1,25(OH)2D3 at 8979 loci with significantly modulated chromatin accessibility. Maximal chromatin opening was observed after 24 h, while after 48 h most sites closed again. The chromatin-organizing protein CTCF bound to 14% of the 1,25(OH)2D3-sensitive chromatin regions. Interestingly, 1,25(OH)2D3 affected the chromatin association of CTCF providing an additional mechanism for the epigenome-wide effects of the VDR ligand. The 1,25(OH)2D3-modulated transcriptome of THP-1 cells comprised 1284 genes, 77.5% of which responded only 24 h after stimulation. During the 1,25(OH)2D3 stimulation time course the proportion of down-regulated genes increased from 0% to 44.9% and the top-ranking physiological function of the respective genes shifted from anti-microbial response to connective tissue disorders. The integration of epigenomic and transcriptomic data identified 165 physiologically important 1,25(OH)2D3 target genes, including HTT and NOD2, whose expression can be predicted primarily from epigenomic data of their genomic loci. Taken together, a large number of 1,25(OH)2D3-triggered epigenome-wide events precede and accompany the transcriptional activation of target genes of the nuclear hormone. PMID:26715761

  3. International news coverage of human trafficking arrests and prosecutions: a content analysis

    OpenAIRE

    Denton, Erin

    2009-01-01

    Trafficking in human beings is a growing phenomenon with an expanding body of literature. However, a gap is evident in this body of literature: original research focusing on specific incidents of human trafficking. At this time, the human trafficking literature is permeated with discussions of the sex trade, sexual exploitation, organized crime, global socioeconomic problems and human rights. While these issues are all pertinent to the human trafficking debate, the literature requires maturat...

  4. Human platelet Fc receptor for immunoglobulin G. Identification as a 40,000-molecular-weight membrane protein shared by monocytes.

    OpenAIRE

    Rosenfeld, S I; Looney, R J; Leddy, J P; Phipps, D C; Abraham, G N; Anderson, C L

    1985-01-01

    We have recently shown that human monocytes and U937 cells possess two molecular classes of Fc gamma receptor. One, a 72,000-mol-wt sialoglycoprotein, has high affinity for certain subclasses of human and murine monomeric IgG. The other is a 40,000-mol-wt protein (p40) with low affinity for monomeric IgG but with the capacity to bind IgG aggregates or IgG-coated particles. In the present study, a 40,000-mol-wt single chain protein, apparently identical to p40 from U937 cells, was isolated fro...

  5. Ganoderma lucidum polysaccharides in human monocytic leukemia cells: from gene expression to network construction

    Directory of Open Access Journals (Sweden)

    Ou Chern-Han

    2007-11-01

    Full Text Available Abstract Background Ganoderma lucidum has been widely used as a herbal medicine for promoting health and longevity in China and other Asian countries. Polysaccharide extracts from Ganoderma lucidum have been reported to exhibit immuno-modulating and anti-tumor activities. In previous studies, F3, the active component of the polysaccharide extract, was found to activate various cytokines such as IL-1, IL-6, IL-12, and TNF-α. This gave rise to our investigation on how F3 stimulates immuno-modulating or anti-tumor effects in human leukemia THP-1 cells. Results Here, we integrated time-course DNA microarray analysis, quantitative PCR assays, and bioinformatics methods to study the F3-induced effects in THP-1 cells. Significantly disturbed pathways induced by F3 were identified with statistical analysis on microarray data. The apoptosis induction through the DR3 and DR4/5 death receptors was found to be one of the most significant pathways and play a key role in THP-1 cells after F3 treatment. Based on time-course gene expression measurements of the identified pathway, we reconstructed a plausible regulatory network of the involved genes using reverse-engineering computational approach. Conclusion Our results showed that F3 may induce death receptor ligands to initiate signaling via receptor oligomerization, recruitment of specialized adaptor proteins and activation of caspase cascades.

  6. Resveratrol oligomers isolated from Carex species inhibit growth of human colon tumorigenic cells mediated by cell cycle arrest.

    Science.gov (United States)

    González-Sarrías, Antonio; Gromek, Samantha; Niesen, Daniel; Seeram, Navindra P; Henry, Geneive E

    2011-08-24

    Research has shown that members of the Carex genus produce biologically active stilbenoids including resveratrol oligomers. This is of great interest to the nutraceutical industry given that resveratrol, a constituent of grape and red wine, has attracted immense research attention due to its potential human health benefits. In the current study, five resveratrol oligomers (isolated from Carex folliculata and Carex gynandra ), along with resveratrol, were evaluated for antiproliferative effects against human colon cancer (HCT-116, HT-29, Caco-2) and normal human colon (CCD-18Co) cells. The resveratrol oligomers included one dimer, two trimers, and two tetramers: pallidol (1); α-viniferin (2) and trans-miyabenol C (3); and kobophenols A (4) and B (5), respectively. Although not cytotoxic, the resveratrol oligomers (1-5), as well as resveratrol, inhibited growth of the human colon cancer cells. Among the six stilbenoids, α-viniferin (2) was most active against the colon cancer cells with IC(50) values of 6-32 μM (>2-fold compared to normal colon cells). Moreover, α-viniferin (at 20 μM) did not induce apoptosis but arrested cell cycle (in the S-phase) for the colon cancer but not the normal colon cells. This study adds to the growing body of knowledge supporting the anticancer effects of resveratrol and its oligomers. Furthermore, Carex species should be investigated for their nutraceutical potential given that they produce biologically active stilbenoids such as α-viniferin. PMID:21761862

  7. Human monocyte-derived dendritic cells expressing both chemotactic cytokines IL-8, MCP-1, RANTES and their receptors,and their selective migration to these chemokines

    Institute of Scientific and Technical Information of China (English)

    2000-01-01

    Objective To characterize the mRNA expression of CXC chemokine IL-8, CC chemokine monocyte chemothractant protein-1 (MCP-1) and regulated on activation,normal T cell expressed and secreted (RANTES), and a newly defined DC chemokine DC- CK1 as well as the expression of IL-8 receptor, MCP-1 receptor and RANTES receptor in human monocyte derived dendritic cells (MoDCs).The migratory responsiveness of MoDC to IL-8, MCP-1 and RANTES was alsso studied. Methods In vitro generated MoDCs were obtained by differentiating monocytes in the presence of GM-CSF and IL-4 for 5 days. The time course of RNA expression was analyzed by RT-PCR and migratoly ability was assessed by a micromultiwell chemotaxis chamber assay. Results IL-8, MCP-1, RANTES and their corres ponding receptors were consistently expressed in MoDCs. DC-CK-1 expression was detectable efter 48 hours of differentiation. MoDC selectively migrated in response to MCP-1 and RANTES but not to IL-8 though transcripts of IL-8 receptor were present. Conclusion Because the capacity of dendritic cells to initiate immune responses depends on their specialized migratory and tissue homing properties, the expression of chemokines and their receptors along with the migratory responsiveness to chemokines of MoDC in our study suggests a potential role of chemokines in the interaction between dendritic cells and T cells and the induction of immune responses.

  8. Development of an osteoblast/osteoclast co-culture derived by human bone marrow stromal cells and human monocytes for biomaterials testing

    Directory of Open Access Journals (Sweden)

    H Worch

    2011-01-01

    Full Text Available The communication of bone-forming osteoblasts and bone-resorbing osteoclasts is a fundamental requirement for balanced bone remodelling. For biomaterial research, development of in vitro models is necessary to investigate this communication. In the present study human bone marrow stromal cells and human monocytes were cultivated in order to differentiate into osteoblasts and osteoclasts, respectively. Finally, a cultivation regime was identified which firstly induces the differentiation of the human bone marrow stromal cells followed by the induction of osteoclastogenesis through the osteoblasts formed – without the external addition of the factors RANKL and M-CSF. As a feedback on osteoblasts enhanced gene expression of BSP II was detected for modifications which facilitated the formation of large multinuclear osteoclasts. Phenotype characterization was performed by biochemical methods (DNA, LDH, ALP, TRAP 5b, gene expression analysis (ALP, BSP II, RANKL, IL-6, VTNR, CTSK, TRAP, OSCAR, CALCR as well as light microscopy, confocal laser scanning microscopy, and scanning electron microscopy. After establishing this model on polystyrene, similar positive results were obtained for cultivation on a relevant bone substitution material – a composite xerogel of silica, collagen, and calcium phosphate.

  9. Phagosomal signaling by Borrelia burgdorferi in human monocytes involves Toll-like receptor (TLR) 2 and TLR8 cooperativity and TLR8-mediated induction of IFN-β

    Science.gov (United States)

    Cervantes, Jorge L.; Dunham-Ems, Star M.; La Vake, Carson J.; Petzke, Mary M.; Sahay, Bikash; Sellati, Timothy J.; Radolf, Justin D.; Salazar, Juan C.

    2011-01-01

    Phagocytosed Borrelia burgdorferi (Bb) induces inflammatory signals that differ both quantitatively and qualitatively from those generated by spirochetal lipoproteins interacting with Toll-like receptor (TLR) 1/2 on the surface of human monocytes. Of particular significance, and in contrast to lipoproteins, internalized spirochetes induce transcription of IFN-β. Using inhibitory immunoregulatory DNA sequences (IRSs) specific to TLR7, TLR8, and TLR9, we show that the TLR8 inhibitor IRS957 significantly diminishes production of TNF-α, IL-6, and IL-10 and completely abrogates transcription of IFN-β in Bb-stimulated monocytes. We demonstrate that live Bb induces transcription of TLR2 and TLR8, whereas IRS957 interferes with their transcriptional regulation. Using confocal and epifluorescence microscopy, we show that baseline TLR expression in unstimulated monocytes is greater for TLR2 than for TLR8, whereas expression of both TLRs increases significantly upon stimulation with live spirochetes. By confocal microscopy, we show that TLR2 colocalization with Bb coincides with binding, uptake, and formation of the phagosomal vacuole, whereas recruitment of both TLR2 and TLR8 overlaps with degradation of the spirochete. We provide evidence that IFN regulatory factor (IRF) 7 is translocated into the nucleus of Bb-infected monocytes, suggesting its activation through phosphorylation. Taken together, these findings indicate that the phagosome is an efficient platform for the recognition of diverse ligands; in the case of Bb, phagosomal signaling involves a cooperative interaction between TLR2 and TLR8 in pro- and antiinflammatory cytokine responses, whereas TLR8 is solely responsible for IRF7-mediated induction of IFN-β. PMID:21321205

  10. Inactivation of normal human fibroblasts by ionising irradiation results to a similar extent from chromosomal damage and p53-dependent G1-arrest

    International Nuclear Information System (INIS)

    After ionizing irradiation, fibroblasts lose clonogenicity (1) by non-repaired DNA double-strand breaks leading to lethal chromosome aberrations and (2) by permanent G1 arrest. The aim of this study was to determine the relative contribution of these two processes. 13 normal human fibroblast strains and 3 cell lines with non-functional p53 (LFS2800, FaDu, CHO). Cells were irradiated in plateau phase followed by immediate or delayed (14 h) plating. Lethal chromosome aberrations (CA) were measured by metaphase technique, the fraction of cells permanently arrested in G1 (fG1arr) by flow cytometry and cell survival by colony assay. For normal human fibroblasts, the number of lethal chromosome aberrations increased with dose but varied substantially among the strains studied. Only for delayed but not immediate plating the surviving fraction was correlated with the number of lethal aberrations (r2 =0.69, p2 =0.19, p=0.16). When survival was converted into lethal events the ratio between these events and the number of lethal aberrations amounted to 2.00±0.05:1, indicating that chromosomal damage accounted on average for only 50% of cell killing. The remainder was attributed to cell inactivation by the p53-dependent permanent G1-arrest, since cells lacking in functional p53 (LFS2800, FaDu, CHO) were characterised by a ratio of 1.01±0.02:1. In addition, there was a negative correlation between the extent of G1-arrest and the number of CA with those cell lines showing the highest G1-arrest having the lowest number of CA indicating that there is an interaction between these two processes. For normal human fibroblasts, cell inactivation results from chromosomal damage and permanent G1 arrest to a similar extent

  11. The innate pattern recognition molecule Ficolin-1 is secreted by monocytes/macrophages and is circulating in human plasma

    DEFF Research Database (Denmark)

    Honoré, Christian; Rørvig, Sara; Munthe-Fog, Lea; Hummelshøj, Tina; Madsen, Hans O; Borregaard, Niels; Garred, Peter

    2008-01-01

    and SDS-PAGE/western blot. Secretion of Ficolin-1 was investigated in cells and plasma from healthy donors through affinity purification using N-acetyl-d-glucosamine-agarose beads and ELISA. Ficolin-1 was found differentially expressed and synthesised by monocytes, macrophages and immature dendritic...

  12. Arecoline induced cell cycle arrest, apoptosis, and cytotoxicity to human endothelial cells.

    Science.gov (United States)

    Tseng, Shuei-Kuen; Chang, Mei-Chi; Su, Cheng-Yao; Chi, Lin-Yang; Chang, Jenny Zwei-Ching; Tseng, Wan-Yu; Yeung, Sin-Yuet; Hsu, Ming-Lun; Jeng, Jiiang-Huei

    2012-08-01

    Betel quid (BQ) chewing is a common oral habit in South Asia and Taiwan. BQ consumption may increase the risk of oral squamous cell carcinoma (OSCC), oral submucous fibrosis (OSF), and periodontitis as well as systemic diseases (atherosclerosis, hypertension, etc.). However, little is known about the toxic effect of BQ components on endothelial cells that play important roles for angiogenesis, carcinogenesis, tissue fibrosis, and cardiovascular diseases. EAhy 926 (EAHY) endothelial cells were exposed to arecoline, a major BQ alkaloid, for various time periods. Cytotoxicity was estimated by 3-(4, 5- dimethylthiazol-2-yl)-2, 5-diphenyl tetrazolium bromide assay. The cell cycle distribution of EAHY cells residing in sub-G0/G1, G0/G1, S-, and G2/M phases was analyzed by propidium iodide staining of cellular DNA content and flow cytometry. Some EAHY cells retracted, became round-shaped in appearance, and even detached from the culture plate after exposure to higher concentrations of arecoline (> 0.4 mM). At concentrations of 0.4 and 0.8 mM, arecoline induced significant cytotoxicity to EAHY cells. At similar concentrations, arecoline induced G2/M cell cycle arrest and increased sub-G0/G1 population, a hallmark of apoptosis. Interestingly, prolonged exposure to arecoline (0.1 mM) for 12 and 21 days significantly suppressed the proliferation of EAHY cells, whereas EAHY cells showed adaptation and survived when exposed to 0.05 mM arecoline. These results suggest that BQ components may contribute to the pathogenesis of OSF and BQ chewing-related cardiovascular diseases via toxicity to oral or systemic endothelial cells, leading to impairment of vascular function. During BQ chewing, endothelial damage may be induced by areca nut components and associate with the pathogenesis of OSF, periodontitis, and cardiovascular diseases. PMID:21847594

  13. Supercritical carbon dioxide extract of Physalis peruviana induced cell cycle arrest and apoptosis in human lung cancer H661 cells.

    Science.gov (United States)

    Wu, Shu-Jing; Chang, Shun-Pang; Lin, Doung-Liang; Wang, Shyh-Shyan; Hou, Fwu-Feuu; Ng, Lean-Teik

    2009-06-01

    Physalis peruviana L. (PP) is a popular folk medicine used for treating cancer, leukemia, hepatitis, rheumatism and other diseases. In this study, our objectives were to examine the total flavonoid and phenol content of different PP extracts (aqueous: HWEPP; ethanolic: EEPP; supercritical carbon dioxide: SCEPP-0, SCEPP-4 and SCEPP-5) and their antiproliferative effects in human lung cancer H661 cells. Among all the extracts tested, results showed that SCEPP-5 possessed the highest total flavonoid (226.19 +/- 4.15 mg/g) and phenol (100.82 +/- 6.25 mg/g) contents. SCEPP-5 also demonstrated the most potent inhibitory effect on H661 cell proliferation. Using DNA ladder and flow cytometry analysis, SCEPP-5 effectively induced H661 cell apoptosis as demonstrated by the accumulation of Sub-G1 peak and fragmentation of DNA. SCEPP-5 not only induced cell cycle arrest at S phase, it also up-regulated the expression of pro-apoptotic protein (Bax) and down-regulated the inhibitor of apoptosis protein (IAP). Furthermore, the apoptotic induction in H661 cells was found to associate with an elevated p53 protein expression, cytochrome c release, caspase-3 activation and PARP cleavage. Taken together, these results conclude that SCEPP-5 induced cell cycle arrest at S phase, and its apoptotic induction could be mediated through the p53-dependent pathway and modification of Bax and XIAP proteins expression. The results have also provided important pharmacological backgrounds for the potential use of PP supercritical fluid extract as products for cancer prevention. PMID:19425186

  14. Nuclear import, virion incorporation, and cell cycle arrest/differentiation are mediated by distinct functional domains of human immunodeficiency virus type 1 Vpr.

    OpenAIRE

    Mahalingam, S.; Ayyavoo, V; M. Patel; Kieber-Emmons, T; Weiner, D B

    1997-01-01

    The vpr gene product of human immunodeficiency virus type 1 (HIV-1) is a virion-associated protein that is essential for efficient viral replication in monocytes/macrophages. Vpr is primarily localized in the nucleus when expressed in the absence of other viral proteins. Vpr is packaged efficiently into viral particles through interactions with the p6 domain of the Gag precursor polyprotein p55gag. We developed a panel of expression vectors encoding Vpr molecules mutated in the amino-terminal...

  15. Functional role of CD11c+ monocytes in atherogenesis associated with hypercholesterolemia

    Science.gov (United States)

    Monocyte activation and migration into the arterial wall are key events in atherogenesis associated with hypercholesterolemia. CD11c/CD18, a beta2 integrin expressed on human monocytes and a subset of mouse monocytes, has been shown to play a distinct role in human monocyte adhesion on endothelial c...

  16. Nonclassical patrolling monocyte function in the vasculature.

    Science.gov (United States)

    Thomas, Graham; Tacke, Robert; Hedrick, Catherine C; Hanna, Richard N

    2015-06-01

    Nonclassical patrolling monocytes are characterized by their unique ability to actively patrol the vascular endothelium under homeostatic and inflammatory conditions. Patrolling monocyte subsets (CX3CR1(high)Ly6C(-) in mouse and CX3CR1(high)CD14(dim)CD16(+) in humans) are distinct from the classical monocyte subsets (CCR2(high)Ly6C(+) in mouse and CCR2(high)CD14(+)CD16(-) in humans) and exhibit unique functions in the vasculature and inflammatory disease. Patrolling monocytes function in several disease settings to remove damaged cells and debris from the vasculature and have been associated with wound healing and the resolution of inflammation in damaged tissues. This review highlights the unique functions of these patrolling monocytes in the vasculature and during inflammation. PMID:25838429

  17. Mannose-binding lectin inhibits monocyte proliferation through transforming growth factor-β1 and p38 signaling pathways.

    Directory of Open Access Journals (Sweden)

    Yan Wang

    Full Text Available Mannose-binding lectin (MBL, a plasma C-type lectin, plays an important role in innate immunity. However, the interaction, and the consequences of it, between MBL and the immune system remain ill defined. We have investigated the contributing mechanisms and effects of MBL on the proliferation of human monocytes. At lower concentrations (≤4 μg/ml MBL was shown to partially enhance monocyte proliferation. By contrast, at higher concentrations (8-20 μg/ml of MBL, cell proliferation was markedly attenuated. MBL-induced growth inhibition was associated with G0/G1 arrest, down-regulation of cyclin D1/D3, cyclin-dependent kinase (Cdk 2/Cdk4 and up-regulation of the Cdk inhibitory protein Cip1/p21. Additionally, MBL induced apoptosis, and did so through caspase-3 activation and poly ADP-ribose polymerase (PARP cleavage. Moreover, transforming growth factor (TGF-β1 levels increased in the supernatants of MBL-stimulated monocyte cultures. We also found that MBL-dependent inhibition of monocyte proliferation could be reversed by the TGF-β receptor antagonist SB-431542, or by anti-TGF-β1 antibody, or by the mitogen-activated protein kinase (MAPK inhibitors specific for p38 (SB203580, but not ERK (U0126 or JNK (SP600125. Thus, at high concentrations, MBL can affect the immune system by inhibiting monocyte proliferation, which suggests that MBL may exhibit anti-inflammatory effects.

  18. Morphine Reciprocally Regulates IL-10 and IL-12 Production by Monocyte-Derived Human Dendritic Cells and Enhances T Cell Activation

    OpenAIRE

    Messmer, Davorka; Hatsukari, Ikusuke; Hitosugi, Naoko; Schmidt-Wolf, Ingo GH; Singhal, Pravin C.

    2006-01-01

    We evaluated the effect of morphine on human dendritic cells (DCs). Interestingly, immature DCs were found to express all 3 (μ, κ, δ) opioid receptors on the cell surface. Chronic morphine treatment (10−8 to 10−12 M) during the development of DCs from monocytes augmented LPS-induced upregulation of HLA-DR, CD86, CD80, and CD83 and increased the T cell stimulatory capacity of DCs, which could be inhibited by naloxone, an opioid receptor antagonist. The change in surface phenotype was parallele...

  19. Platelet-derived growth factor (PDGF-BB-mediated induction of monocyte chemoattractant protein 1 in human astrocytes: implications for HIV-associated neuroinflammation

    Directory of Open Access Journals (Sweden)

    Bethel-Brown Crystal

    2012-12-01

    Full Text Available Abstract Chemokine (C-C motif ligand 2, also known as monocyte chemoattractant protein 1 (MCP-1 is an important factor for the pathogenesis of HIV-associated neurocognitive disorders (HAND. The mechanisms of MCP-1-mediated neuropathogenesis, in part, revolve around its neuroinflammatory role and the recruitment of monocytes into the central nervous system (CNS via the disrupted blood-brain barrier (BBB. We have previously demonstrated that HIV-1/HIV-1 Tat upregulate platelet-derived growth factor (PDGF-BB, a known cerebrovascular permeant; subsequently, the present study was aimed at exploring the regulation of MCP-1 by PDGF-BB in astrocytes with implications in HAND. Specifically, the data herein demonstrate that exposure of human astrocytes to HIV-1 LAI elevated PDGF-B and MCP-1 levels. Furthermore, treating astrocytes with the human recombinant PDGF-BB protein significantly increased the production and release of MCP-1 at both the RNA and protein levels. MCP-1 induction was regulated by activation of extracellular-signal-regulated kinase (ERK1/2, c-Jun N-terminal kinase (JNK and p38 mitogen-activated protein (MAP kinases and phosphatidylinositol 3-kinase (PI3K/Akt pathways and the downstream transcription factor, nuclear factor κB (NFκB. Chromatin immunoprecipitation (ChIP assays demonstrated increased binding of NFκB to the human MCP-1 promoter following PDGF-BB exposure. Conditioned media from PDGF-BB-treated astrocytes increased monocyte transmigration through human brain microvascular endothelial cells (HBMECs, an effect that was blocked by STI-571, a tyrosine kinase inhibitor (PDGF receptor (PDGF-R blocker. PDGF-BB-mediated release of MCP-1 was critical for increased permeability in an in vitro BBB model as evidenced by blocking antibody assays. Since MCP-1 is linked to disease severity, understanding its modulation by PDGF-BB could aid in understanding the proinflammatory responses in HAND. These results suggest that astrocyte

  20. Growth arrest and apoptosis of human hepatocellular carcinoma cells induced by hexamethylene bisacetamide

    OpenAIRE

    Ouyang, Gao-Liang; Cai, Qiu-Feng; Min LIU; Chen, Rui-Chuan; Huang, Zhi; Jiang, Rui-Sheng; Chen, Fu; Hong, Shui-Gen; Bao, Shi-Deng

    2004-01-01

    AIM: To investigate the cellular effects of hybrid polar compound hexamethylene bisacetamide (HMBA) on the growth and apoptosis of human hepatocellular carcinoma cells and to provide the molecular mechanism for potential application of HMBA in the treatment of liver cancer.

  1. H4 histamine receptors mediate cell cycle arrest in growth factor-induced murine and human hematopoietic progenitor cells.

    Directory of Open Access Journals (Sweden)

    Anne-France Petit-Bertron

    Full Text Available The most recently characterized H4 histamine receptor (H4R is expressed preferentially in the bone marrow, raising the question of its role during hematopoiesis. Here we show that both murine and human progenitor cell populations express this receptor subtype on transcriptional and protein levels and respond to its agonists by reduced growth factor-induced cell cycle progression that leads to decreased myeloid, erythroid and lymphoid colony formation. H4R activation prevents the induction of cell cycle genes through a cAMP/PKA-dependent pathway that is not associated with apoptosis. It is mediated specifically through H4R signaling since gene silencing or treatment with selective antagonists restores normal cell cycle progression. The arrest of growth factor-induced G1/S transition protects murine and human progenitor cells from the toxicity of the cell cycle-dependent anticancer drug Ara-C in vitro and reduces aplasia in a murine model of chemotherapy. This first evidence for functional H4R expression in hematopoietic progenitors opens new therapeutic perspectives for alleviating hematotoxic side effects of antineoplastic drugs.

  2. Resibufogenin Induces G1-Phase Arrest through the Proteasomal Degradation of Cyclin D1 in Human Malignant Tumor Cells.

    Directory of Open Access Journals (Sweden)

    Masami Ichikawa

    Full Text Available Huachansu, a traditional Chinese medicine prepared from the dried toad skin, has been used in clinical studies for various cancers in China. Resibufogenin is a component of huachansu and classified as bufadienolides. Resibufogenin has been shown to exhibit the anti-proliferative effect against cancer cells. However, the molecular mechanism of resibufogenin remains unknown. Here we report that resibufogenin induces G1-phase arrest with hypophosphorylation of retinoblastoma (RB protein and down-regulation of cyclin D1 expression in human colon cancer HT-29 cells. Since the down-regulation of cyclin D1 was completely blocked by a proteasome inhibitor MG132, the suppression of cyclin D1 expression by resibufogenin was considered to be in a proteasome-dependent manner. It is known that glycogen synthase kinase-3β (GSK-3β induces the proteasomal degradation of cyclin D1. The addition of GSK-3β inhibitor SB216763 inhibited the reduction of cyclin D1 caused by resibufogenin. These effects on cyclin D1 by resibufogenin were also observed in human lung cancer A549 cells. These findings suggest that the anti-proliferative effect of resibufogenin may be attributed to the degradation of cyclin D1 caused by the activation of GSK-3β.

  3. Phytometabolite Dehydroleucodine Induces Cell Cycle Arrest, Apoptosis, and DNA Damage in Human Astrocytoma Cells through p73/p53 Regulation.

    Directory of Open Access Journals (Sweden)

    Natalia Bailon-Moscoso

    Full Text Available Accumulating evidence supports the idea that secondary metabolites obtained from medicinal plants (phytometabolites may be important contributors in the development of new chemotherapeutic agents to reduce the occurrence or recurrence of cancer. Our study focused on Dehydroleucodine (DhL, a sesquiterpene found in the provinces of Loja and Zamora-Chinchipe. In this study, we showed that DhL displayed cytostatic and cytotoxic activities on the human cerebral astrocytoma D384 cell line. With lactone isolated from Gynoxys verrucosa Wedd, a medicinal plant from Ecuador, we found that DhL induced cell death in D384 cells by triggering cell cycle arrest and inducing apoptosis and DNA damage. We further found that the cell death resulted in the increased expression of CDKN1A and BAX proteins. A marked induction of the levels of total TP73 and phosphorylated TP53, TP73, and γ-H2AX proteins was observed in D384 cells exposed to DhL, but no increase in total TP53 levels was detected. Overall these studies demonstrated the marked effect of DhL on the diminished survival of human astrocytoma cells through the induced expression of TP73 and phosphorylation of TP73 and TP53, suggesting their key roles in the tumor cell response to DhL treatment.

  4. Silencing of AP-4 inhibits proliferation, induces cell cycle arrest and promotes apoptosis in human lung cancer cells

    Science.gov (United States)

    HU, XUANYU; GUO, WEI; CHEN, SHANSHAN; XU, YIZHUO; LI, PING; WANG, HUAQI; CHU, HEYING; LI, JUAN; DU, YUWEN; CHEN, XIAONAN; ZHANG, GUOJUN; ZHAO, GUOQIANG

    2016-01-01

    Activating enhancer-binding protein (AP)-4 is a member of the basic helix-loop-helix transcription factors, and is involved in tumor biology. However, the role of AP-4 in human lung cancer remains to be fully elucidated. In the present study, the expression of AP-4 in human lung cancer tissues and cells was investigated by reverse transcription-quantitative polymerase chain reaction, and it was observed that the level of AP-4 was increased in tumor tissues and cells compared with their normal counterparts. AP-4 expression was knocked down by transfection with a specific small interfering RNA (siRNA) in lung cancer cells, and this indicated that siRNA-mediated silencing of AP-4 inhibited cell proliferation, arrested the cell cycle at the G0/G1 phase and induced apoptosis by modulating the expression of p21 and cyclin D1. The results of the present study suggest that AP-4 may be an oncoprotein that has a significant role in lung cancer, and that siRNA-mediated silencing of AP-4 may have therapeutic potential as a strategy for the treatment of lung cancer.

  5. Piperlongumine Suppresses Proliferation of Human Oral Squamous Cell Carcinoma through Cell Cycle Arrest, Apoptosis and Senescence

    OpenAIRE

    San-Yuan Chen; Geng-Hung Liu; Wen-Ying Chao; Chung-Sheng Shi; Ching-Yen Lin; Yun-Ping Lim; Chieh-Hsiang Lu; Peng-Yeh Lai; Hau-Ren Chen; Ying-Ray Lee

    2016-01-01

    Oral squamous cell carcinoma (OSCC), an aggressive cancer originating in the oral cavity, is one of the leading causes of cancer deaths in males worldwide. This study investigated the antitumor activity and mechanisms of piperlongumine (PL), a natural compound isolated from Piper longum L., in human OSCC cells. The effects of PL on cell proliferation, the cell cycle, apoptosis, senescence and reactive oxygen species (ROS) levels in human OSCC cells were investigated. PL effectively inhibited ...

  6. 3,3'-Diindolylmethane induces G1 arrest and apoptosis in human acute T-cell lymphoblastic leukemia cells.

    Directory of Open Access Journals (Sweden)

    Lyndsey E Shorey

    Full Text Available Certain bioactive food components, including indole-3-carbinol (I3C and 3,3'-diindolylmethane (DIM from cruciferous vegetables, have been shown to target cellular pathways regulating carcinogenesis. Previously, our laboratory showed that dietary I3C is an effective transplacental chemopreventive agent in a dibenzo[def,p]chrysene (DBC-dependent model of murine T-cell lymphoblastic lymphoma. The primary objective of the present study was to extend our chemoprevention studies in mice to an analogous human neoplasm in cell culture. Therefore, we tested the hypothesis that I3C or DIM may be chemotherapeutic in human T-cell acute lymphoblastic leukemia (T-ALL cells. Treatment of the T-ALL cell lines CCRF-CEM, CCRF-HSB2, SUP-T1 and Jurkat with DIM in vitro significantly reduced cell proliferation and viability at concentrations 8- to 25-fold lower than the parent compound I3C. DIM (7.5 µM arrested CEM and HSB2 cells at the G(1 phase of the cell cycle and 15 µM DIM significantly increased the percentage of apoptotic cells in all T-ALL lines. In CEM cells, DIM reduced protein expression of cyclin dependent kinases 4 and 6 (CDK4, CDK6 and D-type cyclin 3 (CCND3; DIM also significantly altered expression of eight transcripts related to human apoptosis (BCL2L10, CD40LG, HRK, TNF, TNFRSF1A, TNFRSF25, TNFSF8, TRAF4. Similar anticancer effects of DIM were observed in vivo. Dietary exposure to 100 ppm DIM significantly decreased the rate of growth of human CEM xenografts in immunodeficient SCID mice, reduced final tumor size by 44% and increased the apoptotic index compared to control-fed mice. Taken together, our results demonstrate a potential for therapeutic application of DIM in T-ALL.

  7. Plectasin shows intracellular activity against Staphylococcus aureus in human THP-1 monocytes and in a mouse peritonitis model

    DEFF Research Database (Denmark)

    Brinch, Karoline Sidelmann; Sandberg, Anne; Baudoux, Pierre; Van Bambeke, Françoise; Tulkens, Paul M; Frimodt-Møller, Niels; Hoiby, Niels; Kristensen, Hans-Henrik

    2009-01-01

    Antimicrobial therapy of infections with Staphylococcus aureus can pose a challenge due to slow response to therapy and recurrence of infection. These treatment difficulties can partly be explained by intracellular survival of staphylococci, which is why the intracellular activity of...... antistaphylococcal compounds has received increased attention within recent years. The intracellular activity of plectasin, an antimicrobial peptide, against S. aureus was determined both in vitro and in vivo. In vitro studies using THP-1 monocytes showed that some intracellular antibacterial activity of plectasin...

  8. Arecoline decreases interleukin-6 production and induces apoptosis and cell cycle arrest in human basal cell carcinoma cells

    International Nuclear Information System (INIS)

    Arecoline, the most abundant areca alkaloid, has been reported to decrease interleukin-6 (IL-6) levels in epithelial cancer cells. Since IL-6 overexpression contributes to the tumorigenic potency of basal cell carcinoma (BCC), this study was designed to investigate whether arecoline altered IL-6 expression and its downstream regulation of apoptosis and the cell cycle in cultured BCC-1/KMC cells. BCC-1/KMC cells and a human keratinocyte cell line, HaCaT, were treated with arecoline at concentrations ranging from 10 to 100 μg/ml, then IL-6 production and expression of apoptosis- and cell cycle progress-related factors were examined. After 24 h exposure, arecoline inhibited BCC-1/KMC cell growth and decreased IL-6 production in terms of mRNA expression and protein secretion, but had no effect on HaCaT cells. Analysis of DNA fragmentation and chromatin condensation showed that arecoline induced apoptosis of BCC-1/KMC cells in a dose-dependent manner, activated caspase-3, and decreased expression of the anti-apoptotic protein Bcl-2. In addition, arecoline induced progressive and sustained accumulation of BCC-1/KMC cells in G2/M phase as a result of reducing checkpoint Cdc2 activity by decreasing Cdc25C phosphatase levels and increasing p53 levels. Furthermore, subcutaneous injection of arecoline led to decreased BCC-1/KMC tumor growth in BALB/c mice by inducing apoptosis. This study demonstrates that arecoline has potential for preventing BCC tumorigenesis by reducing levels of the tumor cell survival factor IL-6, increasing levels of the tumor suppressor factor p53, and eliciting cell cycle arrest, followed by apoptosis. Highlights: ► Arecoline has potential to prevent against basal cell carcinoma tumorigenesis. ► It has more effectiveness on BCC as compared with a human keratinocyte cell line. ► Mechanisms involved including reducing tumor cells’ survival factor IL-6, ► Decreasing Cdc25C phosphatase, enhancing tumor suppressor factor p53, ► Eliciting G2/M

  9. Arecoline decreases interleukin-6 production and induces apoptosis and cell cycle arrest in human basal cell carcinoma cells

    Energy Technology Data Exchange (ETDEWEB)

    Huang, Li-Wen [Department of Medical Laboratory Science and Biotechnology, Kaohsiung Medical University, Kaohsiung 80708, Taiwan (China); Hsieh, Bau-Shan; Cheng, Hsiao-Ling [Department of Biochemistry, Faculty of Medicine, College of Medicine, Kaohsiung Medical University, Kaohsiung 80708, Taiwan (China); Hu, Yu-Chen [Graduate Institute of Medicine, College of Medicine, Kaohsiung Medical University, Kaohsiung 80708, Taiwan (China); Chang, Wen-Tsan [Graduate Institute of Medicine, College of Medicine, Kaohsiung Medical University, Kaohsiung 80708, Taiwan (China); Division of Hepatobiliarypancreatic Surgery, Department of Surgery, Kaohsiung Medical University Hospital, Kaohsiung 80708, Taiwan (China); Chang, Kee-Lung, E-mail: Chang.KeeLung@msa.hinet.net [Department of Biochemistry, Faculty of Medicine, College of Medicine, Kaohsiung Medical University, Kaohsiung 80708, Taiwan (China); Graduate Institute of Medicine, College of Medicine, Kaohsiung Medical University, Kaohsiung 80708, Taiwan (China)

    2012-01-15

    Arecoline, the most abundant areca alkaloid, has been reported to decrease interleukin-6 (IL-6) levels in epithelial cancer cells. Since IL-6 overexpression contributes to the tumorigenic potency of basal cell carcinoma (BCC), this study was designed to investigate whether arecoline altered IL-6 expression and its downstream regulation of apoptosis and the cell cycle in cultured BCC-1/KMC cells. BCC-1/KMC cells and a human keratinocyte cell line, HaCaT, were treated with arecoline at concentrations ranging from 10 to 100 μg/ml, then IL-6 production and expression of apoptosis- and cell cycle progress-related factors were examined. After 24 h exposure, arecoline inhibited BCC-1/KMC cell growth and decreased IL-6 production in terms of mRNA expression and protein secretion, but had no effect on HaCaT cells. Analysis of DNA fragmentation and chromatin condensation showed that arecoline induced apoptosis of BCC-1/KMC cells in a dose-dependent manner, activated caspase-3, and decreased expression of the anti-apoptotic protein Bcl-2. In addition, arecoline induced progressive and sustained accumulation of BCC-1/KMC cells in G2/M phase as a result of reducing checkpoint Cdc2 activity by decreasing Cdc25C phosphatase levels and increasing p53 levels. Furthermore, subcutaneous injection of arecoline led to decreased BCC-1/KMC tumor growth in BALB/c mice by inducing apoptosis. This study demonstrates that arecoline has potential for preventing BCC tumorigenesis by reducing levels of the tumor cell survival factor IL-6, increasing levels of the tumor suppressor factor p53, and eliciting cell cycle arrest, followed by apoptosis. Highlights: ► Arecoline has potential to prevent against basal cell carcinoma tumorigenesis. ► It has more effectiveness on BCC as compared with a human keratinocyte cell line. ► Mechanisms involved including reducing tumor cells’ survival factor IL-6, ► Decreasing Cdc25C phosphatase, enhancing tumor suppressor factor p53, ► Eliciting G2/M

  10. Gypenoside XLIX, a naturally occurring gynosaponin, PPAR-alpha dependently inhibits LPS-induced tissue factor expression and activity in human THP-1 monocytic cells

    International Nuclear Information System (INIS)

    Tissue factor (TF) is involved not only in the progression of atherosclerosis and other cardiovascular diseases, but is also associated with tumor growth, metastasis, and angiogenesis and hence may be an attractive target for directed cancer therapeutics. Gynostemma pentaphyllum (GP) is widely used in the treatment of various cardiovascular diseases including atherosclerosis, as well as cancers. Gypenoside (Gyp) XLIX, a dammarane-type glycoside, is one of the prominent components in GP. We have recently reported Gyp XLIX to be a potent peroxisome proliferator-activated receptor (PPAR)-alpha activator. Here we demonstrate that Gyp XLIX (0-300 μM) concentration dependently inhibited TF promoter activity after induction by the inflammatory stimulus lipopolysaccharide (LPS) in human monocytic THP-1 cells transfected with promoter reporter constructs pTF-LUC. Furthermore, Gyp XLIX inhibited LPS-induced TF mRNA and protein overexpression in THP-1 monocyte cells. Its inhibition of LPS-induced TF hyperactivity was further confirmed by chromogenic enzyme activity assay. The activities of Gyp XLIX reported in this study were similar to those of Wy-14643, a potent synthetic PPAR-alpha activator. Furthermore, the Gyp XLIX-induced inhibitory effect on TF luciferase activity was completely abolished in the presence of the PPAR-alpha selective antagonist MK-886. The present findings suggest that Gyp XLIX inhibits LPS-induced TF overexpression and enhancement of its activity in human THP-1 monocytic cells via PPAR-alpha-dependent pathways. The data provide new insights into the basis of the use of the traditional Chinese herbal medicine G. pentaphyllum for the treatment of cardiovascular and inflammatory diseases, as well as cancers

  11. Impact of 3-D printed PLA- and chitosan-based scaffolds on human monocyte/macrophage responses: unraveling the effect of 3-D structures on inflammation.

    Science.gov (United States)

    Almeida, Catarina R; Serra, Tiziano; Oliveira, Marta I; Planell, Josep A; Barbosa, Mário A; Navarro, Melba

    2014-02-01

    Recent studies have pointed towards a decisive role of inflammation in triggering tissue repair and regeneration, while at the same time it is accepted that an exacerbated inflammatory response may lead to rejection of an implant. Within this context, understanding and having the capacity to regulate the inflammatory response elicited by 3-D scaffolds aimed for tissue regeneration is crucial. This work reports on the analysis of the cytokine profile of human monocytes/macrophages in contact with biodegradable 3-D scaffolds with different surface properties, architecture and controlled pore geometry, fabricated by 3-D printing technology. Fabrication processes were optimized to create four different 3-D platforms based on polylactic acid (PLA), PLA/calcium phosphate glass or chitosan. Cytokine secretion and cell morphology of human peripheral blood monocytes allowed to differentiate on the different matrices were analyzed. While all scaffolds supported monocyte/macrophage adhesion and stimulated cytokine production, striking differences between PLA-based and chitosan scaffolds were found, with chitosan eliciting increased secretion of tumor necrosis factor (TNF)-α, while PLA-based scaffolds induced higher production of interleukin (IL)-6, IL-12/23 and IL-10. Even though the material itself induced the biggest differences, the scaffold geometry also impacted on TNF-α and IL-12/23 production, with chitosan scaffolds having larger pores and wider angles leading to a higher secretion of these pro-inflammatory cytokines. These findings strengthen the appropriateness of these 3-D platforms to study modulation of macrophage responses by specific parameters (chemistry, topography, scaffold architecture). PMID:24211731

  12. Sorafenib induces growth arrest and apoptosis of human glioblastoma cells via dephosphorylation of STAT3

    OpenAIRE

    Yang, Fan; Brown, Christine; Buettner, Ralf; HEDVAT, MICHAEL; Starr, Renate; Scuto, Anna; Schroeder, Anne; Jensen, Michael; Jove, Richard

    2010-01-01

    Glioblastoma is the most common type of primary brain tumor and is rapidly progressive with few treatment options. Here, we report that sorafenib (≤ 10 μM) inhibited cell proliferation and induced apoptosis in two established cell lines (U87, U251) and two primary cultures (PBT015, PBT022) from human glioblastomas. Effects of sorafenib on these tumor cells were associated with inhibiting phosphorylated STAT3 (Tyr705). Expression of a constitutively activated STAT3 mutant partially blocked the...

  13. Involucrin and envelope competence in human keratinocytes: Modulation by hydrocortisone, retinyl acetate and growth arrest

    OpenAIRE

    Rice, Rh; Cline, PR

    1983-01-01

    Involucrin accumulation and ionophore-assisted envelope for mation, markers of keratinocyte differentiation, were found to be highly dependent on culture conditions in the malignant epidermal keratinocyte line, SCC-13, derived from a human squamous cell carcinoma. In confluent cultures, approximately one-half of the cells were competent to form envelopes when grown in medium without hydrocortisone or retinyl acetate supplementation. Ad dition of hydrocortisone to the medi...

  14. The effects of 60Co γ-ray irradiation on the cytoskeleton of mouse peritoneal macrophages and human peripheral blood monocytes in vitro

    International Nuclear Information System (INIS)

    The whole mount cell electron microscopy in combination with selective extraction method for preparing cytoskeletal framework was applied. Cy toskeleton prepared by Triton X-100 treatment of mouse peritoneal macrophages and human peripheral blood monocytes appeared in electron microscopy as a highly organized and interconnected three-dimensional matrix of different fibrous elements. Since such cytoskeletons are open membrane-free system, individual fibrous organizations can be identified by specific antibodies. An indirect immunogold procedure using monoclonal anti-tubulin or anti-actin antibodies was applied to visualize tubulin-or actin-containing structures. The three-dimensional visualization of Triton X-100 resistant cytoskeletons had been used to demonstrate that different doses of 60Co γ-ray caused a distinctive and reproducible alterations of the cytoskeletons of intact mouse peritoneal macrophages and human peripheral blood monocytes in vitro. The results showed that there were some similar alterations with those caused by cytochalasin B and by colchicine. From these observations and other workers' studies, it's likely that 60Co γ-ray irradiation may inhibit cytoplasmic microtubule and microfilament assembling

  15. Exposure of Human Lung Cancer Cells to 8-Chloro-Adenosine Induces G2/M Arrest and Mitotic Catastrophe

    Directory of Open Access Journals (Sweden)

    Hong-Yu Zhang

    2004-11-01

    Full Text Available 8-Chloro-adenosine (8-CI-Ado is a potent chemotherapeutic agent whose cytotoxicity in a variety of tumor cell lines has been widely investigated. However, the molecular mechanisms are uncertain. In this study, we found that exposure of human lung cancer cell lines A549 (p53-wt and H1299 (p53-depleted to 8-CI-Ado induced cell arrest in the G2/M phase, which was accompanied by accumulation of binucleated and polymorphonucleated cells resulting from aberrant mitosis and failed cytokinesis. Western blotting showed the loss of phosphorylated forms of Cdc2 and Cdc25C that allowed progression into mitosis. Furthermore, the increase in Ser10-phosphorylated histone H3-positive cells revealed by fluorescence-activated cell sorting suggested that the agent-targeted cells were able to exit the G2 phase and enter the M phase. Immunocytochemistry showed that microtubule and microfilament arrays were changed in exposed cells, indicating that the dynamic instability of microtubules and microfilaments was lost, which may correlate with mitotic dividing failure. Aberrant mitosis resulted in mitotic catastrophe followed by varying degrees of apoptosis, depending on the cell lines. Thus, 8-CI-Ado appears to exert its cytotoxicity toward cells in culture by inducing mitotic catastrophe.

  16. Tocotrienol-Rich Fraction Prevents Cell Cycle Arrest and Elongates Telomere Length in Senescent Human Diploid Fibroblasts

    Directory of Open Access Journals (Sweden)

    Suzana Makpol

    2011-01-01

    Full Text Available This study determined the molecular mechanisms of tocotrienol-rich fraction (TRF in preventing cellular senescence of human diploid fibroblasts (HDFs. Primary culture of HDFs at various passages were incubated with 0.5 mg/mL TRF for 24 h. Telomere shortening with decreased telomerase activity was observed in senescent HDFs while the levels of damaged DNA and number of cells in G0/G1 phase were increased and S phase cells were decreased. Incubation with TRF reversed the morphology of senescent HDFs to resemble that of young cells with decreased activity of SA-β-gal, damaged DNA, and cells in G0/G1 phase while cells in the S phase were increased. Elongated telomere length and restoration of telomerase activity were observed in TRF-treated senescent HDFs. These findings confirmed the ability of tocotrienol-rich fraction in preventing HDFs cellular ageing by restoring telomere length and telomerase activity, reducing damaged DNA, and reversing cell cycle arrest associated with senescence.

  17. Wogonin induced G1 cell cycle arrest by regulating Wnt/β-catenin signaling pathway and inactivating CDK8 in human colorectal cancer carcinoma cells

    International Nuclear Information System (INIS)

    Highlights: • Wogonin inhibited HCT116 cells growth and arrested at G1 phase of the cell cycle. • Wogonin down-regulated the canonical Wnt/β-catenin signaling pathway. • Wogonin interfered in the combination of β-catenin and TCF/Lef. • Wogonin limited the kinase activity of CDK8. - Abstract: Wogonin, a naturally occurring mono-flavonoid, has been reported to have tumor therapeutic potential and good selectivity both in vitro and in vivo. Herein, we investigated the anti-proliferation effects and associated mechanisms of wogonin in human colorectal cancer in vitro. The flow-cytometric analysis showed that wogonin induced a G1 phase cell cycle arrest in HCT116 cells in a concentration- and time-dependent manner. Meanwhile, the cell cycle-related proteins, such as cyclin A, E, D1, and CDK2, 4 were down-regulated in wogonin-induced G1 cell cycle arrest. Furthermore, we showed that the anti-proliferation and G1 arrest effect of wogonin on HCT116 cells was associated with deregulation of Wnt/β-catenin signaling pathway. Wogonin-treated cells showed decreased intracellular levels of Wnt proteins, and activated degradation complex to phosphorylated and targeted β-catenin for proteasomal degradation. Wogonin inhibited β-catenin-mediated transcription by interfering in the transcriptional activity of TCF/Lef, and repressing the kinase activity of CDK8 which has been considered as an oncogene involving in the development of colorectal cancers. Moreover, CDK8 siRNA-transfected HCT116 cells showed similar results to wogonin treated cells. Thus, our data suggested that wogonin induced anti-proliferation and G1 arrest via Wnt/β-catenin signaling pathway and it can be developed as a therapeutic agent against human colorectal cancer

  18. Paris chinensis dioscin induces G2/M cell cycle arrest and apoptosis in human gastric cancer SGC-7901 cells

    Institute of Scientific and Technical Information of China (English)

    Lin-Lin Gao; Fu-Rong Li; Peng Jiao; Ming-Feng Yang; Xiao-Jun Zhou; Yan-Hong Si; Wen-Jian Jiang; Ting-Ting Zheng

    2011-01-01

    AIM: To investigate the anti-tumor effects of Paris chinensis dioscin (PCD) and mechanisms regarding cell cycle regulation and apoptosis in human gastric cancer SGC-7901 cells.METHODS: Cell viability was analyzed by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide assay. Cell apoptosis was evaluated by flow cytometry and laser scanning confocal microscope (LSCM) using Annexin-V/propidium iodide (PI) staining, and the cell cycle was evaluated using PI staining with flow cytometry. Intracellular calcium ions were detected under fluorescence microscope. The expression of cell cycle and apoptosis-related proteins cyclin B1, CDK1, cytochrome C and caspase-3 was measured by immunohistochemical staining. RESULTS: PCD had an anti-proliferation effect on human gastric cancer SGC-7901 cells in a dose- and time-dependent manner. After treatment of SGC-7901 cells with PCD, apoptosis appeared in SGC-7901 cells. Morphological changes typical of apoptosis were also observed with LSCM by Annexin V/PI staining, and the cell number of the G0/G1 phase was decreased, while the number of cells in the G2/M phase was increased. Cell cycle-related proteins, such as cyclin B1 and CDK1, were all down-regulated, but caspase-3 and cytochrome C were up-regulated. Moreover, intracellular calcium accumulation occurred in PCD-treated cells. CONCLUSION: G2/M phase arrest and apoptosis induced by PCD are associated with the inhibition of CDK-activating kinase activity and the activation of Ca2+-related mitochondrion pathway in SGC-7901 cells.

  19. Peroxisome proliferator-activated receptor γ ligands induce cell cycle arrest and apoptosis in human renal carcinoma cell lines

    Institute of Scientific and Technical Information of China (English)

    Feng-guang YANG; Zhi-wen ZHANG; Dian-qi XIN; Chang-jin SHI; Jie-ping WU; Ying-lu GUO; You-fei GUAN

    2005-01-01

    Aim: To study the effect of peroxisome proliferator-actived receptor γ (PPARγ)ligands on cell proliferation and apoptosis in human renal carcinoma cell lines.Methods: The expression of PPARγ was investigated by reverse transcriptase polymerase chain reaction (RT-PCR), Western blot and immunohistochemistry.The effect of thiazolidinedione (TZD) PPARγ ligands on growth of renal cell carcinoma (RCC) cells was measured by MTT assay and flow cytometric analysis. Cell death ELISA, Hoechst 33342 fluorescent staining and DNA ladder assay were used to observe the effects of PPARγ ligands on apoptosis. Regulatory proteins of cell cycle and apoptosis were detected by Western blot analysis. Results:PPARγ was expressed at much higher levels in renal tumors than in the normal kidney (2.16±0.85 vs 0.90±0.73; P<0.01 ). TZD PPARγ ligands inhibited RCC cell growth in a dose-dependent manner with IC50 values of 7.08 μmol/L and 11.32 μmol/L for pioglitazone, and 5.71 μmol/L and 8.38 μmol/L for troglitazone in 786-O and A498 cells, respectively. Cell cycle analysis showed a G0/G1 arrest in human RCC cells following 24-h exposure to TZD. Analysis of cell cycle regulatory proteins revealed that TZD decreased the protein levels of proliferating cell nuclear antigen, pRb, cyclin D1, and Cdk4 but increased the levels of p21 and p27 in a timedependent manner. Furthermore, high doses of TZD induced massive apoptosis in renal cancer cells, with increased Bax expression and decreased Bcl-2 expression.Conclusion: TZD PPARγ ligands showed potent inhibitory effect on proliferation,and could induce apoptosis in RCC cells. These results suggest that ligands for PPARγ have potential antitumor effects on renal carcinoma cells.

  20. Identification of Siglec-9 as the receptor for MUC16 on human NK cells, B cells, and monocytes

    Directory of Open Access Journals (Sweden)

    Rancourt Claudine

    2010-05-01

    Full Text Available Abstract Background MUC16 is a cell surface mucin expressed at high levels by epithelial ovarian tumors. Following proteolytic cleavage, cell surface MUC16 (csMUC16 is shed in the extracellular milieu and is detected in the serum of cancer patients as the tumor marker CA125. csMUC16 acts as an adhesion molecule and facilitates peritoneal metastasis of ovarian tumors. Both sMUC16 and csMUC16 also protect cancer cells from cytotoxic responses of natural killer (NK cells. In a previous study we demonstrated that sMUC16 binds to specific subset of NK cells. Here, we identify the csMUC16/sMUC16 binding partner expressed on immune cells. Results Analysis of immune cells from the peripheral blood and peritoneal fluid of ovarian cancer patients indicates that in addition to NK cells, sMUC16 also binds to B cells and monocytes isolated from the peripheral blood and peritoneal fluid. I-type lectin, Siglec-9, is identified as the sMUC16 receptor on these immune cells. Siglec-9 is expressed on approximately 30-40% of CD16pos/CD56dim NK cells, 20-30% of B cells and >95% of monocytes. sMUC16 binds to the majority of the Siglec-9pos NK cells, B cells and monocytes. sMUC16 is released from the immune cells following neuraminidase treatment. Siglec-9 transfected Jurkat cells and monocytes isolated from healthy donors bind to ovarian tumor cells via Siglec-9-csMUC16 interaction. Conclusions Recent studies indicate that csMUC16 can act as an anti-adhesive agent that blocks tumor-immune cell interactions. Our results demonstrate that similar to other mucins, csMUC16 can also facilitate cell adhesion by interacting with a suitable binding partner such as mesothelin or Siglec-9. Siglec-9 is an inhibitory receptor that attenuates T cell and NK cell function. sMUC16/csMUC16-Siglec-9 binding likely mediates inhibition of anti-tumor immune responses.

  1. Resistance to ultraviolet-induced apoptosis in DNA repair deficient growth arrested human fibroblasts is not related to recovery from RNA transcription blockage

    International Nuclear Information System (INIS)

    The impact of ultraviolet (UV-C) photoproducts on apoptosis induction was investigated in growth arrested (confluent) and proliferating human primary fibroblasts. Confluent fibroblasts were more resistant to UV-C-induced apoptosis than proliferating cells, and this was observed for normal human cells and for cells from patients with Cockayne and trichothiodystrophy syndromes, deficient in transcription coupled repair. This resistance was sustained for at least seven days and was not due to DNA repair efficiency, as the removal of CPDs in the genome was similar under both growth conditions. There was no correlation between reduced apoptosis and RNA synthesis recovery. Following UV-C treatment, proliferating and confluent fibroblasts showed a similar level of RNA synthesis inhibition and recovery from transcription blockage. These results support the hypothesis that the decrease of DNA replication, in growth arrested cells, protects cell from UV-C-induced apoptosis, even in the presence of DNA lesions

  2. Effects of adenoviral gene transfer of mutated IкBα, a novel inhibitor of NF-кb, on human monocyte-derived dendritic cells

    Institute of Scientific and Technical Information of China (English)

    Lin-fu ZHOU; Ming-shun ZHANG; Kai-sheng YIN; Yong JI; Wei-ping XIE; Xue-fan GUI; Xiao-hui JI

    2006-01-01

    Aim: To investigate the effects of adenoviral gene transfer of IкBα mutant (IкBαM), a novel inhibitor of nuclear factor кB (NF- кB), on apoptosis, phenotype and function of human monocyte-derived dendritic cells (DC). Methods: Monocytes, cocultured with granulocyte/macrophage colony-stimulating factor (GM-CSF; 900 ng/mL) and interleukin (IL)-4 (300 ng/mL) for 5 d, followed by stimulation with lipopolysaccharide (LPS; 100 ng/mL) for 2 d differentiated into mature DC. Monocytes were either left untransfected or were transfected with Ad IкBαM or AdLacZ. The transcription and expression of the IкBαM gene, and the inhibitory effect of IкBαM on tumor necrosis factor (TNF)-α-induced NF-кB activation in mature DC were detected by polymerase chain reaction (PCR), reverse transcription-polymerase chain reaction (RT-PCR), Western blot analysis, and electrophoretic mobility shift assays, respectively. The phenotype, apoptosis, IL-12 secretion level of DC, and ability to stimulate the proliferation of T cells were determined by flow cytometry, enzyme-linked immunosorbent assay and mixed leukocyte reaction. Results: PCR and RT-PCR were used to detect a unique 801 bp band in Ad IкBαM-transfected mature DC, and also a dose- and time-dependent expression of the IкBαM gene, which peaked at a multiplicity of infection of 100 pfu/cell and at 48 h. Furthermore, Ad IкBαM significantly suppressed the TNF-α-induced NF- кB activation, augmented apoptosis, downregulated CD80, CD83, and CD86 surface molecules, IL-12 secretion levels and the ability to stimulate the proliferation of T cells in mature DC. Conclusion: Ad IкBαM effectively transfected and potently inhibited NF-кB activation in monocyte-derived mature DC. Overexpression of the IкBαM gene in mature DC may contribute to T-cell immunosuppression through induction of DC apoptosis and downregulation of B7 molecules, providing a potential strategy for future DC-based immunotherapy of asthma.

  3. Anti-Tumor Effect of Rutin on Human Neuroblastoma Cell Lines through Inducing G2/M Cell Cycle Arrest and Promoting Apoptosis

    OpenAIRE

    Hongyan Chen; Qing Miao; Miao Geng; Jing Liu; Yazhuo Hu; Lei Tian; Jingkun Pan; Yi Yang

    2013-01-01

    Aims. To further investigate the antineuroblastoma effect of rutin which is a type of flavonoid. Methods. The antiproliferation of rutin in human neuroblastoma cells LAN-5 were detected by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Chemotaxis of LAN-5 cells was assessed using transwell migration chambers and scratch wound migration assay. The cell cycle arrest and apoptosis in a dose-dependent manner was measured by flow cytometric and fluorescent microscopy ana...

  4. Chamaejasmine Arrests Cell Cycle, Induces Apoptosis and Inhibits Nuclear NF-κB Translocation in the Human Breast Cancer Cell Line MDA-MB-231

    OpenAIRE

    Yuxian Bai; Guanglu Dong; Li Cai; Hongyang Yu; Tingting Zhang

    2013-01-01

    In this study, the anticancer activity of chamaejasmine was characterized in the human breast cancer cell line, MDA-MB-231. Cell viability and cell cycle distribution were determined by MTT assay and flow cytometry, respectively. Western blotting was performed to determine changes in levels of various proteins. Results showed that treatment with chamaejasmine (4–16 μM) inhibited cell proliferation, which correlated with G2/M phase arrest and apoptosis in MDA-MB-231 cells. C...

  5. Induction of G1-phase cell cycle arrest and apoptosis pathway in MDA-MB-231 human breast cancer cells by sulfated polysaccharide extracted from Laurencia papillosa

    OpenAIRE

    Murad, Hossam; Hawat, Mohammad; Ekhtiar, Adnan; ALJAPAWE, ABDULMUNIM; Abbas, Assef; Darwish, Hussein; Sbenati, Oula; Ghannam, Ahmed

    2016-01-01

    Background Marine algae consumption is linked to law cancer incidences in countries that traditionally consume marine products. Hence, Phytochemicals are considered as potential chemo-preventive and chemotherapeutic agents against cancer. We investigated the effects of the algal sulfated polysaccharide extract (ASPE) from the red marine alga L. papillosa on MDA-MB-231 human breast cancer cell line. Methods Flow cytometry analysis was performed to study the cell viability, cell cycle arrest an...

  6. Reduction of Monocyte Chemoattractant Protein-1 and Interleukin-8 Levels by Ticlopidine in TNF-α Stimulated Human Umbilical Vein Endothelial Cells

    Directory of Open Access Journals (Sweden)

    Chaur-Jong Hu

    2009-01-01

    Full Text Available Atherosclerosis and its associated complications represent major causes of morbidity and mortality in the industrialized or Western countries. Monocyte chemoattractant protein-1 (MCP-1 is critical for the initiating and developing of atherosclerotic lesions. Interleukin-8 (IL-8, a CXC chemokine, stimulates neutrophil chemotaxis. Ticlopidine is one of the antiplatelet drugs used to prevent thrombus formation relevant to the pathophysiology of atherothrombosis. In this study, we found that ticlopidine dose-dependently decreased the mRNA and protein levels of TNF-α-stimulated MCP-1, IL-8, and vascular cell adhesion molecule-1 (VCAM-1 in human umbilical vein endothelial cells (HUVECs. Ticlopidine declined U937 cells adhesion and chemotaxis as compared to TNF-α stimulated alone. Furthermore, the inhibitory effects were neither due to decreased HUVEC viability, nor through NF-kB inhibition. These results suggest that ticlopidine decreased TNF-α induced MCP-1, IL-8, and VCAM-1 levels in HUVECs, and monocyte adhesion. Therefore, the data provide additional therapeutic machinery of ticlopidine in treatment and prevention of atherosclerosis.

  7. Monocyte scintigraphy in rheumatoid arthritis: the dynamics of monocyte migration in immune-mediated inflammatory disease.

    Directory of Open Access Journals (Sweden)

    Rogier M Thurlings

    Full Text Available BACKGROUND: Macrophages are principal drivers of synovial inflammation in rheumatoid arthritis (RA, a prototype immune-mediated inflammatory disease. Conceivably, synovial macrophages are continuously replaced by circulating monocytes in RA. Animal studies from the 1960s suggested that macrophage replacement by monocytes is a slow process in chronic inflammatory lesions. Translation of these data into the human condition has been hampered by the lack of available techniques to analyze monocyte migration in man. METHODS/PRINCIPAL FINDINGS: We developed a technique that enabled us to analyze the migration of labelled autologous monocytes in RA patients using single photon emission computer tomography (SPECT. We isolated CD14+ monocytes by CliniMACS in 8 patients and labeled these with technetium-99m (99mTc-HMPAO. Monocytes were re-infused into the same patient. Using SPECT we calculated that a very small but specific fraction of 3.4 x 10(-3 (0.95-5.1 x 10(-3 % of re-infused monocytes migrated to the inflamed joints, being detectable within one hour after re-infusion. CONCLUSIONS/SIGNIFICANCE: The results indicate monocytes migrate continuously into the inflamed synovial tissue of RA patients, but at a slow macrophage-replacement rate. This suggests that the rapid decrease in synovial macrophages that occurs after antirheumatic treatment might rather be explained by an alteration in macrophage retention than in monocyte influx and that RA might be particularly sensitive to treatments targeting inflammatory cell retention.

  8. Zygote arrest 1 gene in pig, cattle and human: evidence of different transcript variants in male and female germ cells

    Directory of Open Access Journals (Sweden)

    Royere Dominique

    2006-03-01

    Full Text Available Abstract Background Zygote arrest 1 (ZAR1 is one of the few known oocyte-specific maternal-effect genes essential for the beginning of embryo development discovered in mice. This gene is evolutionary conserved in vertebrates and ZAR1 protein is characterized by the presence of atypical plant homeobox zing finger domain, suggesting its role in transcription regulation. This work was aimed at the study of this gene, which could be one of the key regulators of successful preimplantation development of domestic animals, in pig and cattle, as compared with human. Methods Screenings of somatic cell hybrid panels and in silico research were performed to characterize ZAR1 chromosome localization and sequences. Rapid amplification of cDNA ends was used to obtain full-length cDNAs. Spatio-temporal mRNA expression patterns were studied using Northern blot, reverse transcription coupled to polymerase chain reaction and in situ hybridization. Results We demonstrated that ZAR1 is a single copy gene, positioned on chromosome 8 in pig and 6 in cattle, and several variants of correspondent cDNA were cloned from oocytes. Sequence analysis of ZAR1 cDNAs evidenced numerous short inverted repeats within the coding sequences and putative Pumilio-binding and embryo-deadenylation elements within the 3'-untranslated regions, indicating the potential regulation ways. We showed that ZAR1 expressed exclusively in oocytes in pig ovary, persisted during first cleavages in embryos developed in vivo and declined sharply in morulae and blastocysts. ZAR1 mRNA was also detected in testis, and, at lower level, in hypothalamus and pituitary in both species. For the first time, ZAR1 was localized in testicular germ cells, notably in round spermatids. In addition, in pig, cattle and human only shorter ZAR1 transcript variants resulting from alternative splicing were found in testis as compared to oocyte. Conclusion Our data suggest that in addition to its role in early embryo

  9. Both common and specialty mushrooms inhibit adhesion molecule expression and in vitro binding of monocytes to human aortic endothelial cells in a pro-inflammatory environment

    Directory of Open Access Journals (Sweden)

    Martin Keith R

    2010-07-01

    Full Text Available Abstract Background Cardiovascular disease (CVD is a leading cause of mortality in the United States as well as globally. Epidemiological studies show that regular fruit and vegetable consumption reduces CVD risk, in part, due to antioxidant activity and immunomodulation since oxidative stress and inflammation are features of atherogenesis. Accumulating evidence also shows that dietary fungi, viz., mushrooms, can protect against chronic disease by altering inflammatory environments such as those associated with CVD although most research has focused on specialty mushrooms. In this study, we tested the ability of both common and specialty mushrooms to inhibit cellular processes associated with CVD. Methods Human aortic endothelial cells (HAEC were incubated overnight with control media with dimethylsulfoxide (DMSO vehicle (1% v/v or containing DMSO extracts of whole dehydrated mushrooms (0.1 mg/mL, which included Agaricus bisporus (white button and crimini, Lentinula edodes (shiitake, Pleurotus ostreatus (oyster, and Grifola frondosa (maitake. Monolayers were subsequently washed and incubated with medium alone or containing the pro-inflammatory cytokine IL-1β (5 ng/mL for 6 h to upregulate pro-atherosclerotic adhesion molecules (AM. AM expression was assayed by ELISA and binding of U937 human monocytes pre-loaded with fluorescent dye was determined. Results White button mushrooms consistently reduced (p Conclusion These data provide evidence that dietary mushrooms can inhibit cellular processes such as adhesion molecule expression and ultimate binding of monocytes to the endothelium under pro-inflammatory conditions, which are associated with CVD. As a result, these findings support the notion that dietary mushrooms can be protective against CVD.

  10. Knockdown of p54nrb inhibits migration, invasion and TNF-α release of human acute monocytic leukemia THP1 cells.

    Science.gov (United States)

    Zhang, Xiujuan; Wu, Changli; Xiong, Wei; Chen, Chunling; Li, Rong; Zhou, Guangji

    2016-06-01

    54 kDa nuclear RNA- and DNA-binding protein (p54nrb) which is also called non-POU domain-containing octamer-binding protein (NONO) is known to be multifunctional involved in many nuclear processes. It was shown that p54nrb/NONO was closely related to the occurrence of erythroleukemia. Whether p54nrb/NONO plays a role in progress of human acute monocytic leukemia remains unknown. In the present study, we examined the effects of p54nrb/NONO silencing on the biological characteristics of human acute monocytic leukemia THP1 cells. The results showed that p54nrb was strongly expressed in THP1 cells, and knockdown of p54nrb slightly promoted proliferation and strongly inhibited motility and invasion of THP1 cells. Moreover, knockdown of p54nrb strongly decreased the release of TNF-α from THP1 cells by inhibiting certain process of TNF-α secretion, specially for the release of TNF-α induced by lipopolysaccharide (LPS). Notably, the infection of negative control shRNA-containing lentiviruses promoted the migration and the release of TNF-α induced by LPS in THP1 cells. All the above results demonstrated that p54nrb slightly inhibited THP1 cell proliferation, but significantly promoted migration, invasion and release of TNF-α induced by LPS in THP1 cells. The present study indicates that p54nrb is a powerful molecule involved in the regulation of cell motility and promotes the migration and invasion of THP1 cells, and it is more likely to be involved in the release of inflammatory mediators and the motility of inflammatory cells. PMID:27108701

  11. Bacillus anthracis capsule activates caspase-1 and induces interleukin-1beta release from differentiated THP-1 and human monocyte-derived dendritic cells.

    Science.gov (United States)

    Cho, Min-Hee; Ahn, Hae-Jeong; Ha, Hyun-Joon; Park, Jungchan; Chun, Jeong-Hoon; Kim, Bong-Su; Oh, Hee-Bok; Rhie, Gi-Eun

    2010-01-01

    The poly-gamma-d-glutamic acid (PGA) capsule is one of the major virulence factors of Bacillus anthracis, which causes a highly lethal infection. The antiphagocytic PGA capsule disguises the bacilli from immune surveillance and allows unimpeded growth of bacilli in the host. Recently, efforts have been made to include PGA as a component of anthrax vaccine; however, the innate immune response of PGA itself has been poorly investigated. In this study, we characterized the innate immune response elicited by PGA in the human monocytic cell line THP-1, which was differentiated into macrophages with phorbol 12-myristate 13-acetate (PMA) and human monocyte-derived dendritic cells (hMoDCs). PGA capsules were isolated from the culture supernatant of either the pXO1-cured strain of B. anthracis H9401 or B. licheniformis ATCC 9945a. PGA treatment of differentiated THP-1 cells and hMoDCs led to the specific extracellular release of interleukin-1beta (IL-1beta) in a dose-dependent manner. Evaluation of IL-1beta processing by Western blotting revealed that cleaved IL-1beta increased in THP-1 cells and hMoDCs after PGA treatment. Enhanced processing of IL-1beta directly correlated with increased activation of its upstream regulator, caspase-1, also known as IL-1beta-converting enzyme (ICE). The extracellular release of IL-1beta in response to PGA was ICE dependent, since the administration of an ICE inhibitor prior to PGA treatment blocked induction of IL-1beta. These results demonstrate that B. anthracis PGA elicits IL-1beta production through activation of ICE in PMA-differentiated THP-1 cells and hMoDCs, suggesting the potential for PGA as a therapeutic target for anthrax. PMID:19737897

  12. Hemozoin triggers tumor necrosis factor alpha-mediated re-lease of lysozyme by human adherent monocytes:new evi-dences on leukocyte degranulation in P.falciparum malaria

    Institute of Scientific and Technical Information of China (English)

    Prato M; Giribaldi G; Arese P

    2009-01-01

    Objective:Avidly phagocytosed hemozoin (malarial pigment)alters several functions of human monocytes and stimulates generation of several cytokines.Recently,we showed that phagocytosis of hemozoin by human mono-cytes increases expression and activity of matrix metalloproteinase-9,a proteolytic enzyme available in specific gelatinase granules,which contain several enzymes including lysozyme.Present work investigated active lyso-zyme release after phagocytosis of hemozoin and its dependence on production of tumor necrosis factor alpha. Methods:After phagocytosis of hemozoin,hemozoin-containing trophozoites or control meals (opsonized non-parasitized red blood cells and latex particles),monocyte supernatants were monitored for 2 hours,in presence of blocking anti-human tumor necrosis factor alpha antibodies or recombinant human tumor necrosis factor alpha cytokine in selected experiments.Lysozyme release was evaluated by a specific spectrometric assay measuring lysozyme activity after coincubation of cell supernatants with suspensions of Mycrococcus Lysodeikticus,while levels of soluble tumor necrosis factor alpha were analyzed by specific enzyme-linked immunodsorbent assay. Results:Levels of lysozyme activity and soluble tumor necrosis factor alpha protein were increased in hemozo-in-or trophozoites-laden monocytes supernatants.Phagocytosis per se (control meals)also increased lysozyme release,but levels were significantly lower than those obtained after phagocytosis of hemozoin or trophozoites. Interestingly,all effects on lysozyme release observed after phagocytosis were abrogated by blocking anti-human tumor necrosis factor alpha antibodies,while they were mimicked by recombinant human tumor necrosis factor alpha cytokine.Conclusions:Present work shows that phagocytosis of hemozoin promotes monocyte degranula-tion and enhances active lysozyme release.The effect requires tumor necrosis factor alpha mediation.

  13. Interleukin-10 Stimulates Coxiella burnetii Replication in Human Monocytes through Tumor Necrosis Factor Down-Modulation: Role in Microbicidal Defect of Q Fever

    OpenAIRE

    Ghigo, Eric; Capo, Christian; Raoult, Didier; Mege, Jean-Louis

    2001-01-01

    Coxiella burnetii, an obligate intracellular bacterium, is the agent of Q fever. The chronic form of the disease is associated with the overproduction of interleukin-10 and deficient C. burnetii killing by monocytes. We hypothesized that the replication of C. burnetii inside monocytes requires a macrophage-deactivating cytokine such as interleukin-10. In the absence of interleukin-10, C. burnetii survived but did not replicate in monocytes. C. burnetii replication (measured 15 days) was induc...

  14. The role of reactive oxygen species (ROS) production on diallyl disulfide (DADS) induced apoptosis and cell cycle arrest in human A549 lung carcinoma cells

    Energy Technology Data Exchange (ETDEWEB)

    Wu Xinjiang [Institute of Indoor and Environmental Toxicology, Faculty of Medicine, Justus-Liebig-University of Giessen, Aulweg 123, D-35385 Giessen (Germany); Kassie, Fekadu [Institute of Indoor and Environmental Toxicology, Faculty of Medicine, Justus-Liebig-University of Giessen, Aulweg 123, D-35385 Giessen (Germany); Mersch-Sundermann, Volker [Institute of Indoor and Environmental Toxicology, Faculty of Medicine, Justus-Liebig-University of Giessen, Aulweg 123, D-35385 Giessen (Germany)]. E-mail: Volker.mersch-sundermann@uniklinikum-giessen.de

    2005-11-11

    Diallyl disulfide (DADS), an oil soluble constituent of garlic (Allium sativum), has been reported to cause antimutagentic and anticarcinogenic effects in vitro and in vivo by modulating phases I and II enzyme activities. In recent years, several studies suggested that the chemopreventive effects of DADS can also be attributed to induction of cell cycle arrest and apoptosis in cancer cells. In the present study, we reported that DADS-induced cell cycle arrest at G2/M and apoptosis in human A549 lung cancer cells in a time- and dose-dependent manner. Additionally, a significant increase of intracellular reactive oxygen species (ROS) was induced in A549 cells less than 0.5 h after DADS treatment, indicating that ROS may be an early event in DADS-modulated apoptosis. Treatment of A549 cells with N-acetyl cysteine (NAC) completely abrogated DADS-induced cell cycle arrest and apoptosis. The result indicated that oxidative stress modulates cell proliferation and cell death induced by DADS.

  15. Human Herpesvirus-6 U14 Induces Cell-Cycle Arrest in G2/M Phase by Associating with a Cellular Protein, EDD.

    Directory of Open Access Journals (Sweden)

    Junko Mori

    Full Text Available The human herpesvirus-6 (HHV-6 infection induces cell-cycle arrest. In this study, we found that the HHV-6-encoded U14 protein induced cell-cycle arrest at G2/M phase via an association with the cellular protein EDD, a mediator of DNA-damage signal transduction. In the early phase of HHV-6 infection, U14 colocalized with EDD dots in the nucleus, and similar colocalization was also observed in cells transfected with a U14 expression vector. When the carboxyl-terminal region of U14 was deleted, no association of U14 and EDD was observed, and the percentage of cells in G2/M decreased relative to that in cells expressing wild-type U14, indicating that the C-terminal region of U14 and the U14-EDD association are critical for the cell-cycle arrest induced by U14. These results indicate that U14 is a G2/M checkpoint regulator encoded by HHV-6.

  16. The role of reactive oxygen species (ROS) production on diallyl disulfide (DADS) induced apoptosis and cell cycle arrest in human A549 lung carcinoma cells

    International Nuclear Information System (INIS)

    Diallyl disulfide (DADS), an oil soluble constituent of garlic (Allium sativum), has been reported to cause antimutagentic and anticarcinogenic effects in vitro and in vivo by modulating phases I and II enzyme activities. In recent years, several studies suggested that the chemopreventive effects of DADS can also be attributed to induction of cell cycle arrest and apoptosis in cancer cells. In the present study, we reported that DADS-induced cell cycle arrest at G2/M and apoptosis in human A549 lung cancer cells in a time- and dose-dependent manner. Additionally, a significant increase of intracellular reactive oxygen species (ROS) was induced in A549 cells less than 0.5 h after DADS treatment, indicating that ROS may be an early event in DADS-modulated apoptosis. Treatment of A549 cells with N-acetyl cysteine (NAC) completely abrogated DADS-induced cell cycle arrest and apoptosis. The result indicated that oxidative stress modulates cell proliferation and cell death induced by DADS

  17. Inhibitive effect of 3-bromopyruvic acid on human breast cancer MCF-7 cells involves cell cycle arrest and apoptotic induction

    Institute of Scientific and Technical Information of China (English)

    LIU Xiao-hong; ZHENG Xue-fang; WANG Yong-li

    2009-01-01

    Background Breast cancer is one of the most common malignancies in women and is highly resistant to chemotherapy. Due to its high tumour selectivity, 3-bromopyruvic acid (3-BrPA), a well-known inhibitor of energy metabolism has been proposed as a specific anticancer agent. The present study determined the effect of 3-BrPA on proliferation, cell cycle and apoptosis in the human breast cancer MCF-7 cell line and other antitumour mechanisms. Methods MCF-7 cells were treated with various concentrations of 3-BrPA for 1-4 days, and cell growth was measured by 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyl tetrazolium bromide assay. Marked morphological changes in MCF-7 cells after treatment with 3-BrPA were observed using transmission electron microscopy. The distributions of the cell cycle and apoptosis were analyzed by flow cytometry. Immunohistochemistry was used to indicate the changes in the expression of Bcl-2, c-Myc, and mutant p53. Results 3-BrPA (25 μg/ml) significantly inhibited the proliferation of MCF-7 cells in a time-dependent manner. The MCF-7 cells exposed to 3-BrPA showed the typical morphological characteristics of apoptosis, including karyopycnosis, nuclear condensation and oversize cytoplasmic particles. In addition, flow cytometric assay also showed more apoptotic cells after 3-BrPA stimulation. The cells at the GO and G1 phases were dramatically decreased while cells at the S and G2/M phases were increased in response to 3-BrPA treatment after 48 hours. Furthermore, 3-BrPA stimulation decreased the expressions of Bcl-2, c-Myc and mutant p53, which were strongly associated with the programmed cell death signal transduction pathway. Conclusion 3-BrPA inhibits proliferation, induces S phase and G2/M phase arrest, and promotes apoptosis in MCF-7 cells, which processes might be mediated by the downregulation of the expressions of Bcl-2, c-Myc and mutant p53.

  18. Effects of gamma-radiation on cell growth, cycle arrest, death, and superoxide dismutase expression by DU 145 human prostate cancer cells

    Directory of Open Access Journals (Sweden)

    Vucic V.

    2006-01-01

    Full Text Available Gamma-irradiation (gamma-IR is extensively used in the treatment of hormone-resistant prostate carcinoma. The objective of the present study was to investigate the effects of 60Co gamma-IR on the growth, cell cycle arrest and cell death of the human prostate cancer cell line DU 145. The viability of DU 145 cells was measured by the Trypan blue exclusion assay and the 3(4,5-dimethylthiazol-2-yl-2,5,diphenyltetrazolium bromide test. Bromodeoxyuridine incorporation was used for the determination of cell proliferation. Cell cycle arrest and cell death were analyzed by flow cytometry. Superoxide dismutase (SOD, specifically CuZnSOD and MnSOD protein expression, after 10 Gy gamma-IR, was determined by Western immunoblotting analysis. gamma-IR treatment had a significant (P < 0.001 antiproliferative and cytotoxic effect on DU 145 cells. Both effects were time and dose dependent. Also, the dose of gamma-IR which inhibited DNA synthesis and cell proliferation by 50% was 9.7 Gy. Furthermore, gamma-IR induced cell cycle arrest in the G2/M phase and the percentage of cells in the G2/M phase was increased from 15% (control to 49% (IR cells, with a nonsignificant induction of apoptosis. Treatment with 10 Gy gamma-IR for 24, 48, and 72 h stimulated CuZnSOD and MnSOD protein expression in a time-dependent manner, approximately by 3- to 3.5-fold. These data suggest that CuZnSOD and MnSOD enzymes may play an important role in the gamma-IR-induced changes in DU 145 cell growth, cell cycle arrest and cell death.

  19. 5-(2-Carboxyethenyl) isatin derivative induces G2/M cell cycle arrest and apoptosis in human leukemia K562 cells

    International Nuclear Information System (INIS)

    Highlights: • 5-(2-Carboxyethenyl) isatin derivative (HKL 2H) inhibited K562’s proliferation. • HKL 2H caused the morphology change of G2/M phase arrest and typical apoptosis. • HKL 2H induced G2/M cell cycle phase arrest in K562 cells. • HKL 2H induced apoptosis in K562 cells through the mitochondrial pathway. - Abstract: Our previous study successfully identified that the novel isatin derivative (E)-methyl 3-(1-(4-methoxybenzyl)-2,3-dioxoindolin-5-yl) acrylate (HKL 2H) acts as an anticancer agent at an inhibitory concentration (IC50) level of 3 nM. In this study, the molecular mechanism how HKL 2H induces cytotoxic activity in the human chronic myelogenous leukemia K562 cells was investigated. Flow cytometric analysis showed that the cells were arrested in the G2/M phase and accumulated subsequently in the sub-G1 phase in the presence of HKL 2H. HKL 2H treatment down-regulated the expressions of CDK1 and cyclin B but up-regulated the level of phosphorylated CDK1. Annexin-V staining and the classic DNA ladder studies showed that HKL 2H induced the apoptosis of K562 cells. Our study further showed that HKL 2H treatment caused the dissipation of mitochondrial membrane potential, activated caspase-3 and lowered the Bcl-2/Bax ratio in K562 cells, suggesting that the HKL 2H-causing programmed cell death of K562 cells was caused via the mitochondrial apoptotic pathway. Taken together, our data demonstrated that HKL 2H, a 5-(2-carboxyethenyl) isatin derivative, notably induces G2/M cell cycle arrest and mitochondrial-mediated apoptosis in K562 cells, indicating that this compound could be a promising anticancer candidate for further investigation

  20. 5-(2-Carboxyethenyl) isatin derivative induces G{sub 2}/M cell cycle arrest and apoptosis in human leukemia K562 cells

    Energy Technology Data Exchange (ETDEWEB)

    Zhou, Yao; Zhao, Hong-Ye; Han, Kai-Lin; Yang, Yao; Song, Bin-Bin; Guo, Qian-Nan [Key Laboratory of Industrial Microbiology, Ministry of Education, College of Biotechnology, Tianjin University of Science and Technology, Tianjin 300457 (China); Tianjin Key Laboratory of Industry Microbiology, College of Biotechnology, Tianjin University of Science and Technology, Tianjin 300457 (China); Fan, Zhen-Chuan [Key Laboratory of Food Nutrition and Safety (Tianjin University of Science and Technology), Ministry of Education, Tianjin 300457 (China); Obesita and Algaegen LLC, College Station, TX 77845 (United States); Zhang, Yong-Min [Université Pierre et Marie Curie-Paris 6, Institut Parisien de Chimie Moléculaire UMR CNRS 8232, 4 Place Jussieu, 75005 Paris (France); Teng, Yu-Ou, E-mail: tyo201485@tust.edu.cn [Key Laboratory of Industrial Microbiology, Ministry of Education, College of Biotechnology, Tianjin University of Science and Technology, Tianjin 300457 (China); Tianjin Key Laboratory of Industry Microbiology, College of Biotechnology, Tianjin University of Science and Technology, Tianjin 300457 (China); Yu, Peng, E-mail: yupeng@tust.edu.cn [Key Laboratory of Industrial Microbiology, Ministry of Education, College of Biotechnology, Tianjin University of Science and Technology, Tianjin 300457 (China); Tianjin Key Laboratory of Industry Microbiology, College of Biotechnology, Tianjin University of Science and Technology, Tianjin 300457 (China)

    2014-08-08

    Highlights: • 5-(2-Carboxyethenyl) isatin derivative (HKL 2H) inhibited K562’s proliferation. • HKL 2H caused the morphology change of G{sub 2}/M phase arrest and typical apoptosis. • HKL 2H induced G2/M cell cycle phase arrest in K562 cells. • HKL 2H induced apoptosis in K562 cells through the mitochondrial pathway. - Abstract: Our previous study successfully identified that the novel isatin derivative (E)-methyl 3-(1-(4-methoxybenzyl)-2,3-dioxoindolin-5-yl) acrylate (HKL 2H) acts as an anticancer agent at an inhibitory concentration (IC{sub 50}) level of 3 nM. In this study, the molecular mechanism how HKL 2H induces cytotoxic activity in the human chronic myelogenous leukemia K562 cells was investigated. Flow cytometric analysis showed that the cells were arrested in the G{sub 2}/M phase and accumulated subsequently in the sub-G{sub 1} phase in the presence of HKL 2H. HKL 2H treatment down-regulated the expressions of CDK1 and cyclin B but up-regulated the level of phosphorylated CDK1. Annexin-V staining and the classic DNA ladder studies showed that HKL 2H induced the apoptosis of K562 cells. Our study further showed that HKL 2H treatment caused the dissipation of mitochondrial membrane potential, activated caspase-3 and lowered the Bcl-2/Bax ratio in K562 cells, suggesting that the HKL 2H-causing programmed cell death of K562 cells was caused via the mitochondrial apoptotic pathway. Taken together, our data demonstrated that HKL 2H, a 5-(2-carboxyethenyl) isatin derivative, notably induces G{sub 2}/M cell cycle arrest and mitochondrial-mediated apoptosis in K562 cells, indicating that this compound could be a promising anticancer candidate for further investigation.

  1. Cis and trans acting factors involved in human cytomegalovirus experimental and natural latent infection of CD14 (+ monocytes and CD34 (+ cells.

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    Cyprian C Rossetto

    Full Text Available The parameters involved in human cytomegalovirus (HCMV latent infection in CD14 (+ and CD34 (+ cells remain poorly identified. Using next generation sequencing we deduced the transcriptome of HCMV latently infected CD14 (+ and CD34 (+ cells in experimental as well as natural latency settings. The gene expression profile from natural infection in HCMV seropositive donors closely matched experimental latency models, and included two long non-coding RNAs (lncRNAs, RNA4.9 and RNA2.7 as well as the mRNAs encoding replication factors UL84 and UL44. Chromatin immunoprecipitation assays on experimentally infected CD14 (+ monocytes followed by next generation sequencing (ChIP-Seq were employed to demonstrate both UL84 and UL44 proteins interacted with the latent viral genome and overlapped at 5 of the 8 loci identified. RNA4.9 interacts with components of the polycomb repression complex (PRC as well as with the MIE promoter region where the enrichment of the repressive H3K27me3 mark suggests that this lncRNA represses transcription. Formaldehyde Assisted Isolation of Regulatory Elements (FAIRE, which identifies nucleosome-depleted viral DNA, was used to confirm that latent mRNAs were associated with actively transcribed, FAIRE analysis also showed that the terminal repeat (TR region of the latent viral genome is depleted of nucleosomes suggesting that this region may contain an element mediating viral genome maintenance. ChIP assays show that the viral TR region interacts with factors associated with the pre replication complex and a plasmid subclone containing the HCMV TR element persisted in latently infected CD14 (+ monocytes, strongly suggesting that the TR region mediates viral chromosome maintenance.

  2. The two-component adjuvant IC31® boosts type i interferon production of human monocyte-derived dendritic cells via ligation of endosomal TLRs.

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    Attila Szabo

    Full Text Available The aim of this study was to characterize and identify the mode of action of IC31®, a two-component vaccine adjuvant. We found that IC31® was accumulated in human peripheral blood monocytes, MHC class II positive cells and monocyte-derived DCs (moDCs but not in plasmacytoid DCs (pDCs. In the presence of IC31® the differentiation of inflammatory CD1a(+ moDCs and the secretion of chemokines, TNF-α and IL-6 cytokines was inhibited but the production of IFNβ was increased. Sustained addition of IC31® to differentiating moDCs interfered with IκBα phosphorylation, while the level of phospho-IRF3 increased. We also showed that both IC31® and its KLK component exhibited a booster effect on type I IFN responses induced by the specific ligands of TLR3 or TLR7/8, whereas TLR9 ligand induces type I IFN production only in the presence of IC31® or ODN1. Furthermore, long term incubation of moDCs with IC31® caused significantly higher expression of IRF and IFN genes than a single 24 hr treatment. The adjuvant activity of IC31® on the IFN response was shown to be exerted through TLRs residing in the vesicular compartment of moDCs. Based on these results IC31® was identified as a moDC modulatory adjuvant that sets the balance of the NF-κB and IRF3 mediated signaling pathways to the production of IFNβ. Thus IC31® is emerging as a potent adjuvant to increase immune responses against intracellular pathogens and cancer in future vaccination strategies.

  3. Virus-specific HLA-restricted lysis of herpes simplex virus-infected human monocytes and macrophages mediated by cytotoxic T lymphocytes

    International Nuclear Information System (INIS)

    Freshly-isolated peripheral blood human monocytes and 5 day in vitro cultured macrophages were infected with herpes simplex virus type 1 (HSV-1), labeled with 51Cr, and used as target cells in a 12-14 hour cell-mediated cytotoxicity assay. Mononuclear leukocytes (MNL) from HSV-1 non-immune individuals, whether unstimulated or stimulated with HSV-1 antigen, did not mediate significant lysis of either target cell. HSV-immune MNL, both freshly-isolated and cultured for 5 days without antigen, demonstrated only low levels of natural killer (NK) cell-mediate lysis. MNL from HSV-immune individuals incubated for 5 days in vitro with HSV-1 antigen mediated significant virus-specific lysis of both target cells. Mean virus-specific lysis of autologous monocytes was 8.5(/+-/2.0)% compared to a three-fold greater virus-specific lysis of autologous macrophages. Greater than 70% of this lytic activity was mediated by Leu-11-negative, T3-positive cytotoxic T lymphocytes (CTL). Allogeneic target cells lacking a common HLA determinant were not significantly lysed while T8-positive CTL mediated infrequent lysis of target cells sharing a common HLA-A and/or HLA-B determinant. T4-positive lymphocytes were demonstrated to be the predominant cell mediating lysis of autologous target cells and allogeneic target cells sharing both HLA-A and/or HLA-B plus HLA-DR determinants with the CTL; the T4-positive cell was the sole CTL mediator of lysis of allogeneic target cells having a common HLA-DR determinant

  4. Autocrine secretion of tumor necrosis factor under the influence of interferon-γ amplifies HLA-DR gene induction in human monocytes

    International Nuclear Information System (INIS)

    Recombinant interferon-γ (IFN-γ) induced HLA-DR gene expression in both U937 and THP-1 human monocytic cell lines, although the former was only very weakly inducible. Combination of recombinant tumor necrosis factor (TNF) and IFN-γ resulted in a synergistic enhancement of DR mRNA and protein induction in both cell lines. TNF alone increased the constitutive expression of the DR gene in THP-1 cells. In the HLA class II-negative U937 cells, TNF used alone was not able to induce DR gene expression. Such a negative result was not due to a lack of TNF receptor expression in U937 cells, since TNF clearly induced HLA class I and TNF gene expression in this cell line. THP-1, but not U937, cells secreted TNF under the influence of IFN-γ. Neutralization of TNF by a specific antibody decreased IFN-γ-induced DR antigen expression in THP-1 cultures. These observations indicate that TNF is not able to directly induce DR gene expression, but rather amplifies ongoing expression of this gene, whether constitutive or induced by IFN-γ. In the two cell lines tested, the level of DR inducibility under the influence of IFN-γ used alone depended on a different inducibility of TNF secretion by IFN-γ. Altogether, the observations indicate that TNF, whether exogenous or endogenously produced under the influence of IFN-γ, amplifies DR gene expression in monocytes, a phenomenon that may provide to such antigen-presenting cells a selective sensitivity to the DR-inducing effects of IFN-

  5. Electroporated Antigen-Encoding mRNA Is Not a Danger Signal to Human Mature Monocyte-Derived Dendritic Cells

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    Stefanie Hoyer

    2015-01-01

    Full Text Available For therapeutic cancer vaccination, the adoptive transfer of mRNA-electroporated dendritic cells (DCs is frequently performed, usually with monocyte-derived, cytokine-matured DCs (moDCs. However, DCs are rich in danger-sensing receptors which could recognize the exogenously delivered mRNA and induce DC activation, hence influencing the DCs’ immunogenicity. Therefore, we examined whether electroporation of mRNA with a proper cap and a poly-A tail of at least 64 adenosines had any influence on cocktail-matured moDCs. We used 16 different RNAs, encoding tumor antigens (MelanA, NRAS, BRAF, GNAQ, GNA11, and WT1, and variants thereof. None of those RNAs induced changes in the expression of CD25, CD40, CD83, CD86, and CD70 or the secretion of the cytokines IL-8, IL-6, and TNFα of more than 1.5-fold compared to the control condition, while an mRNA encoding an NF-κB-activation protein as positive control induced massive secretion of the cytokines. To determine whether mRNA electroporation had any effect on the whole transcriptome of the DCs, we performed microarray analyses of DCs of 6 different donors. None of 60,000 probes was significantly different between mock-electroporated DCs and MelanA-transfected DCs. Hence, we conclude that no transcriptional programs were induced within cocktail-matured DCs by electroporation of single tumor-antigen-encoding mRNAs.

  6. The ethanol extract of Scutellaria baicalensis and the active compounds induce cell cycle arrest and apoptosis including upregulation of p53 and Bax in human lung cancer cells

    International Nuclear Information System (INIS)

    Despite a lack of scientific authentication, Scutellaria baicalensis is clinically used in Chinese medicine as a traditional adjuvant to chemotherapy of lung cancer. In this study, cytotoxicity assays demonstrated that crude ethanolic extracts of S. baicalensis were selectively toxic to human lung cancer cell lines A549, SK-LU-1 and SK-MES-1 compared with normal human lung fibroblasts. The active compounds baicalin, baicalein and wogonin did not exhibit such selectivity. Following exposure to the crude extracts, cellular protein expression in the cancer cell lines was assessed using 2D gel electrophoresis coupled with MALDI-TOF-MS/Protein Fingerprinting. The altered protein expression indicated that cell growth arrest and apoptosis were potential mechanisms of cytotoxicity. These observations were supported by PI staining cell cycle analysis using flow cytometry and Annexin-V apoptotic analysis by fluorescence microscopy of cancer cells treated with the crude extract and pure active compounds. Moreover, specific immunoblotting identification showed the decreased expression of cyclin A results in the S phase arrest of A549 whereas the G0/G1 phase arrest in SK-MES-1 cells results from the decreased expression of cyclin D1. Following treatment, increased expression in the cancer cells of key proteins related to the enhancement of apoptosis was observed for p53 and Bax. These results provide further insight into the molecular mechanisms underlying the clinical use of this herb as an adjuvant to lung cancer therapy. - Research highlights: → Scutellaria baicalensis is a clinical adjuvant to lung cancer chemotherapy in China. → Scutellaria ethanol extracts selectively toxic to A549, SK-LU-1 and SK-MES-1. → Baicalin, baicalein and wogonin were toxic to all lung cancer cell lines. → Proteomics identified increased p53 and BAX in response to Scutellaria extracts.

  7. HIV-1 Latency in Monocytes/Macrophages

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    Amit Kumar

    2014-04-01

    Full Text Available Human immunodeficiency virus type 1 (HIV-1 targets CD4+ T cells and cells of the monocyte/macrophage lineage. HIV pathogenesis is characterized by the depletion of T lymphocytes and by the presence of a population of cells in which latency has been established called the HIV-1 reservoir. Highly active antiretroviral therapy (HAART has significantly improved the life of HIV-1 infected patients. However, complete eradication of HIV-1 from infected individuals is not possible without targeting latent sources of infection. HIV-1 establishes latent infection in resting CD4+ T cells and findings indicate that latency can also be established in the cells of monocyte/macrophage lineage. Monocyte/macrophage lineage includes among others, monocytes, macrophages and brain resident macrophages. These cells are relatively more resistant to apoptosis induced by HIV-1, thus are important stable hideouts of the virus. Much effort has been made in the direction of eliminating HIV-1 resting CD4+ T-cell reservoirs. However, it is impossible to achieve a cure for HIV-1 without considering these neglected latent reservoirs, the cells of monocyte/macrophage lineage. In this review we will describe our current understanding of the mechanism of latency in monocyte/macrophage lineage and how such cells can be specifically eliminated from the infected host.

  8. Deoxynivalenol (DON) is toxic to human colonic, lung and monocytic cell lines, but does not increase the IgE response in a mouse model for allergy

    International Nuclear Information System (INIS)

    We examined whether the common crop mycotoxin deoxynivalenol (DON) from Fusarium species is toxic to human colonic (Caco-2), lung (A549) and monocytic (U937) cell lines. Moreover, since DON reportedly induces increased levels of Th2 cytokines and total IgE, and we have observed that mould extracts adjuvated allergy development in mice, possible adjuvant effect of DON on allergy was studied in a mouse model. For all the cells, exposure to DON for 24 h reduced cellular protein synthesis, proliferation and survival rate dose-dependently. In addition, production of IL-8 in the U937 cell line increased up to eight-fold at levels of DON just lower than the most toxic one, suggesting that IL-8 can be used as an additional index for cytotoxicity in mononuclear phagocytes. However, DON did not increase levels of allergen-specific IgE or IgG1 in the mouse model for allergy. These results suggest that DON, when inhaled or ingested, may have toxic effect on human alveolar macrophages and epithelial cells in lungs and colon, but does not increase the allergic response to allergens

  9. Evaluation of an optimized protocol using human peripheral blood monocyte derived dendritic cells for the in vitro detection of sensitizers: Results of a ring study in five laboratories.

    Science.gov (United States)

    Reuter, Hendrik; Gerlach, Silke; Spieker, Jochem; Ryan, Cindy; Bauch, Caroline; Mangez, Claire; Winkler, Petra; Landsiedel, Robert; Templier, Marie; Mignot, Aurelien; Gerberick, Frank; Wenck, Horst; Aeby, Pierre; Schepky, Andreas

    2015-08-01

    Allergic contact dermatitis is a delayed T-cell mediated allergic response associated with relevant social and economic impacts. Animal experiments (e.g. the local lymph node assay) are still supplying most of the data used to assess the sensitization potential of new chemicals. However, the 7th amendment to the EU Cosmetic Directive have introduced a testing ban for cosmetic ingredients after March 2013. We have developed and optimized a stable and reproducible in vitro protocol based on human peripheral blood monocyte derived dendritic cells to assess the sensitization potential of chemicals. To evaluate the transferability and the predictivity of this PBMDCs based test protocol, a ring study was organized with five laboratories using seven chemicals with a known sensitization potential (one none-sensitizer and six sensitizers, including one pro-hapten). The results indicated that this optimized test protocol could be successfully transferred to all participating laboratories and allowed a correct assessment of the sensitization potential of the tested set of chemicals. This should allow a wider acceptance of PBMDCs as a reliable test system for the detection of human skin sensitizers and the inclusion of this protocol in the toolbox of in vitro methods for the evaluation of the skin sensitization potential of chemicals. PMID:25868915

  10. Malarial pigment haemozoin, IFN-gamma, TNF-alpha, IL-1beta and LPS do not stimulate expression of inducible nitric oxide synthase and production of nitric oxide in immuno-purified human monocytes

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    Ceretto Monica

    2007-06-01

    Full Text Available Abstract Background Enhanced production of nitric oxide (NO following upmodulation of the inducible isoform of NO synthase (iNOS by haemozoin (HZ, inflammatory cytokines and LPS may provide protection against Plasmodium falciparum malaria by killing hepatic and blood forms of parasites and inhibiting the cytoadherence of parasitized erythrocytes (RBC to endothelial cells. Monocytes and macrophages are considered to contribute importantly to protective upregulation of iNOS and production of NO. Data obtained with murine phagocytes fed with human HZ and synthetic HZ (sHZ indicate that supplemental treatment of those cells with IFN-gamma elicited significant increases in protein and mRNA expression of iNOS and NO production, providing a potential mechanism linking HZ phagocytosis and increased production of NO. Purpose of this study was to analyse the effect of P. falciparum HZ and sHZ supplemental to treatment with IFN-gamma and/or a stimulatory cytokine-LPS mix on iNOS protein and mRNA expression in immuno-purified human monocytes. Methods Adherent immunopurified human monocytes (purity >85%, and murine phagocytic cell lines RAW 264.7, N11 and ANA1 were fed or not with P. falciparum HZ or sHZ and treated or not with IFN-gamma or a stimulatory cytokine-LPS mix. Production of NO was quantified in supernatants, iNOS protein and mRNA expression were measured after immunoprecipitation and Western blotting and quantitative RT-PCT, respectively. Results Phagocytosis of HZ/sHZ by human monocytes did not increase iNOS protein and mRNA expression and NO production either after stimulation by IFN-gamma or the cytokine-LPS mix. By contrast, in HZ/sHZ-laden murine macrophages, identical treatment with IFN-gamma and the cytokine-LPS mix elicited significant increases in protein and mRNA expression of iNOS and NOS metabolites production, in agreement with literature data. Conclusion Results indicate that human monocytes fed or not with HZ/sHZ were constantly

  11. Effects of Cigarette Smoke Condensate on Oxidative Stress, Apoptotic Cell Death, and HIV Replication in Human Monocytic Cells.

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    Pss Rao

    Full Text Available While cigarette smoking is prevalent amongst HIV-infected patients, the effects of cigarette smoke constituents in cells of myeloid lineage are poorly known. Recently, we have shown that nicotine induces oxidative stress through cytochrome P450 (CYP 2A6-mediated pathway in U937 monocytic cells. The present study was designed to examine the effect of cigarette smoke condensate (CSC, which contains majority of tobacco constituents, on oxidative stress, cytotoxicity, expression of CYP1A1, and/or HIV-1 replication in HIV-infected (U1 and uninfected U937 cells. The effects of CSC on induction of CYP1 enzymes in HIV-infected primary macrophages were also analyzed. The results showed that the CSC-mediated increase in production of reactive oxygen species (ROS in U937 cells is dose- and time-dependent. Moreover, CSC treatment was found to induce cytotoxicity in U937 cells through the apoptotic pathway via activation of caspase-3. Importantly, pretreatment with vitamin C blocked the CSC-mediated production of ROS and induction of caspase-3 activity. In U1 cells, acute treatment of CSC increased ROS production at 6H (>2-fold and both ROS (>2 fold and HIV-1 replication (>3-fold after chronic treatment. The CSC mediated effects were associated with robust induction in the expression of CYP1A1 mRNA upon acute CSC treatment of U937 and U1 cells (>20-fold, and upon chronic CSC treatment to U1 cells (>30-fold. In addition, the CYP1A1 induction in U937 cells was mediated through the aromatic hydrocarbon receptor pathway. Lastly, CSC, which is known to increase viral replication in primary macrophages, was also found to induce CYP1 enzymes in HIV-infected primary macrophages. While mRNA levels of both CYP1A1 and CYP1B1 were elevated following CSC treatment, only CYP1B1 protein levels were increased in HIV-infected primary macrophages. In conclusion, these results suggest a possible association between oxidative stress, CYP1 expression, and viral replication in

  12. A sea urchin lectin, SUL-1, from the Toxopneustid sea urchin induces DC maturation from human monocyte and drives Th1 polarization in vitro.

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    Takei, Masao; Nakagawa, Hideyuki

    2006-05-15

    The sea urchin Toxopneustes pileolus belonging to the family Toxopneustidae, they have well-developed globiferous pedicellariae with pharmacologically active substances. We have purified a novel sea urchin lectin-1 (SUL-1) from the large globiferous pedicellariae of T. pileolus. Dendritic cells (DC) are professional APC and play a pivotal role in controlling immune responses. This study investigated whether SUL-1 can drive DC maturation from human immature monocyte-derived DC in vitro. Human monocytes were cultured with GM-CSF and IL-4 for 6 days followed by another 1 day in the presence of SUL-1 or LPS. DC harvested on day 7 were examined using functional assays. The expression levels of CD1a, CD80, CD83, CD86 and HLA-DR as expressed by mean fluorescence intensity (MFI) on DC differentiated from immature DC after culture with 1.0 microg/ml of SUL-1 for 1 day were enhanced and decreased endocytic activity. SUL-1-treated DC also displayed enhanced T cell stimulatory capacity in an MLR, as measured by T cell proliferation. Cell surface expression of CD80, CD83 and CD86 on SUL-1-treated DC was inhibited by anti-DC-SIGN mAb, while anti-DC-SIGN mAb had no influence on allogeneic T cell proliferation by SUL-1-treated DC. DC differentiated with SUL-1 induced the differentiation of naïve T cell towards a helper T cell type 1 (Th1) response at DC/T (1:5) cells ratio depending on IL-12 secretion. In CTL assay, the production of IFN-gamma and 51Cr release on SUL-1-treated DC were more augmented than of immature DC or LPS-treated DC. SUL-1-treated DC expressed CCR7 and had a high migration to MIP-3beta. Intracellular Ca2+ mobilization in SUL-1-treated DC was also induced by MIP-3beta. These results suggest that SUL-1 bindings to DC-SIGN on surface of immature DC may lead to differentiate DC from immature DC. Moreover, it suggests that SUL-1 may be used on DC-based vaccines for cancer immunotherapy. PMID:16197973

  13. A sea urchin lectin, SUL-1, from the Toxopneustid sea urchin induces DC maturation from human monocyte and drives Th1 polarization in vitro

    International Nuclear Information System (INIS)

    The sea urchin Toxopneustes pileolus belonging to the family Toxopneustidae, they have well-developed globiferous pedicellariae with pharmacologically active substances. We have purified a novel sea urchin lectin-1 (SUL-1) from the large globiferous pedicellariae of T. pileolus. Dendritic cells (DC) are professional APC and play a pivotal role in controlling immune responses. This study investigated whether SUL-1 can drive DC maturation from human immature monocyte-derived DC in vitro. Human monocytes were cultured with GM-CSF and IL-4 for 6 days followed by another 1 day in the presence of SUL-1 or LPS. DC harvested on day 7 were examined using functional assays. The expression levels of CD1a, CD80, CD83, CD86 and HLA-DR as expressed by mean fluorescence intensity (MFI) on DC differentiated from immature DC after culture with 1.0 μg/ml of SUL-1 for 1 day were enhanced and decreased endocytic activity. SUL-1-treated DC also displayed enhanced T cell stimulatory capacity in an MLR, as measured by T cell proliferation. Cell surface expression of CD80, CD83 and CD86 on SUL-1-treated DC was inhibited by anti-DC-SIGN mAb, while anti-DC-SIGN mAb had no influence on allogeneic T cell proliferation by SUL-1-treated DC. DC differentiated with SUL-1 induced the differentiation of naive T cell towards a helper T cell type 1 (Th1) response at DC/T (1:5) cells ratio depending on IL-12 secretion. In CTL assay, the production of IFN-γ and 51Cr release on SUL-1-treated DC were more augmented than of immature DC or LPS-treated DC. SUL-1-treated DC expressed CCR7 and had a high migration to MIP-3β. Intracellular Ca2+ mobilization in SUL-1-treated DC was also induced by MIP-3β. These results suggest that SUL-1 bindings to DC-SIGN on surface of immature DC may lead to differentiate DC from immature DC. Moreover, it suggests that SUL-1 may be used on DC-based vaccines for cancer immunotherapy

  14. Interaction between M-CSF and IL-10 on productions of IL-12 and IL-18 and expressions of CD14, CD23, and CD64 by human monocytes

    Institute of Scientific and Technical Information of China (English)

    Xiao-hui JI; Ting YAO; Jun-chuan QIN; Shu-kui WANG; Hui-juan WANG; Kun YAO

    2004-01-01

    AIM: To Study the interaction of macrophage colony-stimulating factor (M-CSF) and interleukin-10 (IL-10) in productions of IL-12 and IL-18 and expressions of CD14, CD23, and CD64 by human monocytes. METHODS:Purified adherent human monocytes were cultured with M-CSF or IL-10 alone, or with M-CSF+IL-10 and 2-3d later, the culture supernatants and cells were separated and collected. IL-12P40 and IL-18 levels in the supernatants were determined by ELISA and the percentages of CD14, CD23, and CD64 positive cells were examined by flow cytometry. RESULTS: (1) IL-10 decreased M-CSF-induced IL-18 levels, while M-CSF further reduced IL-12P40 level in the culture supernatants of IL-10-treated monocytes; (2) IL-10 alone had no effect on the percentage of CD14-positive cells, but further increased the percentage of CD14-positive cells induced by M-CSF; M-CSF alone had no effect on the percentage of CD64-positive cells, but further increased the percentage of CD64-positive cells induced by IL-10; (3) IL-10 decreased the percentage of CD23-positive cells induced by M-CSF.CONCLUSION: Between M-CSF and IL-10, there were antagonistic effects on inducing IL-18 and CD23 expressions by monocytes; there were also synergistic effects on inhibiting IL-12P40 production and inducing CD 14 and CD64 expressions by monocytes.

  15. Artesunate induces G0/G1 cell cycle arrest and iron-mediated mitochondrial apoptosis in A431 human epidermoid carcinoma cells.

    Science.gov (United States)

    Jiang, Zhongyong; Chai, Jin; Chuang, Henry Hon Fung; Li, Shifeng; Wang, Tianran; Cheng, Yi; Chen, Wensheng; Zhou, Deshan

    2012-07-01

    The anticancer effects of artesunate (ART) have been well documented. However, its potential against skin cancer has not been explored yet. Herein we reported that 60 μmol/l ART effectively inhibited A431 (human epidermoid carcinoma cells) growth but not that of HaCaT (normal human keratinocyte cells). Our results revealed that ART induced cell cycle arrest at G0/G1 phase through the downregulation of cyclin A1, cyclin B, cyclin D1, Cdk2, Cdk4, and Cdk6. This correlated with the upregulation of p21 and p27. The 5-bromodeoxyuridine incorporation assay also indicated that ART treatment reduced DNA synthesis in a time-dependent manner. Furthermore, ART induced mitochondrial apoptosis, as evidenced by annexin V/propidium iodide staining and western blot analysis. Interestingly, ART-induced apoptosis diminished under iron-deficient conditions but intensified under iron-overload conditions. Taken together, these findings demonstrated the potential of ART in treating skin cancer through the induction of G0/G1 cell cycle arrest and iron-mediated mitochondrial apoptosis and supported further investigations in other test systems. PMID:22421370

  16. Anti-Tumor Effect of Rutin on Human Neuroblastoma Cell Lines through Inducing G2/M Cell Cycle Arrest and Promoting Apoptosis

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    Hongyan Chen

    2013-01-01

    Full Text Available Aims. To further investigate the antineuroblastoma effect of rutin which is a type of flavonoid. Methods. The antiproliferation of rutin in human neuroblastoma cells LAN-5 were detected by 3-(4,5-dimethylthiazol-2-yl-2,5-diphenyltetrazolium bromide (MTT assay. Chemotaxis of LAN-5 cells was assessed using transwell migration chambers and scratch wound migration assay. The cell cycle arrest and apoptosis in a dose-dependent manner was measured by flow cytometric and fluorescent microscopy analyses. The apoptosis-related proteins BAX and BCL2 as well as MYCN mRNA express were determined by RT-PCR analysis. Secreted TNF-α level were determined using specific enzyme-linked immunosorbent assay kits. Results. Rutin significantly inhibited the growth of LAN-5 cells and chemotactic ability. Flow cytometric analysis revealed that rutin induced G2/M arrest in the cell cycle progression and induced cell apoptosis. The RT-PCR showed that rutin could decrease BCL2 expression and BCL2/BAX ratio. In the meantime, the MYCN mRNA level and the secretion of TNF-α were inhibited. Conclusion. These results suggest that rutin produces obvious antineuroblastoma effects via induced G2/M arrest in the cell cycle progression and induced cell apoptosis as well as regulating the expression of gene related to apoptosis and so on. It supports the viability of developing rutin as a novel therapeutic prodrug for neuroblastoma treatment, as well as providing a new path on anticancer effect of Chinese traditional drug.

  17. Cytotoxicity and Proliferation Studies with Arsenic in Established Human Cell Lines: Keratinocytes, Melanocytes, Dendritic Cells, Dermal Fibroblasts, Microvascular Endothelial Cells, Monocytes and T-Cells

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    Hari H. P. Cohly

    2003-01-01

    Full Text Available Abstract: Based on the hypothesis that arsenic exposure results in toxicity and mitogenecity, this study examined the dose-response of arsenic in established human cell lines of keratinocytes (HaCaT, melanocytes (1675, dendritic cells (THP-1/A23187, dermal fibroblasts (CRL1904, microvascular endothelial cells (HMEC, monocytes (THP-1, and T cells (Jurkat. Cytotoxicity was determined by incubating THP-1, THP-1+ A23187 and JKT cells in RPMI 1640, 1675 in Vitacell, HMEC in EBM, and dermal fibroblasts and HaCaT in DMEM with 10% fetal bovine serum, 1% streptomycin and penicillin for 72 hrs in 96-well microtiter plates, at 37oC in a 5% CO2 incubator with different concentrations of arsenic using fluorescein diacetate (FDA. Cell proliferation in 96-well plates was determined in cultured cells starved by prior incubation for 24 hrs in 1% FBS and exposed for 72 hours, using the 96 cell titer proliferation solution (Promega assay. Cytotoxicity assays yielded LD50s of 9 μg/mL for HaCaT, 1.5 μg/mL for CRL 1675, 1.5 μg/mL for dendritic cells, 37 μg/mL for dermal fibroblasts, 0.48 μg/mL for HMEC, 50 μg/mL for THP-1 cells and 50 μg/mL for JKT-T cells. The peak proliferation was observed at 6 μg/mL for HaCaT and THP-1 cells, 0.19 μg/mL for CRL 1675, dendritic cells, and HMEC, and 1.5 μg/mL for dermal fibroblasts and Jurkat T cells. These results show that arsenic is toxic at high doses to keratinocytes, fibroblasts, monocytes and T cells, and toxic at lower doses to melanocytes, microvascular endothelial cells and dendritic cells. Proliferation studies showed sub-lethal doses of arsenic to be mitogenic.

  18. Modern lineages of Mycobacterium tuberculosis exhibit lineage-specific patterns of growth and cytokine induction in human monocyte-derived macrophages.

    Directory of Open Access Journals (Sweden)

    Rajesh Sarkar

    Full Text Available BACKGROUND: Strains of Mycobacterium tuberculosis vary in virulence. Strains that have caused outbreaks in the United States and United Kingdom have been shown to subvert the innate immune response as a potential immune evasion mechanism. There is, however, little information available as to whether these patterns of immune subversion are features of individual strains or characteristic of broad clonal lineages of M. tuberculosis. METHODS: Strains from two major modern lineages (lineage 2 [East-Asian] and lineage 4 [Euro-American] circulating in the Western Cape in South Africa as well as a comparator modern lineage (lineage 3 [CAS/Delhi] were identified. We assessed two virulence associated characteristics: mycobacterial growth (in liquid broth and monocyte derived macrophages and early pro-inflammatory cytokine induction. RESULTS: In liquid culture, Lineage 4 strains grew more rapidly and reached higher plateau levels than other strains (lineage 4 vs. lineage 2 p=0.0024; lineage 4 vs. lineage 3 p=0.0005. Lineage 3 strains were characterized by low and early plateau levels, while lineage 2 strains showed an intermediate growth phenotype. In monocyte-derived macrophages, lineage 2 strains grew faster than lineage 3 strains (p<0.01 with lineage 4 strains having an intermediate phenotype. Lineage 2 strains induced the lowest levels of pro-inflammatory TNF and IL-12p40 as compared to other lineages (lineage 2: median TNF 362 pg/ml, IL-12p40 91 pg/ml; lineage 3: median TNF 1818 pg/ml, IL-12p40 123 pg/ml; lineage 4: median TNF 1207 pg/ml, IL-12p40 205 pg/ml;. In contrast, lineage 4 strains induced high levels of IL-12p40 and intermediate level of TNF. Lineage 3 strains induced high levels of TNF and intermediate levels of IL-12p40. CONCLUSIONS: Strains of M. tuberculosis from the three major modern strain lineages possess distinct patterns of growth and cytokine induction. Rapid growth and immune subversion may be key characteristics to the success

  19. Silencing of RTKN2 by siRNA suppresses proliferation, and induces G1 arrest and apoptosis in human bladder cancer cells.

    Science.gov (United States)

    Liao, Yi-Xiang; Zeng, Jin-Min; Zhou, Jia-Jie; Yang, Guang-Hua; Ding, Kun; Zhang, Xian-Jue

    2016-06-01

    Human bladder cancer is the most common urological malignancy in China. One of the causes of carcinogenesis in the cancer may be gene mutation. Therefore, the present study investigated the expression levels of Rhotekin 2 (RTKN2), a Rho effector protein, in human bladder cancer tissues and cell lines, and examined the effect of RTKN2 on the proliferation, cell cycle, apoptosis and invasion of human bladder cancer cell lines. The mRNA expression levels of RTKN2 in 30 human bladder cancer tissue samples were significantly higher, compared with those in 30 normal human bladder tissue samples. The protein expression levels of RTKN2 was markedly higher in T24 and 5637 cells, compared with those in four other human bladder cancer cell lines. The silencing of RTKN2 by small interfering (si)RNA inhibited cell proliferation and arrested cell cycle at the G1 phase, via reducing the expression levels of the MCM10, CDK2, CDC24A and CDC6 cell cycle‑associated proteins in the T24 and 5637 cells. Furthermore, RTKN2 knockdown in the cells led to cell apoptosis and the suppression of invasion. These results suggested that RTKN2 is involved in the carcinogenesis and progression of human bladder cancer, indicating that RTKN2 may be a molecular target in cancer therapy. PMID:27082503

  20. Endogenous production of tumour necrosis factor is required for manganese superoxide dismutase expression by irradiation in the human monocytic cell line THP-1

    International Nuclear Information System (INIS)

    Manganese superoxide dismutase (MnSOD) is a mitochondrial enzyme that scavenges superoxide (O2-) ions. We studied the regulation of MnSOD gene expression by irradiation and the mechanisms in human monocytic cell line THP-1. We found that irradiation induced expression of the MnSOD gene through the autocrine mechanism, involving the production of tumour necrosis factor (TNF). Irradiation increased TNF production in THP-1 cells, and TNF increased the levels of MnSOD transcripts. Supernatant from irradiated THP-1 cells induced the expression of MnSOD mRNA, and anti-TNF antibody blocked the induction of MnSOD mRNA. Irradiation also increased the levels of MnSOD mRNA in other myelocytic cell lines, HL60 and KG-1, and the ovarian cancer cell line SK-OV-3. Our results indicate that the endogenous production of TNF is required, at least in part, for the induction of MnSOD mRNA expression by irradiation in THP-1 cells, and the increased levels of MnSOD transcripts on irradiation occur through a pathway involving protein kinase C activation. Our results also indicate that the increase in MnSOD mRNA caused by irradiation is regulated by both transcriptional and post-transcriptional mechanisms. (author)

  1. Comparative DNA microarray analysis of human monocyte derived dendritic cells and MUTZ-3 cells exposed to the moderate skin sensitizer cinnamaldehyde

    International Nuclear Information System (INIS)

    The number of studies involved in the development of in vitro skin sensitization tests has increased since the adoption of the EU 7th amendment to the cosmetics directive proposing to ban animal testing for cosmetic ingredients by 2013. Several studies have recently demonstrated that sensitizers induce a relevant up-regulation of activation markers such as CD86, CD54, IL-8 or IL-1β in human myeloid cell lines (e.g., U937, MUTZ-3, THP-1) or in human peripheral blood monocyte-derived dendritic cells (PBMDCs). The present study aimed at the identification of new dendritic cell activation markers in order to further improve the in vitro evaluation of the sensitizing potential of chemicals. We have compared the gene expression profiles of PBMDCs and the human cell line MUTZ-3 after a 24-h exposure to the moderate sensitizer cinnamaldehyde. A list of 80 genes modulated in both cell types was obtained and a set of candidate marker genes was selected for further analysis. Cells were exposed to selected sensitizers and non-sensitizers for 24 h and gene expression was analyzed by quantitative real-time reverse transcriptase-polymerase chain reaction. Results indicated that PIR, TRIM16 and two Nrf2-regulated genes, CES1 and NQO1, are modulated by most sensitizers. Up-regulation of these genes could also be observed in our recently published DC-activation test with U937 cells. Due to their role in DC activation, these new genes may help to further refine the in vitro approaches for the screening of the sensitizing properties of a chemical.

  2. Effects of 9cis,11trans and 10trans,12cis CLA on osteoclast formation and activity from human CD14+ monocytes

    Directory of Open Access Journals (Sweden)

    El-Sohemy Ahmed

    2009-04-01

    Full Text Available Abstract Background Mixed CLA isomers variably affect bone resorption in animals and decrease osteoclast formation and activity in murine osteoclasts. These variable effects may be due to the different isomers present in commercial preparations of CLA, and the effects of the predominant individual isomers, 9cis,11trans (9,11 and 10trans,12cis (10,12 CLA are not clear. The objectives of this study were to determine the effects of the individual CLA isomers on osteoclast formation and activity from human CD14+ monocytes, and to determine whether any changes are accompanied by changes in cathepsin K, matrix metalloproteinase-9 (MMP-9, receptor activator of NF-κB (RANK and tumour necrosis factor alpha (TNFα gene expression. Osteoclasts were identified as TRAP+ multinucleated cells. Osteoclast activity was quantified by the amount of TRAP in the cultured media. Results At 50 μM, 9,11 CLA inhibited osteoclast formation by ~70%, and both 9,11 and 10,12 CLA decreased osteoclast activity by ~85–90%. Both isomers inhibited cathepsin K (50 μM 9,11 by ~60%; 10,12 by ~50% and RANK (50 μM 9,11 by ~85%; 50 μM 10,12 by ~65% expression, but had no effect on MMP-9 or TNFα expression. Conclusion 9,11 CLA inhibits osteoclast formation and activity from human cells, suggesting that this isomer may prevent bone resorption in humans. Although 10,12 CLA did not significantly reduce osteoclast formation, it reduced osteoclast activity and cathepsin K and RANK expression, suggesting that this isomer may also affect bone resorption.

  3. Human metapneumovirus M2-2 protein inhibits innate immune response in monocyte-derived dendritic cells.

    Directory of Open Access Journals (Sweden)

    Junping Ren

    Full Text Available Human metapneumovirus (hMPV is a leading cause of lower respiratory infection in young children, the elderly and immunocompromised patients. Repeated hMPV infections occur throughout life. However, immune evasion mechanisms of hMPV infection are largely unknown. Recently, our group has demonstrated that hMPV M2-2 protein, an important virulence factor, contributes to immune evasion in airway epithelial cells by targeting the mitochondrial antiviral-signaling protein (MAVS. Whether M2-2 regulates the innate immunity in human dendritic cells (DC, an important family of immune cells controlling antigen presenting, is currently unknown. We found that human DC infected with a virus lacking M2-2 protein expression (rhMPV-ΔM2-2 produced higher levels of cytokines, chemokines and IFNs, compared to cells infected with wild-type virus (rhMPV-WT, suggesting that M2-2 protein inhibits innate immunity in human DC. In parallel, we found that myeloid differentiation primary response gene 88 (MyD88, an essential adaptor for Toll-like receptors (TLRs, plays a critical role in inducing immune response of human DC, as downregulation of MyD88 by siRNA blocked the induction of immune regulatory molecules by hMPV. Since M2-2 is a cytoplasmic protein, we investigated whether M2-2 interferes with MyD88-mediated antiviral signaling. We found that indeed M2-2 protein associated with MyD88 and inhibited MyD88-dependent gene transcription. In this study, we also identified the domains of M2-2 responsible for its immune inhibitory function in human DC. In summary, our results demonstrate that M2-2 contributes to hMPV immune evasion by inhibiting MyD88-dependent cellular responses in human DC.

  4. Ganoderic acid DM, a natural triterpenoid, induces DNA damage, G1 cell cycle arrest and apoptosis in human breast cancer cells.

    Science.gov (United States)

    Wu, Guo-Sheng; Lu, Jin-Jian; Guo, Jia-Jie; Li, Ying-Bo; Tan, Wen; Dang, Yuan-Ye; Zhong, Zhang-Feng; Xu, Zeng-Tao; Chen, Xiu-Ping; Wang, Yi-Tao

    2012-03-01

    Ganoderic acid DM (GADM) is a triterpenoid isolated from Ganoderma lucidum, a well-known edible medicinal mushroom. In the present study, we found that GADM effectively inhibited cell proliferation and colony formation in MCF-7 human breast cancer cells, which was much stronger than that of MDA-MB-231 breast cancer cells. GADM both concentration- and time-dependently mediated G1 cell cycle arrest and significantly decreased the protein level of CDK2, CDK6, cycle D1, p-Rb and c-Myc in MCF-7 cells. Moreover, GADM obviously induced DNA fragmentation and cleavage of PARP which are the characteristics of apoptosis and decreased the mitochondrial membrane potential in MCF-7 cells. Besides, we also showed that GADM elicited DNA damage as measured by comet assay which is a sensitive method for DNA damage detection. γ-H2AX, a marker of DNA damage, was also slightly up-regulated after treated with GADM for 6h, suggesting that the G1 cell cycle arrest and apoptosis induced by GADM may be partially resulted from GADM-induced DNA damage. These results have advanced our current understandings of the anti-cancer mechanisms of GADM. PMID:22178684

  5. Identification of MicroRNAs Involved in Growth Arrest and Apoptosis in Hydrogen Peroxide-Treated Human Hepatocellular Carcinoma Cell Line HepG2

    Directory of Open Access Journals (Sweden)

    Yuan Luo

    2016-01-01

    Full Text Available Although both oxidative stress and microRNAs (miRNAs play vital roles in physiological and pathological processes, little is known about the interactions between them. In this study, we first described the regulation of H2O2 in cell viability, proliferation, cycle, and apoptosis of human hepatocellular carcinoma cell line HepG2. Then, miRNAs expression was profiled after H2O2 treatment. The results showed that high concentration of H2O2 (600 μM could decrease cell viability, inhibit cell proliferation, induce cell cycle arrest, and finally promote cell apoptosis. Conversely, no significant effects could be found under treatment with low concentration (30 μM. miRNAs array analysis identified 131 differentially expressed miRNAs (125 were upregulated and 6 were downregulated and predicted 13504 putative target genes of the deregulated miRNAs. Gene ontology (GO analysis revealed that the putative target genes were associated with H2O2-induced cell growth arrest and apoptosis. The subsequent bioinformatics analysis indicated that H2O2-response pathways, including MAPK signaling pathway, apoptosis, and pathways in cancer and cell cycle, were significantly affected. Overall, these results provided comprehensive information on the biological function of H2O2 treatment in HepG2 cells. The identification of miRNAs and their putative targets may offer new diagnostic and therapeutic strategies for liver cancer.

  6. Enhanced induction of cell cycle arrest and apoptosis via the mitochondrial membrane potential disruption in human U87 malignant glioma cells by aloe emodin.

    Science.gov (United States)

    Ismail, Samhani; Haris, Khalilah; Abdul Ghani, Abdul Rahman Izaini; Abdullah, Jafri Malin; Johan, Muhammad Farid; Mohamed Yusoff, Abdul Aziz

    2013-09-01

    Aloe emodin, one of the active compounds found in Aloe vera leaves, plays an important role in the regulation of cell growth and death. It has been reported to promote the anti-cancer effects in various cancer cells by inducing apoptosis. However, the mechanism of inducing apoptosis by this agent is poorly understood in glioma cells. This research is to investigate the apoptosis and cell cycle arrest inducing by aloe emodin on U87 human malignant glioma cells. Aloe emodin showed a time- and dose-dependent inhibition of U87 cells proliferation and decreased the percentage of viable U87 cells via the induction of apoptosis. Characteristic morphological changes, such as the formation of apoptotic bodies, were observed with confocal microscope by Annexin V-FITC/PI staining, supporting our viability study and flow cytometry analysis results. Our data also demonstrated that aloe emodin arrested the cell cycle in the S phase and promoted the loss of mitochondrial membrane potential in U87 cells that indicated the early event of the mitochondria-induced apoptotic pathway. PMID:23869465

  7. Monocytes mediate metastatic breast tumor cell adhesion to endothelium under flow

    OpenAIRE

    Evani, Shankar J.; Prabhu, Rajesh G.; Gnanaruban, V.; Finol, Ender A.; Anand K. Ramasubramanian

    2013-01-01

    Endothelial adhesion is necessary for the hematogenous dissemination of tumor cells. However, the metastatic breast tumor cell MDA-MB-231 does not bind to the endothelium under physiological flow conditions, suggesting alternate mechanisms of adhesion. Since monocytes are highly represented in the tumor microenvironment, and also bind to endothelium during inflammation, we hypothesized that the monocytes assist in the arrest of MDA-MB-231 on the endothelium. Using in vitro models of the dynam...

  8. Human cytochrome c enters murine J774 cells and causes G1 and G2/M cell cycle arrest and induction of apoptosis

    International Nuclear Information System (INIS)

    Cytochrome c is well known as a carrier of electrons during respiration. Current evidence indicates that cytochrome c also functions as a major component of apoptosomes to induce apoptosis in eukaryotic cells as well as an antioxidant. More recently, a prokaryotic cytochrome c, cytochrome c 551 from Pseudomonas aeruginosa, has been shown to enter in mammalian cells such as the murine macrophage-like J774 cells and causes inhibition of cell cycle progression. Much less is known about such functions by mammalian cytochromes c, particularly the human cytochrome c. We now report that similar to P. aeruginosa cytochrome c 551, the purified human cytochrome c protein can enter J774 cells and induce cell cycle arrest at the G1 to S phase, as well as at the G2/M phase at higher concentrations. Unlike P. aeruginosa cytochrome c 551 which had no effect on the induction of apoptosis, human cytochrome c induces significant apoptosis and cell death in J774 cells, presumably through inhibition of the cell cycle at the G2/M phase. When incubated with human breast cancer MCF-7 and normal mammary epithelial cell line MCF-10A1 cells, human cytochrome c entered in both types of cells but induced cell death only in the normal MCF-10A1 cells. The ability of human cytochrome c to enter J774 cells was greatly reduced at 4 deg. C, suggesting energy requirement in the entry process

  9. Low Doses of Cisplatin Induce Gene Alterations, Cell Cycle Arrest, and Apoptosis in Human Promyelocytic Leukemia Cells.

    Science.gov (United States)

    Velma, Venkatramreddy; Dasari, Shaloam R; Tchounwou, Paul B

    2016-01-01

    Cisplatin is a known antitumor drug, but its mechanisms of action are not fully elucidated. In this research, we studied the anticancer potential of cisplatin at doses of 1, 2, or 3 µM using HL-60 cells as a test model. We investigated cisplatin effects at the molecular level using RNA sequencing, cell cycle analysis, and apoptotic assay after 24, 48, 72, and 96 hours of treatment. The results show that many genes responsible for molecular and cellular functions were significantly altered. Cisplatin treatment also caused the cells to be arrested at the DNA synthesis phase, and as the time increases, the cells gradually accumulated at the sub-G1 phase. Also, as the dose increases, a significant number of cells entered into the apoptotic and necrotic stages. Altogether, the data show that low doses of cisplatin significantly impact the viability of HL-60 cells, through modulation of gene expression, cell cycle, and apoptosis. PMID:27594783

  10. In vivo imaging of monocyte trafficking with 18F-fluorodeoxyglucose labeled monocytes

    International Nuclear Information System (INIS)

    Since the ability to monitor in vivo monocyte trafficking would contribute to our understanding of the pathophysiology of various inflammatory disorders, we investigated the feasibility of labeling human monocytes with 18F-FDG. Human monocytes were separated by Ficoll/Hypaque gradient and purity was assessed by flow cytometry. The influence of insulin and/or glucose on labeling efficiency was evaluated. Cell viability and activation was measured with trypan blue exclusion and hydrogen peroxide assays, respectively. Label stability was measured for up to 18 hr, and the effect of insulin pre-incubation on FDG washout was investigated. PET images were acquired in SD rats at various time points after injection of FDG labeled monocytes. Monocytes were >85% pure, and labeling efficiency was 35% for 1x106 cells after 40 min incubation with 2 mCi 18F-FDG without insulin. Pre-incubation with 10∼100 nM insulin significantly increased FDG uptake which reached 400% of baseline levels, whereas presence of glucose or serum decreased FDG uptake. Labeled cells were >90% viable for up to 22 hr, and the labeling process did appear to significantly activate cells, Washout studies however, demonstrated gradual washout of the FDG from monocytes after initial uptake PET images of FDG labeled monocytes in SD rats showed consistent findings. Utilizing insulin effects on cellular glucose metabolism may be a feasible way of labeling monocytes with 18F-FDG for PET imaging. However, gradual washout of FDG after initial uptake poses as a potential problem which needs to be addressed before practical application

  11. MiR-107 and MiR-185 can induce cell cycle arrest in human non small cell lung cancer cell lines.

    Directory of Open Access Journals (Sweden)

    Yukari Takahashi

    Full Text Available BACKGROUND: MicroRNAs (miRNAs are short single stranded noncoding RNAs that suppress gene expression through either translational repression or degradation of target mRNAs. The annealing between messenger RNAs and 5' seed region of miRNAs is believed to be essential for the specific suppression of target gene expression. One miRNA can have several hundred different targets in a cell. Rapidly accumulating evidence suggests that many miRNAs are involved in cell cycle regulation and consequentially play critical roles in carcinogenesis. METHODOLOGY/PRINCIPAL FINDINGS: Introduction of synthetic miR-107 or miR-185 suppressed growth of the human non-small cell lung cancer cell lines. Flow cytometry analysis revealed these miRNAs induce a G1 cell cycle arrest in H1299 cells and the suppression of cell cycle progression is stronger than that by Let-7 miRNA. By the gene expression analyses with oligonucleotide microarrays, we find hundreds of genes are affected by transfection of these miRNAs. Using miRNA-target prediction analyses and the array data, we listed up a set of likely targets of miR-107 and miR-185 for G1 cell cycle arrest and validate a subset of them using real-time RT-PCR and immunoblotting for CDK6. CONCLUSIONS/SIGNIFICANCE: We identified new cell cycle regulating miRNAs, miR-107 and miR-185, localized in frequently altered chromosomal regions in human lung cancers. Especially for miR-107, a large number of down-regulated genes are annotated with the gene ontology term 'cell cycle'. Our results suggest that these miRNAs may contribute to regulate cell cycle in human malignant tumors.

  12. Zerumbone-loaded nanostructured lipid carrier induces G2/M cell cycle arrest and apoptosis via mitochondrial pathway in a human lymphoblastic leukemia cell line

    Directory of Open Access Journals (Sweden)

    Rahman HS

    2014-01-01

    Full Text Available Heshu Sulaiman Rahman,1–3 Abdullah Rasedee,1,2 Ahmad Bustamam Abdul,2,4 Nazariah Allaudin Zeenathul,1,2 Hemn Hassan Othman,1,3 Swee Keong Yeap,2 Chee Wun How,2 Wan Abd Ghani Wan Nor Hafiza4,51Faculty of Veterinary Medicine, 2Institute of Bioscience, Universiti Putra Malaysia, Selangor, Malaysia; 3Faculty of Veterinary Medicine, University of Sulaimanyah, Sulaimanyah City, Kurdistan Region, Northern Iraq; 4Faculty of Medicine and Health Science, Universiti Putra Malaysia, Selangor, Malaysia; 5College of Medical Laboratory Technology, Institute for Medical Research, Kuala Lumpur, MalaysiaAbstract: This investigation evaluated the antileukemia properties of a zerumbone (ZER-loaded nanostructured lipid carrier (NLC prepared by hot high-pressure homogenization techniques in an acute human lymphoblastic leukemia (Jurkat cell line in vitro. The apoptogenic effect of the ZER-NLC on Jurkat cells was determined by fluorescent and electron microscopy, Annexin V-fluorescein isothiocyanate, Tdt-mediated dUTP nick-end labeling assay, cell cycle analysis, and caspase activity. An MTT (3-(4,5-dimethylthiazol-2-yl-2,5 diphenyltetrazolium bromide assay showed that ZER-NLC did not have adverse effects on normal human peripheral blood mononuclear cells. ZER-NLC arrested the Jurkat cells at G2/M phase with inactivation of cyclin B1 protein. The study also showed that the antiproliferative effect of ZER-NLC on Jurkat cells is through the intrinsic apoptotic pathway via activation of caspase-3 and caspase-9, release of cytochrome c from the mitochondria into the cytosol, and subsequent cleavage of poly (adenosine diphosphate-ribose polymerase (PARP. These findings show that the ZER-NLC is a potentially useful treatment for acute lymphoblastic leukemia in humans.Keywords: zerumbone-loaded nanostructured lipid carrier, cell cycle arrest, apoptosis, mitochondrial pathway

  13. Helicobacter pylori induces IL-1β and IL-18 production in human monocytic cell line through activation of NLRP3 inflammasome via ROS signaling pathway.

    Science.gov (United States)

    Li, Xiang; Liu, Sheng; Luo, Jingjing; Liu, Anyuan; Tang, Shuangyang; Liu, Shuo; Yu, Minjun; Zhang, Yan

    2015-06-01

    This study investigated whether Helicobacter pylori could activate the nucleotide-binding oligomerization domain-like receptor (NLR) family, pyrin domain-containing 3 (NLRP3) inflammasome in human macrophages and the involvement of reactive oxygen species (ROS) in inflammasome activation. Phorbol-12-myristate-13-acetate (PMA)-differentiated human acute monocytic leukemia cell line THP-1 was infected with H. pylori. The levels of pro-inflammatory cytokines interleukin (IL)-1β and IL-18 in supernatant were measured by ELISA. Intracellular ROS level was analyzed by flow cytometry. Quantitative real-time PCR and western blot analysis were employed to determine the mRNA and protein expression levels of NLRP3 and caspase-1 in THP-1 cells, respectively. Our results showed that H. pylori infection could induce IL-1β and IL-18 production in PMA-differentiated THP-1 cells in a dose- and time-dependent manner. Moreover, secretion of IL-1β and IL-18 in THP-1 cells following H. pylori infection was remarkably reduced by NLRP3-specific small interfering RNA treatment. In addition, the intracellular ROS level was elevated by H. pylori infection, which could be eliminated by the ROS scavenger N-acetylcysteine (NAC). Furthermore, NAC treatment could inhibit NLRP3 inflammasome formation and caspase-1 activation and suppress the release of IL-1β and IL-18 from H. pylori-infected THP-1 cells. These findings provide novel insights into the innate immune response against H. pylori infection, which could potentially be used for the prevention and treatment of H. pylori-related diseases. PMID:25834143

  14. Phenolic glycolipid-1 of Mycobacterium leprae binds complement component C3 in serum and mediates phagocytosis by human monocytes

    OpenAIRE

    1991-01-01

    Previous studies from this laboratory have demonstrated that Mycobacterium leprae, an obligate intracellular bacterial parasite, enters human mononuclear phagocytes via complement receptors on these host cells and bacterium-bound C3. The present study investigates the role of M. leprae surface molecules in C3 fixation and phagocytosis. By enzyme-linked immunosorbent assay, C3 binds selectively to phenolic glycolipid-1 (PGL-1), a major surface molecule of the leprosy bacillus. C3 fixation to P...

  15. Gene induction during differentiation of human monocytes into dendritic cells: an integrated study at the RNA and protein levels

    OpenAIRE

    Angénieux, Catherine; Fricker, Dominique; Strub, Jean-Marc; Luche, Sylvie; Bausinger, Huguette; Cazenave, Jean-Pierre; Van Dorsselaer, Alain; Hanau, Daniel; de la Salle, Henri; Rabilloud, Thierry

    2006-01-01

    Changes in gene expression occurring during differentiation of human monoytes into dendritic cells were studied at the RNA and protein levels. These studies showed the induction of several gene classes corresponding to various biological functions. These functions encompass of course antigen processing and presentation, cytoskeleton, cell signalling and signal transduction, but also an increase of mitochondrial function and of the protein synthesis machinery, including some, but not all, chap...

  16. Fatal Monocytic Ehrlichiosis in Woman, Mexico, 2013.

    Science.gov (United States)

    Sosa-Gutierrez, Carolina G; Solorzano-Santos, Fortino; Walker, David H; Torres, Javier; Serrano, Carlos A; Gordillo-Perez, Guadalupe

    2016-05-01

    Human monocytic ehrlichiosis is a febrile illness caused by Ehrlichia chaffeensis, an intracellular bacterium transmitted by ticks. In Mexico, a case of E. chaffeensis infection in an immunocompetent 31-year-old woman without recognized tick bite was fatal. This diagnosis should be considered for patients with fever, leukopenia, thrombocytopenia, and elevated liver enzyme levels. PMID:27088220

  17. Fatal Monocytic Ehrlichiosis in Woman, Mexico, 2013

    Science.gov (United States)

    Sosa-Gutierrez, Carolina G.; Solorzano-Santos, Fortino; Walker, David H.; Torres, Javier; Serrano, Carlos A.

    2016-01-01

    Human monocytic ehrlichiosis is a febrile illness caused by Ehrlichia chaffeensis, an intracellular bacterium transmitted by ticks. In Mexico, a case of E. chaffeensis infection in an immunocompetent 31-year-old woman without recognized tick bite was fatal. This diagnosis should be considered for patients with fever, leukopenia, thrombocytopenia, and elevated liver enzyme levels. PMID:27088220

  18. The soluble recombinant Neisseria meningitidis adhesin NadA(Δ351-405) stimulates human monocytes by binding to extracellular Hsp90.

    Science.gov (United States)

    Cecchini, Paola; Tavano, Regina; Polverino de Laureto, Patrizia; Franzoso, Susanna; Mazzon, Cristina; Montanari, Paolo; Papini, Emanuele

    2011-01-01

    The adhesin NadA favors cell adhesion/invasion by hypervirulent Neisseria meningitidis B (MenB). Its recombinant form NadA(Δ351-405,) devoid of the outer membrane domain, is an immunogenic candidate for an anti-MenB vaccine able to stimulate monocytes, macrophages and dendritic cells. In this study we investigated the molecular mechanism of NadA(Δ351-405) cellular effects in monocytes. We show that NadA(Δ351-405) (against which we obtained polyclonal antibodies in rabbits), binds to hsp90, but not to other extracellular homologous heat shock proteins grp94 and hsp70, in vitro and on the surface of monocytes, in a temperature dependent way. Pre-incubation of monocytes with the MenB soluble adhesin interfered with the binding of anti-hsp90 and anti-hsp70 antibodies to hsp90 and hsp70 at 37°C, a condition in which specific cell-binding occurs, but not at 0°C, a condition in which specific cell-binding is very diminished. Conversely, pre-incubation of monocytes with anti-hsp90 and anti-hsp70 antibodies did not affected NadA(Δ351-405) cell binding in any temperature condition, indicating that it associates to another receptor on their plasma membrane and then laterally diffuses to encounter hsp90. Consistently, polymixin B interfered with NadA(Δ351-405) /hsp90 association, abrogated the decrease of anti-hsp90 antibodies binding to the cell surface due to NadA(Δ351-405) and inhibited adhesin-induced cytokine/chemokine secretion without affecting monocyte-adhesin binding. Co-stimulation of monocytes with anti-hsp90 antibodies and NadA(Δ351-405) determined a stronger but polymixin B insensitive cell activation. This indicated that the formation of a recombinant NadA/hsp90/hsp70 complex, although essential for full monocyte stimulation, can be replaced by anti-hsp90 antibody/hsp90 binding. Finally, the activation of monocytes by NadA(Δ351-405) alone or in the presence of anti-hsp90 antibodies were both inhibited by neutralizing anti-TLR4 antibodies, but not by

  19. Curcumin inhibits the proliferation of a human colorectal cancer cell line Caco-2 partially by both apoptosis and G2/M cell cycle arrest

    Directory of Open Access Journals (Sweden)

    Yohko Fujimoto

    2014-06-01

    Full Text Available The aim of this study was to assess the possible roles of the phytochemical compounds, curcumin, quercetin and resveratrol in the proliferation of human colorectal cancer cell line Caco-2. All three phytochemical compounds inhibited Caco-2 cell proliferation, with curcumin being more effective than quercetin and resveratrol. Investigations concerning DNA fragmentation in the nucleus, Bax and Bcl-2 mRNA expression levels, and caspase-3/7 activity indicated that curcumin induced apoptosis in Caco-2 cells through an increase in the Bax/Bcl-2 ratio and activation of caspase-3/7. Furthermore, the analysis of flow-cytometry showed that curcumin caused an arrest of G2/M phase in Caco-2 cells. These results suggest that curcumin suppresses Caco-2 proliferation partially via activation of the mitochondrial apoptotic pathway and cell cycle retardation.

  20. Characterization of the N-methoxyindole-3-carbinol (NI3C)–Induced Cell Cycle Arrest in Human Colon Cancer Cell Lines

    DEFF Research Database (Denmark)

    Neave, Antje S.; Sarup, Sussi; Seidelin, Michel; Duus, Fritz; Vang, Ole

    2005-01-01

    study was to show the effect of NI3C on cell growth of two human colon cancer cell lines, DLD-1 and HCT-116. For the first time it is shown that NI3C inhibits cellular growth of DLD-1 and HCT-116 and that NI3C is a more potent inhibitor of cell proliferation than I3C. In addition to the inhibition of...... cellular proliferation, NI3C caused an accumulation of HCT-116 cells in the G2/M phase, in contrast to I3C, which led to an accumulation of the colon cells in G0/G1 phase. Furthermore, NI3C delays the G1-S phase transition of synchronized HCT-116 cells. The indole-mediated cell-cycle arrest may be related...

  1. Differential regulation of the human immunodeficiency virus type 2 enhancer in monocytes at various stages of differentiation.

    OpenAIRE

    Hilfinger, J M; Clark, N; Smith, M.; Robinson, K; Markovitz, D M

    1993-01-01

    We have demonstrated that stimulation of the human immunodeficiency virus type 2 (HIV-2) enhancer in T cells is dependent upon at least four cis-acting elements, including two purine-rich binding sites, PuB1 and PuB2, which are capable of binding members of the ets family of proto-oncogenes, the pets (peri-ets) site, which lies just upstream of the PuB2 site, and a single kappa B site (D. M. Markovitz, M. Smith, J. M. Hilfinger, M. C. Hannibal, B. Petryniak, and G. J. Nabel, J. Virol. 66:5479...

  2. Oridonin nanosuspension was more effective than free oridonin on G2/M cell cycle arrest and apoptosis in the human pancreatic cancer PANC-1 cell line

    Directory of Open Access Journals (Sweden)

    Qi XL

    2012-04-01

    Full Text Available Xiaoli Qi1, Dianrui Zhang2, Xia Xu1, Feifei Feng2, Guijie Ren1, Qianqian Chu1, Qiang Zhang3, Keli Tian11Department of Biochemistry and Molecular Biology, Shandong University School of Medicine, Jinan, 2Department of Pharmaceutics, College of Pharmacy, Shandong University, Jinan, 3State Key Laboratory of Natural and Biomimetic Drugs, School of Pharmaceutical Sciences, Peking University, Beijing, People's Republic of ChinaAbstract: Oridonin, a diterpenoid isolated from Rabdosia rubescencs, has been reported to have antitumor effects. However, low solubility has limited its clinical applications. Preparation of drugs in the form of nanosuspensions is an extensively utilized protocol. In this study, we investigated the anticancer activity of oridonin and oridonin nanosuspension on human pancreatic carcinoma PANC-1 cells. 3-(4,5-dimethylthiazol-2-yl-2,5-diphenyltetrazolium bromide assay was performed to investigate the effect of oridonin on cell growth. Propidium iodide and Hoechst 33342 staining were used to detect morphologic changes. The percentage of apoptosis and cell cycle progression was determined by flow cytometric method staining with propidium iodide. Annexin V-fluorescein isothiocyanate (FITC/PI staining was used to evaluate cell apoptosis by flow cytometry. Caspase-3 activity was measured by spectrophotometry. The apoptotic and cell cycle protein expression were determined by Western blot analysis. Both oridonin and oridonin nanosuspension induced apoptosis and G2/M phase cell cycle arrest, and the latter had a more significant cytotoxic effect. The ratio of Bcl-2/Bax protein expression was decreased and caspase-3 activity was stimulated. The expression of cyclin B1 and p-cdc2 (T161 was suppressed. Our results showed that oridonin nanosuspension was more effective than free oridonin on G2/M cell cycle arrest and apoptosis in the human pancreatic cancer PANC-1 cell line.Keywords: cyclin B1, cdc2, caspase-3, Bcl-2, Bax

  3. Sinomenine prevents metastasis of human osteosarcoma cells via S phase arrest and suppression of tumor-related neovascularization and osteolysis through the CXCR4-STAT3 pathway.

    Science.gov (United States)

    Xie, Tao; Ren, Hai-Yong; Lin, Hai-Qing; Mao, Jin-Ping; Zhu, Ting; Wang, Sheng-Dong; Ye, Zhao-Ming

    2016-05-01

    Osteosarcoma is the most common primary malignant tumor of the bone. The long-term survivals continue to be unsatisfactory for patients with metastatic and recurrent disease. Metastasis is still a severe challenge in osteosarcoma treatment. Sinomenine, an alkaloid from traditional Chinese medicine, has been proved to possess potent antitumor and anti-invasion effect on various cancers. However, the effect of sinomenine on human osteosarcoma and the underlying mechanisms remains unknown. We report here that sinomenine inhibited proliferation by inducing S phase arrest and suppressing the clone formation. Significant inhibitory effects were found in invasion and metastasis in osteosarcoma, but little cytotoxicity was observed in tested concentrations. Exposure to sinomenine resulted in suppression of invasion and migration in osteosarcoma cells as well as tube formation ability in the human umbilical vein endothelial cells (HUVEC) and U2OS cells. Furthermore, it demonstrated that CXCR4 played a key role contributing to invasion in osteosarcoma which is considered to be a core target site in sinomenine treatment. Sinomenine inhibited invasion by suppressing CXCR4 and STAT3 phosphorylation then downregulating the expression of MMP-2, MMP-9, RANKL, VEGF downstream. In addition, then RANKL-mediated bone destruction stimulated by osteoclastogenesis and VEGF-related neovascularization were restrained. Importantly, in vivo, sinomenine suppressed proliferation, osteoclastogenesis and bone destruction. Through these various comprehensive means, sinomenine inhibits metastasis in osteosarcoma. Taken together, our results revealed that sinomenine caused S phase arrest, inhibited invasion and metastasis via suppressing the CXCR4-STAT3 pathway and then osteoclastogenesis-mediated bone destruction and neovascularization in osteosarcoma. Sinomenine is therefore a promising adjuvant agent for metastasis control in osteosarcoma. PMID:26983669

  4. The Putative Metal Coordination Motif in the Endonuclease Domain of Human Parvovirus B19 NS1 Is Critical for NS1 Induced S Phase Arrest and DNA Damage

    Directory of Open Access Journals (Sweden)

    Violetta Kivovich, Leona Gilbert, Matti Vuento, Stanley J. Naides

    2012-01-01

    Full Text Available The non-structural proteins (NS of the parvovirus family are highly conserved multi-functional molecules that have been extensively characterized and shown to be integral to viral replication. Along with NTP-dependent helicase activity, these proteins carry within their sequences domains that allow them to bind DNA and act as nucleases in order to resolve the concatameric intermediates developed during viral replication. The parvovirus B19 NS1 protein contains sequence domains highly similar to those previously implicated in the above-described functions of NS proteins from adeno-associated virus (AAV, minute virus of mice (MVM and other non-human parvoviruses. Previous studies have shown that transient transfection of B19 NS1 into human liver carcinoma (HepG2 cells initiates the intrinsic apoptotic cascade, ultimately resulting in cell death. In an effort to elucidate the mechanism of mammalian cell demise in the presence of B19 NS1, we undertook a mutagenesis analysis of the protein's endonuclease domain. Our studies have shown that, unlike wild-type NS1, which induces an accumulation of DNA damage, S phase arrest and apoptosis in HepG2 cells, disruptions in the metal coordination motif of the B19 NS1 protein reduce its ability to induce DNA damage and to trigger S phase arrest and subsequent apoptosis. These studies support our hypothesis that, in the absence of replicating B19 genomes, NS1-induced host cell DNA damage is responsible for apoptotic cell death observed in parvoviral infection of non-permissive mammalian cells.

  5. Α-MMC and MAP30, two ribosome-inactivating proteins extracted from Momordica charantia, induce cell cycle arrest and apoptosis in A549 human lung carcinoma cells.

    Science.gov (United States)

    Fan, Xiang; He, Lingli; Meng, Yao; Li, Gangrui; Li, Linli; Meng, Yanfa

    2015-05-01

    α‑Momorcharin (α‑MMC) and momordica anti‑human immunodeficiency virus protein (MAP30), produced by Momordica charantia, are ribosome‑inactivating proteins, which have been reported to exert inhibitory effects on cultured tumor cells. In order to further elucidate the functions of these agents, the present study aimed to investigate the effects of α‑MMC and MAP30 on cell viability, the induction of apoptosis, cell cycle arrest, DNA integrity and superoxide dismutase (SOD) activity. α‑MMC and MAP30 were purified from bitter melon seeds using ammonium sulfate precipitation in combination with sulfopropyl (SP)‑sepharose fast flow, sephacryl S‑100 and macro‑Cap‑SP chromatography. MTT, flow cytometric and DNA fragmentation analyses were then used to determine the effects of α‑MMC and MAP30 on human lung adenocarcinoma epithelial A549 cells. The results revealed that A549 cells were sensitive to α‑MMC and MAP30 cytotoxicity assays in vitro. Cell proliferation was significantly suppressed following α‑MMC and MAP30 treatment in a dose‑ and time‑dependent manner; in addition, the results indicated that MAP30 had a more potent anti‑tumor activity compared with that of α‑MMC. Cell cycle arrest in S phase and a significantly increased apoptotic rate were observed following treatment with α‑MMC and MAP30. Furthermore, DNA integrity analysis revealed that the DNA of A549 cells was degraded following treatment with α‑MMC and MAP30 for 48 h. The pyrogallol autoxidation method and nitrotetrazolium blue chloride staining were used to determine SOD activity, the results of which indicated that α‑MMC and MAP30 did not possess SOD activity. In conclusion, the results of the present study indicated that α‑MMC and MAP30 may have potential as novel therapeutic agents for the prophylaxis and treatment of cancer. PMID:25573293

  6. In Vitro Evidence for Immune-Modulatory Properties of Non-Digestible Oligosaccharides: Direct Effect on Human Monocyte Derived Dendritic Cells.

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    Sarah Lehmann

    Full Text Available Infant formulas containing non-digestible oligosaccharides (NDO similar to the composition in breast milk or a combination of lactic acid bacteria (LAB and NDO have been shown to harbor preventive effects towards immune-regulatory disorders. The aim of this study was to investigate the immune-modulatory potential of non-digestible short chain galacto- and long chain fructo-oligosaccharides (scGOS/lcFOS mimicking the natural distribution of oligosaccharides in human breast milk in presence or absence of certain LAB strains in human monocyte derived dendritic cells (MoDC. Immature human MoDC prepared from peripheral blood of healthy non-atopic volunteers were screened in vitro after stimulation with specific scGOS/lcFOS in presence or absence of LAB. IL-10 and IL-12p70 release was analyzed after 24 hours in cell-free supernatants by enzyme-linked immunosorbent assay (ELISA. A luminex-based assay was conducted to assess further cytokine and chemokine release by MoDC. To investigate the resulting T cell response, stimulated MoDC were co-incubated with naïve T cells in allogeneic stimulation assays and intracellular Foxp3 expression, as well as immune-suppressive capacity was determined. Oligosaccharides did not induce relevant amounts of IL-12p70 production, but did promote IL-10 release by MoDC. Furthermore, scGOS/lcFOS mixtures exerted a significant enhancing effect on LAB induced IL-10 secretion by MoDC while no increase in IL-12p70 production was observed. Blocking toll like receptor (TLR4 abrogated the increase in IL-10 in both the direct stimulation and the LAB stimulation of MoDC, suggesting that scGOS/lcFOS act via TLR4. Finally, scGOS/lcFOS-treated MoDC were shown to upregulate the number of functional suppressive Foxp3 positive T cells following allogeneic stimulation. Our results indicate anti-inflammatory and direct, microbiota independent, immune-modulatory properties of scGOS/lcFOS mixtures on human MoDC suggesting a possible

  7. Monocyte functions in diabetes mellitus

    DEFF Research Database (Denmark)

    Geisler, C; Almdal, T; Bennedsen, J;

    1982-01-01

    The aim of this study was to investigate the functions of monocytes obtained from 14 patients with diabetes mellitus (DM) compared with those of monocytes from healthy individuals. It was found that the total number of circulating monocytes in the 14 diabetic patients was lower than that from the...... elucidation of concomitant infections in diabetic patients are discussed....

  8. Interaction of angiogenically stimulated intermediate CD163+ monocytes/macrophages with soft hydrophobic poly(n-butyl acrylate) networks with elastic moduli matched to that of human arteries.

    Science.gov (United States)

    Mayer, Anke; Kratz, Karl; Hiebl, Bernhard; Lendlein, Andreas; Jung, Friedrich

    2012-03-01

    The cell population of peripheral blood monocytes/macrophages (MO) is heterogeneous: The majority of the MO are CD14++ CD16- and named "classical" (= MO1). Furthermore, two other subpopulations were described: CD14++ CD16+ ("intermediate" = MO2) and CD14+ CD16++ ("non-classical" = MO3). It is reported that MO2 possess anti-inflammatory properties and express the MO lineage marker CD163. On a hydrophilic neutrally charged acrylamide-based hydrogel human intermediate (CD14++ CD16+ ), angiogenically stimulated CD163++ monocytes/macrophages (aMO2) maintained a proangiogenic and noninflammatory status for at least 14 days. Here, we explored whether this aMO2 subset adhered to hydrophobic poly(n-butyl acrylate) networks (cPnBA) and also remained in its proangiogenic and noninflammatory status. Because substrate elasticity can impact adherence, morphology, and function of cells, cPnBAs with different Young's modulus (250 and 1100 kPa) were investigated, whereby their elasticity was tailored by variation of the cross-linker content and matched to the elasticity of human arteries. The cPnBAs exhibited similar surface properties (e.g., surface roughness), which were maintained after ethylene oxide sterilization and exposure in serum-free cell culture medium for 18 h at 37°C. aMO2 were seeded on cPnBA samples (1.7 × 10(5) cells/1.33 cm(2) ) in Dulbecco's modified Eagle medium (DMEM high glucose) supplemented with vascular endothelial growth factor 165 (VEGF-A(165) , 10 ng/mL) and fetal calf serum (10 vol%) for 3 and 72 h. On both polymeric samples (n = 3 each), the numbers of adherent cells per unit area were significantly higher (P < 0.01; cPnBA0250: 3 h 13 ± 5 cells/mm(2) , 72 h 234 ± 106 cells/mm(2) ; cPnBA1100: 3 h 14 ± 3 cells/mm(2) , 72 h 198 ± 113 cells/mm(2) ) compared to control cultures (glass, 3 h: 6 ± 3 cells/mm(2) , 72 h: 130 ± 83 cells/mm(2) ) and showed a typically spread morphology. The

  9. Production of proinflammatory cytokines in the human THP-1 monocyte cell line following induction by Tp0751,a recombinant protein of Treponema pallidum

    Institute of Scientific and Technical Information of China (English)

    2010-01-01

    The tissue destruction characteristic of syphilis infection may be caused by inflammation due to Treponema pallidum and the ensuing immune responses to the pathogen.T.pallidum membrane proteins are thought to be potent inducers of inflammation during the early stages of infection.However,the actual membrane proteins that induce inflammatory cytokine production are not known,nor are the molecular mechanisms responsible for triggering and sustaining the inflammatory cascades.In the present study,Tp0751 recombinant protein from T.pallidum was found to induce the production of proinflammatory cytokines,including TNF-α,IL-1βand IL-6,in a THP-1 human monocyte cell line.The signal transduction pathways involved in the production of these cytokines were then further investigated.No inhibition of TNF-a,IL-1β,or IL-6 production was observed following treatment with the SAPK/JNK specific inhibitor SP600125 or with an ERK inhibitor PD98059.By contrast,anti-TLR2 mAb,anti-CD14 mAb,and the p38 inhibitor SB203580 significantly inhibited the production of all three cytokines.In addition,pyrrolidine dithiocarbamate (PDTC),a specific inhibitor of NF-κB,profoundly inhibited the production of these cytokines.Tp0751 treatment strongly activated NF-κB,as revealed by Western blotting.However,NF-κB translocation was significantly inhibited by treatment with PDTC.These results indicated that TLR2,CD14,MAPKs/p38,and NF-κB might be implicated in the inflammatory reaction caused by T.pallidum infection.

  10. Regulation of the Expression of Heparan Sulfate 3-O-Sulfotransferase 3B (HS3ST3B) by Inflammatory Stimuli in Human Monocytes.

    Science.gov (United States)

    Sikora, Anne-Sophie; Delos, Maxime; Martinez, Pierre; Carpentier, Mathieu; Allain, Fabrice; Denys, Agnès

    2016-07-01

    Heparan sulfate (HS) is recognized as an important player in a wide range of dynamic steps of inflammatory reactions. Thereby, structural HS remodeling is likely to play an important role in the regulation of inflammatory and immune responses; however, little is known about underlying mechanism. In this study, we analyzed the regulation of expression of HS 3-O-sulfotransferases (HS3STs) in response to inflammatory stimuli. We found that among the seven HS3ST isoenzymes, only the expression of HS3ST3B was markedly up-regulated in human primary monocytes and the related cell line THP1 after exposure to TLR agonists. TNF-α was also efficient, to a lesser extent, to increase HS3ST3B expression, while IL-6, IL-4, and IFN-γ were poor inducers. We then analyzed the molecular mechanisms that regulate the high expression of HS3ST3B in response to LPS. Based on the expression of HS3ST3B transcripts and on the response of a reporter gene containing the HS3ST3B1 promoter, we provide evidence that LPS induces a rapid and strong transcription of HS3ST3B1 gene, which was mainly dependent on the activation of NF-κB and JNK signaling pathways. Additionally, active p38 MAPK and de novo synthesized proteins are involved in post-transcriptional mechanisms to maintain a high level of HS3ST3B mRNA to a steady state. Altogether, our findings indicate that HS3ST3B1 gene behaves as a primary response gene, suggesting that it may play an important role in making 3-O-sulfated HS with specific functions in the regulation of inflammatory and immune responses. J. Cell. Biochem. 117: 1529-1542, 2016. © 2015 Wiley Periodicals, Inc. PMID:26575945

  11. Enterovirus-71 virus-like particles induce the activation and maturation of human monocyte-derived dendritic cells through TLR4 signaling.

    Directory of Open Access Journals (Sweden)

    Yu-Li Lin

    Full Text Available Enterovirus 71 (EV71 causes seasonal epidemics of hand-foot-and-mouth disease and has a high mortality rate among young children. We recently demonstrated potent induction of the humoral and cell-mediated immune response in monkeys immunized with EV71 virus-like particles (VLPs, with a morphology resembling that of infectious EV71 virions but not containing a viral genome, which could potentially be safe as a vaccine for EV71. To elucidate the mechanisms through which EV71 VLPs induce cell-mediated immunity, we studied the immunomodulatory effects of EV71 VLPs on human monocyte-derived dendritic cells (DCs, which bind to and incorporate EV71 VLPs. DC treatment with EV71 VLPs enhanced the expression of CD80, CD86, CD83, CD40, CD54, and HLA-DR on the cell surface; increased the production of interleukin (IL-12 p40, IL-12 p70, and IL-10 by DCs; and suppressed the capacity of DCs for endocytosis. Treatment with EV71 VLPs also enhanced the ability of DCs to stimulate naïve T cells and induced secretion of interferon (IFN-γ by T cells and Th1 cell responses. Neutralization with antibodies against Toll-like receptor (TLR 4 suppressed the capacity of EV71 VLPs to induce the production of IL-12 p40, IL-12 p70, and IL-10 by DCs and inhibited EV71 VLPs binding to DCs. Our study findings clarified the important role for TLR4 signaling in DCs in response to EV71 VLPs and showed that EV71 VLPs induced inhibitor of kappaB alpha (IκBα degradation and nuclear factor of kappaB (NF-κB activation.

  12. Peroxisome Proliferator-activated Receptor γ Induces Apoptosis and Inhibits Autophagy of Human Monocyte-derived Macrophages via Induction of Cathepsin L

    Science.gov (United States)

    Mahmood, Dler Faieeq Darweesh; Jguirim-Souissi, Imene; Khadija, El-Hadri; Blondeau, Nicolas; Diderot, Vimala; Amrani, Souliman; Slimane, Mohamed-Naceur; Syrovets, Tatiana; Simmet, Thomas; Rouis, Mustapha

    2011-01-01

    Macrophages play a pivotal role in the pathophysiology of atherosclerosis. These cells express cathepsin L (CatL), a cysteine protease that has been implicated in atherogenesis and the associated arterial remodeling. In addition, macrophages highly express peroxisome proliferator-activated receptor (PPAR) γ, a transcription factor that regulates numerous genes important for lipid and lipoprotein metabolism, for glucose homeostasis, and inflammation. Hence, PPARγ might affect macrophage function in the context of chronic inflammation such as atherogenesis. In the present study, we examined the effect of PPARγ activation on the expression of CatL in human monocyte-derived macrophages (HMDM). Activation of PPARγ by the specific agonist GW929 concentration-dependently increased the levels of CatL mRNA and protein in HMDM. By promoter analysis, we identified a functional PPAR response element-like sequence that positively regulates CatL expression. In addition, we found that PPARγ-induced CatL promotes the degradation of Bcl2 without affecting Bax protein levels. Consistently, degradation of Bcl2 could be prevented by a specific CatL inhibitor, confirming the causative role of CatL. PPARγ-induced CatL was found to decrease autophagy through reduction of beclin 1 and LC3 protein levels. The reduction of these proteins involved in autophagic cell death was antagonized either by the CatL inhibitor or by CatL knockdown. In conclusion, our data show that PPARγ can specifically induce CatL, a proatherogenic protease, in HMDM. In turn, CatL inhibits autophagy and induces apoptosis. Thus, the proatherogenic effect of CatL could be neutralized by apoptosis, a beneficial phenomenon, at least in the early stages of atherosclerosis. PMID:21700710

  13. Silica Nanoparticles Sensitize Human Multiple Myeloma Cells to Snake (Walterinnesia aegyptia Venom-Induced Apoptosis and Growth Arrest

    Directory of Open Access Journals (Sweden)

    Douaa Sayed

    2012-01-01

    Full Text Available Background. Multiple myeloma (MM, an almost incurable disease, is the second most common blood cancer. Initial chemotherapeutic treatment could be successful; however, resistance development urges the use of higher toxic doses accompanied by hematopoietic stem cell transplantation. The establishment of more effective treatments that can overcome or circumvent chemoresistance has become a priority. We recently demonstrated that venom extracted from Walterinnesia aegyptia (WEV either alone or in combination with silica nanoparticles (WEV+NPs mediated the growth arrest and apoptosis of prostate cancer cells. In the present study, we evaluated the impact of WEV alone and WEV+NP on proliferation and apoptosis of MM cells. Methods. The impacts of WEV alone and WEV+NP were monitored in MM cells from 70 diagnosed patients. The influences of WEV and WEV+NP were assessed with flow cytometry analysis. Results. WEV alone and WEV+NP decreased the viability of MM cells. Using a CFSE proliferation assay, we found that WEV+NP strongly inhibited MM cell proliferation. Furthermore, analysis of the cell cycle using the propidium iodide (PI staining method indicated that WEV+NP strongly altered the cell cycle of MM cells and enhanced the induction of apoptosis. Conclusions. Our data reveal the biological effects of WEV and WEV+NP on MM cells that enable these compounds to function as effective treatments for MM.

  14. Formyl-peptide Receptor Agonists and Amorphous SiO2-NPs Synergistically and Selectively Increase the Inflammatory Responses of Human Monocytes and PMNs

    Directory of Open Access Journals (Sweden)

    Regina Tavano

    2016-02-01

    ligands, but not toward FPR2-specific ones. Conversely, the chemotaxis of monocytes toward FPR2-specific peptides was inhibited by SiO2-NPs. NADPH-oxidase activation triggered by FPR1- and FPR2-specific ligands in both cell types was not altered by SiO2-NPs. Microbial and tissue danger signals sensed by FPRs selectively amplified the functional responses of monocytes and PMNS to SiO2-NPs, and should be carefully considered in the assessment of the risk associated with nanoparticle exposure.

  15. Toll-like receptors, chemokine receptors and death receptor ligands responses in SARS coronavirus infected human monocyte derived dendritic cells

    Directory of Open Access Journals (Sweden)

    Law Helen KW

    2009-06-01

    Full Text Available Abstract Background The SARS outbreak in 2003 provides a unique opportunity for the study of human responses to a novel virus. We have previously reported that dendritic cells (DCs might be involved in the immune escape mechanisms for SARS-CoV. In this study, we focussed on the gene expression of toll-like receptors (TLRs, chemokine receptors (CCRs and death receptor ligands in SARS-CoV infected DCs. We also compared adult and cord blood (CB DCs to find a possible explanation for the age-dependent severity of SARS. Results Our results demonstrates that SARS-CoV did not modulate TLR-1 to TLR-10 gene expression but significantly induced the expression of CCR-1, CCR-3, and CCR-5. There was also strong induction of TNF-related apoptosis-inducing ligand (TRAIL, but not Fas ligand gene expression in SARS-CoV infected DCs. Interestingly, the expressions of most genes studied were higher in CB DCs than adult DCs. Conclusion The upregulation of chemokines and CCRs may facilitate DC migration from the infection site to the lymph nodes, whereas the increase of TRAIL may induce lymphocyte apoptosis. These findings may explain the increased lung infiltrations and lymphoid depletion in SARS patients. Further explorations of the biological significance of these findings are warranted.

  16. Jaridonin-induced G2/M phase arrest in human esophageal cancer cells is caused by reactive oxygen species-dependent Cdc2-tyr15 phosphorylation via ATM–Chk1/2–Cdc25C pathway

    Energy Technology Data Exchange (ETDEWEB)

    Ma, Yong-Cheng [Clinical Pharmacology Laboratory, Henan Province People' s Hospital, No. 7, Wei Wu Road, Zhengzhou, Henan (China); Su, Nan [Department of Quality Detection and Management, Henan University of Animal Husbandry and Economy, Zhengzhou, Henan (China); Shi, Xiao-Jing; Zhao, Wen; Ke, Yu [School of Pharmaceutical Sciences, Zhengzhou University, No. 100, Science Avenue, Zhengzhou, Henan (China); Zi, Xiaolin [Department of Urology, University of California, Irvine, Orange, CA (United States); Department of Pharmacology, University of California, Irvine, Orange, CA (United States); Department of Pharmaceutical Sciences, University of California, Irvine, Orange, CA (United States); Zhao, Ning-Min; Qin, Yu-Hua; Zhao, Hong-Wei [Clinical Pharmacology Laboratory, Henan Province People' s Hospital, No. 7, Wei Wu Road, Zhengzhou, Henan (China); Liu, Hong-Min, E-mail: liuhm@zzu.edu.cn [School of Pharmaceutical Sciences, Zhengzhou University, No. 100, Science Avenue, Zhengzhou, Henan (China)

    2015-01-15

    Jaridonin, a novel diterpenoid from Isodon rubescens, has been shown previously to inhibit proliferation of esophageal squamous cancer cells (ESCC) through G2/M phase cell cycle arrest. However, the involved mechanism is not fully understood. In this study, we found that the cell cycle arrest by Jaridonin was associated with the increased expression of phosphorylation of ATM at Ser1981 and Cdc2 at Tyr15. Jaridonin also resulted in enhanced phosphorylation of Cdc25C via the activation of checkpoint kinases Chk1 and Chk2, as well as in increased phospho-H2A.X (Ser139), which is known to be phosphorylated by ATM in response to DNA damage. Furthermore, Jaridonin-mediated alterations in cell cycle arrest were significantly attenuated in the presence of NAC, implicating the involvement of ROS in Jaridonin's effects. On the other hand, addition of ATM inhibitors reversed Jaridonin-related activation of ATM and Chk1/2 as well as phosphorylation of Cdc25C, Cdc2 and H2A.X and G2/M phase arrest. In conclusion, these findings identified that Jaridonin-induced cell cycle arrest in human esophageal cancer cells is associated with ROS-mediated activation of ATM–Chk1/2–Cdc25C pathway. - Highlights: • Jaridonin induced G2/M phase arrest through induction of redox imbalance. • Jaridonin increased the level of ROS through depleting glutathione in cell. • ATM–Chk1/2–Cdc25C were involved in Jaridonin-induced cell cycle arrest. • Jaridonin selectively inhibited cancer cell viability and cell cycle progression.

  17. Jaridonin-induced G2/M phase arrest in human esophageal cancer cells is caused by reactive oxygen species-dependent Cdc2-tyr15 phosphorylation via ATM–Chk1/2–Cdc25C pathway

    International Nuclear Information System (INIS)

    Jaridonin, a novel diterpenoid from Isodon rubescens, has been shown previously to inhibit proliferation of esophageal squamous cancer cells (ESCC) through G2/M phase cell cycle arrest. However, the involved mechanism is not fully understood. In this study, we found that the cell cycle arrest by Jaridonin was associated with the increased expression of phosphorylation of ATM at Ser1981 and Cdc2 at Tyr15. Jaridonin also resulted in enhanced phosphorylation of Cdc25C via the activation of checkpoint kinases Chk1 and Chk2, as well as in increased phospho-H2A.X (Ser139), which is known to be phosphorylated by ATM in response to DNA damage. Furthermore, Jaridonin-mediated alterations in cell cycle arrest were significantly attenuated in the presence of NAC, implicating the involvement of ROS in Jaridonin's effects. On the other hand, addition of ATM inhibitors reversed Jaridonin-related activation of ATM and Chk1/2 as well as phosphorylation of Cdc25C, Cdc2 and H2A.X and G2/M phase arrest. In conclusion, these findings identified that Jaridonin-induced cell cycle arrest in human esophageal cancer cells is associated with ROS-mediated activation of ATM–Chk1/2–Cdc25C pathway. - Highlights: • Jaridonin induced G2/M phase arrest through induction of redox imbalance. • Jaridonin increased the level of ROS through depleting glutathione in cell. • ATM–Chk1/2–Cdc25C were involved in Jaridonin-induced cell cycle arrest. • Jaridonin selectively inhibited cancer cell viability and cell cycle progression

  18. Potentially probiotic bacteria induce efficient maturation but differential cytokine production in human monocyte-derived dendritic cells

    Institute of Scientific and Technical Information of China (English)

    Sinikka Latvala; Taija E Pietil(a); Ville Veckman; Riina A Kekkonen; Soile Tynkkynen; Riitta Korpela; Ilkka Julkunen

    2008-01-01

    MM: To analyze the ability of nine different potentially probiotic bacteria to induce maturation and cytokine production in human monocyLe-derived dendritic cells (moDCs).METHODS: Cytokine production and maturation of moDCs in response to bacterial stimulation was analyzed with enzyme-linked immunosorbent assay (ELISA) and flow cytometric analysis (FACS),respectively.The kinetics of mRNA expression of cytokine genes was determined by Northern blotting.The involvement of different signaling pathways in cytokine gene expression was studied using specific pharmacological signaling inhibitors.RESULTS: All studied bacteria induced the maturation of moDCs in a dose-dependent manner.More detailed analysis with S.thermophilus THS,B.breve Bb99,and L.lactis subsp,cremoris ARH74 indicated that these bacteria induced the expression of moDC maturation markers HLA class II and CD86 as efficiently as pathogenic bacteria.However,these bacteria differed in their ability to induce moDC cytokine gene expression.S.therrnophilus induced the expression of pro-inflammatory (TNF-a,IL-12,IL-6,and CCL20)and Th1 type (IL-12 and IFN-y) cytokines,while B.breve and L.lactis were also potent inducers of antiinflammatory IL-10.Mitogen-activated protein kinase (MAPK) p38,phosphatidylinositol 3 (PI3) kinase,and nuclear factor-kappa B (NF-κB) signaling pathways were shown to be involved in bacteria-induced cytokine production.CONCLUSION: Our results indicate that potentially probiotic bacteria are able to induce moDC maturation,but their ability to induce cytokine gene expression varies significantly from one bacterial strain to another.

  19. Deterrence and arrest ratios.

    Science.gov (United States)

    Carmichael, Stephanie E; Piquero, Alex R

    2006-02-01

    In the limited research on the origins of sanction threat perceptions, researchers have focused on either the effects of actively engaging in crime or the effects of formal sanctioning but rarely on both (i.e., the arrest ratio or the number of arrests relative to the number of crimes committed). This article extends this line of research by using a sample of Colorado inmates and measures arrest ratios and sanction perceptions for eight different crime types. Analyses reveal that the offenders report both significant experiential and arrest ratio effects. Theoretical and policy implications, limitations, and directions for future research are outlined. PMID:16397123

  20. Honokiol arrests cell cycle, induces apoptosis, and potentiates the cytotoxic effect of gemcitabine in human pancreatic cancer cells.

    Directory of Open Access Journals (Sweden)

    Sumit Arora

    Full Text Available Survival rates for patients with pancreatic cancer are extremely poor due to its asymptomatic progression to advanced and metastatic stage for which current therapies remain largely ineffective. Therefore, novel therapeutic agents and treatment approaches are desired to improve the clinical outcome. In this study, we determined the effects of honokiol, a biologically active constituent of oriental medicinal herb Magnolia officinalis/grandiflora, on two pancreatic cancer cell lines, MiaPaCa and Panc1, alone and in combination with the standard chemotherapeutic drug, gemcitabine. Honokiol exerted growth inhibitory effects on both the pancreatic cancer cell lines by causing cell cycle arrest at G₁ phase and induction of apoptosis. At the molecular level, honokiol markedly decreased the expression of cyclins (D1 and E and cyclin-dependent kinases (Cdk2 and Cdk4, and caused an increase in Cdk inhibitors, p21 and p27. Furthermore, honokiol treatment led to augmentation of Bax/Bcl-2 and Bax/Bcl-xL ratios to favor apoptosis in pancreatic cancer cells. These changes were accompanied by enhanced cytoplasmic accumulation of NF-κB with a concomitant decrease in nuclear fraction and reduced transcriptional activity of NF-κB responsive promoter. This was associated with decreased phosphorylation of inhibitor of kappa B alpha (IκB-α causing its stabilization and thus increased cellular levels. Importantly, honokiol also potentiated the cytotoxic effects of gemcitabine, in part, by restricting the gemcitabine-induced nuclear accumulation of NF-κB in the treated pancreatic cancer cell lines. Altogether, these findings demonstrate, for the first time, the growth inhibitory effects of honokiol in pancreatic cancer and indicate its potential usefulness as a novel natural agent in prevention and therapy.

  1. Adenosine induces cell cycle arrest and apoptosis via cyclinD1/Cdk4 and Bcl-2/Bax pathways in human ovarian cancer cell line OVCAR-3.

    Science.gov (United States)

    Shirali, Saeid; Aghaei, Mahmoud; Shabani, Mahdi; Fathi, Mojtaba; Sohrabi, Majid; Moeinifard, Marzieh

    2013-04-01

    Adenosine is a regulatory molecule with widespread physiological effects in almost every cells and acts as a potent regulator of cell growth. Adenosine has been shown to inhibit cell growth and induce apoptosis in the several cancer cells via caspase activation and Bcl-2/Bax pathway. The present study was designed to understand the mechanism underlying adenosine-induced apoptosis in the OVCAR-3 human ovarian cancer cells. MTT viability, BrdU and cell counting assays were used to study the cell proliferation effect of adenosine in presence of adenosine deaminase inhibitor and the nucleoside transporter inhibitor. Cell cycle analysis, propidium iodide and annexin V staining, caspase-3 activity assay, cyclinD1, Cdk4, Bcl-2 and Bax protein expressions were assessed to detect apoptosis. Adenosine significantly inhibited cell proliferation in a concentration-dependent manner in OVCAR-3 cell line. Adenosine induced cell cycle arrest in G0/G1 phase via Cdk4/cyclinD1-mediated pathway. Adenosine induced apoptosis, which was determined by Annexin V-FITC staining and increased sub-G1 population. Moreover, down-regulation of Bcl-2 protein expression, up-regulation of Bax protein expression and activation of caspase-3 were observed in response to adenosine treatment. The results of this study suggest that extracellular adenosine induced G1 cell cycle arrest and apoptosis in ovarian cancer cells via cyclinD1/ Cdk4 and Bcl-2/Bax pathways and caspase-3 activation. These data might suggest that adenosine could be used as an agent for the treatment of ovarian cancer. PMID:23345014

  2. 4β-Hydroxywithanolide E from Physalis peruviana (golden berry) inhibits growth of human lung cancer cells through DNA damage, apoptosis and G2/M arrest

    International Nuclear Information System (INIS)

    The crude extract of the fruit bearing plant, Physalis peruviana (golden berry), demonstrated anti-hepatoma and anti-inflammatory activities. However, the cellular mechanism involved in this process is still unknown. Herein, we isolated the main pure compound, 4β-Hydroxywithanolide (4βHWE) derived from golden berries, and investigated its antiproliferative effect on a human lung cancer cell line (H1299) using survival, cell cycle, and apoptosis analyses. An alkaline comet-nuclear extract (NE) assay was used to evaluate the DNA damage due to the drug. It was shown that DNA damage was significantly induced by 1, 5, and 10 μg/mL 4βHWE for 2 h in a dose-dependent manner (p < 0.005). A trypan blue exclusion assay showed that the proliferation of cells was inhibited by 4βHWE in both dose- and time-dependent manners (p < 0.05 and 0.001 for 24 and 48 h, respectively). The half maximal inhibitory concentrations (IC50) of 4βHWE in H1299 cells for 24 and 48 h were 0.6 and 0.71 μg/mL, respectively, suggesting it could be a potential therapeutic agent against lung cancer. In a flow cytometric analysis, 4βHWE produced cell cycle perturbation in the form of sub-G1 accumulation and slight arrest at the G2/M phase with 1 μg/mL for 12 and 24 h, respectively. Using flow cytometric and annexin V/propidium iodide immunofluorescence double-staining techniques, these phenomena were proven to be apoptosis and complete G2/M arrest for H1299 cells treated with 5 μg/mL for 24 h. In this study, we demonstrated that golden berry-derived 4βHWE is a potential DNA-damaging and chemotherapeutic agent against lung cancer

  3. 4β-Hydroxywithanolide E from Physalis peruviana (golden berry inhibits growth of human lung cancer cells through DNA damage, apoptosis and G2/M arrest

    Directory of Open Access Journals (Sweden)

    Guo Zong-Lun

    2010-02-01

    Full Text Available Abstract Background The crude extract of the fruit bearing plant, Physalis peruviana (golden berry, demonstrated anti-hepatoma and anti-inflammatory activities. However, the cellular mechanism involved in this process is still unknown. Methods Herein, we isolated the main pure compound, 4β-Hydroxywithanolide (4βHWE derived from golden berries, and investigated its antiproliferative effect on a human lung cancer cell line (H1299 using survival, cell cycle, and apoptosis analyses. An alkaline comet-nuclear extract (NE assay was used to evaluate the DNA damage due to the drug. Results It was shown that DNA damage was significantly induced by 1, 5, and 10 μg/mL 4βHWE for 2 h in a dose-dependent manner (p p 50 of 4βHWE in H1299 cells for 24 and 48 h were 0.6 and 0.71 μg/mL, respectively, suggesting it could be a potential therapeutic agent against lung cancer. In a flow cytometric analysis, 4βHWE produced cell cycle perturbation in the form of sub-G1 accumulation and slight arrest at the G2/M phase with 1 μg/mL for 12 and 24 h, respectively. Using flow cytometric and annexin V/propidium iodide immunofluorescence double-staining techniques, these phenomena were proven to be apoptosis and complete G2/M arrest for H1299 cells treated with 5 μg/mL for 24 h. Conclusions In this study, we demonstrated that golden berry-derived 4βHWE is a potential DNA-damaging and chemotherapeutic agent against lung cancer.

  4. 17-Allylamino-17-demethoxygeldanamycin induces downregulation of critical Hsp90 protein clients and results in cell cycle arrest and apoptosis of human urinary bladder cancer cells

    International Nuclear Information System (INIS)

    17-Allylamino-17-demethoxygeldanamycin (17-AAG), a benzoquinone ansamycin antibiotic, specifically targets heat shock protein 90 (Hsp90) and interferes with its function as a molecular chaperone that maintains the structural and functional integrity of various protein clients involved in cellular signaling. In this study, we have investigated the effect of 17-AAG on the regulation of Hsp90-dependent signaling pathways directly implicated in cell cycle progression, survival and motility of human urinary bladder cancer cell lines. We have used MTT-based assays, FACS analysis, Western blotting, semi-quantitative RT-PCR, immunocytochemistry and scratch-wound assay in RT4, RT112 and T24 human urinary bladder cancer cell lines. We have demonstrated that, upon 17-AAG treatment, bladder cancer cells are arrested in the G1 phase of the cell cycle and eventually undergo apoptotic cell death in a dose-dependent manner. Furthermore, 17-AAG administration was shown to induce a pronounced downregulation of multiple Hsp90 protein clients and other downstream effectors, such as IGF-IR, Akt, IKK-α, IKK-β, FOXO1, ERK1/2 and c-Met, resulting in sequestration-mediated inactivation of NF-κB, reduced cell proliferation and decline of cell motility. In total, we have clearly evinced a dose-dependent and cell type-specific effect of 17-AAG on cell cycle progression, survival and motility of human bladder cancer cells, due to downregulation of multiple Hsp90 clients and subsequent disruption of signaling integrity

  5. Two new isoquinoline alkaloids from Scolopendra subspinipes mutilans induce cell cycle arrest and apoptosis in human glioma cancer U87 cells.

    Science.gov (United States)

    Ding, Dan; Guo, Ya-Ru; Wu, Rui-Ling; Qi, Wei-Yan; Xu, Han-Mei

    2016-04-01

    Two new isoquinoline alkaloids 1-2 and seven known compounds 3-9 were isolated from the ethanol extract of centipede Scolopendra subspinipes mutilans L. Koch. The structures were elucidated by a combination of spectroscopic analyses including 1D and 2D NMR, and HRESIMS. Compounds 1-2 exhibited good cytotoxicity with IC50 values ranging from 1.19 to 31.28μM against six human cancer cell lines and low cytotoxicity against human normal liver L-02 cell lines, suggesting that compounds 1-2 had high specific cytotoxicity on human cancer cell lines. Further analyses showed that compounds 1-2 inhibited U87 cells proliferation by arresting cell cycle progress at G0/G1 phase and inducing apoptosis through loss of mitochondrial membrane potential (MMP), activation of caspase 9/3 and down-regulation of the Bcl-2/Bax protein ratio. The results suggest that compounds 1-2 induce apoptosis in U87 cells through the mitochondria apoptosis pathway, and they deserve further research as potential anti-glioma cancer agents. PMID:26947248

  6. Crude extract of Rheum palmatum L. Induces cell cycle arrest S phase and apoptosis through mitochondrial-dependent pathways in U-2 OS human osteosarcoma cells.

    Science.gov (United States)

    Lin, Chin-Chung; Lee, Ming-Huei; Lin, Ju-Hwa; Lin, Meng-Liang; Chueh, Fu-Shin; Yu, Chien-Chih; Lin, Jing-Pin; Chou, Yu-Cheng; Hsu, Shu-Chun; Chung, Jing-Gung

    2016-08-01

    Cancer is the second cause of death in children. Osteosarcoma is the most common primary malignancy of solid bone cancer primarily affecting adolescents and young adults. In the Chinese population, the crude extract of Rheum palmatum L. (CERP) has been used for treating different diseases, including SARS, rheumatoid arthritis, coxsackievirus B3, and human colon cancer cell, pancreatic cancer. There are no reports on CERP and human osteosarcoma cells. The present study examined effects of CERP on cytotoxicity including cell cycle distribution and cell death (apoptosis) in U-2 OS human osteosarcoma cells. CERP significantly induced S phase arrest in U-2 OS cells in a dose-dependent. CERP produced DNA damage and DNA condensation. Other effects of CERP were stimulation of ROS and Ca(2+) , mitochondria impairment, and activation of caspase-3, -8, and -9. CERP increased the levels of Bax, Bak, Bad, cyclin B, Fas, PARP, GRP78, GADD153, AIF, Endo G, Calpain-2, p21, and p27, but decreased the levels of Bcl-2, BCL-X, XIAP, Akt, CDC25A, CDK2, Cyclin A, and Cyclin E of U-2 OS cells. It was also observed that CERP promoted the expression of AIF, Endo G, GADD153, and cytochrome c. These results indicate that CERP has anticancer effects in vitro and provide the foundation for in vivo studies of animal models of osteosarcoma. © 2015 Wiley Periodicals, Inc. Environ Toxicol 31: 957-969, 2016. PMID:25689151

  7. Respective roles and interactions of T-lymphocyte and PGE2-mediated monocyte suppressive activities in human newborns and mothers at the time of delivery

    International Nuclear Information System (INIS)

    Recently the concept of a poorly functional humoral immune response in the newborn was proposed. Data have been presented indicating that the impaired newborn B cell maturation, as shown in vitro in a pokeweed mitogen-induced B cell maturation system, is due both to an immaturity of lymphocyte subsets and to an increased suppressive T activity. In the present work, we present evidence that there exists a predominance of a naturally occurring T lymphocyte suppressive activity in the cord blood in that the removal of the suppressive activity by irradiation allows a normal maturation of newborn B cells. Such normal maturation of newborn B cells can also be obtained using mixed cultures of adult T cells and newborn B cells. Newborn suppressor T cells belong to both EA gamma (+) and EA gamma (-) fractions, and it is not known whether these two groups do or do not belong to different subsets. The PGE2-dependent monocyte suppressive activity does not play any role in the suppression observed in newborns since newborn monocytes are poorly suppressive and since they produce a smaller amount of PGE2 than adult monocytes. Some observations suggest, on the contrary, that the suppressive T lymphocytes can regulate the level of the PGE2-dependent monocyte suppressive activity. It should be noticed that similar observations about T lymphocyte and PGE2-dependent monocyte suppressive activities have been made at the same time using mothers' cells. These observations suggest the possibility that such changes in B cell immune regulation may result from an interaction between maternal and fetal lymphoid cells

  8. Paclitaxel Arrests Growth of Intracellular Toxoplasma gondii

    OpenAIRE

    Estes, Randee; Vogel, Nicolas; Mack, Douglas; McLeod, Rima

    1998-01-01

    Addition of paclitaxel (Taxol) at a concentration of 1 μM to Toxoplasma gondii-infected human foreskin fibroblasts arrested parasite multiplication. Division of the T. gondii tachyzoite nucleus was inhibited, leading to syncytium-like parasite structures within the fibroblasts by 24 h after infection and treatment of the cultures. By 4 days after infection and treatment of the cultures with paclitaxel, this inhibition was irreversible, since the arrested intracellular form was incapable of le...

  9. Fucoidan induces G1 arrest of the cell cycle in EJ human bladder cancer cells through down-regulation of pRB phosphorylation

    Directory of Open Access Journals (Sweden)

    Hye Young Park

    2015-06-01

    Full Text Available AbstractFucoidan, a sulfated polysaccharide found in marine algae and brown seaweeds, has been shown to inhibit the in vitro growth of human cancer cells. This study was conducted in cultured human bladder cancer EJ cells to elucidate the possible mechanisms by which fucoidan exerts its anti-proliferative activity, which until now has remained poorly understood. Fucoidan treatment of EJ cells resulted in dose-dependent inhibition of cell growth and induced apoptotic cell death. Flow cytometric analysis revealed that fucoidan led to G1 arrest in cell cycle progression. It was associated with down-regulation of cyclin D1, cyclin E, and cyclin-dependent-kinases (Cdks in a concentration-dependent manner, without any change in Cdk inhibitors, such as p21 and p27. Furthermore, dephosphorylation of retinoblastoma protein (pRB by this compound was associated with enhanced binding of pRB with the transcription factors E2F-1 and E2F-4. Overall, our results demonstrate that fucoidan possesses anticancer activity potential against bladder cancer cells by inhibiting pRB phosphorylation.

  10. Ethanol extract of Innotus obliquus (Chaga mushroom) induces G1 cell cycle arrest in HT-29 human colon cancer cells

    OpenAIRE

    Lee, Hyun Sook; Kim, Eun Ji; Kim, Sun Hyo

    2015-01-01

    BACKGROUND/OBJECTIVES Inonotus obliquus (I. obliquus, Chaga mushroom) has long been used as a folk medicine to treat cancer. In the present study, we examined whether or not ethanol extract of I. obliquus (EEIO) inhibits cell cycle progression in HT-29 human colon cancer cells, in addition to its mechanism of action. MATERIALS/METHODS To examine the effects of Inonotus obliquus on the cell cycle progression and the molecular mechanism in colon cancer cells, HT-29 human colon cancer cells were...

  11. Growth arrest and apoptosis via caspase activation of dioscoreanone in human non-small-cell lung cancer A549 cells

    OpenAIRE

    Hansakul, Pintusorn; Aree, Kalaya; Tanuchit, Sermkiat; Itharat, Arunporn

    2014-01-01

    Background Dioscoreanone (DN) isolated from Dioscorea membranacea Pierre has been reported to exert potent cytotoxic effects against particular types of cancer. The present study was carried out to investigate the cytotoxicity of DN against a panel of different human lung cancer cell lines. The study further examined the underlying mechanisms of its anticancer activity in the human lung adenocarcinoma cell line A549. Methods Antiproliferative effects of DN were determined by SRB and CFSE assa...

  12. Early interleukin-6 and slope of monocyte human leukocyte antigen-DR: a powerful association to predict the development of sepsis after major trauma.

    Directory of Open Access Journals (Sweden)

    Aurélie Gouel-Chéron

    Full Text Available OBJECTIVE: Major trauma is characterized by a pro-inflammatory response, followed by an immunosuppression. Recently, in trauma patients, the lack of recovery of monocyte Human Leukocyte Antigen DR (mHLA-DR, a biomarker of ICU-acquired immunosuppression between days 1-2 and days 3-4 has been demonstrated to be independently associated with sepsis development. The main objective of this study was to determine whether early measurements of IL-6 (interleukin-6 and IL-10 plasma concentrations (as markers of initial severity could improve, in association with mHLA-DR recovery, the prediction of sepsis occurrence in severe trauma patients. DESIGN: Prospective observational study over 24 months in a Trauma ICU at university hospital. PATIENTS: Trauma patients with an ISS over 25 and age over 18 were included. MEASUREMENTS AND MAIN RESULTS: mHLA-DR was assessed by flow cytometry, IL-6 and IL-10 concentrations by ELISA. 100 consecutive severely injured patients were monitored (mean ISS 37±10. 37 patients developed sepsis. IL-6 concentrations and slope of mHLA-DR expression between days 1-2 and days 3-4 were significantly different between septic and non-septic patients. IL-10 was not detectable in most patients. After adjustment for usual clinical confounders, when assessed as a pair, multivariate logistic regression analysis revealed that a slope of mHLA-DR expression (days 3-4/days 1-2≤1.1 and a IL-6 concentration ≥ 67.1 pg/ml remained highly associated with the development of sepsis (adjusted OR 18.4, 95% CI 4.9; 69.4, p = .00002. CONCLUSIONS: After multivariate regression logistic analysis, when assessed as a pair, a high IL-6 concentration and a persistent mHLA-DR decreased expression were found to be in relation with the development of sepsis with the best predictive value. This study underlines the usefulness of daily monitoring of immune function to identify trauma patients at a high risk of infection.

  13. A matrix of cholesterol crystals, but not cholesterol alone, primes human monocytes/macrophages for excessive endotoxin-induced production of tumor necrosis factor-alpha. Role in atherosclerotic inflammation?

    DEFF Research Database (Denmark)

    Bendtzen, Klaus; Christensen, Ole; Nielsen, Claus Henrik;

    2014-01-01

    When exposed to small amounts of bacterial endotoxin, matrices of cholesterol crystals, but not cholesterol itself, primed human monocytes/macrophages to a highly augmented (>10-fold) production of inflammatory tumor necrosis factor-α. Priming also sensitized the cells, as 10- to 100-fold lower...... levels of endotoxin were needed for TNF-α production equivalent to that of unprimed cells. The pro-inflammatory effect was selective as endotoxin-induced production of other pro-inflammatory cytokines was unaffected while production of anti-inflammatory interleukin-10 was diminished. These findings...

  14. Morphological alterations and G0/G1 cell cycle arrest induced by curcumin in human SK-MEL-37 melanoma cells

    Directory of Open Access Journals (Sweden)

    Marcella Lemos Brettas Carneiro

    2010-04-01

    Full Text Available The aim of this work was to study the effect of curcumin on cell cycle in the human SK-MEL-37 melanoma cell line. In addition, morphological and structural analyses were also performed. Flow cytometric analysis showed a G0/G1 arrest at 5 µM after 24 h exposure and a concentration-dependent increase in the proportion of sub-G0 hypodiploid cells. Typical apoptotic events were also observed by the fluorescence microscopy, transmission and scanning electronic microscopy. Loss of mitochondrial membrane potential was not detected. Results suggested that curcumin could arrest human melanoma cells at G0/G1 phase and induce a mitochondrial-independent apoptotic pathway.O melanoma é um tipo agressivo de câncer cujo tratamento culmina com o estabelecimento de resistência aos quimioterápicos empregados. Portanto, é importante o desenvolvimento de novos agentes farmacológicos que sejam menos tóxicos e que não provoquem quimiorresistência. As inúmeras propriedades terapêuticas da curcumina vêm sendo confirmadas através de estudos sobre o seu mecanismo de ação em células cultivadas. No presente estudo, empregamos células de melanoma humano da linhagem SK-MEL-37, que desenvolveram resistência in vitro à doxorubicina e cisplatina, drogas normalmente utilizadas na clínica. Investigamos o efeito da curcumina sobre o ciclo celular através de citometria de fluxo. Além disso, análises morfológicas e estruturais também foram realizadas. Os resultados demonstraram que o tratamento com uma concentração de 5 ?M de curcumina provocou uma parada na subfase G0/G1. Além disso, observou-se um aumento dose-dependente na proporção de células hipodiplóides em sub-G0. Eventos apoptóticos típicos foram observados por microscopia de fluorescência, microscopia eletrônica de transmissão e microscopia eletrônica de varredura. Não foi detectada alteração no potencial de membrana mitocondrial. Os resultados indicam que futuros estudos poder

  15. Labeling monocytes for imaging chronic inflammation

    International Nuclear Information System (INIS)

    With growing interest in cell-based scintigraphic diagnosis or therapy monitoring, there is an increasing demand for non-invasive observation and quantification of cell trafficking in the preclinical and clinical setting. Monocytes are members of the human mononuclear phagocyte system originating from a myeloid precursor in the bone. Labeled monocytes are being used for investigation of pathogenesis like atherosclerosis and for monitoring of therapeutic intervention in inflammatory diseases like rheumatoid arthritis. Labeling mononuclear cells at high specific activity without affecting their biological functions allows (delayed) non-invasive imaging with g or PET cameras. Monocytes labeled before their final differentiation into macrophages or dendritic cells may reveal centers of inflammation in a patient and, thereby, contribute to scintigraphic diagnosis. Macrophages or dendritic cells may be in vitro cultured and by means of genetic transformation specified towards specific targets prior to re-injection, an approach with therapeutic potency. This review addresses issues on autologous monocytes, particularly their properties and labeling for non-invasive in vivo radionuclide imaging of chronic inflammation.

  16. Organophosphates and monocyte esterase deficiency.

    OpenAIRE

    1995-01-01

    AIMS--To examine the possibility that monocyte esterase deficiency (MED) could be caused by exposure to organophosphates. METHODS--Pseudocholinesterase, paraoxonase and arylesterase activities were measured in the serum and acetylcholinesterase activity was measured in the red cells of a group of monocyte esterase deficient subjects and compared with the enzyme activities of a control group of monocyte esterase positive subjects. RESULTS--No significant difference was found between the enzyme...

  17. Functional Langerinhigh-Expressing Langerhans-like Cells Can Arise from CD14highCD16- Human Blood Monocytes in Serum-Free Condition.

    Science.gov (United States)

    Picarda, Gaëlle; Chéneau, Coraline; Humbert, Jean-Marc; Bériou, Gaëlle; Pilet, Paul; Martin, Jérôme; Duteille, Franck; Perrot, Pierre; Bellier-Waast, Frédérique; Heslan, Michèle; Haspot, Fabienne; Guillon, Fabien; Josien, Regis; Halary, Franck Albert

    2016-05-01

    Langerhans cells (LCs) are epithelial APCs that sense danger signals and in turn trigger specific immune responses. In steady-state, they participate in the maintenance of peripheral tolerance to self-antigens whereas under inflammation LCs efficiently trigger immune responses in secondary lymphoid organs. It has been demonstrated in mice that LC-deprived epithelia are rapidly replenished by short half-life langerin-expressing monocyte-derived LCs (MDLCs). These surrogate LCs are thought to be progressively replaced by langerin(high) LCs arising from self-renewing epithelial precursors of hematopoietic origin. How LCs arise from blood monocytes is not fully understood. Hence, we sought to characterize key factors that induce differentiation of langerin(high)-expressing monocyte-derived Langerhans-like cells. We identified GM-CSF and TGF-β1 as key cytokines to generate langerin(high)-expressing cells but only in serum-free conditions. These cells were shown to express the LC-specific TROP-2 and Axl surface markers and contained Birbeck granules. Surprisingly, E-cadherin was not spontaneously expressed by these cells but required a direct contact with keratinocytes to be stably induced. MDLCs induced stronger allogeneic T cell proliferations but released low amounts of inflammatory cytokines upon TLR stimulation compared with donor-paired monocyte-derived dendritic cells. Immature langerin(high) MDLCs were responsive to MIP-3β/CCL20 and CTAC/CCL27 chemokine stimulations. Finally, we demonstrated that those cells behaved as bona fide LCs when inserted in a three-dimensional rebuilt epithelium by becoming activated upon TLR or UV light stimulations. Collectively, these results prompt us to propose these langerin(high) MDLCs as a relevant model to address LC biology-related questions. PMID:27016604

  18. Activation of human monocytes by live Borrelia burgdorferi generates TLR2-dependent and -independent responses which include induction of IFN-beta.

    Directory of Open Access Journals (Sweden)

    Juan C Salazar

    2009-05-01

    Full Text Available It is widely believed that innate immune responses to Borrelia burgdorferi (Bb are primarily triggered by the spirochete's outer membrane lipoproteins signaling through cell surface TLR1/2. We recently challenged this notion by demonstrating that phagocytosis of live Bb by peripheral blood mononuclear cells (PBMCs elicited greater production of proinflammatory cytokines than did equivalent bacterial lysates. Using whole genome microarrays, we show herein that, compared to lysates, live spirochetes elicited a more intense and much broader transcriptional response involving genes associated with diverse cellular processes; among these were IFN-beta and a number of interferon-stimulated genes (ISGs, which are not known to result from TLR2 signaling. Using isolated monocytes, we demonstrated that cell activation signals elicited by live Bb result from cell surface interactions and uptake and degradation of organisms within phagosomes. As with PBCMs, live Bb induced markedly greater transcription and secretion of TNF-alpha, IL-6, IL-10 and IL-1beta in monocytes than did lysates. Secreted IL-18, which, like IL-1beta, also requires cleavage by activated caspase-1, was generated only in response to live Bb. Pro-inflammatory cytokine production by TLR2-deficient murine macrophages was only moderately diminished in response to live Bb but was drastically impaired against lysates; TLR2 deficiency had no significant effect on uptake and degradation of spirochetes. As with PBMCs, live Bb was a much more potent inducer of IFN-beta and ISGs in isolated monocytes than were lysates or a synthetic TLR2 agonist. Collectively, our results indicate that the enhanced innate immune responses of monocytes following phagocytosis of live Bb have both TLR2-dependent and -independent components and that the latter induce transcription of type I IFNs and ISGs.

  19. MMP-2, MMP-9 and their inhibitors TIMP-2 and TIMP-1 production by human monocytes in vitro in the presence of different forms of hydroxyapatite particles.

    OpenAIRE

    Laquerriere, Patrice; Grandjean-Laquerriere, Alexia; Addadi-Rebbah, Salima; Jallot, Edouard; Laurent-Maquin, Dominique; Frayssinet, Patrick; Guenounou, Moncef

    2004-01-01

    After calcium-phosphates biomaterials based implantation like hydroxyapatite (HA) coating, particles are released in the periprosthetic tissues. Wear-debris induced fibrous membranes contain macrophage subsets that can produce metalloproteinases (MMPs), which are considered to be key enzymes in extra-cellular matrix turnover. Tissue inhibitors of metalloproteinases (TIMPs) are important regulator of MMPs activity. Interleukin-1 mainly produced by monocytes can also regulate MMPs production. I...

  20. Induction of S-Phase Arrest in Human Glioma Cells by Selenocysteine, a Natural Selenium-Containing Agent Via Triggering Reactive Oxygen Species-Mediated DNA Damage and Modulating MAPKs and AKT Pathways.

    Science.gov (United States)

    Wang, Kun; Fu, Xiao-Ting; Li, Yuan; Hou, Ya-Jun; Yang, Ming-Feng; Sun, Jing-Yi; Yi, Shu-Ying; Fan, Cun-Dong; Fu, Xiao-Yan; Zhai, Jing; Sun, Bao-Liang

    2016-06-01

    Selenocysteine (SeC) a natural available selenoamino acid exhibits novel anticancer activities against human cancer cell lines. However, the growth inhibitory effect and mechanism of SeC in human glioma cells remain unclear. The present study reveals that SeC time- and dose-dependently inhibited U251 and U87 human glioma cells growth by induction of S-phase cell cycle arrest, followed by the marked decrease of cyclin A. SeC-induced S-phase arrest was achieved by inducing DNA damage through triggering generation of reactive oxygen species (ROS) and superoxide anion, with concomitant increase of TUNEL-positive cells and induction of p21waf1/Cip1 and p53. SeC treatment also caused the activation of p38MAPK, JNK and ERK, and inactivation of AKT. Four inhibitors of MAPKs and AKT pathways further confirmed their roles in SeC-induced S-phase arrest in human glioma cells. Our findings advance the understanding on the molecular mechanisms of SeC in human glioma management. PMID:26846141

  1. An evidence on G2/M arrest, DNA damage and caspase mediated apoptotic effect of biosynthesized gold nanoparticles on human cervical carcinoma cells (HeLa)

    Energy Technology Data Exchange (ETDEWEB)

    Jeyaraj, M. [Department of Biotechnology and Genetic Engineering, School of Biotechnology, Bharathidasan University, Tiruchirappalli 620024, Tamil Nadu (India); Arun, R. [Department of Biomedical Sciences, Bharathidasan University, Tiruchirappalli 620024 (India); Sathishkumar, G. [Department of Biotechnology and Genetic Engineering, School of Biotechnology, Bharathidasan University, Tiruchirappalli 620024, Tamil Nadu (India); MubarakAli, D. [Central Inter-Disciplinary Research Facility, Mahatma Gandhi Medical College and Research Institute Campus, Pondicherry 607402 (India); Rajesh, M.; Sivanandhan, G.; Kapildev, G.; Manickavasagam, M. [Department of Biotechnology and Genetic Engineering, School of Biotechnology, Bharathidasan University, Tiruchirappalli 620024, Tamil Nadu (India); Thajuddin, N. [Department of Microbiology, Bharathidasan University, Tiruchirappalli 620024 (India); Ganapathi, A., E-mail: aganapathi2007@gmail.com [Department of Biotechnology and Genetic Engineering, School of Biotechnology, Bharathidasan University, Tiruchirappalli 620024, Tamil Nadu (India)

    2014-04-01

    Highlights: • Gold nanoparticles (AuNPs) have been synthesized using Podophyllum hexandrum L. • AuNPs induces the oxidative stress to cell death in human cervical carcinoma cells. • It activates the caspase-cascade to cellular death. • It is actively blocks G2/M phase of cell cycle. - Abstract: Current prospect of nanobiotechnology involves in the greener synthesis of nanostructured materials particularly noble metal nanoparticles for various biomedical applications. In this study, biologically (Podophyllum hexandrum L.) synthesized crystalline gold nanoparticles (AuNPs) with the size range between 5 and 35 nm were screened for its anticancereous potential against human cervical carcinoma cells (HeLa). Stoichiometric proportion of the reaction mixture and conditions were optimized to attain stable nanoparticles with narrow size range. Different high throughput techniques like transmission electron microscope (TEM), X-ray diffraction (XRD) and UV–vis spectroscopy were adopted for the physio-chemical characterization of AuNPs. Additionally, Fourier transform infrared spectroscopy (FTIR) study revealed that the water soluble fractions present in the plant extract solely influences the reduction of AuNPs. Sublimely, synthesized AuNPs exhibits an effective in vitro anticancer activity against HeLa cells via induction of cell cycle arrest and DNA damage. Furthermore, it was evidenced that AuNPs treated cells are undergone apoptosis through the activation of caspase cascade which subsequently leads to mitochondrial dysfunction. Thereby, this study proves that biogenic colloidal AuNPs can be developed as a promising drug candidature for human cervical cancer therapy.

  2. Inhibition of Oncogenic Transcription Factor REL by the Natural Product Derivative Calafianin Monomer 101 Induces Proliferation Arrest and Apoptosis in Human B-Lymphoma Cell Lines

    Science.gov (United States)

    Yeo, Alan T.; Chennamadhavuni, Spandan; Whitty, Adrian; Porco, John A.; Gilmore, Thomas D.

    2016-01-01

    Increased activity of transcription factor NF-κB has been implicated in many B-cell lymphomas. We investigated effects of synthetic compound calafianin monomer (CM101) on biochemical and biological properties of NF-κB. In human 293 cells, CM101 selectively inhibited DNA binding by overexpressed NF-κB subunits REL (human c-Rel) and p65 as compared to NF-κB p50, and inhibition of REL and p65 DNA binding by CM101 required a conserved cysteine residue. CM101 also inhibited DNA binding by REL in human B-lymphoma cell lines, and the sensitivity of several B-lymphoma cell lines to CM101-induced proliferation arrest and apoptosis correlated with levels of cellular and nuclear REL. CM101 treatment induced both phosphorylation and decreased expression of anti-apoptotic protein Bcl-XL, a REL target gene product, in sensitive B-lymphoma cell lines. Ectopic expression of Bcl-XL protected SUDHL-2 B-lymphoma cells against CM101-induced apoptosis, and overexpression of a transforming mutant of REL decreased the sensitivity of BJAB B-lymphoma cells to CM101-induced apoptosis. Lipopolysaccharide-induced activation of NF-κB signaling upstream components occurred in RAW264.7 macrophages at CM101 concentrations that blocked NF-κB DNA binding. Direct inhibitors of REL may be useful for treating B-cell lymphomas in which REL is active, and may inhibit B-lymphoma cell growth at doses that do not affect some immune-related responses in normal cells. PMID:25915462

  3. Dactylone inhibits epidermal growth factor-induced transformation and phenotype expression of human cancer cells and induces G1-S arrest and apoptosis.

    Science.gov (United States)

    Fedorov, Sergey N; Shubina, Larisa K; Bode, Ann M; Stonik, Valentin A; Dong, Zigang

    2007-06-15

    The marine natural chamigrane-type sesquiterpenoid, dactylone, is closely related to secondary metabolites of some edible species of red algae. In the present study, the effect of dactylone was tested on the mouse skin epidermal JB6 P+ Cl41 cell line and its stable transfectants as well as on several human tumor cell lines, including lung (H460), colon (HCT-116), and skin melanomas (SK-MEL-5 and SK-MEL-28). This natural product was effective at nontoxic doses as a cancer-preventive agent, which exerted its actions, at least in part, through the inhibition of cyclin D3 and Cdk4 expression and retinoblastoma tumor suppressor protein (Rb) phosphorylation. The inhibition of these cell cycle components was followed by cell cycle arrest at the G1-S transition with subsequent p53-independent apoptosis. Therefore, these data showed that application of dactylone and related compounds may lead to decreased malignant cell transformation and/or decreased tumor cell proliferation. PMID:17575161

  4. G protein-coupled receptor 30 ligand G-1 increases aryl hydrocarbon receptor signalling by inhibition of tubulin assembly and cell cycle arrest in human MCF-7 cells.

    Science.gov (United States)

    Tarnow, Patrick; Tralau, Tewes; Luch, Andreas

    2016-08-01

    Regulatory crosstalk between the aryl hydrocarbon receptor (AHR) and oestrogen receptor α (ERα) is well established. Apart from the nuclear receptors ERα and ERβ, oestrogen signalling further involves an unrelated G protein-coupled receptor termed GPR30. In order to investigate potential regulatory crosstalk, this study investigated the influence of G-1 as one of the few GPR30-specific ligands on the AHR regulon in MCF-7 cells. As a well-characterised model system, these human mammary carcinoma cells co-express all three receptors (AHR, ERα and GPR30) and are thus ideally suited to study corresponding regulatory pathway interactions on transcript level. Indeed, treatment with micromolar concentrations of the GPR30-specific agonist G-1 resulted in up-regulation of AHR as well as the transcripts for cytochromes P450 1A1 and 1B1, two well-known targets of the AHR regulon. While this was partly attributable to G-1-mediated inhibition of tubulin assembly and subsequent cell cycle arrest in the G2/M phase, the effects nevertheless required functional AHR. However, G-1-induced up-regulation of CYP 1A1 was not mediated by GPR30, as G15 antagonist treatment as well as a knockdown of GPR30 and AHR failed to inhibit this effect. PMID:26475489

  5. Effects of maple (Acer) plant part extracts on proliferation, apoptosis and cell cycle arrest of human tumorigenic and non-tumorigenic colon cells.

    Science.gov (United States)

    González-Sarrías, Antonio; Li, Liya; Seeram, Navindra P

    2012-07-01

    Phenolic-enriched extracts of maple sap and syrup, obtained from the sugar and red maple species (Acer saccharum Marsh, A. rubrum L., respectively), are reported to show anticancer effects. Despite traditional medicinal uses of various other parts of these plants by Native Americans, they have not been investigated for anticancer activity. Here leaves, stems/twigs, barks and sapwoods of both maple species were evaluated for antiproliferative effects against human colon tumorigenic (HCT-116, HT-29, Caco-2) and non-tumorigenic (CCD-18Co) cells. Extracts were standardized to total phenolic and ginnalin-A (isolated in our laboratory) levels. Overall, the extracts inhibited the growth of the colon cancer more than normal cells (over two-fold), their activities increased with their ginnalin-A levels, with red > sugar maple extracts. The red maple leaf extract, which contained the highest ginnalin-A content, was the most active extract (IC₅₀  = 35 and 16 µg/mL for extract and ginnalin-A, respectively). The extracts were not cytotoxic nor did they induce apoptosis of the colon cancer cells. However, cell cycle analyses revealed that the antiproliferative effects of the extracts were mediated through cell cycle arrest in the S-phase. The results from the current study suggest that these maple plant part extracts may have potential anticolon cancer effects. PMID:22147441

  6. Sudden Cardiac Arrest

    Science.gov (United States)

    ... scan, or MUGA, which shows how well your heart is pumping blood. Magnetic resonance imaging (MRI) which gives doctors detailed pictures of your heart. How is SCA treated? Sudden cardiac arrest should ...

  7. Cardiac arrest in children

    Directory of Open Access Journals (Sweden)

    Tress Erika

    2010-01-01

    Full Text Available Major advances in the field of pediatric cardiac arrest (CA were made during the last decade, starting with the publication of pediatric Utstein guidelines, the 2005 recommendations by the International Liaison Committee on Resuscitation, and culminating in multicenter collaborations. The epidemiology and pathophysiology of in-hospital and out-of-hospital CA are now well described. Four phases of CA are described and the term "post-cardiac arrest syndrome" has been proposed, along with treatment goals for each of its four phases: immediate post-arrest, early post-arrest, intermediate and recovery phase. Hypothermia is recommended to be considered as a therapy for post-CA syndrome in comatose patients after CA, and large multicenter prospective studies are underway. We reviewed landmark articles related to pediatric CA published during the last decade. We present the current knowledge of epidemiology, pathophysiology and treatment of CA relevant to pre-hospital and acute care health practitioners.

  8. Cardiac arrest in children.

    Science.gov (United States)

    Tress, Erika E; Kochanek, Patrick M; Saladino, Richard A; Manole, Mioara D

    2010-07-01

    Major advances in the field of pediatric cardiac arrest (CA) were made during the last decade, starting with the publication of pediatric Utstein guidelines, the 2005 recommendations by the International Liaison Committee on Resuscitation, and culminating in multicenter collaborations. The epidemiology and pathophysiology of in-hospital and out-of-hospital CA are now well described. Four phases of CA are described and the term "post-cardiac arrest syndrome" has been proposed, along with treatment goals for each of its four phases: immediate post-arrest, early post-arrest, intermediate and recovery phase. Hypothermia is recommended to be considered as a therapy for post-CA syndrome in comatose patients after CA, and large multicenter prospective studies are underway. We reviewed landmark articles related to pediatric CA published during the last decade. We present the current knowledge of epidemiology, pathophysiology and treatment of CA relevant to pre-hospital and acute care health practitioners. PMID:20930971

  9. Sudden Cardiac Arrest

    Science.gov (United States)

    ... Heart Risk Factors & Prevention Heart Diseases & Disorders Atrial Fibrillation (AFib) Sudden Cardiac Arrest (SCA) SCA: Who's At Risk? Prevention of SCA What Causes SCA? SCA Awareness Atrial Flutter Heart Block Heart Failure Sick Sinus Syndrome Substances & Heart Rhythm Disorders Symptoms & ...

  10. Sudden Cardiac Arrest

    Science.gov (United States)

    ... often are found in public places, such as shopping malls, golf courses, businesses, airports, airplanes, casinos, ... arrest In a study published online today in the New England Journal of Medicine , ...

  11. Cardiac arrest - cardiopulmonary resuscitation

    Institute of Scientific and Technical Information of China (English)

    Basri Lenjani; Besnik Elshani; Nehat Baftiu; Kelmend Pallaska; Kadir Hyseni; Njazi Gashi; Nexhbedin Karemani; Ilaz Bunjaku; Taxhidin Zaimi; Arianit Jakupi

    2014-01-01

    Objective:To investigate application of cardiopulmonary resuscitation(CPR) measures within the golden minutes inEurope.Methods:The material was taken from theUniversityClinical Center ofKosovo -EmergencyCentre inPristina, during the two(2) year period(2010-2011).The collected date belong to the patients with cardiac arrest have been recorded in the patients' log book protocol at the emergency clinic.Results:During the2010 to2011 in the emergency center of theCUCK inPristina have been treated a total of269 patients with cardiac arrest, of whom159 or59.1% have been treated in2010, and110 patients or40.9% in2011.Of the269 patients treated in the emergency centre,93 or34.6% have exited lethally in the emergency centre, and176 or 65.4% have been transferred to other clinics.In the total number of patients with cardiac arrest, males have dominated with186 cases, or69.1%.The average age of patients included in the survey was56.7 year oldSD±16.0 years.Of the269 patients with cardiac arrest, defibrillation has been applied for93 or34.6% of patients.In the outpatient settings defibrillation has been applied for3 or3.2% of patients.Patients were defibrillated with application of one to four shocks. Of27 cases with who have survived cardiac arrest, none of them have suffered cardiac arrest at home,3 or11.1% of them have suffered cardiac arrest on the street, and24 or88.9% of them have suffered cardiac arrest in the hospital.5 out of27 patients survived have ended with neurological impairment.Cardiac arrest cases were present during all days of the week, but frequently most reported cases have been onMonday with32.0% of cases, and onFriday with24.5% of cases. Conclusions:All survivors from cardiac arrest have received appropriate medical assistance within10 min from attack, which implies that if cardiac arrest occurs near an institution health care(with an opportunity to provide the emergent health care) the rate of survival is higher.

  12. Inflammatory monocytes and the pathogenesis of viral encephalitis

    Directory of Open Access Journals (Sweden)

    Terry Rachael L

    2012-12-01

    Full Text Available Abstract Monocytes are a heterogeneous population of bone marrow-derived cells that are recruited to sites of infection and inflammation in many models of human diseases, including those of the central nervous system (CNS. Ly6Chi/CCR2hi inflammatory monocytes have been identified as the circulating precursors of brain macrophages, dendritic cells and arguably microglia in experimental autoimmune encephalomyelitis; Alzheimer’s disease; stroke; and more recently in CNS infection caused by Herpes simplex virus, murine hepatitis virus, Theiler’s murine encephalomyelitis virus, Japanese encephalitis virus and West Nile virus. The precise differentiation pathways and functions of inflammatory monocyte-derived populations in the inflamed CNS remains a contentious issue, especially in regard to the existence of monocyte-derived microglia. Furthermore, the contributions of monocyte-derived subsets to viral clearance and immunopathology are not well-defined. Thus, understanding the pathways through which inflammatory monocytes migrate to the brain and their functional capacity within the CNS is critical to inform future therapeutic strategies. This review discusses some of the key aspects of inflammatory monocyte trafficking to the brain and addresses the role of these cells in viral encephalitis.

  13. Activation of protein tyrosine phosphatase non-receptor type 2 by spermidine exerts anti-inflammatory effects in human THP-1 monocytes and in a mouse model of acute colitis.

    Directory of Open Access Journals (Sweden)

    Belén Morón

    Full Text Available BACKGROUND: Spermidine is a dietary polyamine that is able to activate protein tyrosine phosphatase non-receptor type 2 (PTPN2. As PTPN2 is known to be a negative regulator of interferon-gamma (IFN-γ-induced responses, and IFN-γ stimulation of immune cells is a critical process in the immunopathology of inflammatory bowel disease (IBD, we wished to explore the potential of spermidine for reducing pro-inflammatory effects in vitro and in vivo. METHODS: Human THP-1 monocytes were treated with IFN-γ and/or spermidine. Protein expression and phosphorylation were analyzed by Western blot, cytokine expression by quantitative-PCR, and cytokine secretion by ELISA. Colitis was induced in mice by dextran sodium sulfate (DSS administration. Disease severity was assessed by recording body weight, colonoscopy and histology. RESULTS: Spermidine increased expression and activity of PTPN2 in THP-1 monocytes and reduced IFN-γ-induced phosphorylation of signal transducer and activator of transcription (STAT 1 and 3, as well as p38 mitogen-activated protein kinase (MAPK in a PTPN2 dependent manner. Subsequently, IFN-γ-induced expression/secretion of intracellular cell adhesion molecule (ICAM-1 mRNA, monocyte chemoattractant protein (MCP-1, and interleukin (IL-6 was reduced in spermidine-treated cells. The latter effects were absent in PTPN2-knockdown cells. In mice with DSS-induced colitis, spermidine treatment resulted in ameliorated weight loss and decreased mucosal damage indicating reduced disease severity. CONCLUSIONS: Activation of PTPN2 by spermidine ameliorates IFN-γ-induced inflammatory responses in THP-1 cells. Furthermore, spermidine treatment significantly reduces disease severity in mice with DSS-induced colitis; hence, spermidine supplementation and subsequent PTPN2 activation may be helpful in the treatment of chronic intestinal inflammation such as IBD.

  14. Immunomodulatory effects of human umbilical cord Wharton's jelly-derived mesenchymal stem cells on differentiation, maturation and endocytosis of monocyte-derived dendritic cells.

    Science.gov (United States)

    Saeidi, Mohsen; Masoud, Ahmad; Shakiba, Yadollah; Hadjati, Jamshid; Mohyeddin Bonab, Mandana; Nicknam, Mohammad Hossein; Latifpour, Mostafa; Nikbin, Behrooz

    2013-03-01

    The Wharton's jelly of the umbilical cord is believed to be a source of mesenchymal stem cells (MSCs) which can be therapeutically applied in degenerative diseases.In this study, we investigated the immunomodulatory effect of umbilical cord derived-mesenchymal stem cells (UC-MSCs) and bone marrow-derived-mesenchymal stem cells (BM-MSCs) on differentiation, maturation, and endocytosis of monocyte-derived dendritic cells in a transwell culture system under laboratory conditions. Monocytes were differentiated into immature dendritic cells (iDCs) in the presence of GM-CSF and IL-4 for 6 days and then differentiated into mature dendritic cells (mDCs) in the presence of TNF-α for 2 days. In every stage of differentiation, immature and mature dendritic cells were separately co-cultured with UC-MSCs and BM-MSCs. The findings showed that UC-MSCs and BM-MSCs inhibited strongly differentiation and maturation of dendritic cells at higher dilution ratios (1:1). The BM-MSCs and UC-MSCs showed more inhibitory effect on CD1a, CD83, CD86 expression, and dendritic cell endocytic activity, respectively. On the other hand, these cells severely up-regulated CD14 marker expression. We concluded that UC-MSCs and BM-MSCs could inhibit differentiation, maturation and endocytosis in monocyte-derived DCs through the secreted factors and free of any cell-cell contacts under laboratory conditions. As DCs are believed to be the main antigen presenting cells for naïve T cells in triggering immune responses, it would be logical that their inhibitory effect on differentiation, maturation and function can decrease or modulate immune and inflammatory responses. PMID:23454777

  15. Immunomodulatory effects of human umbilical cord Wharton's jelly-derived mesenchymal stem cells on differentiation, maturation and endocytosis of monocyte-derived dendritic cells.

    Directory of Open Access Journals (Sweden)

    Mohsen Saeidi

    2013-03-01

    Full Text Available The Wharton’s jelly of the umbilical cord is believed to be a source of mesenchymal stem cells (MSCs which can be therapeutically applied in degenerative diseases.In this study, we investigated the immunomodulatory effect of umbilical cord derived- mesenchymal stem  cells (UC-MSCs and  bone  marrow-derived-mesenchymal stem  cells (BM-MSCs on differentiation, maturation, and endocytosis of monocyte-derived dendritic cells in a transwell culture system under laboratory conditions. Monocytes were differentiated into immature dendritic cells (iDCs in the presence of GM-CSF and IL-4 for 6 days and then differentiated into mature dendritic cells (mDCs in the presence of TNF-α for 2 days. In every stage of differentiation, immature and mature dendritic cells were separately co- cultured with UC-MSCs and BM-MSCs.The findings showed that UC-MSCs and BM-MSCs inhibited strongly differentiation and maturation of dendritic cells at higher dilution ratios (1:1. The BM-MSCs and UC-MSCs showed more inhibitory effect on CD1a, CD83, CD86 expression, and dendritic cell endocytic activity, respectively. On the other hand, these cells severely up-regulated CD14 marker expression. We concluded that UC-MSCs and BM-MSCs could inhibit differentiation, maturation and endocytosis in monocyte-derived DCs through the secreted factors and free of any cell- cell contacts  under  laboratory conditions. As DCs  are believed to  be the  main antigen presenting cells for naïve T cells in triggering immune responses, it would be logical that their inhibitory effect on differentiation, maturation and function can decrease or modulate immune and inflammatory responses.

  16. Dual involvement of growth arrest-specific gene 6 in the early phase of human IgA nephropathy.

    Directory of Open Access Journals (Sweden)

    Kojiro Nagai

    Full Text Available BACKGROUND: Gas6 is a growth factor that causes proliferation of mesangial cells in the development of glomerulonephritis. Gas6 can bind to three kinds of receptors; Axl, Dtk, and Mer. However, their expression and functions are not entirely clear in the different glomerular cell types. Meanwhile, representative cell cycle regulatory protein p27 has been reported to be expressed in podocytes in normal glomeruli with decreased expression in proliferating glomeruli, which inversely correlated with mesangial proliferation in human IgA nephropathy (IgAN. METHODS: The aim of this study is to clarify Gas6 involvement in the progression of IgAN. Expression of Gas6/Axl/Dtk was examined in 31 biopsy proven IgAN cases. We compared the expression levels with histological severity or clinical data. Moreover, we investigated the expression of Gas6 and its receptors in cultured podocytes. RESULTS: In 28 of 31 cases, Gas6 was upregulated mainly in podocytes. In the other 3 cases, Gas6 expression was induced in endothelial and mesangial cells, which was similar to animal nephritis models. Among 28 podocyte type cases, the expression level of Gas6 correlated with the mesangial hypercellularity score of IgAN Oxford classification and urine protein excretion. It also inversely correlated with p27 expression in glomeruli. As for the receptors, Axl was mainly expressed in endothelial and mesangial cells, while Dtk was expressed in podocytes. In vitro, Dtk was expressed in cultured murine podocytes, and the expression of p27 was decreased by Gas6 stimulation. CONCLUSIONS: Gas6 was uniquely upregulated in either endothelial/mesangial cells or podocytes in IgAN. The expression pattern can be used as a marker to classify IgAN. Gas6 has a possibility to be involved in not only mesangial proliferation via Axl, but also podocyte injury via Dtk in IgAN.

  17. SKLB70326, a novel small-molecule inhibitor of cell-cycle progression, induces G{sub 0}/G{sub 1} phase arrest and apoptosis in human hepatic carcinoma cells

    Energy Technology Data Exchange (ETDEWEB)

    Han, Yuanyuan; He, Haiyun [State Key Laboratory of Biotherapy and Cancer Center, West China Hospital, West China Medical School, Sichuan University, Chengdu 610041 (China); Peng, Feng [Department of Thoracic Oncology of the Cancer Center and State Key Laboratory of Biotherapy, West China Hospital, West China Medical School, Sichuan University, Chengdu 610041 (China); Liu, Jiyan; Dai, Xiaoyun; Lin, Hongjun; Xu, Youzhi; Zhou, Tian; Mao, Yongqiu; Xie, Gang; Yang, Shengyong; Yu, Luoting; Yang, Li [State Key Laboratory of Biotherapy and Cancer Center, West China Hospital, West China Medical School, Sichuan University, Chengdu 610041 (China); Zhao, Yinglan, E-mail: alancenxb@sina.com [State Key Laboratory of Biotherapy and Cancer Center, West China Hospital, West China Medical School, Sichuan University, Chengdu 610041 (China)

    2012-05-18

    Highlights: Black-Right-Pointing-Pointer SKLB70326 is a novel compound and has activity of anti-HCC. Black-Right-Pointing-Pointer SKLB70326 induces cell cycle arrest and apoptosis in HepG2 cells. Black-Right-Pointing-Pointer SKLB70326 induces G{sub 0}/G{sub 1} phase arrest via inhibiting the activity of CDK2, CDK4 and CDK6. Black-Right-Pointing-Pointer SKLB70326 induces apoptosis through the intrinsic pathway. -- Abstract: We previously reported the potential of a novel small molecule 3-amino-6-(3-methoxyphenyl)thieno[2.3-b]pyridine-2-carboxamide (SKLB70326) as an anticancer agent. In the present study, we investigated the anticancer effects and possible mechanisms of SKLB70326 in vitro. We found that SKLB70326 treatment significantly inhibited human hepatic carcinoma cell proliferation in vitro, and the HepG2 cell line was the most sensitive to its treatment. The inhibition of cell proliferation correlated with G{sub 0}/G{sub 1} phase arrest, which was followed by apoptotic cell death. The SKLB70326-mediated cell-cycle arrest was associated with the downregulation of cyclin-dependent kinase (CDK) 2, CDK4 and CDK6 but not cyclin D1 or cyclin E. The phosphorylation of the retinoblastoma protein (Rb) was also observed. SKLB70326 treatment induced apoptotic cell death via the activation of PARP, caspase-3, caspase-9 and Bax as well as the downregulation of Bcl-2. The expression levels of p53 and p21 were also induced by SKLB70326 treatment. Moreover, SKLB70326 treatment was well tolerated. In conclusion, SKLB70326, a novel cell-cycle inhibitor, notably inhibits HepG2 cell proliferation through the induction of G{sub 0}/G{sub 1} phase arrest and subsequent apoptosis. Its potential as a candidate anticancer agent warrants further investigation.

  18. TELOMERE SHORTENING IN MONOCYTES OF THE PATIENTS WITH RHEUMATOID ARTHRITIS

    Directory of Open Access Journals (Sweden)

    V. I. Borisov

    2006-01-01

    Full Text Available Abstract. Present study deals with size measurements of telomeric DNA from the human peripheral mononuclear immune cells in rheumatoid arthritis (RA. A method for measuring the relative telomere length by in situ hybridization followed by flow cytometric analysis (flow-FISH was used. Relative telomere length (RTL in monocytes was estimated as mean fluorescence intensity (MFI of test cells divided by MFI values of internal control cells. Hybridization conditions for analysis of telomere length in monocytes have been optimized in advance. It has been shown that RTL of monocytes was significantly lower in RA patients compared to donors. Significant differences in telomere length of monocytes between RA patients and donors were revealed for the young persons under 30 years old. The findings obtained may be considered as an additional argument confirming the hypothesis on genetic defects of hematopoietic stem cells determining RA development.

  19. Statins attenuate polymethylmethacrylate-mediated monocyte activation.

    LENUS (Irish Health Repository)

    Laing, Alan J

    2012-02-03

    BACKGROUND: Periprosthetic osteolysis precipitates aseptic loosening of components, increases the risk of periprosthetic fracture and, through massive bone loss, complicates revision surgery and ultimately is the primary cause for failure of joint arthroplasty. The anti-inflammatory properties of HMG-CoA reductase inhibitors belonging to the statin family are well recognized. We investigated a possible role for status in initiating the first stage of the osteolytic cycle, namely monocytic activation. METHODS: We used an in vitro model of the human monocyte\\/macrophage inflammatory response to poly-methylmethacrylate (PMMA) particles after pretreat-ing cells with cerivastatin, a potent member of the statin family. Cell activation based upon production of TNF-alpha and MCP-1 cytokines was analyzed and the intracellular Raf-MEK-ERK signal transduction pathway was evaluated using western blot analysis, to identify its role in cell activation and in any cerivastatin effects observed. RESULTS: We found that pretreatment with cerivastatin significantly abrogates the production of inflammatory cytokines TNF-alpha and MCP-1 by human monocytes in response to polymethylmethacrylate particle activation. This inflammatory activation and attenuation appear to be mediated through the intracellular Raf-MEK-ERK pathway. INTERPRETATION: We propose that by intervening at the upstream activation stage, subsequent osteoclast activation and osteolysis can be suppressed. We believe that the anti-inflammatory properties of statins may potentially play a prophylactic role in the setting of aseptic loosening, and in so doing increase implant longevity.

  20. Cellular pharmacokinetics and intracellular activity of the novel peptide deformylase inhibitor GSK1322322 against Staphylococcus aureus laboratory and clinical strains with various resistance phenotypes: studies with human THP-1 monocytes and J774 murine macrophages.

    Science.gov (United States)

    Peyrusson, Frédéric; Butler, Deborah; Tulkens, Paul M; Van Bambeke, Françoise

    2015-09-01

    GSK1322322 is a peptide deformylase inhibitor active against Staphylococcus aureus strains resistant to currently marketed antibiotics. Our aim was to assess the activity of GSK1322322 against intracellular S. aureus using an in vitro pharmacodynamic model and, in parallel, to examine its cellular pharmacokinetics and intracellular disposition. For intracellular activity analysis, we used an established model of human THP-1 monocytes and tested one fully susceptible S. aureus strain (ATCC 25923) and 8 clinical strains with resistance to oxacillin, vancomycin, daptomycin, macrolides, clindamycin, linezolid, or moxifloxacin. Uptake, accumulation, release, and subcellular distribution (cell fractionation) of [(14)C]GSK1322322 were examined in uninfected murine J774 macrophages and uninfected and infected THP-1 monocytes. GSK1322322 demonstrated a uniform activity against the intracellular forms of all S. aureus strains tested, disregarding their resistance phenotypes, with a maximal relative efficacy (E max) of a 0.5 to 1 log10 CFU decrease compared to the original inoculum within 24 h and a static concentration (C s) close to its MIC in broth. Influx and efflux were very fast (gemfibrozil and verapamil. GSK1322322 was recovered in the cell-soluble fraction and was dissociated from the main subcellular organelles and from bacteria (in infected cells). The results of this study show that GSK1322322, as a typical novel deformylase inhibitor, may act against intracellular forms of S. aureus. They also suggest that GSK1322322 has the ability to freely diffuse into and out of eukaryotic cells as well as within subcellular compartments. PMID:26169402

  1. P53-mediated cell cycle arrest and apoptosis through a caspase-3-independent, but caspase-9-dependent pathway in oridonin-treated MCF-7 human breast cancer cells

    Institute of Scientific and Technical Information of China (English)

    Qiao CUI; Jing-hua YU; Jin-nan WU; Shin-ichi TASHIRO; Satoshi ONODERA; Mutsuhiko MINAMI; Takashi IKEJIMA

    2007-01-01

    Aim: To study the caspase-3-independent mechanisms in oridonin-induced MCF-7 human breast cancer cell apoptosis in vitro. Methods: The viability of oridonin-treated MCF-7 cells was measured by MTT (thiazole blue) assay. Apoptotic cells with condensed nuclei were visualized by phase contrast microscopy. Nucleoso-mal DNA fragmentation was assayed by agarose gel electrophoresis. The apoptotic ratio was determined by lactate dehydrogenase assay. Cell cycle alternation and mitochondrial membrane potential were measured by flow cytometric analysis. Bax, Bcl-2, caspase-3, caspase-9, heat shock protein (Hsp)90, p53, p-p53, p21, Poly (ADP-ribose) polymerase (PARP), and the inhibitor of caspase-activated Dnase (ICAD) protein expressions were detected by Western blot analysis. Results: Oridonin inhibited cell growth in a time- and dose-dependent manner. Cell cycle was altered through the upregulation of p53 and p21 protein expressions. Pan-caspase inhibitor Z-VAD-fmk and calpain inhibitor Ⅱ both decreased cell death ratio. Nucleosomal DNA fragmentation and the downregulation of △ψmit were detected in oridonin-induced MCF-7 cell apoptosis, which was involved in a postmitochondrial caspase-9-dependent pathway. Decreased Bcl-2 and Hsp90 expression levels and increased Bax and p21 expression levels were positively correlated with elevated levels of phosphorylated p53 phosphorylation. Moreover, PARP was partially cleaved by calpain rather than by capase-3. Conclusion: DNA damage provoked alternations in the mitochondrial and caspase-9 pathways as well as p53-mediated cell cycle arrest, but was not related to caspase-3 activity in oridonin-induced MCF-7 cells.

  2. Induction of G1 and G2/M cell cycle arrests by the dietary compound 3,3'-diindolylmethane in HT-29 human colon cancer cells

    Directory of Open Access Journals (Sweden)

    Choi Hyun

    2009-05-01

    Full Text Available Abstract Background 3,3'-Diindolylmethane (DIM, an indole derivative produced in the stomach after the consumption of broccoli and other cruciferous vegetables, has been demonstrated to exert anti-cancer effects in both in vivo and in vitro models. We have previously determined that DIM (0 – 30 μmol/L inhibited the growth of HT-29 human colon cancer cells in a concentration-dependent fashion. In this study, we evaluated the effects of DIM on cell cycle progression in HT-29 cells. Methods HT-29 cells were cultured with various concentrations of DIM (0 – 30 μmol/L and the DNA was stained with propidium iodide, followed by flow cytometric analysis. [3H]Thymidine incorporation assays, Western blot analyses, immunoprecipitation and in vitro kinase assays for cyclin-dependent kinase (CDK and cell division cycle (CDC2 were conducted. Results The percentages of cells in the G1 and G2/M phases were dose-dependently increased and the percentages of cells in S phase were reduced within 12 h in DIM-treated cells. DIM also reduced DNA synthesis in a dose-dependent fashion. DIM markedly reduced CDK2 activity and the levels of phosphorylated retinoblastoma proteins (Rb and E2F-1, and also increased the levels of hypophosphorylated Rb. DIM reduced the protein levels of cyclin A, D1, and CDK4. DIM also increased the protein levels of CDK inhibitors, p21CIP1/WAF1 and p27KIPI. In addition, DIM reduced the activity of CDC2 and the levels of CDC25C phosphatase and cyclin B1. Conclusion Here, we have demonstrated that DIM induces G1 and G2/M phase cell cycle arrest in HT-29 cells, and this effect may be mediated by reduced CDK activity.

  3. Apoptosis induced by genipin in human leukemia K562 cells:involvement of C-Jun N-terminal kinase in G2/M arrest

    Institute of Scientific and Technical Information of China (English)

    Qian FENG; Hou-li CAO; Wei XU; Xiao-rong LI; Yan-qin REN; Lin-fang DU

    2011-01-01

    Aim:To investigate the effect of genipin on apoptosis in human leukemia K562 cells in vitro and elucidate the underlying mechanisms.Methods:The effect of genipin on K562 cell viability was measured using trypan blue dye exclusion and cell counting.Morphological changes were detected using phase-contrast microscopy.Apoptosis was analyzed using DNA ladder, propidium iodide(PI)-labeled flow cytometry(FCM)and Hoechst 33258 staining.The infiuence of genipin on cell cycle distribution was determined using Plstaining.Caspase 3 activity was analyzed to detect apoptosis at different time points.Protein levels of phospho-c-Jun,phosphor-C-Jun N-terminal kinase(p-JNK).phosphor-p38-Fas-L,p63,and Bax and the release of cytochrome c were detected using Western blot analysis.Results:Genipin reduced the viability of K562 cells with an IC50 value of approximately 250 μmol/L.Genipin 200-400 μmol/L induced formation of typicaI apoptotic bodies and DNA fragmentation.Additionally,genipin 400 μmol/L significantly increased the caspase 3activity from 8-24 h and arrested the cells in the G2/M phase.After stimulation with genipin 500 μmol/L, the levels of p-JNK, p-c-Jun.Fas-L,Bax.and cytochrome c were remarkably upregulated,but there were no obvious changes of p-p38.Genipin 200-500 μmol/Lsignificantly upregulated the Fas-L expression and downregulated p63 expression.Dicoumarol 100 μmol/L.a JNK1/2 inhibitor,markedly suppressed the formation of apoptotic bodies and JNK activation induced by genipin 400 μmol/L.Conclusion:These results suggest that genipin inhibits the proliferation of K562 cells and induces apoptosis through the activation of JNK and induction of the Fas ligand.

  4. Gallic Acid Is the Major Active Component of Cortex Moutan in Inhibiting Immune Maturation of Human Monocyte-Derived Dendritic Cells

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    Ben Chung Lap Chan

    2015-09-01

    Full Text Available Atopic dermatitis (AD is a widely prevalent and chronically relapsing inflammatory skin disease. Penta Herbs Formula (PHF is efficacious in improving the quality of life and reducing topical corticosteroid used in children with AD and one of the active herbs it contains is Cortex Moutan. Recent studies showed that altered functions of dendritic cells (DC were observed in atopic individuals, suggesting that DC might play a major role in the generation and maintenance of inflammation by their production of pro-inflammatory cytokines. Hence, the aims of the present study were to identify the major active component(s of Cortex Moutan, which might inhibit DC functions and to investigate their possible interactions with conventional corticosteroid on inhibiting the development of DC from monocytes. Monocyte-derived dendritic cells (moDC culture model coupled with the high-speed counter-current chromatography (HSCCC, high pressure liquid chromatography (HPLC and Liquid Chromatography-Mass Spectrometry (LCMS analyses were used. Gallic acid was the major active component from Cortex Moutan which could dose dependently inhibit interleukin (IL-12 p40 and the functional cluster of differentiation (CD surface markers CD40, CD80, CD83 and CD86 expression from cytokine cocktail-activated moDC. Gallic acid could also lower the concentration of hydrocortisone required to inhibit the activation of DC.

  5. Activation of the canonical Wnt/β-catenin pathway enhances monocyte adhesion to endothelial cells

    International Nuclear Information System (INIS)

    Monocyte adhesion to vascular endothelium has been reported to be one of the early processes in the development of atherosclerosis. In an attempt to develop strategies to prevent or delay atherosclerosis progression, we analyzed effects of the Wnt/β-catenin signaling pathway on monocyte adhesion to various human endothelial cells. Adhesion of fluorescein-labeled monocytes to various human endothelial cells was analyzed under a fluorescent microscope. Unlike sodium chloride, lithium chloride enhanced monocyte adhesion to endothelial cells in a dose-dependent manner. We further demonstrated that inhibitors for glycogen synthase kinase (GSK)-3β or proteosome enhanced monocyte-endothelial cell adhesion. Results of semi-quantitative reverse transcriptase polymerase chain reaction (RT-PCR) indicated that activation of Wnt/β-catenin pathway did not change expression levels of mRNA for adhesion molecules. In conclusion, the canonical Wnt/β-catenin pathway enhanced monocyte-endothelial cell adhesion without changing expression levels of adhesion molecules

  6. A novel site contributing to growth-arrest-specific gene 6 binding to its receptors as revealed by a human monoclonal antibody

    Science.gov (United States)

    2004-01-01

    Gas6 (growth-arrest-specific gene 6) is a vitamin K-dependent protein known to activate the Axl family of receptor tyrosine kinases. It is an important regulator of thrombosis and many other biological functions. The C-terminus of Gas6 binds to receptors and consists of two laminin-like globular domains LG1 and LG2. It has been reported that a Ca2+-binding site at the junction of LG1 and LG2 domains and a hydrophobic patch at the LG2 domain are important for receptor binding [Sasaki, Knyazev, Cheburkin, Gohring, Tisi, Ullrich, Timpl and Hohenester (2002) J. Biol. Chem. 277, 44164–44170]. In the present study, we developed a neutralizing human monoclonal antibody, named CNTO300, for Gas6. The antibody was generated by immunization of human IgG-expressing transgenic mice with recombinant human Gas6 protein and the anti-Gas6 IgG sequences were rescued from an unstable hybridoma clone. Binding of Gas6 to its receptors was partially inhibited by the CNTO300 antibody in a dose-dependent manner. To characterize further the interaction between Gas6 and this antibody, the binding kinetics of CNTO300 for recombinant Gas6 were compared with independently expressed LG1 and LG2. The CNTO300 antibody showed comparable binding affinity, yet different dependence on Ca2+, to Gas6 and LG1. No binding to LG2 was detected. In the presence of EDTA, binding of the antibody to Gas6 was disrupted, but no significant effect of EDTA on LG1 binding was evident. Further epitope mapping identified a Gas6 peptide sequence recognized by the CNTO300 antibody. This peptide sequence was found to be located at the LG1 domain distant from the Ca2+-binding site and the hydrophobic patch. Co-interaction of Gas6 with its receptor and CNTO300 antibody was detected by BIAcore analysis, suggesting a second receptor-binding site on the LG1 domain. This hypothesis was further supported by direct binding of Gas6 receptors to an independently expressed LG1 domain. Our results revealed, for the first time, a

  7. Transcriptome analysis of monocyte-HIV interactions

    Directory of Open Access Journals (Sweden)

    Tran Huyen

    2010-06-01

    Full Text Available Abstract Background During HIV infection and/or antiretroviral therapy (ART, monocytes and macrophages exhibit a wide range of dysfunctions which contribute significantly to HIV pathogenesis and therapy-associated complications. Nevertheless, the molecular components which contribute to these dysfunctions remain elusive. We therefore applied a parallel approach of genome-wide microarray analysis and focused gene expression profiling on monocytes from patients in different stages of HIV infection and/or ART to further characterise these dysfunctions. Results Processes involved in apoptosis, cell cycle, lipid metabolism, proteasome function, protein trafficking and transcriptional regulation were identified as areas of monocyte dysfunction during HIV infection. Individual genes potentially contributing to these monocyte dysfunctions included several novel factors. One of these is the adipocytokine NAMPT/visfatin, which we show to be capable of inhibiting HIV at an early step in its life cycle. Roughly half of all genes identified were restored to control levels under ART, while the others represented a persistent dysregulation. Additionally, several candidate biomarkers (in particular CCL1 and CYP2C19 for the development of the abacavir hypersensitivity reaction were suggested. Conclusions Previously described areas of monocyte dysfunction during HIV infection were confirmed, and novel themes were identified. Furthermore, individual genes associated with these dysfunctions and with ART-associated disorders were pinpointed. These genes form a useful basis for further functional studies concerning the contribution of monocytes/macrophages to HIV pathogenesis. One such gene, NAMPT/visfatin, represents a possible novel restriction factor for HIV. Background Both macrophages and T lymphocyte subsets express the CD4 receptor and either the CXCR4 and/or the CCR5 coreceptor which confer susceptibility to infection with the Human Immunodeficiency Virus

  8. Polypeptide structure of a human dermal fibroblast-activating factor (FAF) derived from the U937 cultured line of human monocyte-like cells

    International Nuclear Information System (INIS)

    Six liter batches of 1 x 106 U937 cells/ml of serum-free RPMI medium were incubated with 100 ng/ml of phorbol myristate acetate for 48 hr at 370C in 5% CO2 in air to generate FAFs, as quantified by the stimulation of uptake of [3H]thymidine by quiescent human dermal fibroblasts. Filtration of the supernatants on Sephadex G-75 resolved two FAFs of approximately 40,000 and 10-13,000 daltons. The latter principle was purified to homogeneity by sequential Sephadex G-50 filtration, revealing an apparent m.w. of 7-8000, Mono-Q FPLC anion-exchange chromatography with a linear gradient from 20 mM Tris-HCl (pH 8.3) to 0.5 M NaCl-20 mM Tris-HCl in 30 min, and two cycles of high-performance liquid chromatography (HPLC) on a 300 A pore 10 μm C4 column at 1 ml/min with 0.05% trifluoroacetic acid (TFA) in water to 30:70 (v:v) and then to 60:40 (v:v) acetonitrile: 0.05% TFA linearly in 15 min and 30 min, respectively, The FAF activity eluted from HPLC in a sharp peak of O.D. 215 nm at 45% acetonitrile. Analyses of amino acid composition of the highly purified 7-8000 dalton FAF-U937 revealed 37% hydrophobic, 14% basic, and 21% acidic or amide residues, as well as one tryrosine and one methionine. This U937 cell-derived FAF appears to be a unique acidic polypeptide growth factor

  9. Monocytes in inflammatory bowel disease: absolute monocyte counts.

    OpenAIRE

    Mee, A. S.; Berney, J.; Jewell, D P

    1980-01-01

    Using a cytochemical staining technique, peripheral blood monocytes have been precisely identified and enumerated in patients with inflammatory bowel disease and compared with healthy and disease control subjects. For ulcerative colitis there was a significant monocytosis, which was closely correlated with the total white cell count and with the activity of the disease. For patients with Crohn's disease, the peripheral blood monocyte count was also raised compared with that of the control gro...

  10. Growth arrest by the antitumor steroidal lactone withaferin A in human breast cancer cells is associated with down-regulation and covalent binding at cysteine 303 of β-tubulin.

    Science.gov (United States)

    Antony, Marie L; Lee, Joomin; Hahm, Eun-Ryeong; Kim, Su-Hyeong; Marcus, Adam I; Kumari, Vandana; Ji, Xinhua; Yang, Zhen; Vowell, Courtney L; Wipf, Peter; Uechi, Guy T; Yates, Nathan A; Romero, Guillermo; Sarkar, Saumendra N; Singh, Shivendra V

    2014-01-17

    Withaferin A (WA), a C5,C6-epoxy steroidal lactone derived from a medicinal plant (Withania somnifera), inhibits growth of human breast cancer cells in vitro and in vivo and prevents mammary cancer development in a transgenic mouse model. However, the mechanisms underlying the anticancer effect of WA are not fully understood. Herein, we report that tubulin is a novel target of WA-mediated growth arrest in human breast cancer cells. The G2 and mitotic arrest resulting from WA exposure in MCF-7, SUM159, and SK-BR-3 cells was associated with a marked decrease in protein levels of β-tubulin. These effects were not observed with the naturally occurring C6,C7-epoxy analogs of WA (withanone and withanolide A). A non-tumorigenic normal mammary epithelial cell line (MCF-10A) was markedly more resistant to mitotic arrest by WA compared with breast cancer cells. Vehicle-treated control cells exhibited a normal bipolar spindle with chromosomes aligned along the metaphase plate. In contrast, WA treatment led to a severe disruption of normal spindle morphology. NMR analyses revealed that the A-ring enone in WA, but not in withanone or withanolide A, was highly reactive with cysteamine and rapidly succumbed to irreversible nucleophilic addition. Mass spectrometry demonstrated direct covalent binding of WA to Cys(303) of β-tubulin in MCF-7 cells. Molecular docking indicated that the WA-binding pocket is located on the surface of β-tubulin and characterized by a hydrophobic floor, a hydrophobic wall, and a charge-balanced hydrophilic entrance. These results provide novel insights into the mechanism of growth arrest by WA in breast cancer cells. PMID:24297176

  11. Prion protein induced signaling cascades in monocytes

    International Nuclear Information System (INIS)

    Prion proteins play a central role in transmission and pathogenesis of transmissible spongiform encephalopathies. The cellular prion protein (PrPC), whose physiological function remains elusive, is anchored to the surface of a variety of cell types including neurons and cells of the lymphoreticular system. In this study, we investigated the response of a mouse monocyte/macrophage cell line to exposure with PrPC fusion proteins synthesized with a human Fc-tag. PrPC fusion proteins showed an attachment to the surface of monocyte/macrophages in nanomolar concentrations. This was accompanied by an increase of cellular tyrosine phosphorylation as a result of activated signaling pathways. Detailed investigations exhibited activation of downstream pathways through a stimulation with PrP fusion proteins, which include phosphorylation of ERK1,2 and Akt kinase. Macrophages opsonize and present antigenic structures, contact lymphocytes, and deliver cytokines. The findings reported here may become the basis of understanding the molecular function of PrPC in monocytes and macrophages

  12. Review: the Multiple Roles of Monocytic Microparticles.

    Science.gov (United States)

    Halim, Ahmad Tarmizi Abdul; Ariffin, Nur Azrah Fazera Mohd; Azlan, Maryam

    2016-08-01

    Monocytic microparticles (mMP) are microparticles derived from human monocytes either under in vivo or in vitro conditions. The size of mMP is between 0.1 and 1.0 μm. Apart from the size range, mMPs are also identified based on phosphatidylserine and CD14 expression on their surface, though this is not always the case. Monocytic MP are critical players in inflammation, endothelial cell function, and blood coagulation. They exhibit dual function by either helping the progression of such conditions or limiting it, depending on certain factors. Furthermore, the numbers of mMP are elevated in some autoimmune diseases, infectious diseases, and metabolic disorders. However, it is unknown whether mMP play an active role in these diseases or are simply biomarkers. The mechanism of mMP modulation is yet to be identified. In this review, we highlight the mechanism of mMP formation and the roles that they play in inflammation, blood coagulation, and different disease settings. PMID:27216803

  13. Lipopolysaccharide regulated protein expression is only partly impaired in monocytes from patients with type I diabetes

    Directory of Open Access Journals (Sweden)

    Abke Sabine

    2006-03-01

    Full Text Available Abstract Background Monocytes play an important role in innate immunity and atherosclerosis. A disturbed secretion of cytokines in lipopolysaccharide (LPS activated monocytes from type 1 diabetes (T1D patients has been described and may contribute to the impaired inflammatory response in these individuals. In the present study the influence of LPS on five different proteins with a function in immunity and atherosclerosis was analyzed in monocytes from controls and T1D patients. Methods Monocytes were isolated from controls and T1D patients and the LPS-stimulated increase of IL-6, CXCL8, monocyte chemotactic protein 1 (CCL2, MCP-1 and superoxide dismutase (SOD 2, as well as the LPS-mediated decrease of apolipoprotein E (Apo E in primary human monocytes from controls and T1D patients was determined. Results CCL2 and IL-6 secretion in response to LPS was found significantly reduced in monocytes from T1D patients when compared to controls whereas basal CCL2 release was similar in control and T1D cells. In contrast, CXCL8 and apolipoprotein E secretion and SOD 2 expression upon LPS stimulation is similar from T1D and control monocytes. Conclusion These data indicate that LPS-mediated protein expression is only partly disturbed in monocytes from T1D patients. Reduced secretion of IL-6 and CCL2 in activated monocytes of these patients may contribute to an impaired inflammatory response and vascular disease.

  14. Palmitic Acid-Induced Neuron Cell Cycle G2/M Arrest and Endoplasmic Reticular Stress through Protein Palmitoylation in SH-SY5Y Human Neuroblastoma Cells

    Directory of Open Access Journals (Sweden)

    Yung-Hsuan Hsiao

    2014-11-01

    Full Text Available Obesity-related neurodegenerative diseases are associated with elevated saturated fatty acids (SFAs in the brain. An increase in SFAs, especially palmitic acid (PA, triggers neuron cell apoptosis, causing cognitive function to deteriorate. In the present study, we focused on the specific mechanism by which PA triggers SH-SY5Y neuron cell apoptosis. We found that PA induces significant neuron cell cycle arrest in the G2/M phase in SH-SY5Y cells. Our data further showed that G2/M arrest is involved in elevation of endoplasmic reticular (ER stress according to an increase in p-eukaryotic translation inhibition factor 2α, an ER stress marker. Chronic exposure to PA also accelerates beta-amyloid accumulation, a pathological characteristic of Alzheimer’s disease. Interestingly, SFA-induced ER stress, G2/M arrest and cell apoptosis were reversed by treatment with 2-bromopalmitate, a protein palmitoylation inhibitor. These findings suggest that protein palmitoylation plays a crucial role in SFA-induced neuron cell cycle G2/M arrest, ER stress and apoptosis; this provides a novel strategy for preventing SFA-induced neuron cell dysfunction.

  15. Crack-arrest technology

    International Nuclear Information System (INIS)

    Over the last several years, the Heavy Section Steel Technology (HSST) Program has conducted several fracture mechanics experiments on large specimens that produced crack-arrest fracture-toughness values above 220 MPa·√m, which is the limit imposed by the ASME Code and the limit included in the Issues on Pressurized Thermal Shock studies. It is therefore appropriate and timely to investigate the influence that these high crack-arrest data have on the integrity assessment of nuclear Reactor Pressure Vessels (RPVs). A review of the evolution of the Pressurized Thermal Shock (PTS) issue and current methods of analysis provides insight into the motivation for the HSST Program performing the large-specimen fracture mechanics experiments. During the early 1970s, it was recognized that RPVs could be subjected to severe thermal shock as the result of a large-break loss-of-coolant accident (LBLOCA). Analyses performed at that time indicated that thermal shock alone would not result in failure (through-wall cracking) of the vessel. However, a combination of pressure and a less severe thermal shock, the result of some postulated transients, could result in vessel failure. In March 1978, such a transient occurred at the Rancho Seco nuclear power plant. As a result of these events, parametric PTS studies were undertaken. Because of the apparent need for and the existence of high-temperature crack-arrest capability, the NRC HSST Program and others began to investigate the effect of higher crack-arrest values on the probability of failure and to determine if these values actually exist for prototypical RPV materials. This report describes the results of HSST Program large-specimen crack-arrest testing

  16. TNF Drives Monocyte Dysfunction with Age and Results in Impaired Anti-pneumococcal Immunity

    OpenAIRE

    Puchta, Alicja; Naidoo, Avee; Verschoor, Chris P.; Loukov, Dessi; Thevaranjan, Netusha; Mandur, Talveer S.; Nguyen, Phuong-son; Jordana, Manel; Loeb, Mark; Xing, Zhou; Kobzik, Lester; Larché, Maggie J.; Bowdish, Dawn M. E.

    2016-01-01

    Monocyte phenotype and output changes with age, but why this occurs and how it impacts anti-bacterial immunity are not clear. We found that, in both humans and mice, circulating monocyte phenotype and function was altered with age due to increasing levels of TNF in the circulation that occur as part of the aging process. Ly6C+ monocytes from old (18–22 mo) mice and CD14+CD16+ intermediate/inflammatory monocytes from older adults also contributed to this “age-associated inflammation” as they p...

  17. RBP-J-interacting and tubulin-associated protein induces apoptosis and cell cycle arrest in human hepatocellular carcinoma by activating the p53–Fbxw7 pathway

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Haihe [The Key Laboratory of Molecular Diagnosis in Laboratory Medicine, Department of Pathogenobiology, Daqing Branch of Harbin Medical University, Daqing 163319 (China); Yang, Zhanchun [Department of General Surgery of Fifth Clinical Hospital of Harbin Medical University, Daqing 163319 (China); Liu, Chunbo; Huang, Shishun; Wang, Hongzhi; Chen, Yingli [The Key Laboratory of Molecular Diagnosis in Laboratory Medicine, Department of Pathogenobiology, Daqing Branch of Harbin Medical University, Daqing 163319 (China); Chen, Guofu, E-mail: zhangyanjie3@aliyun.com [Department of General Surgery of Fifth Clinical Hospital of Harbin Medical University, Daqing 163319 (China)

    2014-11-07

    Highlights: • RITA overexpression increased protein expression of p53 and Fbxw7 and downregulated the expression of cyclin D1, cyclin E, CDK2, Hes-1 and NF-κB p65. • RITA can significantly inhibit the in vitro growth of SMMC7721 and HepG2 cells. • RITA exerts tumor-suppressive effects in hepatocarcinogenesis through induction of G0/G1 cell cycle arrest and apoptosis and suggest a therapeutic application of RITA in HCC. - Abstract: Aberrant Notch signaling is observed in human hepatocellular carcinoma (HCC) and has been associated with the modulation of cell growth. However, the role of Notch signaling in HCC and its underlying mechanism remain elusive. RBP-J-interacting and tubulin-associated (RITA) mediates the nuclear export of RBP-J to tubulin fibers and downregulates Notch-mediated transcription. In this study, we found that RITA overexpression increased protein expression of p53 and Fbxw7 and downregulated the expression of cyclin D1, cyclin E, CDK2, Hes-1 and NF-κB p65. These changes led to growth inhibition and induced G0/G1 cell cycle arrest and apoptosis in SMMC7721 and HepG2 cells. Our findings indicate that RITA exerts tumor-suppressive effects in hepatocarcinogenesis through induction of G0/G1 cell cycle arrest and apoptosis and suggest a therapeutic application of RITA in HCC.

  18. Ex vivo expanded human regulatory T cells delay islet allograft rejection via inhibiting islet-derived monocyte chemoattractant protein-1 production in CD34+ stem cells-reconstituted NOD-scid IL2rγnull mice.

    Directory of Open Access Journals (Sweden)

    Fang Xiao

    Full Text Available Type 1 diabetes mellitus (T1DM is an autoimmune disease caused by immune-mediated destruction of insulin-secreting β cells of the pancreas. Near complete dependence on exogenous insulin makes T1DM very difficult to control, with the result that patients are exposed to high blood glucose and risk of diabetic complications and/or intermittent low blood glucose that can cause unconsciousness, fits and even death. Allograft transplantation of pancreatic islets restores normoglycemia with a low risk of surgical complications. However, although successful immediately after transplantation, islets are progressively lost, with most of the patients requiring exogenous insulin within 2 years post-transplant. Therefore, there is an urgent requirement for the development of new strategies to prevent islet rejection. In this study, we explored the importance of human regulatory T cells in the control of islets allograft rejection. We developed a pre-clinical model of human islet transplantation by reconstituting NOD-scid IL2rγnull mice with cord blood-derived human CD34+ stem cells and demonstrated that although the engrafted human immune system mediated the rejection of human islets, their survival was significantly prolonged following adoptive transfer of ex vivo expanded human Tregs. Mechanistically, Tregs inhibited the infiltration of innate immune cells and CD4+ T cells into the graft by down-regulating the islet graft-derived monocyte chemoattractant protein-1. Our findings might contribute to the development of clinical strategies for Treg therapy to control human islet rejection. We also show for the first time that CD34+ cells-reconstituted NOD-scid IL2rγnull mouse model could be beneficial for investigating human innate immunity in vivo.

  19. The purinergic receptor P2X7 role in control of Dengue virus-2 infection and cytokine/chemokine production in infected human monocytes.

    Science.gov (United States)

    Corrêa, Gladys; de A Lindenberg, Carolina; Fernandes-Santos, Caroline; Gandini, Mariana; Petitinga Paiva, Fabienne; Coutinho-Silva, Robson; F Kubelka, Claire

    2016-07-01

    Purinergic signaling has a crucial role in intracellular pathogen elimination. The P2X7 purinergic receptor (P2X7R), once activated by ATP, leads to pro-inflammatory responses including reactive oxygen species production. ATP can be released by injured cells, as endogenous danger signals. Dengue fever may evolve to a severe disease, leading to hypovolemic shock and coagulation dysfunctions as a result of a cytokine storm. Our aim was to evaluate the role of P2X7R activation during Dengue virus (DENV) infection. Extracellular ATP inhibited viral load in pretreated monocytes, as measured by NS1 secretion and by decrease in DENV(+) P2X7(+) cell frequencies, suggesting that P2X7R is involved in the antiviral response. Nitric oxide (NO) has anti-DENV properties and is decreased after DENV infection. NO production after ATP stimulation is abrogated by KN62 treatment, a specific P2X7R inhibitor, indicating that P2X7R likely is acting in the virus containment process. Additionally, TNF, CXCL8, CCL2 and CXCL10 factors that are associated with dengue severity were modulated by the P2X7R activation. We conclude that P2X7R is directly involved in the modulation of the antiviral and inflammatory process that occurs during DENV infection in vitro, and may have an important role in patient recovery in a first moment. PMID:26969484

  20. Immunologic effector mechanisms of a standardized mistletoe extract on the function of human monocytes and lymphocytes in vitro, ex vivo, and in vivo.

    Science.gov (United States)

    Heinzerling, Lucie; von Baehr, Volker; Liebenthal, Christa; von Baehr, Rüdiger; Volk, Hans-Dieter

    2006-07-01

    Even though mistletoe extracts have been in clinical use for centuries their exact mode of action is still unknown. Currently, the application scheme for registered preparations is a dose-escalating scheme to thus reduce side effects. In this study, healthy controls and patients were evaluated for their immunologic response to treatment with a standardized mistletoe extract (Iscador). It shows a strong effect as adjuvant that induces TNF-alpha and IL-12, which was partly mediated via CD14. Desensitization of the TNF-alpha response could be shown after repeated application in vitro and in vivo. Furthermore, Iscador induces a specific lymphocyte sensitization upon multiple injections and production of IgG1- and IgG3 -mistletoe antibodies. Remarkably, a systemic bystander effect (heterologous immunity against other recall antigens) was observed after long-term treatment. In conclusion, dose-escalation reduces the monocyte-related clinical side effects. A T-lymphocyte sensitization stimulates mainly a specific Th1 response. The most interesting clinical long-term effect is the bystander stimulation of various memory T cells that might mediate in vivo antitumor and antiinfectious T-cell response under mistletoe-extract immunization. PMID:16705487

  1. High Intracellular Concentrations of Posaconazole Do Not Impact on Functional Capacities of Human Polymorphonuclear Neutrophils and Monocyte-Derived Macrophages In Vitro.

    Science.gov (United States)

    Farowski, Fedja; Cornely, Oliver A; Hartmann, Pia

    2016-06-01

    Posaconazole is a commonly used antifungal for the prophylaxis and treatment of invasive fungal infections. We previously demonstrated that the intracellular concentration of posaconazole in peripheral blood mononuclear cells (PBMCs) and polymorphonuclear neutrophils (PMNs) was greatly increased compared to the plasma concentration. As these professional phagocytes are crucial to combat fungal infections, we set out to investigate if and how, beneficial or deleterious, this high loading of intracellular posaconazole impacts the functional capacities of these cells. Here, we show that high intracellular concentrations of posaconazole do not significantly impact PMN and monocyte-derived macrophage function in vitro In particular, killing capacity and cytoskeletal features of PMN, such as migration, are not affected, indicating that these cells serve as vehicles for posaconazole to the site of infection. Moreover, since posaconazole as such slowed the germination of Aspergillus fumigatus conidia, infected neutrophils released less reactive oxygen species (ROS). Based on these findings, we propose that the delivery of posaconazole by neutrophils to the site of Aspergillus species infection warrants control of the pathogen and preservation of tissue integrity at the same time. PMID:27021317

  2. (4-Methoxyphenyl)(3,4,5-trimethoxyphenyl)methanone inhibits tubulin polymerization, induces G2/M arrest, and triggers apoptosis in human leukemia HL-60 cells

    International Nuclear Information System (INIS)

    (4-Methoxyphenyl)(3,4,5-trimethoxyphenyl)methanone (PHT) is a known cytotoxic compound belonging to the phenstatin family. However, the exact mechanism of action of PHT-induced cell death remains to be determined. The aim of this study was to investigate the mechanisms underlying PHT-induced cytotoxicity. We found that PHT displayed potent cytotoxicity in different tumor cell lines, showing IC50 values in the nanomolar range. Cell cycle arrest in G2/M phase along with the augmented metaphase cells was found. Cells treated with PHT also showed typical hallmarks of apoptosis such as cell shrinkage, chromatin condensation, phosphatidylserine exposure, increase of the caspase 3/7 and 8 activation, loss of mitochondrial membrane potential, and internucleosomal DNA fragmentation without affecting membrane integrity. Studies conducted with isolated tubulin and docking models confirmed that PHT binds to the colchicine site and interferes in the polymerization of microtubules. These results demonstrated that PHT inhibits tubulin polymerization, arrests cancer cells in G2/M phase of the cell cycle, and induces their apoptosis, exhibiting promising anticancer therapeutic potential. - Highlights: • PHT inhibits tubulin polymerization. • PHT arrests cancer cells in G2/M phase of the cell cycle. • PHT induces caspase-dependent apoptosis

  3. (4-Methoxyphenyl)(3,4,5-trimethoxyphenyl)methanone inhibits tubulin polymerization, induces G{sub 2}/M arrest, and triggers apoptosis in human leukemia HL-60 cells

    Energy Technology Data Exchange (ETDEWEB)

    Magalhães, Hemerson I.F. [Departamento de Fisiologia e Farmacologia, Faculdade de Medicina, Universidade Federal do Ceará, Fortaleza, Ceará (Brazil); Centro de Ciências da Saúde, Departamento de Ciências Farmacêuticas, Universidade Federal da Paraíba, João Pessoa, Paraíba (Brazil); Wilke, Diego V. [Departamento de Fisiologia e Farmacologia, Faculdade de Medicina, Universidade Federal do Ceará, Fortaleza, Ceará (Brazil); Bezerra, Daniel P., E-mail: danielpbezerra@gmail.com [Centro de Pesquisa Gonçalo Moniz, Fundação Oswaldo Cruz, Salvador, Bahia (Brazil); Cavalcanti, Bruno C. [Departamento de Fisiologia e Farmacologia, Faculdade de Medicina, Universidade Federal do Ceará, Fortaleza, Ceará (Brazil); Rotta, Rodrigo; Lima, Dênis P. de; Beatriz, Adilson [Centro de Ciências Exatas e Tecnológicas (Laboratório LP4), Universidade Federal do Mato Grosso do Sul, Campo Grande, Mato Grosso do Sul (Brazil); Moraes, Manoel O.; Diniz-Filho, Jairo [Departamento de Fisiologia e Farmacologia, Faculdade de Medicina, Universidade Federal do Ceará, Fortaleza, Ceará (Brazil); Pessoa, Claudia, E-mail: c_pessoa@yahoo.com [Departamento de Fisiologia e Farmacologia, Faculdade de Medicina, Universidade Federal do Ceará, Fortaleza, Ceará (Brazil)

    2013-10-01

    (4-Methoxyphenyl)(3,4,5-trimethoxyphenyl)methanone (PHT) is a known cytotoxic compound belonging to the phenstatin family. However, the exact mechanism of action of PHT-induced cell death remains to be determined. The aim of this study was to investigate the mechanisms underlying PHT-induced cytotoxicity. We found that PHT displayed potent cytotoxicity in different tumor cell lines, showing IC{sub 50} values in the nanomolar range. Cell cycle arrest in G{sub 2}/M phase along with the augmented metaphase cells was found. Cells treated with PHT also showed typical hallmarks of apoptosis such as cell shrinkage, chromatin condensation, phosphatidylserine exposure, increase of the caspase 3/7 and 8 activation, loss of mitochondrial membrane potential, and internucleosomal DNA fragmentation without affecting membrane integrity. Studies conducted with isolated tubulin and docking models confirmed that PHT binds to the colchicine site and interferes in the polymerization of microtubules. These results demonstrated that PHT inhibits tubulin polymerization, arrests cancer cells in G{sub 2}/M phase of the cell cycle, and induces their apoptosis, exhibiting promising anticancer therapeutic potential. - Highlights: • PHT inhibits tubulin polymerization. • PHT arrests cancer cells in G{sub 2}/M phase of the cell cycle. • PHT induces caspase-dependent apoptosis.

  4. Human peripheral blood monocyte separation with Nycodenz-NaCl density osmotic pressure medium centrifugation%Nycodenz-密度渗透压介质离心分离人外周血单核细胞研究

    Institute of Scientific and Technical Information of China (English)

    尹建平; 余谨; 王先广; 陈禹潭; 马威; 赵云斌

    2011-01-01

    目的 研究人外周血单核细胞分离方法.方法 采集24名健康志愿献血者外周血,分别以EDTA-K2、肝素钠、枸橼酸钠抗凝,将抗凝全血或单个核细胞悬液缓慢加入不同密度和渗透压组合的Nycodenz-NaCI溶液上层,离心分离单核细胞;EDTA抗凝血经Ficoll-Hypaque密度梯度离心获得的单个核细胞,再分别用Nycodenz-NaCl密度渗透压介质离心法、Percoll密度梯度离心法、贴壁法、MACS(抗CD-14磁珠)分离单核细胞.结果 如上4种方法 获得的单核细胞纯度和收获率中位数(M)分别为98.9%和68.5%、89.6%和64.5%、64.8%和60.5%、94.1%和73.5%,比较显示Nycodenz法获得的单核细胞纯度最高(P<0.01);吞噬指数和趋化指数分别为188.8±11.2和26.8±5.9、187.8±12.2和26.1±5.1、144.4 ±24.8和15.4±7.3、177.1 ±18.3和18.9±6.7.统计表明Percoll法和Nycodenz法获得的单核细胞的吞噬指数、趋化指数高于贴壁法和MACS法(P<0.05);单核细胞受LPS刺激分泌细胞因子IL-12p70、IL-10、TNF-α水平(pg/ml)分别为2 545±341、1 216±397、3.999±418,2 489±425、1080±277、3 891±446,2 147±223、794±180、3268±411,35±12、142±81、407±199,比较得知MACS的单核细胞受LPS刺激分泌细胞因子IL-12p70、IL-10、TNF-α水平非常显著低于其他方法 (P<0.01);单核细胞经典亚群CD14+CD16-HLA-DR+百分比和非经典亚群CD14+CD16+HLA-DR+百分比(M)分别为89%和5%、88%和1%、73%和1%、51%和36%,统计表明MACS法的单核细胞经典亚群百分比明显低于其他分离方法 (P<0.01),而非经典亚群百分比明显高于其他方法 (P<0.01).结论 Nycodenz-NaCl密度渗透压介质离心法能够分离获得较高纯度、较多经典型的单核细胞.%Objective To study separation methods of human peripheral blood monocytes. Methods Peripheral whole blood collected from 24 healthy volunteer donors wercare each anticoagulated respectively with EDTA-K2, heparin, and citrate sodium. The a

  5. Pleiotropic effects of Blastocystis spp. Subtypes 4 and 7 on ligand-specific toll-like receptor signaling and NF-κB activation in a human monocyte cell line.

    Directory of Open Access Journals (Sweden)

    Joshua D W Teo

    Full Text Available Blastocystis spp. is a common enteric stramenopile parasite that colonizes the colon of hosts of a diverse array of species, including humans. It has been shown to compromise intestinal epithelial cell barrier integrity and mediate the production of pro-inflammatory cytokines and chemokines. Mucosal epithelial surfaces, including the intestinal epithelium, are increasingly recognized to perform a vital surveillance role in the context of innate immunity, through the expression of pathogen recognition receptors, such as Toll-like receptors (TLRs. In this study, we use the human TLR reporter monocytic cell line, THP1-Blue, which expresses all human TLRs, to investigate effects of Blastocystis on TLR activation, more specifically the activation of TLR-2, -4 and -5. We have observed that live Blastocystis spp. parasites and whole cell lysate (WCL alone do not activate TLRs in THP1-Blue. Live ST4-WR1 parasites inhibited LPS-mediated NF-κB activation in THP1-Blue. In contrast, ST7-B WCL and ST4-WR1 WCL induced pleiotropic modulation of ligand-specific TLR-2 and TLR-4 activation, with no significant effects on flagellin-mediated TLR-5 activation. Real time-qPCR analysis on SEAP reporter gene confirmed the augmenting effect of ST7-B on LPS-mediated NF-κB activation in THP1-Blue. Taken together, this is the first study to characterize interactions between Blastocystis spp. and host TLR activation using an in vitro reporter model.

  6. Induction of IL-12 Production in Human Peripheral Monocytes by Trypanosoma cruzi Is Mediated by Glycosylphosphatidylinositol-Anchored Mucin-Like Glycoproteins and Potentiated by IFN-γ and CD40-CD40L Interactions

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    Lúcia Cristina Jamli Abel

    2014-01-01

    Full Text Available Chagas disease, caused by the protozoan parasite Trypanosoma cruz