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  1. Decreased cisplatin uptake by resistant L1210 leukemia cells

    International Nuclear Information System (INIS)

    Hromas, R.A.; North, J.A.; Burns, C.P.

    1987-01-01

    Cisplatin resistance remains poorly understood compared to other forms of anti-neoplastic drug resistance. In this report radiolabelled cisplatin and rapid separation techniques were used to compare drug uptake by L1210 leukemia cells that are sensitive (K25) or resistant (SCR9) to cisplatin. Uptake of cisplatin by both cell lines was linear without saturation kinetics up to 100 μM. The resistant ZCR9 cells had 36-60% reduced drug uptake as compared to its sensitive parent line, K25. In contrast, there was no difference in the rate of efflux. We conclude that a decreased rate of uptake is one possible mechanism of cellular cisplatin resistance. (Author)

  2. Decreased stability of DNA in cells treated with alkylating agents

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    Frankfurt, O.S. (Cedars Medical Center, Miami, FL (United States))

    1990-12-01

    A modified highly sensitive procedure for the evaluation of DNA damage in individual cells treated with alkylating agents is reported. The new methodology is based on the amplification of single-strandedness in alkylated DNA by heating in the presence of Mg{sup 2+}. Human ovarian carcinoma cells A2780 were treated with nitrogen mustard (HN2), fixed in methanol, and stained with monoclonal antibody (MOAB) F7-26 generated against HN2-treated DNA. Binding of MOAB was measured by flow cytometry with indirect immunofluorescence. Intensive binding of MOAB to control and drug-treated cells was observed after heating in Tris buffer supplemented with MgCl{sub 2}. Thus, the presence of phosphates and MgCl{sub 2} during heating was necessary for the detection of HN2-induced changes in DNA stability. Fluorescence of HN2-treated cells decreased to background levels after treatment with single-strand-specific S{sub 1} nuclease. MOAB F7-26 interacted with single-stranded regions in DNA and did not bind to dsDNA or other cellular antigens. It is suggested that alkylation of guanines decreased the stability of the DNA molecule and increased the access of MOAB F7-26 to deoxycytidines on the opposite DNA strand.

  3. Glucocorticoids decrease Treg cell numbers in lungs of allergic mice.

    Science.gov (United States)

    Olsen, P C; Kitoko, J Z; Ferreira, T P; de-Azevedo, C T; Arantes, A C; Martins, Μ A

    2015-01-15

    Glucocorticoids have been the hallmark anti-inflammatory drug used to treat asthma. It has been shown that glucocorticoids ameliorate asthma by increasing numbers and activity of Tregs, in contrast recent data show that glucocorticoid might have an opposite effect on Treg cells from normal mice. Since Tregs are target cells that act on the resolution of asthma, the aim of this study was to elucidate the effect of glucocorticoid treatment on lung Tregs in mouse models of asthma. Allergen challenged mice were treated with either oral dexamethasone or nebulized budesonide. Broncoalveolar lavage and airway hyperresponsiveness were evaluated after allergenic challenge. Lung, thymic and lymph node cells were phenotyped on Treg through flow cytometry. Lung cytokine secretion was detected by ELISA. Although dexamethasone inhibited airway inflammation and hyperresponsiveness, improving resolution, we have found that both dexamethasone and budesonide induce a reduction of Treg numbers on lungs and lymphoid organs of allergen challenged mice. The reduction of lung Treg levels was independent of mice strain or type of allergen challenge. Our study also indicates that both glucocorticoids do not increase Treg activity through production of IL-10. Glucocorticoid systemic or localized treatment induced thymic atrophy. Taken together, our results demonstrate that glucocorticoids decrease Treg numbers and activity in different asthma mouse models, probably by reducing thymic production of T cells. Therefore, it is possible that glucocorticoids do not have beneficial effects on lung populations of Treg cells from asthmatic patients. Copyright © 2014 Elsevier B.V. All rights reserved.

  4. Fluid flow plate for decreased density of fuel cell assembly

    Science.gov (United States)

    Vitale, Nicholas G.

    1999-01-01

    A fluid flow plate includes first and second outward faces. Each of the outward faces has a flow channel thereon for carrying respective fluid. At least one of the fluids serves as reactant fluid for a fuel cell of a fuel cell assembly. One or more pockets are formed between the first and second outward faces for decreasing density of the fluid flow plate. A given flow channel can include one or more end sections and an intermediate section. An interposed member can be positioned between the outward faces at an interface between an intermediate section, of one of the outward faces, and an end section, of that outward face. The interposed member can serve to isolate the reactant fluid from the opposing outward face. The intermediate section(s) of flow channel(s) on an outward face are preferably formed as a folded expanse.

  5. Inhibition of Geranylgeranyl Transferase-I Decreases Cell Viability of HTLV-1-Transformed Cells

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    Cynthia A. Pise-Masison

    2011-10-01

    Full Text Available Human T-cell leukemia virus type-1 (HTLV-1 is the etiological agent of adult T-cell leukemia (ATL, an aggressive and highly chemoresistant malignancy. Rho family GTPases regulate multiple signaling pathways in tumorigenesis: cytoskeletal organization, transcription, cell cycle progression, and cell proliferation. Geranylgeranylation of Rho family GTPases is essential for cell membrane localization and activation of these proteins. It is currently unknown whether HTLV-1-transformed cells are preferentially sensitive to geranylgeranylation inhibitors, such as GGTI-298. In this report, we demonstrate that GGTI-298 decreased cell viability and induced G2/M phase accumulation of HTLV-1-transformed cells, independent of p53 reactivation. HTLV-1-LTR transcriptional activity was inhibited and Tax protein levels decreased following treatment with GGTI-298. Furthermore, GGTI-298 decreased activation of NF-κB, a downstream target of Rho family GTPases. These studies suggest that protein geranylgeranylation contributes to dysregulation of cell survival pathways in HTLV-1-transformed cells.

  6. [Thyroid C cells are decreased in experimental CDH].

    Science.gov (United States)

    Martínez, L; De Ceano-Vivas, M; González-Reyes, S; Fernández-Dumont, V; Calonge, W M; Ruiz, E; Rodríguez, J I; Tovar, J A

    2006-04-01

    Experimental CDH is often associated with malformations of neural crest origin. Several of these features are present in human CDH and therefore likely similar pathogenic mechanisms should be explored. The aim of the present study is to examine whether thyroid C-cells, another neural crest derivative, are abnormal in this rat model. Pregnant rats were exposed either to 100 mg of 2-4-dichlorophenyl-p-nitrophenyl ether (nitrofén) or vehicle (controls) on 9.5 day of gestation. Fetuses were recovered on day 21st and the thyroids of those with CDH (68%) were immuno-histochemically stained with anti-calcitonin antibody. The number of positively stained cells per high power field were counted using a computer-assisted image analysis method in at least 5 sections per thyroid. The distribution of the cells within the gland was assessed as well. Comparisons between CDH and control rats were made by non-parametric tests with a significance threshold of pCDH animals in comparison with controls (101.2 +/- 61.3 vs 23.1 +/- 37, pCDH rats with the cells tending to be located at the periphery rather than at the core of the lobes. Nitrofén induces a severe decrease in thyroid C cells accompanied by abnormal distribution patterns. These results add further evidence of the involvement of a neural crest dysregulation as a component of the pathogenesis of experimental CDH. Whether there is or not a clinical counterpart to these findings is still unknown, but the nature of the cardiovascular and craneo-facial malformations in some babies with CDH strongly support further research in this field.

  7. Differentiation of chronic lymphocytic leukemia B cells into immunoglobulin secreting cells decreases LEF-1 expression.

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    Albert Gutierrez

    Full Text Available Lymphocyte enhancer binding factor 1 (LEF-1 plays a crucial role in B lineage development and is only expressed in B cell precursors as B cell differentiation into mature B and plasma cells silences its expression. Chronic lymphocytic leukemia (CLL cells aberrantly express LEF-1 and its expression is required for cellular survival. We hypothesized that modification of the differentiation status of CLL cells would result in loss of LEF-1 expression and eliminate the survival advantage provided by its aberrant expression. In this study, we first established a methodology that induces CLL cells to differentiate into immunoglobulin (Ig secreting cells (ISC using the TLR9 agonist, CpG, together with cytokines (CpG/c. CpG/c stimulation resulted in dramatic CLL cell phenotypic and morphologic changes, expression of cytoplasmic Ig, and secretion of light chain restricted Ig. CpG/c stimulation also resulted in decreased CLL cell LEF-1 expression and increased Blimp-1 expression, which is crucial for plasma cell differentiation. Further, Wnt pathway activation and cellular survival were impaired in differentiated CLL cells compared to undifferentiated CLL cells. These data support the notion that CLL can differentiate into ISC and that this triggers decreased leukemic cell survival secondary to the down regulation of LEF-1 and decreased Wnt pathway activation.

  8. Quercetin decrease somatic cells count in mastitis of dairy cows.

    Science.gov (United States)

    Burmańczuk, Artur; Hola, Piotr; Milczak, Andrzej; Piech, Tomasz; Kowalski, Cezary; Wojciechowska, Beata; Grabowski, Tomasz

    2018-01-09

    Quercetin is a dietary flavonoid which has an effect on inflammation, angiogenesis and vascular inflammation. In several other flavonoids (e.g. kaempferol, astragalin, alpinetin, baicalein, indirubin), anti-inflammatory mechanism was proven by using mice mastitis model. The aim of the current study was pilot analysis of quercetin tolerability and its impact on somatic cells count (SCC) after multiple intramammary treatment on dairy cows with clinical mastitis. Based on SCC and clinical investigation, 9 dairy cows with clinical mastitis of one quarter were selected for the pilot study. Baseline analysis (hematology, TNFα, SCC) was performed every 24h among all cows three days before the first dose (B1-B3). After the baseline monitoring (B1-B3) eight days treatment (D1-D8) was performed with a high and low dose. Selected blood parameters were analyzed. Starting from D1 to D8, a decrease of SCC in relation to baseline was characterized by declining trend. The presented results allowed the confirmation of the significant influence of quercetin on the reduction of SCC in mastitis in dairy cows after 8days of therapy. Copyright © 2018. Published by Elsevier Ltd.

  9. Resveratrol causes cell cycle arrest, decreased collagen synthesis, and apoptosis in rat intestinal smooth muscle cells.

    Science.gov (United States)

    Garcia, Patricia; Schmiedlin-Ren, Phyllissa; Mathias, Jason S; Tang, Huaijing; Christman, Gregory M; Zimmermann, Ellen M

    2012-02-01

    One of the most difficult and treatment-resistant complications of Crohn's disease is the development of fibrotic intestinal strictures due to mesenchymal cell hyperplasia and collagen deposition. Resveratrol, a phytoalexin found in berries, peanuts, grapes, and red wine, has been shown to inhibit fibrosis in vasculature, heart, lung, kidney, liver, and esophagus in animal models. Resveratrol has also been shown to inhibit oxidation, inflammation, and cell proliferation and to decrease collagen synthesis in several cell types or animal models. The aim of this study was to determine whether resveratrol has antifibrotic effects on intestinal smooth muscle cells. Responses to resveratrol by cultured smooth muscle cells isolated from colons of untreated Lewis rats were examined; this rat strain is used in a model of Crohn's disease with prominent intestinal fibrosis. A relative decrease in cell numbers following treatment with 50 and 100 μM resveratrol was evident at 24 h (P ≤ 0.005). This effect was largely due to cell cycle arrest, with an increase in the percent of cells in S phase from 8 to 25-35% (P intestinal smooth muscle cell numbers through its effects on cell cycle arrest and apoptosis and also decreases collagen synthesis by the cells. These effects could be useful in preventing the smooth muscle cell hyperplasia and collagen deposition that characterize stricture formation in Crohn's disease.

  10. Adipose Tissue Inflammation Induces B Cell Inflammation and Decreases B Cell Function in Aging

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    Daniela Frasca

    2017-08-01

    Full Text Available Aging is the greatest risk factor for developing chronic diseases. Inflamm-aging, the age-related increase in low-grade chronic inflammation, may be a common link in age-related diseases. This review summarizes recent published data on potential cellular and molecular mechanisms of the age-related increase in inflammation, and how these contribute to decreased humoral immune responses in aged mice and humans. Briefly, we cover how aging and related inflammation decrease antibody responses in mice and humans, and how obesity contributes to the mechanisms for aging through increased inflammation. We also report data in the literature showing adipose tissue infiltration with immune cells and how these cells are recruited and contribute to local and systemic inflammation. We show that several types of immune cells infiltrate the adipose tissue and these include macrophages, neutrophils, NK cells, innate lymphoid cells, eosinophils, T cells, B1, and B2 cells. Our main focus is how the adipose tissue affects immune responses, in particular B cell responses and antibody production. The role of leptin in generating inflammation and decreased B cell responses is also discussed. We report data published by us and by other groups showing that the adipose tissue generates pro-inflammatory B cell subsets which induce pro-inflammatory T cells, promote insulin resistance, and secrete pathogenic autoimmune antibodies.

  11. Tumor necrosis factor (cachetin) decreases adipose cell differentiation in primary cell culture

    International Nuclear Information System (INIS)

    Martin, R.J.; Jones, D.D.; Jewell, D.E.; Hausman, G.J.

    1986-01-01

    Cachetin has been shown to effect gene product expression in the established adipose cell line 3T3-L1. Expression of messenger RNA for lipoprotein lipase is suppressed in cultured adipocytes. The purpose of this study was to determine the effect of Cachetin on adipose cell differentiation in primary cell culture. Stromalvascular cells obtained from the inguinal fat pad of 4-5 week old Sprague-Dawley rats were grown in culture for two weeks. During the proliferative growth phase all cells were grown on the same medium and labelled with 3 H-thymidine. Cachetin treatment (10 -6 to 10 -10 M) was initiated on day 5, the initial phase of preadipocyte differentiation. Adipocytes and stromal cells were separated using density gradient, and 3 H-thymidine was determined for both cell types. Thymidine incorporation into adipose cells was decreased maximally (∼ 50%) at 10 -10 M. Stromalvascular cells were not influenced at any of the doses tested. Adipose cell lipid content as indicated by oil red-O staining was decreased by Cachetin. Esterase staining by adipose cells treated with Cachetin was increased indicating an increase in intracellular lipase. These studies show that Cachetin has specific effects on primary adipose cell differentiation

  12. Decreased Immunity to Varicella Zoster Virus in Giant Cell Arteritis

    NARCIS (Netherlands)

    Rondaan, Christien; van der Geest, Kornelis S. M.; Eelsing, Elisabeth; Boots, Annemieke M. H.; Bos, Nicolaas A.; Westra, Johanna; Brouwer, Elisabeth

    2017-01-01

    Introduction: Herpes zoster, which can have a major impact on quality of life, results from reactivation of a latent varicella zoster virus (VZV) infection. We hypothesized that giant cell arteritis (GCA) patients are at increased risk of herpes zoster because of treatment with high-dose

  13. Arecoline decreases interleukin-6 production and induces apoptosis and cell cycle arrest in human basal cell carcinoma cells

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    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

  14. Reduction of angiocidin contributes to decreased HepG2 cell ...

    African Journals Online (AJOL)

    Angiocidin protein production in HepG2 cells were reduced significantly by siRNA. When HepG2 cells were transfected with siRNA-angiocidin, these cells showed very low proliferation activity compared with control cells. Our study suggests that reduction of angiocidin may contribute to decreased proliferation activity in liver ...

  15. Decreased natural killer cell activity is associated with atherosclerosis in elderly humans

    DEFF Research Database (Denmark)

    Bruunsgaard, H.; Pedersen, Agnes Nadelmann; Schroll, M.

    2001-01-01

    -related inflammatory diseases such as atherosclerosis. Elderly people had decreased cytotoxicity per NK cell in short-term but not in long-term assays. Ca2+ independent cytotoxicity was unaltered, and NK cells maintained their cytotoxic responses to interleukin-2 and interferon-alpha signals. The decreased...... cytotoxicity per NK cell was not completely counteracted by increased circulating numbers of NK cells in the blood. Elderly people with severe medical disorders had low numbers of circulating NK cells. Furthermore, elderly people with atherosclerosis had low cytotoxicity per NK cell and a high number...

  16. Nonspecific suppressor T cells cause decreased mixed lymphocyte culture reactivity in bone marrow transplant patients

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    Harada, M.; Ueda, M.; Nakao, S.; Kondo, K.; Odaka, K.; Shiobara, S.; Matsue, K.; Mori, T.; Matsuda, T.

    1986-07-15

    Decreased reactivity in mixed lymphocyte culture (MLC) was observed in patients within 1 yr after allogeneic and autologous bone marrow transplantation. Suppressor activity of peripheral blood mononuclear cells (PBMC) from transplant patients was studied by adding these cells as modulator cells to a bidirectional MLC with cells from normal individuals. PBMC from transplant patients markedly suppressed MLC reactivity in a dose-dependent manner. Suppressor activity was present in cells forming rosettes with sheep erythrocytes. Treatment of modulator cells with monoclonal antibodies against T cell differentiation antigens (OKT8, OKIa1) and complement completely abolished suppression of MLC. Suppressor activity was unaffected by 30 Gy irradiation. Suppressor activity declined gradually after transplantation and was inversely correlated with MLC reactivity of each patient at a significant level (p less than 0.01). These observations suggest that OKT8+ Ia+ radioresistant suppressor T cells play a role in the development of decreased MLC reactivity observed during the early post-transplant period.

  17. Hypoxia Decreases Invasin-Mediated Yersinia enterocolitica Internalization into Caco-2 Cells.

    Science.gov (United States)

    Zeitouni, Nathalie E; Dersch, Petra; Naim, Hassan Y; von Köckritz-Blickwede, Maren

    2016-01-01

    Yersinia enterocolitica is a major cause of human yersiniosis, with enterocolitis being a typical manifestation. These bacteria can cross the intestinal mucosa, and invade eukaryotic cells by binding to host β1 integrins, a process mediated by the bacterial effector protein invasin. This study examines the role of hypoxia on the internalization of Y. enterocolitica into intestinal epithelial cells, since the gastrointestinal tract has been shown to be physiologically deficient in oxygen levels (hypoxic), especially in cases of infection and inflammation. We show that hypoxic pre-incubation of Caco-2 cells resulted in significantly decreased bacterial internalization compared to cells grown under normoxia. This phenotype was absent after functionally blocking host β1 integrins as well as upon infection with an invasin-deficient Y. enterocolitica strain. Furthermore, downstream phosphorylation of the focal adhesion kinase was also reduced under hypoxia after infection. In good correlation to these data, cells grown under hypoxia showed decreased protein levels of β1 integrins at the apical cell surface whereas the total protein level of the hypoxia inducible factor (HIF-1) alpha was elevated. Furthermore, treatment of cells with the HIF-1 α stabilizer dimethyloxalylglycine (DMOG) also reduced invasion and decreased β1 integrin protein levels compared to control cells, indicating a potential role for HIF-1α in this process. These results suggest that hypoxia decreases invasin-integrin-mediated internalization of Y. enterocolitica into intestinal epithelial cells by reducing cell surface localization of host β1 integrins.

  18. Down-regulation of Rab5 decreases characteristics associated with maintenance of cell transformation

    Energy Technology Data Exchange (ETDEWEB)

    Silva, Patricio; Soto, Nicolás [Institute for Research in Dental Sciences, Faculty of Dentistry, Universidad de Chile, Santiago (Chile); Díaz, Jorge [Institute for Research in Dental Sciences, Faculty of Dentistry, Universidad de Chile, Santiago (Chile); Center for Molecular Studies of the Cell, Institute of Biomedical Sciences (ICBM), Faculty of Medicine, Universidad de Chile, Santiago (Chile); Mendoza, Pablo [Institute for Research in Dental Sciences, Faculty of Dentistry, Universidad de Chile, Santiago (Chile); Díaz, Natalia [Institute for Research in Dental Sciences, Faculty of Dentistry, Universidad de Chile, Santiago (Chile); Center for Molecular Studies of the Cell, Institute of Biomedical Sciences (ICBM), Faculty of Medicine, Universidad de Chile, Santiago (Chile); Quest, Andrew F.G. [Center for Molecular Studies of the Cell, Institute of Biomedical Sciences (ICBM), Faculty of Medicine, Universidad de Chile, Santiago (Chile); Advanced Center for Chronic Diseases (ACCDiS), Faculty of Medicine, Universidad de Chile, Santiago (Chile); Torres, Vicente A., E-mail: vatorres@med.uchile.cl [Institute for Research in Dental Sciences, Faculty of Dentistry, Universidad de Chile, Santiago (Chile); Advanced Center for Chronic Diseases (ACCDiS), Faculty of Medicine, Universidad de Chile, Santiago (Chile)

    2015-08-21

    The early endosomal protein Rab5 is highly expressed in tumor samples, although a causal relationship between Rab5 expression and cell transformation has not been established. Here, we report the functional effects of targeting endogenous Rab5 with specific shRNA sequences in different tumor cell lines. Rab5 down-regulation in B16-F10 cells decreased tumor formation by subcutaneous injection into C57/BL6 mice. Accordingly, Rab5 targeting in B16-F10 and A549, but not MDA-MB-231 cells was followed by decreased cell proliferation, increased apoptosis and decreased anchorage-independent growth. These findings suggest that Rab5 expression is required to maintain characteristics associated with cell transformation. - Highlights: • Rab5 is important to the maintenance of cell transformation characteristics. • Down-regulation of Rab5 decreases cell proliferation and increases apoptosis in different cancer cells. • Rab5 is required for anchorage-independent growth and tumorigenicity in-vivo.

  19. Down-regulation of Rab5 decreases characteristics associated with maintenance of cell transformation

    International Nuclear Information System (INIS)

    Silva, Patricio; Soto, Nicolás; Díaz, Jorge; Mendoza, Pablo; Díaz, Natalia; Quest, Andrew F.G.; Torres, Vicente A.

    2015-01-01

    The early endosomal protein Rab5 is highly expressed in tumor samples, although a causal relationship between Rab5 expression and cell transformation has not been established. Here, we report the functional effects of targeting endogenous Rab5 with specific shRNA sequences in different tumor cell lines. Rab5 down-regulation in B16-F10 cells decreased tumor formation by subcutaneous injection into C57/BL6 mice. Accordingly, Rab5 targeting in B16-F10 and A549, but not MDA-MB-231 cells was followed by decreased cell proliferation, increased apoptosis and decreased anchorage-independent growth. These findings suggest that Rab5 expression is required to maintain characteristics associated with cell transformation. - Highlights: • Rab5 is important to the maintenance of cell transformation characteristics. • Down-regulation of Rab5 decreases cell proliferation and increases apoptosis in different cancer cells. • Rab5 is required for anchorage-independent growth and tumorigenicity in-vivo

  20. Aging is associated with decreased maximal life span and accelerated senescence of bone marrow stromal cells

    DEFF Research Database (Denmark)

    Dokkedahl, Karin Stenderup; Justesen, Jeannette; Clausen, Christian

    2003-01-01

    Age-related decrease in bone formation is well described. However, the cellular causes are not known. Thus, we have established cultures of bone marrow stromal cells (MSC) from young (aged 18-29 years, n = 6) and old (aged 68-81 years, n = 5) donors. MSC were serially passaged until reaching...... donors were able to form similar amounts of mineralized matrix in vitro and of normal lamellar bone in vivo. In adipogenic medium similar numbers of adipocytes formed in cultures of young and old donors. In conclusion, aging is associated with decreased proliferative capacity of osteoprogenitor cells......, suggesting that decreased osteoblastic cell number, and not function, leads to age-related decrease in bone formation....

  1. Methamphetamine decreases dentate gyrus stem cell self-renewal and shifts the differentiation towards neuronal fate

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    Sofia Baptista

    2014-09-01

    Full Text Available Methamphetamine (METH is a highly addictive psychostimulant drug of abuse that negatively interferes with neurogenesis. In fact, we have previously shown that METH triggers stem/progenitor cell death and decreases neuronal differentiation in the dentate gyrus (DG. Still, little is known regarding its effect on DG stem cell properties. Herein, we investigate the impact of METH on mice DG stem/progenitor cell self-renewal functions. METH (10 nM decreased DG stem cell self-renewal, while 1 nM delayed cell cycle in the G0/G1-to-S phase transition and increased the number of quiescent cells (G0 phase, which correlated with a decrease in cyclin E, pEGFR and pERK1/2 protein levels. Importantly, both drug concentrations (1 or 10 nM did not induce cell death. In accordance with the impairment of self-renewal capacity, METH (10 nM decreased Sox2+/Sox2+ while increased Sox2−/Sox2− pairs of daughter cells. This effect relied on N-methyl-d-aspartate (NMDA signaling, which was prevented by the NMDA receptor antagonist, MK-801 (10 μM. Moreover, METH (10 nM increased doublecortin (DCX protein levels consistent with neuronal differentiation. In conclusion, METH alters DG stem cell properties by delaying cell cycle and decreasing self-renewal capacities, mechanisms that may contribute to DG neurogenesis impairment followed by cognitive deficits verified in METH consumers.

  2. Human mast cells decrease SLPI levels in type II – like alveolar cell model, in vitro

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    Nyström Max

    2003-08-01

    Full Text Available Abstract Background Mast cells are known to accumulate at sites of inflammation and upon activation to release their granule content, e.g. histamine, cytokines and proteases. The secretory leukocyte protease inhibitor (SLPI is produced in the respiratory mucous and plays a role in regulating the activity of the proteases. Result We have used the HMC-1 cell line as a model for human mast cells to investigate their effect on SLPI expression and its levels in cell co-culture experiments, in vitro. In comparison with controls, we found a significant reduction in SLPI levels (by 2.35-fold, p Conclusion These results indicate that SLPI-producing cells may assist mast cell migration and that the regulation of SLPI release and/or consumption by mast cells requires interaction between these cell types. Therefore, a "local relationship" between mast cells and airway epithelial cells might be an important step in the inflammatory response.

  3. Loss of Foxd3 Results in Decreased β-Cell Proliferation and Glucose Intolerance During Pregnancy

    Science.gov (United States)

    Plank, Jennifer L.; Frist, Audrey Y.; LeGrone, Alison W.; Magnuson, Mark A.

    2011-01-01

    A complete molecular understanding of β-cell mass expansion will be useful for the improvement of therapies to treat diabetic patients. During normal periods of metabolic challenges, such as pregnancy, β-cells proliferate, or self-renew, to meet the new physiological demands. The transcription factor Forkhead box D3 (Foxd3) is required for maintenance and self-renewal of several diverse progenitor cell lineages, and Foxd3 is expressed in the pancreatic primordium beginning at 10.5 d postcoitum, becoming localized predominantly to β-cells after birth. Here, we show that mice carrying a pancreas-specific deletion of Foxd3 have impaired glucose tolerance, decreased β-cell mass, decreased β-cell proliferation, and decreased β-cell size during pregnancy. In addition, several genes known to regulate proliferation, Foxm1, Skp2, Ezh2, Akt2, and Cdkn1a, are misregulated in islets isolated from these Foxd3 mutant mice. Together, these data place Foxd3 upstream of several pathways critical for β-cell mass expansion in vivo. PMID:21952247

  4. Decreased CD8+ T cell response to Epstein-Barr virus infected B cells in multiple sclerosis is not due to decreased HLA class I expression on B cells or monocytes

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    Csurhes Peter A

    2011-08-01

    Full Text Available Abstract Background Patients with multiple sclerosis (MS have a decreased frequency of CD8+ T cells reactive to their own Epstein-Barr virus (EBV infected B cells. We have proposed that this might predispose to the development of MS by allowing EBV-infected autoreactive B cells to accumulate in the central nervous system. The decreased CD8+ T cell response to EBV results from a general CD8+ T cell deficiency and also a decreased proportion of EBV-specific T cells within the total CD8+ T cell population. Because decreased HLA class I expression on monocytes and B cells has been reported in MS and could influence the generation and effector function of EBV-specific CD8+ T cells, the present study was undertaken to measure the expression of HLA molecules on B cells and monocytes in patients with MS. Methods We used flow cytometry to determine the proportions of T cells, natural killer cells, B cells and monocytes in peripheral blood mononuclear cells (PBMC and to quantify the expression of HLA molecules on T cells, B cells and monocytes of 59 healthy subjects and 62 patients with MS who had not received corticosteroids or immunomodulatory therapy in the previous 3 months. Results The levels of HLA class I and class II molecules expressed on T cells, B cells and monocytes were normal in patients with MS, with the exception of two patients with secondary progressive MS with very low class II expression on B cells. In confirmation of previous studies we also found that the percentage of CD8+ T cells was significantly decreased whereas the percentage of CD4+ T cells and the CD4:CD8 ratio were significantly increased in patients with MS compared to healthy subjects. Conclusions The decreased CD8+ T cell response to EBV-infected B cells in MS patients is not due to decreased HLA class I expression on monocytes or B cells. In a small proportion of patients decreased HLA class II expression on B cells might impair the CD8+ T cell response to EBV by

  5. Density increment and decreased survival of rat red blood cells induced by cadmium

    International Nuclear Information System (INIS)

    Kunimoto, M.; Miura, T.

    1986-01-01

    Male Wistar rats were injected with CdCl 2 subcutaneously to examine in vivo effects of Cd on density and survival of red blood cells. During the 7 days after administration of 1.0 mg Cd/kg, the following sequence of events occurred: (1) a progressive increase in the amount of more dense red blood cells concomitant with a decrease in that of light red blood cells from the first to the third day; (2) an increase in the spleen weight at the third day; (3) a decrease in the hematocrit value and an increase in the amount of light red blood cells at the fifth day; and (4) a recovery of the hematocrit value at the seventh day. Five days after administration, the hematocrit value decreased in a dose-dependent mode and the decrease was significant at the 1% level at 1.0 and 1.5 mg Cd/kg. A highly significant splenomegaly was also observed at 0.5 to 1.5 mg Cd/kg. In order to label red blood cells in vivo, [ 3 H] diisopropylfluorophosphate ([ 3 H]DFP) was injected into rats. At Day 11, Cd at either 0.5 or 1.0 mg/kg was administered to [ 3 H]DFP-prelabeled animals. Cd administration accelerated 3 H-labeled red cell clearance from the blood. Six days after Cd administration, the radioactivity of red blood cells was 76 and 68% of the control at 0.5 and 1.0 mg Cd/kg, respectively. In vitro treatment of rat red density and accelerated in vivo clearance of red blood cells from the recipient circulation. These results show that Cd at low dose can cause anemia by increasing red cell density and by accelerating red cell sequestration, presumably in the spleen

  6. Nocodazole treatment decreases expression of pluripotency markers Nanog and Oct4 in human embryonic stem cells

    DEFF Research Database (Denmark)

    Kallas, Ade; Pook, Martin; Maimets, Martti

    2011-01-01

    in the expression of transcription markers Nanog and Oct4 as well as SSEA-3 and SSEA-4 in human embryonic cells after their treatment with nocodazole. Multivariate permeabilised-cell flow cytometry was applied for characterising the expression of Nanog and Oct4 during different cell cycle phases. Among untreated h......ESC we detected Nanog-expressing cells, which also expressed Oct4, SSEA-3 and SSEA-4. We also found another population expressing SSEA-4, but without Nanog, Oct4 and SSEA-3 expression. Nocodazole treatment resulted in a decrease of cell population positive for all four markers Nanog, Oct4, SSEA-3, SSEA-4....... Nocodazole-mediated cell-cycle arrest was accompanied by higher rate of apoptosis and upregulation of p53. Twenty-four hours after the release from nocodazole block, the cell cycle of hESC normalised, but no increase in the expression of transcription markers Nanog and Oct4 was detected. In addition...

  7. Poly-lactic-glycolic-acid surface nanotopographies selectively decrease breast adenocarcinoma cell functions

    Science.gov (United States)

    Zhang, Lijuan; Webster, Thomas J.

    2012-04-01

    The ability of poly(lactic-co-glycolic acid) (PLGA, 50:50 PLG/PGA, wt%) nanotopographies to decrease lung epithelial carcinoma cell functions (including adhesion, proliferation, apoptosis and vascular endothelial growth factor (VEGF) secretion) has been previously reported. Specifically, results demonstrated decreased lung epithelial carcinoma cell VEGF synthesis on 23 nm surface-featured PLGA compared to traditional nanosmooth PLGA. However, clearly, different cell lines could have different behaviors on similar biomaterials. Thus, to investigate the universality of nanopatterned PLGA substrates to inhibit numerous cancer cell functions, here, breast epithelial adenocarcinoma cell (MCF-7) adhesion, proliferation, apoptosis and VEGF secretion were determined on different PLGA nanometer surface topographies. To isolate surface nanotopographical effects from all other surface properties, PLGA surfaces with various nanotopographies but similar chemistry and hydrophobicity were fabricated here. Atomic force microscopy (AFM) verified the varied nanotopographies on the PLGA surfaces prepared in this study. Importantly, results demonstrated for the first time significantly decreased breast adenocarcinoma cell functions (including decreased proliferation rate, increased apoptosis and decreased VEGF synthesis) on 23 nm featured PLGA surfaces compared to all other PLGA surface topographies fabricated (specifically, nanosmooth, 300 and 400 nm surface-featured PLGA surfaces). In contrast, healthy breast epithelial cells proliferated more (24%) on the 23 nm featured PLGA surfaces compared to all other PLGA samples. In summary, these results provided further insights into understanding the role PLGA surface nanotopographies can have on cancer cell functions and, more importantly, open the possibility of using polymer nanotopographies for a wide range of anticancer regenerative medicine applications (without resorting to the use of chemotherapeutics).

  8. ACh-induced hyperpolarization and decreased resistance in mammalian type II vestibular hair cells.

    Science.gov (United States)

    Poppi, Lauren A; Tabatabaee, Hessam; Drury, Hannah R; Jobling, Phillip; Callister, Robert J; Migliaccio, Americo A; Jordan, Paivi M; Holt, Joseph C; Rabbitt, Richard D; Lim, Rebecca; Brichta, Alan M

    2018-01-01

    In the mammalian vestibular periphery, electrical activation of the efferent vestibular system (EVS) has two effects on afferent activity: 1) it increases background afferent discharge and 2) decreases afferent sensitivity to rotational stimuli. Although the cellular mechanisms underlying these two contrasting afferent responses remain obscure, we postulated that the reduction in afferent sensitivity was attributed, in part, to the activation of α9- containing nicotinic acetylcholine (ACh) receptors (α9*nAChRs) and small-conductance potassium channels (SK) in vestibular type II hair cells, as demonstrated in the peripheral vestibular system of other vertebrates. To test this hypothesis, we examined the effects of the predominant EVS neurotransmitter ACh on vestibular type II hair cells from wild-type (wt) and α9-subunit nAChR knockout (α9 -/- ) mice. Immunostaining for choline acetyltransferase revealed there were no obvious gross morphological differences in the peripheral EVS innervation among any of these strains. ACh application onto wt type II hair cells, at resting potentials, produced a fast inward current followed by a slower outward current, resulting in membrane hyperpolarization and decreased membrane resistance. Hyperpolarization and decreased resistance were due to gating of SK channels. Consistent with activation of α9*nAChRs and SK channels, these ACh-sensitive currents were antagonized by the α9*nAChR blocker strychnine and SK blockers apamin and tamapin. Type II hair cells from α9 -/- mice, however, failed to respond to ACh at all. These results confirm the critical importance of α9nAChRs in efferent modulation of mammalian type II vestibular hair cells. Application of exogenous ACh reduces electrical impedance, thereby decreasing type II hair cell sensitivity. NEW & NOTEWORTHY Expression of α9 nicotinic subunit was crucial for fast cholinergic modulation of mammalian vestibular type II hair cells. These findings show a multifaceted

  9. Downregulation of integrin β4 decreases the ability of airway epithelial cells to present antigens.

    Directory of Open Access Journals (Sweden)

    Chi Liu

    Full Text Available Airway epithelial cells have been demonstrated to be accessory antigen presentation cells (APC capable of activating T cells and may play an important role in the development of allergic airway inflammation of asthma. In asthmatic airways, loss of expression of the adhesion molecule integrin β4 (ITGB4 and an increase in Th2 inflammation bias has been observed in our previous study. Given that ITGB4 is engaged in multiple signaling pathways, we studied whether disruption of ITGB4-mediated cell adhesion may contribute to the adaptive immune response of epithelial cells, including their ability to present antigens, induce the activate and differentiate of T cells. We silenced ITGB4 expression in bronchial epithelial cells with an effective siRNA vector and studied the effects of ITGB4 silencing on the antigen presentation ability of airway epithelial cells. T cell proliferation and cytokine production was investigated after co-culturing with ITGB4-silenced epithelial cells. Surface expression of B7 homologs and the major histocompatibility complex (MHC class II was also detected after ITGB4 was silenced. Our results demonstrated that silencing of ITGB4 resulted in impaired antigen presentation processes and suppressed T cell proliferation. Meanwhile, decrease in Th1 cytokine production and increase in Th17 cytokine production was induced after co-culturing with ITGB4-silenced epithelial cells. Moreover, HLA-DR was decreased and the B7 homologs expression was different after ITGB4 silencing. Overall, this study suggested that downregulation of ITGB4 expression in airway epithelial cells could impair the antigen presentation ability of these cells, which further regulate airway inflammation reaction in allergic asthma.

  10. FK506 BINDING PROTEIN 12 DEFICIENCY IN ENDOTHELIAL AND HEMATOPOIETIC CELLS DECREASES REGULATORY T CELLS AND CAUSES HYPERTENSION

    Science.gov (United States)

    Chiasson, Valorie L.; Talreja, Deepa; Young, Kristina J.; Chatterjee, Piyali; Banes-Berceli, Amy K.; Mitchell, Brett M.

    2011-01-01

    Patients treated with the immunosuppressive drug tacrolimus (FK506), which binds FK506 Binding Protein 12 (FKBP12) then inhibits the calcium-dependent phosphatase calcineurin, exhibit decreased regulatory T cells, endothelial dysfunction, and hypertension; however the mechanisms and whether altered T cell polarization play a role are unknown. Tacrolimus treatment of mice for 1 week dose-dependently decreased CD4+/FoxP3+ (regulatory T cells) and increased CD4+/IL-17+ (T helper 17) cells in the spleen, and caused endothelial dysfunction and hypertension. To determine the mechanisms, we crossed floxed FKBP12 mice with Tie2-Cre mice to generate offspring lacking FKBP12 in endothelial and hematopoietic cells only (FKBP12EC KO). Given FKBP12’s role in inhibiting TGF-β receptor activation, Tie2-Cre-mediated deletion of FKBP12 increased TGF-β receptor activation and SMAD2/3 signaling. FKBP12EC KO mice exhibited increased vascular expression of genes and proteins related to endothelial cell activation and inflammation. Serum levels of the pro-inflammatory cytokines IL-2, IL-6, IFNγ, IL-17a, IL-21, and IL-23 were increased significantly suggesting a Th17 cell-mediated inflammatory state. Flow cytometry studies confirmed this as splenocyte levels of CD4+/IL-17+ cells were increased significantly while CD4+/FoxP3+ cells were decreased in FKBP12EC KO mice. Furthermore, spleens from FKBP12EC KO mice showed increased STAT3 activation, involved in Th17 cell induction, and decreased STAT5 activation, involved in regulatory T cell induction. FKBP12EC KO mice also exhibited endothelial dysfunction and hypertension. These data suggest that tacrolimus, through its activation of TGF-β receptors in endothelial and hematopoietic cells, may cause endothelial dysfunction and hypertension by activating endothelial cells, reducing Tregs, and increasing Th17 cell polarization and inflammation. PMID:21518963

  11. Decreased accessory cell function by human monocytic cells after infection with HIV

    NARCIS (Netherlands)

    Petit, A. J.; Tersmette, M.; Terpstra, F. G.; de Goede, R. E.; van Lier, R. A.; Miedema, F.

    1988-01-01

    We studied the effect of HIV infection on the human monocytic cell line U937. The cell line was infected with cellfree HIV, strain HTLV-IIIB. After 3 wk, a high reverse transcriptase activity was continuously detected in the supernatant of the cell line. Neither cytopathic effects nor changes in

  12. Hyaluronan-CD44 Interactions Decrease the Metastatic Potential of Breast Cancer Cells

    Science.gov (United States)

    2008-06-01

    cancer cells in vivo. This study focuses on understanding the mechanisms by which CD44 inhibits breast cancer cell invasion. We have found that the differential interaction between CD44 and soluble or embedded Hyaluronan leads to differential phosphorylation of ERK and Pak. These differentially activated signaling pathways molecularly govern a phenotypic switch between invasiveness or proliferation. Additionally, this interactions also leads to decreased transcription of the metalloprotease MMP9 when on collagen IV gels but not collagen I gels. Together, these mechanisms

  13. Deferoxamine Compensates for Decreases in B Cell Counts and Reduces Mortality in Enterovirus 71-Infected Mice

    OpenAIRE

    Yang, Yajun; Ma, Jing; Xiu, Jinghui; Bai, Lin; Guan, Feifei; Zhang, Li; Liu, Jiangning; Zhang, Lianfeng

    2014-01-01

    Enterovirus 71 is one of the major causative agents of hand, foot and mouth disease in children under six years of age. No vaccine or antiviral therapy is currently available. In this work, we found that the number of B cells was reduced in enterovirus 71-infected mice. Deferoxamine, a marine microbial natural product, compensated for the decreased levels of B cells caused by enterovirus 71 infection. The neutralizing antibody titer was also improved after deferoxamine treatment. Furthermore...

  14. Simvastatin decreases steroid production in the H295R cell line and decreases steroids and FSH in female rats

    DEFF Research Database (Denmark)

    Jensen, Anna Guldvang; Hansen, Cecilie Hurup; Weisser, Johan J

    2015-01-01

    .3 (L), 5.0 (M), and 20.0 (H)mg SV/kg bw/day for 14 days. 10 Steroids were investigated in H295R growth media, and in tissues and plasma from rats using GC-MS/MS. Plasma LH and FSH were quantified by ELISA. In the H295R assay, SV and SVA particularly decreased progestagens with IC50-values from 0...

  15. Estradiol decreases iodide uptake by rat thyroid follicular FRTL-5 cells

    Directory of Open Access Journals (Sweden)

    Furlanetto T.W.

    2001-01-01

    Full Text Available Estradiol has well-known indirect effects on the thyroid. A direct effect of estradiol on thyroid follicular cells, increasing cell growth and reducing the expression of the sodium-iodide symporter gene, has been recently reported. The aim of the present investigation was to study the effect of estradiol on iodide uptake by thyroid follicular cells, using FRTL-5 cells as a model. Estradiol decreased basal iodide uptake by FRTL-5 cells from control levels of 2.490 ± 0.370 to 2.085 ± 0.364 pmol I-/µg DNA at 1 ng/ml (P<0.02, to 1.970 ± 0.302 pmol I-/µg DNA at 10 ng/ml (P<0.003, and to 2.038 ± 0.389 pmol I-/µg DNA at 100 ng/ml (P<0.02. In addition, 4 ng/ml estradiol decreased iodide uptake induced by 0.02 mIU/ml thyrotropin from 8.678 ± 0.408 to 7.312 ± 0.506 pmol I-/µg DNA (P<0.02. A decrease in iodide uptake by thyroid cells caused by estradiol has not been described previously and may have a role in goiter pathogenesis.

  16. Autophagy Alleviates Melamine-Induced Cell Death in PC12 Cells Via Decreasing ROS Level.

    Science.gov (United States)

    Wang, Hui; Gao, Na; Li, Zhigui; Yang, Zhuo; Zhang, Tao

    2016-04-01

    Since melamine was illegally added to raw milk for increasing the apparent protein content, such a scandal has not been quite blown out. Previous studies showed that melamine induced apoptosis and oxidative damage in both in vivo and in vitro experiments. It is well known that autophagy is closely related to oxidative stress. In the present study, we examined whether autophagy played an important role in protecting PC12 cells, which were damaged by melamine. Immunofluorescence assay showed that melamine enhanced the number of punctuate dot, indicating the increase of autophagosomes. Western blot assay presented that melamine significantly elevated the expression level of autophagy markers including LC3-II/LC3-I ratio, beclin-1, and Atg 7. Rapamycin further enhanced the effect, whereas 3-methyadenine (3-MA) inhibited it. MTT assay exhibited that rapamycin significantly enhanced the cell viability (P PC12 cells (P cells (P PC12 cells (P cells (P PC12 cells from melamine-induced cell death via inhibiting the excessive generation of ROS. Regulating autophagy may become a new targeted therapy to relieve the damage induced by melamine.

  17. MDMA (Ecstasy) Decreases the Number of Neurons and Stem Cells in Embryonic Cortical Cultures

    DEFF Research Database (Denmark)

    Kindlundh-Högberg, Anna M S; Pickering, Chris; Wicher, Grzegorz

    2010-01-01

    Ecstasy, 3,4-methylenedioxymetamphetamine (MDMA), is a recreational drug used among adolescents, including young pregnant women. MDMA passes the placental barrier and may therefore influence fetal development. The aim was to investigate the direct effect of MDMA on cortical cells using dissociated...... CNS cortex of rat embryos, E17. The primary culture was exposed to a single dose of MDMA and collected 5 days later. MDMA caused a dramatic, dose-dependent (100 and 400 muM) decrease in nestin-positive stem cell density, as well as a significant reduction (400 muM) in NeuN-positive cells. By q...

  18. Deferoxamine compensates for decreases in B cell counts and reduces mortality in enterovirus 71-infected mice.

    Science.gov (United States)

    Yang, Yajun; Ma, Jing; Xiu, Jinghui; Bai, Lin; Guan, Feifei; Zhang, Li; Liu, Jiangning; Zhang, Lianfeng

    2014-07-07

    Enterovirus 71 is one of the major causative agents of hand, foot and mouth disease in children under six years of age. No vaccine or antiviral therapy is currently available. In this work, we found that the number of B cells was reduced in enterovirus 71-infected mice. Deferoxamine, a marine microbial natural product, compensated for the decreased levels of B cells caused by enterovirus 71 infection. The neutralizing antibody titer was also improved after deferoxamine treatment. Furthermore, deferoxamine relieved symptoms and reduced mortality and muscle damage caused by enterovirus 71 infection. This work suggested that deferoxamine has the potential for further development as a B cell-immunomodulator against enterovirus 71.

  19. Glucose-6-phosphate dehydrogenase activity decreases during storage of leukoreduced red blood cells

    NARCIS (Netherlands)

    Peters, Anna L.; van Bruggen, Robin; de Korte, Dirk; van Noorden, Cornelis J. F.; Vlaar, Alexander P. J.

    2016-01-01

    During storage, the activity of the red blood cell (RBC) antioxidant system decreases. Glucose-6-phosphate dehydrogenase (G6PD) is essential for protection against oxidative stress by producing NADPH. G6PD function of RBC transfusion products is reported to remain stable during storage, but activity

  20. Role of BK channels in the apoptotic volume decrease in native eel intestinal cells

    DEFF Research Database (Denmark)

    Lionetto, Maria Giulia; Giordano, Maria Elena; Calisi, Antonio

    2010-01-01

    of these channels in the Apoptotic Volume Decrease (AVD) of isolated eel enterocytes, and the possible interaction between BK channels and the progression of apoptosis. The detection of apoptosis was performed by confocal microscopy and annexin V and propidium iodide labelling; cell volume changes were monitored...

  1. Age-related decrease in rod bipolar cell density of the human retina ...

    Indian Academy of Sciences (India)

    PRAKASH

    Rod bipolar cells in ageing human retina. 293. J. Biosci. ... During normal ageing, the rods (and other neurones) undergo a significant decrease in density in the human retina ..... Brain Res. 84 293–300. Figure 3. Immunohistochemical demonstration of protein kinase C-α labelling in the macula of the 91-year-old donor. The.

  2. Age-related decrease in rod bipolar cell density of the human retina ...

    Indian Academy of Sciences (India)

    PRAKASH

    Age-related decrease in rod bipolar cell density of the human retina: an immunohistochemical study. P AGGARWAL, T C NAG and S WADHWA*. Department of Anatomy, All India Institute of Medical Sciences, New Delhi 110029, India. *Corresponding author (Fax, 91-11-26588663; Email, shashiwadhwa@hotmail.com).

  3. Decreased cell proliferation and higher oxidative stress in fibroblasts from Down Syndrome fetuses. Preliminary study.

    Science.gov (United States)

    Gimeno, Amparo; García-Giménez, José Luis; Audí, Laura; Toran, Nuria; Andaluz, Pilar; Dasí, Francisco; Viña, José; Pallardó, Federico V

    2014-01-01

    Down Syndrome is the most common chromosomal disease and is also known for its decreased incidence of solid tumors and its progeroid phenotype. Cellular and systemic oxidative stress has been considered as one of the Down Syndrome phenotype causes. We correlated, in a preliminary study, the fibroblast proliferation rate and different cell proliferation key regulators, like Rcan1 and the telomere length from Down Syndrome fetuses, with their oxidative stress profile and the Ribonucleic acid and protein expression of the main antioxidant enzymes together with their activity. Increased oxidized glutathione/glutathione ratio and high peroxide production were found in our cell model. These results correlated with a distorted antioxidant shield. The messenger RNA (SOD1) and protein levels of copper/zinc superoxide dismutase were increased together with a decreased mRNA expression and protein levels of glutathione peroxidase (GPx). As a consequence the [Cu/ZnSOD/(catalase+GPx)] activity ratio increases which explains the oxidative stress generated in the cell model. In addition, the expression of thioredoxin 1 and glutaredoxin 1 is decreased. The results obtained show a decreased antioxidant phenotype that correlates with increased levels of Regulator of calcineurin 1 and attrition of telomeres, both related to oxidative stress and cell cycle impairment. Our preliminary results may explain the proneness to a progeroid phenotype. © 2013.

  4. Ubiquitin-specific protease 2 decreases p53-dependent apoptosis in cutaneous T-cell lymphoma

    Science.gov (United States)

    Wei, Tianling; Biskup, Edyta; Rahbek Gjerdrum, Lise Mette; Niazi, Omid; Ødum, Niels; Gniadecki, Robert

    2016-01-01

    Treatment of advanced cutaneous T-cell lymphomas (CTCL) is challenging because they are resistant to conventional chemotherapy. USP2 has been shown to promote resistance to chemotherapeutic agents in several cancer models. We show here USP2 is expressed in quiescent and activated T-cells and its expression is 50% lower in CTCL cell lines (MyLa2000, SeAx and Hut-78) than in normal T-cells. USP2 is expressed in neoplastic cells in early, plaque-stage mycosis fungoides (MF) and is downregulated in advanced tumor stages. Upon treatment with psoralen with UVA (PUVA) or a p53 activator, nutlin3a, USP2 expression is significantly increased in MyLa2000 (p53wt/wt), but not in SeAx (p53mut) or Hut-78 (p53−/−). USP2 knockdown decreases MyLa2000 cell viability after PUVA by 50% but not Hut-78, suggesting that the function of USP2 in CTCL cells is p53-dependent. Furthermore, USP2 knockdown results in a decreased Mdm2 expression and upregulation of p53. Taken together, our findings suggest that USP2 stabilizes Mdm2 which antagonizes pro-apoptotic activity of p53 and possibly contributes to therapeutic resistance in CTCL. PMID:27351221

  5. Eltrombopag modulates reactive oxygen species and decreases acute myeloid leukemia cell survival.

    Directory of Open Access Journals (Sweden)

    Anna Kalota

    Full Text Available Previous studies have demonstrated that the small molecule thrombopoietin (TPO mimetic, eltrombopag (E, induces apoptosis in acute myeloid leukemia (AML cells. Here, we sought to define the mechanism of the anti-leukemic effect of eltrombopag. Our studies demonstrate that, at a concentration of 5 μM E in 2% serum, E induces apoptosis in leukemia cells by triggering PARP cleavage and activation of caspase cascades within 2-6 hours. The induction of apoptotic enzymes is critically dependent on drug concentration and the concentration of serum. This effect is not associated with an alteration in mitochondrial potential but is associated with a rapid decrease in a reactive oxygen species (ROS in particular hydrogen peroxide (H2O2. Interestingly, E also decreases mitochondrial maximal and spare respiratory capacities suggesting an induced mitochondrial dysfunction that may not be readily apparent under basal conditions but becomes manifest only under stress. Co-treatment of MOLM14 AML cells with E plus Tempol or H2O2 provides a partial rescue of cell toxicity. Ferric ammonioum citrate (FAC also antagonized the E induced toxicity, by inducing notable increase in ROS level. Overall, we propose that E dramatically decreases ROS levels leading to a disruption of AML intracellular metabolism and rapid cell death.

  6. NK cell terminal differentiation: correlated stepwise decrease of NKG2A and acquisition of KIRs.

    Directory of Open Access Journals (Sweden)

    Vivien Béziat

    Full Text Available BACKGROUND: Terminal differentiation of NK cells is crucial in maintaining broad responsiveness to pathogens and discriminating normal cells from cells in distress. Although it is well established that KIRs, in conjunction with NKG2A, play a major role in the NK cell education that determines whether cells will end up competent or hyporesponsive, the events underlying the differentiation are still debated. METHODOLOGY/PRINCIPAL FINDINGS: A combination of complementary approaches to assess the kinetics of the appearance of each subset during development allowed us to obtain new insights into these terminal stages of differentiation, characterising their gene expression profiles at a pan-genomic level, their distinct surface receptor patterns and their prototypic effector functions. The present study supports the hypothesis that CD56dim cells derive from the CD56bright subset and suggests that NK cell responsiveness is determined by persistent inhibitory signals received during their education. We report here the inverse correlation of NKG2A expression with KIR expression and explore whether this correlation bestows functional competence on NK cells. We show that CD56dimNKG2A-KIR+ cells display the most differentiated phenotype associated to their unique ability to respond against HLA-E+ target cells. Importantly, after IL-12+IL-18 stimulation, reacquisition of NKG2A strongly correlates with IFN-gamma production in CD56dimNKG2A- NK cells. CONCLUSIONS/SIGNIFICANCE: Together, these findings call for the reclassification of mature human NK cells into distinct subsets and support a new model, in which the NK cell differentiation and functional fate are based on a stepwise decrease of NKG2A and acquisition of KIRs.

  7. Increased circulating follicular helper T cells with decreased programmed death-1 in chronic renal allograft rejection.

    Science.gov (United States)

    Shi, Jian; Luo, Fengbao; Shi, Qianqian; Xu, Xianlin; He, Xiaozhou; Xia, Ying

    2015-11-03

    Chronic antibody-mediated rejection is a major issue that affects long-term renal allograft survival. Since follicular helper T (Tfh) cells promote the development of antigen-specific B cells in alloimmune responses, we investigated the potential roles of Tfh cells, B cells and their alloimmune-regulating molecules in the pathogenesis of chronic renal allograft rejection in this study. The frequency of Tfh, B cells and the levels of their alloimmune-regulating molecules including chemokine receptor type 5 (CXCR5), inducible T cell co-stimulator (ICOS), programmed death-1 (PD-1), ICOSL, PDL-1 and interleukin-21 (IL-21), of peripheral blood were comparatively measured in 42 primary renal allograft recipients within 1-3 years after transplantation. Among them, 24 patients had definite chronic rejection, while other 18 patients had normal renal function. Tfh-cell ratio was significantly increased with PD-1 down-regulation in the patients with chronic renal allograft rejection, while B cells and the alloimmune-regulating molecules studied did not show any appreciable change in parallel. The patients with chronic renal allograft rejection have a characteristic increase in circulating Tfh cells with a decrease in PD-1 expression. These pathological changes may be a therapeutic target for the treatment of chronic renal allograft rejection and can be useful as a clinical index for monitoring conditions of renal transplant.

  8. Exogenous p53 upregulated modulator of apoptosis (PUMA) decreases growth of lung cancer A549 cells.

    Science.gov (United States)

    Liu, Chun-Ju; Zhang, Xia-Li; Luo, Da-Ya; Zhu, Wei-Feng; Wan, Hui-Fang; Yang, Jun-Ping; Yang, Xiao-Jun; Wan, Fu-Sheng

    2015-01-01

    To investigate the influence of exogenous p53 upregulated modulator of apoptosis (PUMA) expression on cell proliferation and apoptosis in human non-small cell lung cancer A549 cells and transplanted tumor cell growth in nude mice. A549 cells were divided into the following groups: control, non- carrier (NC), PUMA (transfected with pCEP4- (HA) 2-PUMA plasmid), DDP (10 μg/mL cisplatin treatment) and PUMA+DDP (transfected with pCEP4-(HA)2-PUMA plasmid and 10 μg/mL cisplatin treatment). The MTT method was used to detect the cell survival rate. Cell apoptosis rates were measured by flow cytometry, and PUMA, Bax and Bcl-2 protein expression levels were measured by Western blotting. Compared to the control group, the PUMA, DDP and PUMA+DDP groups all had significantly decreased A549 cell proliferation (pPUMA+DDP group. Conversely, the apoptosis rates of the three groups were significantly increased (PPUMA and DDP treatments were synergistic. Moreover, Bax protein levels significantly increased (pPUMA+DDP group was significantly higher than in the single DDP or PUMA groups. Exogenous PUMA effectively inhibited lung cancer A549 cell proliferation and transplanted tumor growth by increasing Bax protein levels and reducing Bcl-2 protein levels.

  9. Upregulating Nonneuronal Cholinergic Activity Decreases TNF Release from Lipopolysaccharide-Stimulated RAW264.7 Cells

    Directory of Open Access Journals (Sweden)

    Yi Lv

    2014-01-01

    Full Text Available Nonneuronal cholinergic system plays a primary role in maintaining homeostasis. It has been proved that endogenous neuronal acetylcholine (ACh could play an anti-inflammatory role, and exogenous cholinergic agonists could weaken macrophages inflammatory response to lipopolysaccharide (LPS stimulation through activation of α7 subunit-containing nicotinic acetylcholine receptor (α7nAChR. We assumed that nonneuronal cholinergic system existing in macrophages could modulate inflammation through autocrine ACh and expressed α7nAChR on the cells. Therefore, we explored whether LPS continuous stimulation could upregulate the nonneuronal cholinergic activity in macrophages and whether increasing autocrine ACh could decrease TNF release from the macrophages. The results showed that, in RAW264.7 cells incubated with LPS for 20 hours, the secretion of ACh was significantly decreased at 4 h and then gradually increased, accompanied with the enhancement of α7nAChR expression level. The release of TNF was greatly increased from RAW264.7 cells at 4 h and 8 h exposure to LPS; however, it was suppressed at 20 h. Upregulating choline acetyltransferase (ChAT expression through ChAT gene transfection could enhance ACh secretion and reduce TNF release from the infected RAW264. 7cells. The results indicated that LPS stimulation could modulate the activity of nonneuronal cholinergic system of RAW264.7 cells. Enhancing autocrine ACh production could attenuate TNF release from RAW264.7 cells.

  10. Professional killer cell deficiencies and decreased survival in pulmonary arterial hypertension.

    Science.gov (United States)

    Edwards, Adrienne L; Gunningham, Sarah P; Clare, Geoffrey C; Hayman, Matthew W; Smith, Mark; Frampton, Christopher M A; Robinson, Bridget A; Troughton, Richard W; Beckert, Lutz E L

    2013-11-01

    Increasing evidence implicates lymphocytes in pulmonary arterial hypertension (PAH) pathogenesis. Rats deficient in T-lymphocytes show increased propensity to develop PAH but when injected with endothelial progenitor cells are protected from PAH (a mechanism dependent on natural killer (NK) cells). A decreased quantity of circulating cytotoxic CD8+ T-lymphocytes and NK cells are now reported in PAH patients; however, the effect of lymphocyte depletion on disease outcome is unknown. This prospective study analysed the lymphocyte profile and plasma brain natriuretic peptide (BNP) levels of patients with idiopathic PAH (IPAH), connective tissue disease-associated PAH (CTD-APAH) and matched healthy controls. Lymphocyte surface markers studied include: CD4+ (helper T-cell marker), CD8+ (cytotoxic T-cell marker), CD56/CD16 (NK cell marker) and CD19+ (mature B-cell marker). Lymphocyte deficiencies and plasma BNP levels were then correlated with clinical outcome. Fourteen patients with PAH (9 IPAH, 5CTD) were recruited. Three patients were deceased at 1-year follow-up; all had elevated CD4 : CD8 ratios and deficiencies of NK cells and cytotoxic CD8+ T-lymphocytes at recruitment. Patients with normal lymphocyte profiles at recruitment were all alive a year later, and none were on the active transplant list. As univariate markers, cytotoxic CD8+ T-cell and NK cell counts were linked to short-term survival. Deficiencies in NK cells and cytotoxic CD8+ T-cells may be associated with an increased risk of death in PAH patients. Further research is required in larger numbers of patients and to elucidate the mechanism of these findings. © 2013 The Authors. Respirology © 2013 Asian Pacific Society of Respirology.

  11. SBDS-deficiency results in deregulation of reactive oxygen species leading to increased cell death and decreased cell growth.

    Science.gov (United States)

    Ambekar, Chhaya; Das, Bikul; Yeger, Herman; Dror, Yigal

    2010-12-01

    Shwachman-Diamond syndrome (SDS) is characterized by reduced hematopoietic and exocrine pancreatic cell numbers and a marked propensity for leukemia. Most patients have mutations in the SBDS gene. We previously reported that SBDS-deficient cells overexpress Fas, undergo accelerated spontaneous and Fas-mediated apoptosis and grow slowly. However the mechanism of how SBDS regulates apoptosis remains unknown. Several studies have shown that reactive oxygen species (ROS) regulate cell growth and spontaneous and Fas-mediated cell death. Therefore, we hypothesized that SBDS-deficiency disrupts ROS regulation and subsequently increases sensitivity to Fas stimulation and reduced cell growth. SBDS was knocked down in HeLa cervical cancer cells and TF-1 myeloid cells using short hairpin RNA. ROS levels were evaluated by oxidation of 2',7'-dichlorodihydrofluorescein diacetate. Apoptosis and cell growth were evaluated with and without antioxidants by annexin V/propidium iodide and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assays, respectively. We found that shRNA mediated SBDS-knockdown resulted in a significant increase in ROS levels compared to control cells. Fas stimulation further increased ROS levels in the SBDS-knockdown HeLa cells more than in the controls. Importantly, balancing ROS levels by antioxidants rescued SBDS-deficient cells from spontaneous and Fas-mediated apoptosis and reduced cell growth. ROS levels are increased in SBDS-deficient cells, which leads to increased apoptosis and decreased cell growth. Increased baseline and Fas-mediated ROS levels in SBDS-deficient cells can enhance the sensitivity to Fas stimulation. By balancing ROS levels, antioxidants can improve cell growth and survival in SBDS-deficient cells.

  12. Hypoxia-activated prodrug TH-302 decreased survival rate of canine lymphoma cells under hypoxic condition.

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    Yamazaki, Hiroki; Lai, Yu-Chang; Tateno, Morihiro; Setoguchi, Asuka; Goto-Koshino, Yuko; Endo, Yasuyuki; Nakaichi, Munekazu; Tsujimoto, Hajime; Miura, Naoki

    2017-01-01

    We tested the hypotheses that hypoxic stimulation enhances growth potentials of canine lymphoma cells by activating hypoxia-inducible factor 1α (HIF-1α), and that the hypoxia-activated prodrug (TH-302) inhibits growth potentials in the cells. We investigated how hypoxic culture affects the growth rate, chemoresistance, and invasiveness of canine lymphoma cells and doxorubicin (DOX)-resistant lymphoma cells, and influences of TH-302 on survival rate of the cells under hypoxic conditions. Our results demonstrated that hypoxic culture upregulated the expression of HIF-1α and its target genes, including ATP-binding cassette transporter B1 (ABCB1), ATP-binding cassette transporter G2 (ABCG2), platelet-derived growth factor (PDGF), vascular endothelial growth factor (VEGF), and survivin, and enhanced the growth rate, DOX resistance, and invasiveness of the cells. Additionally, TH-302 decreased the survival rate of the cells under hypoxic condition. Our studies suggest that hypoxic stimulation may advance the tumorigenicity of canine lymphoma cells, favoring malignant transformation. Therefore, the data presented may contribute to the development of TH-302-based hypoxia-targeting therapies for canine lymphoma.

  13. Cadmium affects osmotic phase and regulatory volume decrease in cultured human embryonic kidney cells

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    Rossana Morabito

    2016-06-01

    Full Text Available The present investigation aims to verify whether cadmium (Cd2+, a metal possibly accumulated in body tissues from air and food, affects cell volume regulation capability in cultured human embryonic kidney (HEK 293 Phoenix cells. The osmotic phase (OP, which is the expected cell swelling due to aquaporins involvement after hyposmotic challenge, and regulatory volume decrease (RVD, bringing cell volume back to control values through Ca2+-dependent ion efflux (K+ and Cl–, have been monitored in HEK 293 cells treated with Cd2+ (1-10-100 μM for different time intervals (30 min, 3 h, overnight and then submitted to 15 % hyposmotic shock. The results show that both 1 and 10 μM Cd2+ significantly reduced OP, whereas 100 μM impaired Cd2+ RVD mechanisms. The use of glutathione (GSH, 200 μM confirmed that Cd2+ elicited its effect via oxidative damage, being RVD inhibition after Cd2+ treatment prevented by this antioxidant compound. Our findings show that: i HEK 293 cells are a suitable model to assay the effect of xenobiotics on cell homeostasis; ii Cd2+, depending on its concentration, affects cell homeostasis at different levels, i.e. water and ion permeability, responsible for, respectively, OP and RVD mechanism, adding thus more information to the knowledge of Cd2+ toxicology.

  14. Inhibition of myeloperoxidase decreases vascular oxidative stress and increases vasodilatation in sickle cell disease mice.

    Science.gov (United States)

    Zhang, Hao; Xu, Hao; Weihrauch, Dorothee; Jones, Deron W; Jing, Xigang; Shi, Yang; Gourlay, David; Oldham, Keith T; Hillery, Cheryl A; Pritchard, Kirkwood A

    2013-11-01

    Activated leukocytes and polymorphonuclear neutrophils (PMN) release myeloperoxidase (MPO), which binds to endothelial cells (EC), is translocated, and generates oxidants that scavenge nitric oxide (NO) and impair EC function. To determine whether MPO impairs EC function in sickle cell disease (SCD), control (AA) and SCD mice were treated with N-acetyl-lysyltyrosylcysteine-amide (KYC). SCD humans and mice have high plasma MPO and soluble L-selectin (sL-selectin). KYC had no effect on MPO but decreased plasma sL-selectin and malondialdehyde in SCD mice. MPO and 3-chlorotyrosine (3-ClTyr) were increased in SCD aortas. KYC decreased MPO and 3-ClTyr in SCD aortas to the levels in AA aortas. Vasodilatation in SCD mice was impaired. KYC increased vasodilatation in SCD mice more than 2-fold, to ∼60% of levels in AA mice. KYC inhibited MPO-dependent 3-ClTyr formation in EC proteins. SCD mice had high plasma alanine transaminase (ALT), which tended to decrease in KYC-treated SCD mice (P = 0.07). KYC increased MPO and XO/XDH and decreased 3-ClTyr and 3-nitrotyrosine (3-NO₂Tyr) in SCD livers. These data support the hypothesis that SCD increases release of MPO, which generates oxidants that impair EC function and injure livers. Inhibiting MPO is an effective strategy for decreasing oxidative stress and liver injury and restoring EC function in SCD.

  15. Gestational lead exposure selectively decreases retinal dopamine amacrine cells and dopamine content in adult mice

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    Fox, Donald A., E-mail: dafox@uh.edu [College of Optometry, University of Houston, Houston, TX (United States); Department of Biology and Biochemistry, University of Houston, Houston, TX (United States); Department of Pharmacology and Pharmaceutical Sciences, University of Houston, Houston, TX (United States); Hamilton, W. Ryan [Department of Biology and Biochemistry, University of Houston, Houston, TX (United States); Johnson, Jerry E. [Department of Natural Sciences, University of Houston-Downtown, Houston, TX (United States); Xiao, Weimin [College of Optometry, University of Houston, Houston, TX (United States); Chaney, Shawntay; Mukherjee, Shradha [Department of Biology and Biochemistry, University of Houston, Houston, TX (United States); Miller, Diane B.; O' Callaghan, James P. [Toxicology and Molecular Biology Branch, Health Effects Research Laboratory, Centers for Disease Control and Prevention-NIOSH, Morgantown, WV USA (United States)

    2011-11-15

    Gestational lead exposure (GLE) produces supernormal scotopic electroretinograms (ERG) in children, monkeys and rats, and a novel retinal phenotype characterized by an increased number of rod photoreceptors and bipolar cells in adult mice and rats. Since the loss of dopaminergic amacrine cells (DA ACs) in GLE monkeys and rats contributes to supernormal ERGs, the retinal DA system was analyzed in mice following GLE. C57BL/6 female mice were exposed to low (27 ppm), moderate (55 ppm) or high (109 ppm) lead throughout gestation and until postnatal day 10 (PN10). Blood [Pb] in control, low-, moderate- and high-dose GLE was {<=} 1, {<=} 10, {approx} 25 and {approx} 40 {mu}g/dL, respectively, on PN10 and by PN30 all were {<=} 1 {mu}g/dL. At PN60, confocal-stereology studies used vertical sections and wholemounts to characterize tyrosine hydroxylase (TH) expression and the number of DA and other ACs. GLE dose-dependently and selectively decreased the number of TH-immunoreactive (IR) DA ACs and their synaptic plexus without affecting GABAergic, glycinergic or cholinergic ACs. Immunoblots and confocal revealed dose-dependent decreases in retinal TH protein expression and content, although monoamine oxidase-A protein and gene expression were unchanged. High-pressure liquid chromatography showed that GLE dose-dependently decreased retinal DA content, its metabolites and DA utilization/release. The mechanism of DA selective vulnerability is unknown. However, a GLE-induced loss/dysfunction of DA ACs during development could increase the number of rods and bipolar cells since DA helps regulate neuronal proliferation, whereas during adulthood it could produce ERG supernormality as well as altered circadian rhythms, dark/light adaptation and spatial contrast sensitivity. -- Highlights: Black-Right-Pointing-Pointer Peak [BPb] in control, low-, moderate- and high-dose newborn mice with gestational lead exposure: {<=} 1, {<=} 10, 25 and 40 {mu}g/dL Black

  16. Metformin decreases glucose oxidation and increases the dependency of prostate cancer cells on reductive glutamine metabolism.

    Science.gov (United States)

    Fendt, Sarah-Maria; Bell, Eric L; Keibler, Mark A; Davidson, Shawn M; Wirth, Gregory J; Fiske, Brian; Mayers, Jared R; Schwab, Matthias; Bellinger, Gary; Csibi, Alfredo; Patnaik, Akash; Blouin, Marie Jose; Cantley, Lewis C; Guarente, Leonard; Blenis, John; Pollak, Michael N; Olumi, Aria F; Vander Heiden, Matthew G; Stephanopoulos, Gregory

    2013-07-15

    Metformin inhibits cancer cell proliferation, and epidemiology studies suggest an association with increased survival in patients with cancer taking metformin; however, the mechanism by which metformin improves cancer outcomes remains controversial. To explore how metformin might directly affect cancer cells, we analyzed how metformin altered the metabolism of prostate cancer cells and tumors. We found that metformin decreased glucose oxidation and increased dependency on reductive glutamine metabolism in both cancer cell lines and in a mouse model of prostate cancer. Inhibition of glutamine anaplerosis in the presence of metformin further attenuated proliferation, whereas increasing glutamine metabolism rescued the proliferative defect induced by metformin. These data suggest that interfering with glutamine may synergize with metformin to improve outcomes in patients with prostate cancer. ©2013 AACR.

  17. PUMA decreases the growth of prostate cancer PC-3 cells independent of p53.

    Science.gov (United States)

    Shan, Zhengfei; Liu, Qingzuo; Li, Yuling; Wu, Jitao; Sun, Dekang; Gao, Zhenli

    2017-03-01

    PUMA (p53 upregulated modulator of apoptosis), a member of the B-cell lymphoma 2 (Bcl-2) protein family, is a pro-apoptotic protein. PUMA expression is modulated by the tumor suppressor p53. PUMA has a role in rapid cell death via p53-dependent and -independent mechanisms. To evaluate whether p53 is required for PUMA-mediated apoptosis in prostate cancer cells, p53 protein was silenced in human prostate cancer PC-3 cells by using p53 small interfering RNA (siRNA). The interference efficiency of p53 on RNA and protein levels was detected by reverse transcription-quantitative polymerase chain reaction and western blotting. Cell proliferation and p21 expression were subsequently examined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and western blot analysis, respectively. p53-silenced or control PC-3 cells were transfected with pCEP4-(hemagglutinin)-PUMA plasmid, or non-carrier plasmid. Enzyme-linked immunosorbent assay was used to determine cell apoptosis by measuring histone release and caspase-3 activation, and MTT assay was used to measure cell viability. In addition, the expression of pro-apoptosis protein Bax and anti-apoptosis protein Bcl-2 were evaluated. The results of the present study revealed that p53 siRNA significantly suppressed p53 RNA and protein expression in PC-3 cells. Deficiency of p53 increased the cell growth rate and decreased p21 expression. However, PUMA overexpression remained able to induce apoptosis in p53-silenced and control cells by increasing Bax expression and decreasing Bcl-2 expression, leading to the activation of caspase-3. These results suggest that PUMA may mediate apoptosis of prostate cancer PC-3 cells, potentially independently of p53. Furthermore, PUMA gene treatment to induce cancer cell apoptosis may be more efficient compared with p53-dependent apoptosis, where loss of p53 expression or function may lead to limited efficacy of PUMA expression. Therefore, the present study proposes the

  18. Combination of azacitidine and trichostatin A decreased the tumorigenic potential of lung cancer cells

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    Yang Y

    2017-06-01

    Full Text Available Yang Yang,1,* Wei Yin,2,* Fengying Wu,3,* Jiang Fan1 1Department of Thoracic Surgery, Shanghai Pulmonary Hospital, Tongji University, Shanghai, China; 2Key Laboratory of Oral Biomedical Engineering of Ministry of Education, Hospital of Stomatology, School of Stomatology, Wuhan University, Wuhan, China; 3Oncology Department, Shanghai Pulmonary Hospital, Tongji University, Shanghai, China *These authors contributed equally to this work Purpose: This study aims to investigate the possibility of using epigenetic inhibitors against lung cancer. Methods: The changes in the proliferation of human lung cancer cells, NCI-H1975 and NCI-H1299 cells, treated with various doses of inhibitors of DNA methyltransferase (azacitidine [5-AZA] or histone deacetylase inhibitors (trichostatin A [TSA] were determined by cell counting. The cell viability of NCI-H1975 and NCI-H1299 cells treated with 5-AZA and/or TSA was measured by the MTT assay. The changes in expression of the AKT signaling pathway molecules caused by the application of 5-AZA and TSA were analyzed through their protein and mRNA levels. A xenograft model was used to observe the effects of 5-AZA and TSA on tumor growth in vivo. Results: 5-AZA and TSA inhibited the proliferation and viability of NCI-H1975 and NCI-H1299 cells. Their joint application significantly influenced the expression of key molecules in AKT signaling pathway in vitro, and inhibited the growth of xenograft tumors in vivo. Furthermore, TSA and 5-AZA decreased the tumorigenic ability of NCI-H1975 cells in vivo. Conclusion: The decreased cell viability and tumorigenic ability, as well as increased anti-oncogene expression following the joint application of 5-AZA and TSA, make these epigenetic inhibitors prospective therapeutic agents for lung cancer. Keywords: lung cancer, epigenetic inhibitor, azacitidine (5-AZA, trichostatin A

  19. L-carnitine significantly decreased aging of rat adipose tissue-derived mesenchymal stem cells.

    Science.gov (United States)

    Mobarak, Halimeh; Fathi, Ezzatollah; Farahzadi, Raheleh; Zarghami, Nosratollah; Javanmardi, Sara

    2017-03-01

    Mesenchymal stem cells are undifferentiated cells that have the ability to divide continuously and tissue regeneration potential during the transplantation. Aging and loss of cell survival, is one of the main problems in cell therapy. Since the production of free radicals in the aging process is effective, the use of antioxidant compounds can help in scavenging free radicals and prevent the aging of cells. The aim of this study is evaluate the effects of L-carnitine (LC) on proliferation and aging of rat adipose tissue-derived mesenchymal stem cells (rADSC). rADSCs were isolated from inguinal region of 5 male Rattus rats. Oil red-O, alizarin red-S and toluidine blue staining were performed to evaluate the adipogenic, osteogenic and chondrogenic differentiation of rADSCs, respectively. Flow cytometric analysis was done for investigating the cell surface markers. The methyl thiazol tetrazolium (MTT) method was used to determine the cell proliferation of rADSCs following exposure to different concentrations of LC. rADSCs aging was evaluated by beta-galactosidase staining. The results showed significant proliferation of rADSCs 48 h after treatment with concentrations of 0.2 mM LC. In addition, in the presence of 0.2 mM LC, rADSCs appeared to be growing faster than control group and 0.2 mM LC supplementation could significantly decrease the population doubling time and aging of rADSCs. It seems that LC would be a good antioxidant to improve lifespan of rADSCs due to the decrease in aging.

  20. Cell Volume Regulation and Apoptotic Volume Decrease in Rat Distal Colon Superficial Enterocytes

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    Stefania Antico

    2013-12-01

    Full Text Available Background: The colon epithelium is physiologically exposed to osmotic stress, and the activation of cell volume regulation mechanisms is essential in colonocyte physiology. Moreover, colon is characterized by a high apoptotic rate of mature cells balancing the high division rate of stem cells. Aim: The aim of the present work was to investigate the main cell volume regulation mechanisms in rat colon surface colonocytes and their role in apoptosis. Methods: Cell volume changes were measured by light microscopy and video imaging on colon explants; apoptosis sign appearance was monitored by confocal microscopy on annexin V/propidium iodide labeled explants. Results: Superficial colonocytes showed a dynamic regulation of their cell volume during anisosmotic conditions with a Regulatory Volume Increase (RVI response following hypertonic shrinkage and Regulatory Volume Decrease (RVD response following hypotonic swelling. RVI was completely inhibited by bumetanide, while RVD was completely abolished by high K+ or iberiotoxin treatment and by extracellular Ca2+ removal. DIDS incubation was also able to affect the RVD response. When colon explants were exposed to H2O2 as apoptotic inducer, colonocytes underwent an isotonic volume decrease ascribable to Apoptotic Volume Decrease (AVD within about four hours of exposure. AVD was shown to precede annexin V positivity. It was also inhibited by high K+ or iberiotoxin treatment. Interestingly, treatment with iberiotoxin significantly inhibited apoptosis progression. Conclusions: In rat superficial colonocytes K+ efflux through high conductance Ca2+-activated K+ channels (BK channels was demonstrated to be the main mechanism of RVD and to plays also a crucial role in the AVD process and in the progression of apoptosis.

  1. Bee Venom Decreases LPS-Induced Inflammatory Responses in Bovine Mammary Epithelial Cells.

    Science.gov (United States)

    Jeong, Chang Hee; Cheng, Wei Nee; Bae, Hyojin; Lee, Kyung Woo; Han, Sang Mi; Petriello, Michael C; Lee, Hong Gu; Seo, Han Geuk; Han, Sung Gu

    2017-10-28

    The world dairy industry has long been challenged by bovine mastitis, an inflammatory disease, which causes economic loss due to decreased milk production and quality. Attempts have been made to prevent or treat this disease with multiple approaches, primarily through increased abuse of antibiotics, but effective natural solutions remain elusive. Bee venom (BV) contains a variety of peptides ( e.g. , melittin) and shows multiple bioactivities, including prevention of inflammation. Thus, in the current study, it was hypothesized that BV can reduce inflammation in bovine mammary epithelial cells (MAC-T). To examine the hypothesis, cells were treated with LPS (1 μg/ml) to induce an inflammatory response and the anti-inflammatory effects of BV (2.5 and 5 μg/ml) were investigated. The cellular mechanisms of BV against LPS-induced inflammation were also investigated. Results showed that BV can attenuate expression of an inflammatory protein, COX2, and pro-inflammatory cytokines such as IL-6 and TNF-α. Activation of NF-κB, an inflammatory transcription factor, was significantly downregulated by BV in cells treated with LPS, through dephosphorylation of ERK1/2. Moreover, pretreatment of cells with BV attenuated LPS-induced production of intracellular reactive oxygen species ( e.g. , superoxide anion). These results support our hypothesis that BV can decrease LPS-induced inflammatory responses in bovine mammary epithelial cells through inhibition of oxidative stress, NF-κB, ERK1/2, and COX-2 signaling.

  2. Decreasing lncRNA HOTAIR expression inhibits human colorectal cancer stem cells.

    Science.gov (United States)

    Dou, Jun; Ni, Yaoyao; He, Xiangfeng; Wu, Di; Li, Miao; Wu, Songyan; Zhang, Rong; Guo, Mei; Zhao, Fengsu

    2016-01-01

    Research on the relationship between aberrant long non-coding RNA (lncRNA) and cancer stem cell (CSC) biology in cancer patients has been recently gaining attention. The goal of this study was to investigate whether the decreasing lncRNA HOTAIR expression would inhibit human colorectal cancer (CRC) stem cells. CD133(+)CSCs were isolated from human CRC LoVo cell line by using a magnetic-activated cell sorting system, and were transfected with the expression vector-based small hairpin RNA targeting HOTAIR (shHOTAIR). The ability of cellular proliferation, migration, invasion, colony-forming, and the epithelial-mesenchymal transition (EMT)-associated molecule expression as well as the tumorigenicity of CD133(+)-shHOTAIR were evaluated by the MTT, wound-healing, cellular invasion, colony formation and Western blot assays, respectively. This study found that, when compared with control cells in vitro, CD133(+)-shHOTAIR exhibited the decreased HOTAIR expression, suppressed cellular proliferation, migration, invasion, colony-forming, and inhibited the Vimentin expression with increased E-cadherin expression. In particular, the down-regulation of the HOTAIR expression in CD133(+)CSCs markedly attenuated the tumor growth and lung metastasis in xenograft nude mice. Taken together, this study found that down-regulating the HOTAIR expression in CD133(+)CSCs could serve as a potential anti-cancer regimen to inhibit the invasiveness and metastasis of CRC CSCs.

  3. Thyroid Autoimmunity is Associated with Decreased Cytotoxicity T Cells in Women with Repeated Implantation Failure

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    Chunyu Huang

    2015-08-01

    Full Text Available Thyroid autoimmunity (TAI, which is defined as the presence of autoantibodies against thyroid peroxidase (TPO and/or thyroglobulin (TG, is related to repeated implantation failure (RIF. It is reported that TAI was involved in reproductive failure not only through leading thyroid function abnormality, but it can also be accompanied with immune imbalance. Therefore, this study was designed to investigate the association of thyroid function, immune status and TAI in women with RIF. Blood samples were drawn from 72 women with RIF to evaluate the prevalence of TAI, the thyroid function, the absolute numbers and percentages of lymphocytes. The prevalence of thyroid function abnormality in RIF women with TAI was not significantly different from that in RIF women without TAI (c2 = 0.484, p > 0.05. The absolute number and percentage of T cells, T helper (Th cells, B cells and natural killer (NK cells were not significantly different in RIF women with TAI compared to those without TAI (all p > 0.05. The percentage of T cytotoxicity (Tc cells was significantly decreased in RIF women with TAI compared to those without TAI (p < 0.05. Meanwhile, Th/Tc ratio was significantly increased (p < 0.05. These results indicated that the decreased Tc percentage and increased Th/Tc ratio may be another influential factor of adverse pregnancy outcomes in RIF women with TAI.

  4. Decreased Sp1 Expression Mediates Downregulation of SHIP2 in Gastric Cancer Cells

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    Yan Ye

    2017-01-01

    Full Text Available Past studies have shown that the Src homology 2-containing inositol 5-phosphatase 2 (SHIP2 is commonly downregulated in gastric cancer, which contributes to elevated activation of PI3K/Akt signaling, proliferation and tumorigenesis of gastric cancer cells. However, the mechanisms underlying the reduced expression of SHIP2 in gastric cancer remain unclear. While gene copy number variation analysis and exon sequencing indicated the absence of genomic alterations of SHIP2, bisulfite genomic sequencing (BGS showed promoter hypomethylation of SHIP2 in gastric cancer cells. Analysis of transcriptional activity of SHIP2 promoter revealed Specificity protein 1 (Sp1 was responsible for the regulation of SHIP2 expression in gastric cancer cells. Furthermore, Sp1 expression, but not Sp3, was frequently downregulated in gastric cancer compared with normal gastric mucosa, which was associated with a paralleled reduction in SHIP2 levels in gastric cancer. Moreover, overexpression of Sp1 inhibited cell proliferation, induced apoptosis, suppressed cell motility and invasion in gastric cancer cells in vitro, which was, at least in part, due to transcriptional activation of SHIP2 mediated by Sp1, thereby inactivating Akt. Collectively, these results indicate that decreased expression of transcription factor Sp1 contributes to suppression of SHIP2 in gastric cancer cells.

  5. Bone marrow mesenchymal stem cells decrease CHOP expression and neuronal apoptosis after spinal cord injury.

    Science.gov (United States)

    Gu, Chuanlong; Li, Heyangzi; Wang, Chao; Song, Xinghui; Ding, Yuemin; Zheng, Mingzhi; Liu, Wei; Chen, Yingying; Zhang, Xiaoming; Wang, Linlin

    2017-01-01

    Spinal cord injury (SCI) leads to irreversible neuronal loss and ultimately leads to paralysis. Bone marrow derived mesenchymal stem cells (BMSCs) have been demonstrated to be an effective approach to treat SCI. The present study was designed to investigate the role of BMSCs in rats with spinal cord injury and in oxygen-glucose deprivation (OGD) treated motor neurons. The results demonstrated that BMSCs could improve locomotor function and decrease expression of pro-apoptotic transcription factor C/EBP homologous protein (CHOP) and apoptosis after SCI. Furthermore, co-culture with BMSCs or conditioned medium from BMSCs could also decrease the expression of CHOP and apoptosis in post-OGD motor neurons, supporting that BMSCs exerts protective effects by decreasing the expression of CHOP in injured motor neurons. Our findings provide a potential novel mechanism for BMSCs treatments in patients with SCI. Copyright © 2016. Published by Elsevier Ireland Ltd.

  6. Knockdown of XBP1 by RNAi in Mouse Granulosa Cells Promotes Apoptosis, Inhibits Cell Cycle, and Decreases Estradiol Synthesis

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    Nan Wang

    2017-05-01

    Full Text Available Granulosa cells are crucial for follicular growth, development, and follicular atresia. X-box binding protein 1 (XBP1, a basic region-leucine zipper protein, is widely involved in cell differentiation, proliferation, apoptosis, cellular stress response, and other signaling pathways. In this study, RNA interference, flow cytometry, western blot, real-time PCR, Cell Counting Kit (CCK8, and ELISA were used to investigate the effect of XBP1 on steroidogenesis, apoptosis, cell cycle, and proliferation of mouse granulosa cells. ELISA analysis showed that XBP1 depletion significantly decreased the concentrations of estradiol (E2. Additionally, the expression of estrogen synthesis enzyme Cyp19a1 was sharply downregulated. Moreover, flow cytometry showed that knockdown of XBP1 increased the apoptosis rate and arrests the cell cycle in S-phase in granulosa cells (GCs. Further study confirmed these results. The expression of CCAAT-enhancer-binding protein homologous protein (CHOP, cysteinyl aspartate specific proteases-3 (caspase-3, cleaved caspase-3, and Cyclin E was upregulated, while that of Bcl-2, Cyclin A1, and Cyclin B1 was downregulated. Simultaneously, CCK8 analysis indicated that XBP1 disruption inhibited cell proliferation. In addition, XBP1 knockdown also alters the expression of Has2 and Ptgs2, two essential genes for folliculogenesis. Collectively, these data reveal a novel critical role of XBP1 in folliculogenesis by regulating the cell cycle, apoptosis, and steroid synthesis of mouse granulosa cells.

  7. Methylene blue photodynamic therapy in malignant melanoma decreases expression of proliferating cell nuclear antigen and heparanases.

    Science.gov (United States)

    Wagner, M; Suarez, E R; Theodoro, T R; Machado Filho, C D A S; Gama, M F M; Tardivo, J P; Paschoal, F M; Pinhal, M A S

    2012-07-01

    Malignant melanoma (MM) is a very aggressive tumour. Although surgical excision of MM in the early stages has a very good prognosis, it often fails to completely inhibit tumour progression. Methylene blue photodynamic therapy (MB-PDT) is a technique that induces tissue damage by reactive oxygen species (ROS). To investigate the efficacy of and potential use of MB-PDT in restraining the aggressiveness of MM by analysing levels of proliferating cell nuclear antigen (PCNA) and heparanase (HPSE, a molecular marker of cell invasion) in a mouse model. Expression of PCNA and two HPSE isoforms were analysed using immunohistochemistry (IHC) after MB-PDT in mice. Tumour volume and weight were also measured. Two treatments with MB-PDT promoted a decrease of 99% decrease in tumour volume and 75% in tumour weight compared with untreated mice (P < 0.05). Using IHC, a decrease in expression of 75% for PCNA and 95% for both HPSE isoforms (P < 0.05) was found. MB-PDT is a cheap and efficient method of decreasing MM volume and thus disease progression. This reduction is mediated by downregulation of PCNA and heparanases. © The Author(s). CED © 2012 British Association of Dermatologists.

  8. Decreased soluble cell adhesion molecules after tirofiban infusion in patients with unstable angina pectoris

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    Aliyev Emil

    2004-04-01

    Full Text Available Abstract Aim The inflammatory response, initiated by neutrophil and monocyte adhesion to endothelial cells, is important in the pathogenesis of acute coronary syndromes. Platelets play an important role in inflammatory process by interacting with monocytes and neutrophils. In this study, we investigated the effect of tirofiban on the levels of cell adhesion molecules (soluble intercellular adhesion molecule-1, sICAM-1, and vascular cell adhesion molecule-1, sVCAM-1 in patients with unstable angina pectoris (AP. Methods Thirty-five patients with unstable AP (Group I, ten patients with stable AP (Group II and ten subjects who had angiographycally normal coronary arteries (Group III were included the study. Group I was divided into two subgroups for the specific treatment regimens: Group IA (n = 15 received tirofiban and Group IB (n = 20 did not. Blood samples for investigating the cell adhesion molecules were drawn at zero time (baseline; 0 h in all patients and at 72 h in Group I. Results The baseline levels of sICAM-1 and sVCAM-1 were higher in Group I than in Groups II and III. They were higher in Group IA than in Group IB. However, the sICAM-1 and sVCAM-1 levels decreased significantly in Group IA after tirofiban infusion. In contrast, these levels remained unchanged or were increased above the baseline value in Group IB at 72 h. Conclusion The levels of cell adhesion molecules in patients with unstable AP decreased significantly after tirofiban infusion. Inhibition of platelet function by specific glycoprotein IIb/IIIa antagonists may decrease platelet-mediated inflammation and the ischemic end-point.

  9. Pulmonary artery endothelial cell dysfunction and decreased populations of highly proliferative endothelial cells in experimental congenital diaphragmatic hernia

    Science.gov (United States)

    Seedorf, Gregory J.; Abman, Steven H.; Nozik-Grayck, Eva; Partrick, David A.; Gien, Jason

    2013-01-01

    Decreased lung vascular growth and pulmonary hypertension contribute to poor outcomes in congenital diaphragmatic hernia (CDH). Mechanisms that impair angiogenesis in CDH are poorly understood. We hypothesize that decreased vessel growth in CDH is caused by pulmonary artery endothelial cell (PAEC) dysfunction with loss of a highly proliferative population of PAECs (HP-PAEC). PAECs were harvested from near-term fetal sheep that underwent surgical disruption of the diaphragm at 60–70 days gestational age. Highly proliferative potential was measured via single cell assay. PAEC function was assessed by assays of growth and tube formation and response to known proangiogenic stimuli, vascular endothelial growth factor (VEGF), and nitric oxide (NO). Western blot analysis was used to measure content of angiogenic proteins, and superoxide production was assessed. By single cell assay, the proportion of HP-PAEC with growth of >1,000 cells was markedly reduced in the CDH PAEC, from 29% (controls) to 1% (CDH) (P CDH PAEC growth and tube formation were decreased by 31% (P = 0.012) and 54% (P CDH PAEC growth and tube formation. VEGF and VEGF-R2 proteins were increased in CDH PAEC; however, eNOS and extracellular superoxide dismutase proteins were decreased by 29 and 88%, respectively. We conclude that surgically induced CDH in fetal sheep causes endothelial dysfunction and marked reduction of the HP-PAEC population. We speculate that this CDH PAEC phenotype contributes to impaired vascular growth in CDH. PMID:24124189

  10. Increased Mesenchymal Stem Cell Response and Decreased Staphylococcus aureus Adhesion on Titania Nanotubes without Pharmaceuticals

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    Zhiqiang Xu

    2015-01-01

    Full Text Available Titanium (Ti implants with enhanced biocompatibility and antibacterial property are highly desirable and characterized by improved success rates. In this study, titania nanotubes (TNTs with various tube diameters were fabricated on Ti surfaces through electrochemical anodization at 10, 30, and 60 V (denoted as NT10, NT30, and NT60, resp.. Ti was also investigated and used as a control. NT10 with a diameter of 30 nm could promote the adhesion and proliferation of bone marrow mesenchymal stem cells (BMSCs without noticeable differentiation. NT30 with a diameter of 100 nm could support the adhesion and proliferation of BMSCs and induce osteogenesis. NT60 with a diameter of 200 nm demonstrated the best ability to promote cell spreading and osteogenic differentiation; however, it clearly impaired cell adhesion and proliferation. As the tube diameter increased, bacterial adhesion on the TNTs decreased and reached the lowest value on NT60. Therefore, NT30 without pharmaceuticals could be used to increase mesenchymal stem cell response and decrease Staphylococcus aureus adhesion and thus should be further studied for improving the efficacy of Ti-based orthopedic implants.

  11. Decreased memory B cells and increased CD8 memory T cells in blood of breastfed children: the generation R study.

    Science.gov (United States)

    Jansen, Michelle A E; van den Heuvel, Diana; van Zelm, Menno C; Jaddoe, Vincent W V; Hofman, Albert; de Jongste, Johan C; Hooijkaas, Herbert; Moll, Henriette A

    2015-01-01

    Breastfeeding provides a protective effect against infectious diseases in infancy. Still, immunological evidence for enhanced adaptive immunity in breastfed children remains inconclusive. To determine whether breastfeeding affects B- and T-cell memory in the first years of life. We performed immunophenotypic analysis on blood samples within a population-based prospective cohort study. Participants included children at 6 months (n=258), 14 months (n=166), 25 months (n=112) and 6 years of age (n=332) with both data on breastfeeding and blood lymphocytes. Total B- and T-cell numbers and their memory subsets were determined with 6-color flow cytometry. Mothers completed questionnaires on breastfeeding when their children were aged 2, 6, and 12 months. Multiple linear regression models with adjustments for potential confounders were performed. Per month continuation of breastfeeding, a 3% (95% CI -6, -1) decrease in CD27+IgM+, a 2% (95 CI % -5, -1) decrease in CD27+IgA+ and a 2% (95% CI -4, -1) decrease in CD27-IgG+ memory B cell numbers were observed at 6 months of age. CD8 T-cell numbers at 6 months of age were 20% (95% CI 3, 37) higher in breastfed than in non-breastfed infants. This was mainly found for central memory CD8 T cells and associated with exposure to breast milk, rather than duration. The same trend was observed at 14 months, but associations disappeared at older ages. Longer breastfeeding is associated with increased CD8 T-cell memory, but not B-cell memory numbers in the first 6 months of life. This transient skewing towards T cell memory might contribute to the protective effect against infectious diseases in infancy.

  12. Decreased memory B cells and increased CD8 memory T cells in blood of breastfed children: the generation R study.

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    Michelle A E Jansen

    Full Text Available Breastfeeding provides a protective effect against infectious diseases in infancy. Still, immunological evidence for enhanced adaptive immunity in breastfed children remains inconclusive.To determine whether breastfeeding affects B- and T-cell memory in the first years of life.We performed immunophenotypic analysis on blood samples within a population-based prospective cohort study. Participants included children at 6 months (n=258, 14 months (n=166, 25 months (n=112 and 6 years of age (n=332 with both data on breastfeeding and blood lymphocytes. Total B- and T-cell numbers and their memory subsets were determined with 6-color flow cytometry. Mothers completed questionnaires on breastfeeding when their children were aged 2, 6, and 12 months. Multiple linear regression models with adjustments for potential confounders were performed.Per month continuation of breastfeeding, a 3% (95% CI -6, -1 decrease in CD27+IgM+, a 2% (95 CI % -5, -1 decrease in CD27+IgA+ and a 2% (95% CI -4, -1 decrease in CD27-IgG+ memory B cell numbers were observed at 6 months of age. CD8 T-cell numbers at 6 months of age were 20% (95% CI 3, 37 higher in breastfed than in non-breastfed infants. This was mainly found for central memory CD8 T cells and associated with exposure to breast milk, rather than duration. The same trend was observed at 14 months, but associations disappeared at older ages.Longer breastfeeding is associated with increased CD8 T-cell memory, but not B-cell memory numbers in the first 6 months of life. This transient skewing towards T cell memory might contribute to the protective effect against infectious diseases in infancy.

  13. Physical exercise decreases the number of fetal cells in maternal blood

    DEFF Research Database (Denmark)

    Schlütter, Jacob Mørup; Kirkegaard, Ida; Christensen, Connie Britta

    Physical exercise decreases the number of fetal cells in maternal blood J. M. Schlütter1, I. Kirkegaard1, B. Christensen2, S. Kølvraa3, N. Uldbjerg1 1. Department of Gynecology and Obstetrics, Aarhus University Hospital, Skejby, Aarhus N, Denmark. 2. FCMB ApS, Vejle, Denmark. 3. Department...... and 10-15 % have no cells in 30 mL, which is a challenge for the implementation of the method for prenatal diagnostic purposes. Considering the assumed fragility of circulating fetal cells we wondered if physical exercise prior to blood sampling might influence the number of fcmb. The objective...... in maternal blood. The explanation might be increased clearance of fcmb from the circulation during exercise. If analysis of fcmb is introduced as a clinical parameter pregnant women should avoid extensive physical activity 24 hours prior to blood sampling....

  14. Tyrosine kinase/phosphatase inhibitors decrease dengue virus production in HepG2 cells.

    Science.gov (United States)

    Limjindaporn, Thawornchai; Panaampon, Jutatip; Malakar, Shilu; Noisakran, Sansanee; Yenchitsomanus, Pa-Thai

    2017-01-29

    Dengue virus is the causative agent of dengue fever, dengue hemorrhagic fever, and dengue shock syndrome. High rates of dengue virus replication and virion production are related to disease severity. To identify anti-DENV compounds, we performed cell-based ELISA testing to detect the level of DENV E protein expression. Among a total of 83 inhibitors, eight were identified as inhibitors with antiviral activity. Epidermal growth factor receptor inhibitor II (EGFR/ErbB-2/ErbB-4 inhibitor II) and protein tyrosine phosphatase inhibitor IV (PTP inhibitor IV) significantly inhibited dengue virus production and demonstrated low toxicity in hepatocyte cell lines. Our results suggest the efficacy of tyrosine kinase/phosphatase inhibitors in decreasing dengue virus production in HepG2 cells. Copyright © 2017 Elsevier Inc. All rights reserved.

  15. Dichloroacetate Decreases Cell Health and Activates Oxidative Stress Defense Pathways in Rat Alveolar Type II Pneumocytes

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    Alexis Valauri-Orton

    2015-01-01

    Full Text Available Dichloroacetate (DCA is a water purification byproduct that is known to be hepatotoxic and hepatocarcinogenic and to induce peripheral neuropathy and damage macrophages. This study characterizes the effects of the haloacetate on lung cells by exposing rat alveolar type II (L2 cells to 0–24 mM DCA for 6–24 hours. Increasing DCA concentration and the combination of increasing DCA concentration plus longer exposures decrease measures of cellular health. Length of exposure has no effect on oxidative stress biomarkers, glutathione, SOD, or CAT. Increasing DCA concentration alone does not affect total glutathione or its redox ratio but does increase activity in the SOD/CAT oxidative stress defense pathway. These data suggest that alveolar type II cells rely on SOD and CAT more than glutathione to combat DCA-induced stress.

  16. Deferoxamine Compensates for Decreases in B Cell Counts and Reduces Mortality in Enterovirus 71-Infected Mice

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    Yajun Yang

    2014-07-01

    Full Text Available Enterovirus 71 is one of the major causative agents of hand, foot and mouth disease in children under six years of age. No vaccine or antiviral therapy is currently available. In this work, we found that the number of B cells was reduced in enterovirus 71-infected mice. Deferoxamine, a marine microbial natural product, compensated for the decreased levels of B cells caused by enterovirus 71 infection. The neutralizing antibody titer was also improved after deferoxamine treatment. Furthermore, deferoxamine relieved symptoms and reduced mortality and muscle damage caused by enterovirus 71 infection. This work suggested that deferoxamine has the potential for further development as a B cell-immunomodulator against enterovirus 71.

  17. Doxycycline decreases production of interleukin-8 in a549 human lung epithelial cells

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    Hoyt JC

    2013-03-01

    Full Text Available Doxycycline is an antibiotic that possess anti-inflammatory properties. These anti-inflammatory properties make doxycycline an attractive candidate for possible treatments for a variety of common chronic obstructive airway diseases. Interleukin-8 (IL-8 is a major inflammatory chemokine and a powerful chemo-attractant for both neutrophils and monocytes. We hypothesized that doxycycline might exert its anti-inflammatory effects, at least in part, by modulating IL-8 production. To test this hypothesis, A549 human lung epithelial cells were stimulated with cytomix (IL-1beta, TNF-alpha and gamma-IFN in the presence or absence of varying concentrations of doxycycline. Doxycycline decreased IL-8 protein production in a concentration- and time-dependent manner. In the presence of 30 microg/ml doxycycline IL-8 protein production was decreased by 63% through out a 30 hr time course. In chemotaxis assays monocyte and neutrophil migration was decreased by 55% and 57% respectively. Reverse transcriptase-polymerase chain reaction (RT-PCR experiments suggest that doxycycline does not decrease expression of IL-8 mRNA and that use of the RNA polymerase II inhibitor DRB indicates that doxycycline does not effect stability of this mRNA. In the presence of doxycycline p38-alpha mitogen-activated protein kinase (MAPK expression is decreased by 36% in cytomix-stimulated cells. These data demonstrate that doxycycline can modulate IL-8 release and suggest that it has potential as an anti-inflammatory in those disorders where IL-8 is an important inflammatory mediator.

  18. Cell-free supernatants from probiotic Lactobacillus casei and Lactobacillus rhamnosus GG decrease colon cancer cell invasion in vitro.

    Science.gov (United States)

    Escamilla, Juanita; Lane, Michelle A; Maitin, Vatsala

    2012-08-01

    Probiotics have been shown to have a preventative role in colorectal carcinogenesis but research concerning their prophylactic potential in the later stages of colorectal cancer, specifically metastasis is limited. This study explored the potential of cell-free supernatants (CFS) from 2 probiotic Lactobacillus sp., Lactobacillus casei and Lactobacillus rhamnosus GG, to inhibit colon cancer cell invasion by influencing matrix metalloproteinase-9 (MMP-9) activity and levels of the tight junction protein zona occludens-1 (ZO-1) in cultured metastatic human colorectal carcinoma cells. HCT-116 cells were treated with CFS from L. casei, L. rhamnosus, or Bacteroides thetaiotaomicron (a gut commensal); or with uninoculated bacterial growth media. Treatment with CFS from both Lactobacillus sp. decreased colorectal cell invasion but treatment with CFS from B. thetaiotaomicron did not. CFS from both Lactobacillus sp. decreased MMP-9 and increased ZO-1 protein levels. L. rhamnosus CFS also lowered MMP-9 activity. To begin elucidating the secreted bacterial factor conveying these responses, Lactobacillus sp. CFS were fractionated into defined molecular weight ranges and cell invasion assessed. Fractionation revealed that the inhibitory activity was contained primarily in the >100 kDa and 50-100 kDa fractions, suggesting the inhibitory compound may be a macromolecule such as a protein, nucleic acid, or a polysaccharide.

  19. Mitofusin2 decreases intracellular cholesterol of oxidized LDL-induced foam cells from rat vascular smooth muscle cells.

    Science.gov (United States)

    He, Chao; Chen, Ying; Liu, Chun; Cao, Ming; Fan, Yu-jin; Guo, Xiao-mei

    2013-04-01

    Mitofusin2 (Mfn2) plays a pivotal role in the proliferation and apoptosis of vascular smooth muscle cells (VSMCs). The purpose of this study was to investigate the effects of Mfn2 on the trafficking of intracellular cholesterol in the foam cells derived from rat VSMCs (rVSMCs) and also to investigate the effects of Mfn2 on the expression of adenosine triphosphate-binding cassette subfamily A member 1 (ABCA1), adenosine triphosphate-binding cassette subfamily G member 1 (ABCG1) and peroxisome proliferator-activated receptor gamma (PPARγ). The rVSMCs were co-cultured with oxidized low density lipoprotein (LDL, 80 μg/mL) to produce foam cells and cholesterol accumulation in cells. Before oxidized LDL treatment, different titers (20, 40 and 60 pfu/cell) of recombinant adenovirus containing Mfn2 gene (Adv-Mfn2) were added into the culture medium for 24 h to transfect the Mfn2 gene into the rVSMCs. Then the cells were harvested for analyses. The protein expression of Mfn2 was significantly higher in Adv-Mfn2-transfected group than in untransfected group (PLDL treatment, rVSMCs became irregular and their nuclei became larger, and their plasma abounded with red lipid droplets. However, the number of red lipid droplets was significantly decreased in Adv-Mfn2-transfected group as compared with untransfected group. At 48 h after oxidized LDL treatment, the intracellular cholesterol in rVSMCs was significantly increased (P0.05), the phosporylation levels of PPARγ were significantly decreased in Adv-Mfn2-transfected group as compared with untransfected group (Pcholesterol in oxidized LDL-induced rVSMCs possibly by decreasing PPARγ phosporylation and then increasing protein expression levels of ABCA1 and ABCG1, which may be helpful to suppress the formation of foam cells.

  20. Bmi1 loss produces an increase in astroglial cells and a decrease in neural stem cell population and proliferation.

    Science.gov (United States)

    Zencak, Dusan; Lingbeek, Merel; Kostic, Corinne; Tekaya, Meriem; Tanger, Ellen; Hornfeld, Dana; Jaquet, Muriel; Munier, Francis L; Schorderet, Daniel F; van Lohuizen, Maarten; Arsenijevic, Yvan

    2005-06-15

    The polycomb transcriptional repressor Bmi1 promotes cell cycle progression, controls cell senescence, and is implicated in brain development. Loss of Bmi1 leads to a decreased brain size and causes progressive ataxia and epilepsy. Recently, Bmi1 was shown to control neural stem cell (NSC) renewal. However, the effect of Bmi1 loss on neural cell fate in vivo and the question whether the action of Bmi1 was intrinsic to the NSCs remained to be investigated. Here, we show that Bmi1 is expressed in the germinal zone in vivo and in NSCs as well as in progenitors proliferating in vitro, but not in differentiated cells. Loss of Bmi1 led to a decrease in proliferation in zones known to contain progenitors: the newborn cortex and the newborn and adult subventricular zone. This decrease was accentuated in vitro, where we observed a drastic reduction in NSC proliferation and renewal because of NSC-intrinsic effects of Bmi1 as shown by the means of RNA interference. Bmi1(-/-) mice also presented more astrocytes at birth, and a generalized gliosis at postnatal day 30. At both stages, colocalization of bromodeoxyuridine and GFAP demonstrated that Bmi1 loss did not prevent astrocyte precursor proliferation. Supporting these observations, Bmi1(-/-) neurospheres generate preferentially astrocytes probably attributable to a different responsiveness to environmental factors. Bmi1 is therefore necessary for NSC renewal in a cell-intrinsic mode, whereas the altered cell pattern of the Bmi1(-/-) brain shows that in vivo astrocyte precursors can proliferate in the absence of Bmi1.

  1. Three-dimensional culture conditions lead to decreased radiation induced cytotoxicity in human mammary epithelial cells

    International Nuclear Information System (INIS)

    Sowa, Marianne B.; Chrisler, William B.; Zens, Kyra D.; Ashjian, Emily J.; Opresko, Lee K.

    2010-01-01

    For both targeted and non-targeted exposures, the cellular responses to ionizing radiation have predominantly been measured in two-dimensional monolayer cultures. Although convenient for biochemical analysis, the true interactions in vivo depend upon complex interactions between cells themselves and the surrounding extracellular matrix. This study directly compares the influence of culture conditions on radiation induced cytotoxicity following exposure to low-LET ionizing radiation. Using a three-dimensional (3D) human mammary epithelial tissue model, we have found a protective effect of 3D cell culture on cell survival after irradiation. The initial state of the cells (i.e., 2D versus 3D culture) at the time of irradiation does not alter survival, nor does the presence of extracellular matrix during and after exposure to dose, but long term culture in 3D which offers significant reduction in cytotoxicity at a given dose (e.g. ∼4-fold increased survival at 5 Gy). The cell cycle delay induced following exposure to 2 and 5 Gy was almost identical between 2D and 3D culture conditions and cannot account for the observed differences in radiation responses. However the amount of apoptosis following radiation exposure is significantly decreased in 3D culture relative to the 2D monolayer after the same dose. A likely mechanism of the cytoprotective effect afforded by 3D culture conditions is the down regulation of radiation induced apoptosis in 3D structures.

  2. Decreased Helios Expression in Regulatory T Cells in Acute Coronary Syndrome

    Directory of Open Access Journals (Sweden)

    Lili Jiang

    2017-01-01

    Full Text Available Regulatory T cells (Tregs play an essential role in acute coronary syndrome (ACS. However, there is debate about which Treg subsets are truly critical to ACS. Helios, a transcription factor, was recently reported to be a bona fide marker for natural Tregs or activated Tregs with a suppression function, but little is known about its role in ACS. We therefore examined Helios+ Tregs in patients with ACS, patients with stable angina, and control subjects. 73 patients with ACS, 30 patients with stable angina, and 48 control subjects were enrolled. The frequencies and estimated absolute numbers of different Treg subsets in peripheral blood were measured by flow cytometry. Plasma cytokine level was measured by ELISA. The mRNA expression of Foxp3 and Helios in purified CD4+ T cells was determined by RT-PCR. Helios+ Tregs was decreased significantly in patients with ACS. The frequency and estimated absolute numbers of CD4+Foxp3+Helios+ Tregs were negatively correlated with IL-6 and positively correlated with circulating level of TGF-beta1 and HDL-C. The mRNA expression of Foxp3 and Helios was decreased in CD4+ T cells from patients with ACS. In summary, Helios+ Tregs was downregulated in patients with ACS and may play a role in ACS.

  3. Tripterygium regelii decreases the biosynthesis of triacylglycerol and cholesterol in HepG2 cells.

    Science.gov (United States)

    Kang, Myung-Ji; Kwon, Eun-Bin; Yuk, Heung Joo; Ryu, Hyung Won; Kim, Soo-Yeon; Lee, Mi-Kyeong; Moon, Dong-Oh; Lee, Su Ui; Oh, Sei-Ryang; Lee, Hyun-Sun; Kim, Mun-Ock

    2017-12-01

    In the course of screening to find a plant material decreasing the activity of triacylglycerol and cholesterol, we identified Tripterygium regelii (TR). The methanol extract of TR leaves (TR-LM) was shown to reduce the intracellular lipid contents consisting of triacylglycerol (TG) and cholesterol in HepG2 cells. TR-LM also downregulated the mRNA and protein expression of the lipogenic genes such as SREBP-1 and its target enzymes. Consequently, TR-LM reduced the TG biosynthesis in HepG2 cells. In addition, TR-LM decreased SREBP2 and its target enzyme HMG-CoA reductase, which is involved in cholesterol synthesis. In this study, we evaluated that TR-LM attenuated cellular lipid contents through the suppression of de novo TG and cholesterol biosynthesis in HepG2 cells. All these taken together, TR-LM could be beneficial in regulating lipid metabolism and useful preventing the hyperlipidemia and its complications, in that liver is a crucial tissue for the secretion of serum lipids.

  4. Phytoncide Extracted from Pinecone Decreases LPS-Induced Inflammatory Responses in Bovine Mammary Epithelial Cells.

    Science.gov (United States)

    Kang, Sukyung; Lee, Jae Sung; Lee, Hai Chon; Petriello, Michael C; Kim, Bae Yong; Do, Jeong Tae; Lim, Dae-Seog; Lee, Hong Gu; Han, Sung Gu

    2016-03-01

    Mastitis is a prevalent inflammatory disease that remains one of the main causes of poor quality of milk. Phytoncides are naturally occurring anti-inflammatory compounds derived from plants and trees. To determine if treatment with phytoncide could decrease the severity of lipopolysaccharide (LPS)-induced inflammatory responses, mammary alveolar epithelial cells (MAC-T) were pretreated with phytoncide (0.02% and 0.04% (v/v)) followed by LPS treatment (1 and 25 μg/ml). The results demonstrated that phytoncide downregulated LPS-induced pro-inflammatory cyclooxygenase-2 (COX-2) expression. Additionally, LPS-induced activation of ERK1/2, p38, and Akt was attenuated by phytoncide. Treatment of cells with known pharmacological inhibitors of ERK1/2 (PD98059), p38 (SB203580), and Akt (LY294002) confirmed the association of these signaling pathways with the observed alterations in COX-2 expression. Moreover, phytoncide attenuated LPS-induced NF-κB activation and superoxide production, and, finally, treatment with phytoncide increased Nrf2 activation. Results suggest that phytoncide can decrease LPS-induced inflammation in MAC-T cells.

  5. GPR30 decreases cardiac chymase/angiotensin II by inhibiting local mast cell number

    Energy Technology Data Exchange (ETDEWEB)

    Zhao, Zhuo [Department of Anesthesiology, Wake Forest School of Medicine, Medical Center Boulevard, Winston-Salem, NC 27159-1009 (United States); Department of Cardiology, Jinan Central Hospital, Affiliated with Shandong University, 105 Jiefang Road, Jinan, 250013 (China); Wang, Hao; Lin, Marina [Department of Anesthesiology, Wake Forest School of Medicine, Medical Center Boulevard, Winston-Salem, NC 27159-1009 (United States); Groban, Leanne, E-mail: lgroban@wakehealth.edu [Department of Anesthesiology, Wake Forest School of Medicine, Medical Center Boulevard, Winston-Salem, NC 27159-1009 (United States); Hypertension and Vascular Disease Center, Wake Forest School of Medicine, Medical Center Boulevard, Winston-Salem, NC 27157 (United States); Office of Women in Medicine and Science, Wake Forest School of Medicine, Medical Center Boulevard, Winston-Salem, NC 27157 (United States)

    2015-03-27

    Chronic activation of the novel estrogen receptor GPR30 by its agonist G1 mitigates the adverse effects of estrogen (E2) loss on cardiac structure and function. Using the ovariectomized (OVX) mRen2.Lewis rat, an E2-sensitive model of diastolic dysfunction, we found that E2 status is inversely correlated with local cardiac angiotensin II (Ang II) levels, likely via Ang I/chymase-mediated production. Since chymase is released from cardiac mast cells during stress (e.g., volume/pressure overload, inflammation), we hypothesized that GPR30-related cardioprotection after E2 loss might occur through its opposing actions on cardiac mast cell proliferation and chymase production. Using real-time quantitative PCR, immunohistochemistry, and immunoblot analysis, we found mast cell number, chymase expression, and cardiac Ang II levels were significantly increased in the hearts of OVX-compared to ovary-intact mRen2.Lewis rats and the GPR30 agonist G1 (50 mg/kg/day, s.c.) administered for 2 weeks limited the adverse effects of estrogen loss. In vitro studies revealed that GPR30 receptors are expressed in the RBL-2H3 mast cell line and G1 inhibits serum-induced cell proliferation in a dose-dependent manner, as determined by cell counting, BrdU incorporation assay, and Ki-67 staining. Using specific antagonists to estrogen receptors, blockage of GPR30, but not ERα or ERβ, attenuated the inhibitory effects of estrogen on BrdU incorporation in RBL-2H3 cells. Further study of the mechanism underlying the effect on cell proliferation showed that G1 inhibits cyclin-dependent kinase 1 (CDK1) mRNA and protein expression in RBL-2H3 cells in a dose-dependent manner. - Highlights: • GPR30 activation limits mast cell number in hearts from OVX mRen2.Lewis rats. • GPR30 activation decreases cardiac chymase/angiotensin II after estrogen loss. • GPR30 activation inhibits RBL-2H3 mast cell proliferation and CDK1 expression.

  6. Dehydropachymic acid decreases bafilomycin A1 induced β-Amyloid accumulation in PC12 cells.

    Science.gov (United States)

    Yu, Mengyao; Xu, Xiaoyan; Jiang, Nan; Wei, Wei; Li, Fang; He, Liming; Luo, Xia

    2017-02-23

    Fuling, the sclerotium of Poria cocos, was frequently used in traditional Chinese medicine (TCM) formulae for Alzheimer's disease (AD) intervention over the past 10 centuries. And its extracts exhibited significant effects in both cellular and animal models of AD in previous studies. However, its mechanisms on prevention and treatment of AD have not been well elucidated yet. To investigate the effect and corresponding mechanisms of dehydropachymic acid, which is one of the major triterpenes in P. cocos, on the clearance of β-amyloid accumulation in bafilomycin A1 induced PC12 cells. MTT assay was used to examine the DPA effect on the viability of PC12 cells stable transfected with pCB6-APP (PC12-APP). PC12-APP cells were treated with DPA at the concentration of 6.25, 12.5, 25μg/mL for 4h, and then co-treated with 50nmol/L bafilomycin A1 for 48h except the controls. The Aβ 1-42 content in culture medium was determined by ELISA. The intracellular amount of APP, Aβ 1-42 , LC3, cathepsin D was measured by Western blotting and normalized to GAPDH loading control. The PC12 cells stable transfected with pSelect-LC3-GFP (PC12-LC3-GFP) was used in the fluorescence microscopy estimation of autophagosomes accumulation. The internal pH in lysosome was detected by LysoTracker Red staining. DPA had no significant effect on the cell viability but could significantly decrease Aβ 1-42 content in culture medium and eliminate the intracellular accumulation of APP and Aβ 1-42 in bafilomycin A1 induced PC12-APP cells. Furthermore, DPA lowered the LC3-II/LC3-I ratio and reduced the GFP-labeled LC3 puncta which were elevated by bafilomycin A1. And the increase in internal pH of lysosome and decrease in mCatD amount in Bafilomycin A1 induced PC12-APP cells were restored by DPA treatment. These results indicated that DPA could restore the lysosomal acidification and recover the autophgic flux which is impaired by bafilomycin A1. DPA could effectively clear the accumulation of Aβ 1

  7. Ketone supplementation decreases tumor cell viability and prolongs survival of mice with metastatic cancer.

    Science.gov (United States)

    Poff, A M; Ari, C; Arnold, P; Seyfried, T N; D'Agostino, D P

    2014-10-01

    Cancer cells express an abnormal metabolism characterized by increased glucose consumption owing to genetic mutations and mitochondrial dysfunction. Previous studies indicate that unlike healthy tissues, cancer cells are unable to effectively use ketone bodies for energy. Furthermore, ketones inhibit the proliferation and viability of cultured tumor cells. As the Warburg effect is especially prominent in metastatic cells, we hypothesized that dietary ketone supplementation would inhibit metastatic cancer progression in vivo. Proliferation and viability were measured in the highly metastatic VM-M3 cells cultured in the presence and absence of β-hydroxybutyrate (βHB). Adult male inbred VM mice were implanted subcutaneously with firefly luciferase-tagged syngeneic VM-M3 cells. Mice were fed a standard diet supplemented with either 1,3-butanediol (BD) or a ketone ester (KE), which are metabolized to the ketone bodies βHB and acetoacetate. Tumor growth was monitored by in vivo bioluminescent imaging. Survival time, tumor growth rate, blood glucose, blood βHB and body weight were measured throughout the survival study. Ketone supplementation decreased proliferation and viability of the VM-M3 cells grown in vitro, even in the presence of high glucose. Dietary ketone supplementation with BD and KE prolonged survival in VM-M3 mice with systemic metastatic cancer by 51 and 69%, respectively (p < 0.05). Ketone administration elicited anticancer effects in vitro and in vivo independent of glucose levels or calorie restriction. The use of supplemental ketone precursors as a cancer treatment should be further investigated in animal models to determine potential for future clinical use. © 2014 The Authors Published by Wiley Periodicals, Inc. on behalf of UICC.

  8. Total glucosides of paeony inhibits Th1/Th17 cells via decreasing dendritic cells activation in rheumatoid arthritis.

    Science.gov (United States)

    Lin, Jinpiao; Xiao, Lianbo; Ouyang, Guilin; Shen, Yu; Huo, Rongfen; Zhou, Zhou; Sun, Yue; Zhu, Xianjin; Zhang, Jie; Shen, Baihua; Li, Ningli

    2012-12-01

    Total glucoside of paeony (TGP), an active compound extracted from paeony root, has been used in therapy for rheumatoid arthritis (RA). Th1 and Th17 cells are now believed to play crucial roles in the lesions of RA. However, the molecular mechanism of TGP in inhibition of Th1 and Th17 cells remains unclear. In this study, we found that TGP treatment significantly decreased percentage and number of Th1 and Th17 cells in collagen induced arthritis (CIA) mice. Consistently, treatment with TGP decreased expression of T-bet and RORγt as well as phosphorylation of STAT1 and STAT3. In particular, TGP treatment inhibited dendritic cells (DCs) maturation and reduced production of IL-12 and IL-6. Moreover, TGP-treatment RA patients showed shank population of matured DCs and IFN-γ-, IL-17-producing cells. Taken together, our results demonstrated that TGP inhibited maturation and activation of DCs, which led to impaired Th1 and Th17 differentiation in vivo. Copyright © 2013 Elsevier Inc. All rights reserved.

  9. Murine Lung Cancer Increases CD4+ T Cell Apoptosis and Decreases Gut Proliferative Capacity in Sepsis.

    Science.gov (United States)

    Lyons, John D; Mittal, Rohit; Fay, Katherine T; Chen, Ching-Wen; Liang, Zhe; Margoles, Lindsay M; Burd, Eileen M; Farris, Alton B; Ford, Mandy L; Coopersmith, Craig M

    2016-01-01

    Mortality is significantly higher in septic patients with cancer than in septic patients without a history of cancer. We have previously described a model of pancreatic cancer followed by sepsis from Pseudomonas aeruginosa pneumonia in which cancer septic mice have higher mortality than previously healthy septic mice, associated with increased gut epithelial apoptosis and decreased T cell apoptosis. The purpose of this study was to determine whether this represents a common host response by creating a new model in which both the type of cancer and the model of sepsis are altered. C57Bl/6 mice received an injection of 250,000 cells of the lung cancer line LLC-1 into their right thigh and were followed three weeks for development of palpable tumors. Mice with cancer and mice without cancer were then subjected to cecal ligation and puncture and sacrificed 24 hours after the onset of sepsis or followed 7 days for survival. Cancer septic mice had a higher mortality than previously healthy septic mice (60% vs. 18%, p = 0.003). Cancer septic mice had decreased number and frequency of splenic CD4+ lymphocytes secondary to increased apoptosis without changes in splenic CD8+ numbers. Intestinal proliferation was also decreased in cancer septic mice. Cancer septic mice had a higher bacterial burden in the peritoneal cavity, but this was not associated with alterations in local cytokine, neutrophil or dendritic cell responses. Cancer septic mice had biochemical evidence of worsened renal function, but there was no histologic evidence of renal injury. Animals with cancer have a significantly higher mortality than previously healthy animals following sepsis. The potential mechanisms associated with this elevated mortality differ significantly based upon the model of cancer and sepsis utilized. While lymphocyte apoptosis and intestinal integrity are both altered by the combination of cancer and sepsis, the patterns of these alterations vary greatly depending on the models used.

  10. Radiation-induced decrease of CD8+ dendritic cells contributes to Th1/Th2 shift.

    Science.gov (United States)

    Liu, Hu; Li, Bailong; Jia, Xiaojing; Ma, Yan; Gu, Yifeng; Zhang, Pei; Wei, Qun; Cai, Jianming; Cui, Jianguo; Gao, Fu; Yang, Yanyong

    2017-05-01

    Exposure to ionizing radiation (IR) often reduce the helper T (Th) 1 like function, resulting in a Th1/Th2 imbalance, which could affect the efficacy of cancer radiotherapy. As the most potent antigen presenting cells, dendritic cells (DC) can be divided into several subsets with specialized function. However, there is no literature covering the changes of DC subsets and their roles in immune regulation in response to IR. In the present study, we were aimed to investigate the changes of DC subsets after IR and its relationship with Th1/Th2 immunity. We found a significant decrease of BDCA3+DC in the blood of patients treated with radiotherapy. CD8+DC, a mouse equivalent of human BDCA3+DC, was also found decreased in mice spleen, peripheral blood and lymph node tissues after irradiation. As CD8+DC mainly induce Th1 immunity, we tested the changes of Th1/Th2 response and found that IR caused a repression of Th1 immunity, indicating a possible role of CD8+DC in radiation-induced Th1/Th2 imbalance. We also found that a CD8+DC-inducing cytokine, Fms-like tyrosine kinase 3 ligand (FLT3 ligand), restored CD8+DC and reversed Th1/Th2 shift. And then we found that bone marrow cells from irradiated mice differentiated into less CD8+DC, which was also protected by FLT3 ligand. In conclusion, our data showed that IR induced a decrease of CD8+DC and Th1/Th2 shift, which was reversed by Flt3 ligand treatment, suggesting a novel mechanism for radiation-induced immunosuppression. Copyright © 2017. Published by Elsevier B.V.

  11. Murine Lung Cancer Increases CD4+ T Cell Apoptosis and Decreases Gut Proliferative Capacity in Sepsis.

    Directory of Open Access Journals (Sweden)

    John D Lyons

    Full Text Available Mortality is significantly higher in septic patients with cancer than in septic patients without a history of cancer. We have previously described a model of pancreatic cancer followed by sepsis from Pseudomonas aeruginosa pneumonia in which cancer septic mice have higher mortality than previously healthy septic mice, associated with increased gut epithelial apoptosis and decreased T cell apoptosis. The purpose of this study was to determine whether this represents a common host response by creating a new model in which both the type of cancer and the model of sepsis are altered.C57Bl/6 mice received an injection of 250,000 cells of the lung cancer line LLC-1 into their right thigh and were followed three weeks for development of palpable tumors. Mice with cancer and mice without cancer were then subjected to cecal ligation and puncture and sacrificed 24 hours after the onset of sepsis or followed 7 days for survival.Cancer septic mice had a higher mortality than previously healthy septic mice (60% vs. 18%, p = 0.003. Cancer septic mice had decreased number and frequency of splenic CD4+ lymphocytes secondary to increased apoptosis without changes in splenic CD8+ numbers. Intestinal proliferation was also decreased in cancer septic mice. Cancer septic mice had a higher bacterial burden in the peritoneal cavity, but this was not associated with alterations in local cytokine, neutrophil or dendritic cell responses. Cancer septic mice had biochemical evidence of worsened renal function, but there was no histologic evidence of renal injury.Animals with cancer have a significantly higher mortality than previously healthy animals following sepsis. The potential mechanisms associated with this elevated mortality differ significantly based upon the model of cancer and sepsis utilized. While lymphocyte apoptosis and intestinal integrity are both altered by the combination of cancer and sepsis, the patterns of these alterations vary greatly depending on

  12. The cell-free fetal DNA fraction in maternal blood decreases after physical activity

    DEFF Research Database (Denmark)

    Schlütter, Jacob Mørup; Hatt, Lotte; Bach, Cathrine

    2014-01-01

    OBJECTIVE: If noninvasive prenatal testing using next generation sequencing is to be effective for pregnant women, a cell-free fetal DNA (cffDNA) fraction above 4% is essential unless the depth of sequencing is increased. This study's objective is to determine whether physical activity has...... an effect on the proportion of cell-free DNA (cfDNA) arising from the fetus (fetal fraction). METHODS: Nine pregnant women carrying male fetuses at gestational age 12(+0)  weeks to 14(+6)  weeks were included. Plasma from nine pregnant women was drawn prior to, immediately after, and 30 min after 30 min...... of cycling with a pulse-rate of 150 beats per minute. The concentrations of cffDNA (DYS14) and cfDNA (RASSF1A) were assessed using quantitative real-time polymerase chain reaction. RESULTS: The fetal fraction decreased significantly in all participants after physical activity (p 

  13. Chemotherapy-induced cognitive impairment is associated with decreases in cell proliferation and histone modifications

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    Briones Teresita L

    2011-12-01

    Full Text Available Abstract Background In this study, we examined the effects of cyclophosphamide, methothrexate, and 5-Fluorouracil (CMF drug combination on various aspects of learning and memory. We also examined the effects of CMF on cell proliferation and chromatin remodeling as possible underlying mechanisms to explain chemotherapy-associated cognitive dysfunction. Twenty-four adult female Wistar rats were included in the study and had minimitter implantation for continuous activity monitoring two weeks before the chemotherapy regimen was started. Once baseline activity data were collected, rats were randomly assigned to receive either CMF or saline injections given intraperitoneally. Treatments were given once a week for a total of 4 weeks. Two weeks after the last injection, rats were tested in the water maze for spatial learning and memory ability as well as discrimination learning. Bromodeoxyuridine (BrdU injection was given at 100 mg/Kg intraperitoneally 4 hours prior to euthanasia to determine hippocampal cell proliferation while histone acetylation and histone deacetylase activity was measured to determine CMF effects on chromatin remodeling. Results Our data showed learning and memory impairment following CMF administration independent of the drug effects on physical activity. In addition, CMF-treated rats showed decreased hippocampal cell proliferation, associated with increased histone acetylation and decreased histone deacetylase activity. Conclusions These results suggest the negative consequences of chemotherapy on brain function and that anti-cancer drugs can adversely affect the self-renewal potential of neural progenitor cells and also chromatin remodeling in the hippocampus. The significance of our findings lie on the possible usefulness of animal models in addressing the clinical phenomenon of 'chemobrain.'

  14. Paradoxical sleep deprivation decreases serum testosterone and Leydig cells in male rats

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    Fitranto Arjadi

    2014-04-01

    Full Text Available Background Chronic stress increases glucocorticoid levels and accelerates reduction in Leydig cells functions and numbers. Chronic stress models in the working place comprise sleep deprivation, sedentary stress, and physical stress. The aim of this study was to evaluate the effect of various work stress models, such as stress from paradoxical sleep deprivation (PSD, immobilization, and footshock, on serum testosterone levels and number of Leydig cells in male albino rats. Methods This study was of experimental randomized post-test only with control group design using 24 male Wistar albino rats (Rattus norvegicus. The sample was divided into 4 groups: K1 (control, K2 (PSD, K3 (immobilization and K4 (footshock, receiving treatment for 25 days. Measured parameters were serum testosterone level and Leydig cell number. Analysis of variance (ANOVA was used for statistical analysis, followed by post hoc LSD. Results Mean serum testosterone levels (0.07 ± 0.08 ng/mL and Leydig cell numbers (4.22 ± l0.96 were lowest in the PSD stress model. Serum testosterone levels differed significantly between controls and PSD group (p=0.014, while there was a significant difference in numbers of Leydig cells between footshock stress and PSD (p=0.011 and between the three stress groups and controls (p=0.006. Conclusion This study demonstrated that PSD, immobilization and footshock stress significantly decreased serum testosterone levels and number of Leydig cells in male albino rats (Rattus norvegicus. The mechanism by which PSD affects serum testosterone is still unclear.

  15. Losartan therapy decreases albuminuria with stable glomerular filtration and permselectivity in sickle cell anemia.

    Science.gov (United States)

    Yee, Marianne E; Lane, Peter A; Archer, David R; Joiner, Clinton H; Eckman, James R; Guasch, Antonio

    2018-03-01

    Sickle cell nephropathy begins with hyperfiltration and microalbuminuria and may progress to renal failure. The aim of this study was to determine the effects of losartan on glomerular function and albumin excretion in sickle cell anemia (SCA). Individuals with SCA on hydroxyurea with persistent albuminuria were enrolled in a 1-year study of losartan. Glomerular filtration rate (GFR) measured by iohexol clearance, albumin excretion rate (AER), and fractional clearance of dextran were assessed at baseline, short-term (1-2month), and long-term (≥12month) intervals. Twelve subjects (6 microalbuminuria, 6 macroalbuminuria) completed short-term studies; 8 completed long-term studies. Baseline GFR was 112ml/min/1.73m 2 (71-147ml/min/1.73m 2 ). AER decreased significantly at the short-term (median decrease -134 mcg/min, p=0.0063). GFR was not significantly-different at short-term or long-term intervals. Dextran clearance improved for diameters smaller than albumin (therapy for ≥1year in sickle nephropathy results in lower albumin excretion with stable GFR. Filtration of neutral molecules ≥36Å was not changed by losartan, suggesting that the effect of losartan is a mechanism other than alteration of glomerular filtration size-selectivity. Copyright © 2017 Elsevier Inc. All rights reserved.

  16. Valproic acid decreases urothelial cancer cell proliferation and induces thrombospondin-1 expression

    Science.gov (United States)

    2012-01-01

    Background Prevention of bladder cancer recurrence is a central challenge in the management of this highly prevalent disease. The histone deacetylase inhibitor valproic acid (sodium valproate) has anti-angiogenic properties and has been shown to decrease bladder cancer growth in model systems. We have previously shown reduced expression of thrombospondin-1 in a mouse model and in human bladder cancer relative to normal urothelium. We speculated that inhibition of angiogenesis by valproate might be mediated by this anti-angiogenic protein. Methods Bladder cancer cell lines UMUC3 and T24 were treated with valproate or another histone deacetylase inhibitor, vorinostat, in culture for a period of three days. Proliferation was assessed by alamar blue reduction. Gene expression was evaluated by reverse transcription of RNA and quantitative PCR. Results Proliferation assays showed treatment with valproate or vorinostat decreased proliferation in both cell lines. Histone deacetylase inhibition also increased relative expression of thrombospondin-1 up to 8 fold at 5 mM valproate. Conclusions Histone deacetylase inhibitors warrant further study for the prevention or treatment of bladder cancer. PMID:22898175

  17. Valproic acid decreases urothelial cancer cell proliferation and induces thrombospondin-1 expression

    Directory of Open Access Journals (Sweden)

    Byler Timothy K

    2012-08-01

    Full Text Available Abstract Background Prevention of bladder cancer recurrence is a central challenge in the management of this highly prevalent disease. The histone deacetylase inhibitor valproic acid (sodium valproate has anti-angiogenic properties and has been shown to decrease bladder cancer growth in model systems. We have previously shown reduced expression of thrombospondin-1 in a mouse model and in human bladder cancer relative to normal urothelium. We speculated that inhibition of angiogenesis by valproate might be mediated by this anti-angiogenic protein. Methods Bladder cancer cell lines UMUC3 and T24 were treated with valproate or another histone deacetylase inhibitor, vorinostat, in culture for a period of three days. Proliferation was assessed by alamar blue reduction. Gene expression was evaluated by reverse transcription of RNA and quantitative PCR. Results Proliferation assays showed treatment with valproate or vorinostat decreased proliferation in both cell lines. Histone deacetylase inhibition also increased relative expression of thrombospondin-1 up to 8 fold at 5 mM valproate. Conclusions Histone deacetylase inhibitors warrant further study for the prevention or treatment of bladder cancer.

  18. Fluoxetine Decreases the Proliferation and Adipogenic Differentiation of Human Adipose-Derived Stem Cells

    Directory of Open Access Journals (Sweden)

    Bo Kyung Sun

    2015-07-01

    Full Text Available Fluoxetine was originally developed as an antidepressant, but it has also been used to treat obesity. Although the anti-appetite effect of fluoxetine is well-documented, its potential effects on human adipose-derived stem cells (ASCs or mature adipocytes have not been investigated. Therefore, we investigated the mechanisms underlying the inhibitory effects of fluoxetine on the proliferation of ASCs. We also investigated its inhibitory effect on adipogenic differentiation. Fluoxetine significantly decreased ASC proliferation, and signal transduction PCR array analysis showed that it increased expression of autophagy-related genes. In addition, fluoxetine up-regulated SQSTM1 and LC3B protein expression as detected by western blotting and immunofluorescence. The autophagy inhibitor, 3-methyladenine (3-MA, significantly attenuated fluoxetine-mediated effects on ASC proliferation and SQSTM1/LC3B expression. In addition, 3-MA decreased the mRNA expression of two autophagy-related genes, beclin-1 and Atg7, in ASCs. Fluoxetine also significantly inhibited lipid accumulation and down-regulated the levels of PPAR-γ and C/EBP-α in ASCs. Collectively, these results indicate that fluoxetine decreases ASC proliferation and adipogenic differentiation. This is the first in vitro evidence that fluoxetine can reduce fat accumulation by inhibiting ASC proliferation and differentiation.

  19. Cabergoline decreases alcohol drinking and seeking behaviors via glial cell line-derived neurotrophic factor.

    Science.gov (United States)

    Carnicella, Sebastien; Ahmadiantehrani, Somayeh; He, Dao-Yao; Nielsen, Carsten K; Bartlett, Selena E; Janak, Patricia H; Ron, Dorit

    2009-07-15

    Cabergoline is an ergotamine derivative that increases the expression of glial cell line-derived neurotrophic factor (GDNF) in vitro. We recently showed that GDNF in the ventral tegmental area (VTA) reduces the motivation to consume alcohol. We therefore set out to determine whether cabergoline administration decreases alcohol-drinking and -seeking behaviors via GDNF. Reverse transcription polymerase chain reaction (RT-PCR) and Enzyme-Linked ImmunoSorbent Assay (ELISA) were used to measure GDNF levels. Western blot analysis was used for phosphorylation experiments. Operant self-administration in rats and a two-bottle choice procedure in mice were used to assess alcohol-drinking behaviors. Instrumental performance tested during extinction was used to measure alcohol-seeking behavior. The [35S]GTPgammaS binding assay was used to assess the expression and function of the dopamine D2 receptor (D2R). We found that treatment of the dopaminergic-like cell line SH-SY5Y with cabergoline and systemic administration of cabergoline in rats resulted in an increase in GDNF level and in the activation of the GDNF pathway. Cabergoline treatment decreased alcohol-drinking and -seeking behaviors including relapse, and its action to reduce alcohol consumption was localized to the VTA. Finally, the increase in GDNF expression and the decrease in alcohol consumption by cabergoline were abolished in GDNF heterozygous knockout mice. Together, these findings suggest that cabergoline-mediated upregulation of the GDNF pathway attenuates alcohol-drinking behaviors and relapse. Alcohol abuse and addiction are devastating and costly problems worldwide. This study puts forward the possibility that cabergoline might be an effective treatment for these disorders.

  20. Nanomolar Caffeic Acid Decreases Glucose Uptake and the Effects of High Glucose in Endothelial Cells.

    Directory of Open Access Journals (Sweden)

    Lucia Natarelli

    Full Text Available Epidemiological studies suggest that moderate and prolonged consumption of coffee is associated with a reduced risk of developing type 2 diabetes but the molecular mechanisms underlying this effect are not known. In this study, we report the effects of physiological concentrations of caffeic acid, easily achievable by normal dietary habits, in endothelial cells cultured in 25 mM of glucose (high glucose, HG. In HG, the presence of 10 nM caffeic acid was associated with a decrease of glucose uptake but not to changes of GLUT-1 membrane localization or mRNA levels. Moreover, caffeic acid countered HG-induced loss of barrier integrity, reducing actin rearrangement and FITC-dextran passage. The decreased flux of glucose associated to caffeic acid affected HG induced apoptosis by down-regulating the expression of initiator (caspase 8 and 9 and effector caspases (caspase 7 and 3 and by increasing the levels of phosphorylated Bcl-2. We also observed that caffeic acid in HG condition was associated to a reduction of p65 subunit nuclear levels with respect to HG alone. NF-κB activation has been shown to lead to apoptosis in HG treated cells and the analysis of the expression of a panel of about 90 genes related to NF-κB signaling pathway revealed that caffeic acid significantly influenced gene expression changes induced by HG. In conclusion, our results suggest that caffeic acid, decreasing the metabolic stress induced by HG, allows the activation of survival mechanisms mediated by a different modulation of NF-κB-related signaling pathways and to the activation of anti-apoptotic proteins.

  1. Childhood tuberculosis is associated with decreased abundance of T cell gene transcripts and impaired T cell function.

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    Cheryl Hemingway

    Full Text Available The WHO estimates around a million children contract tuberculosis (TB annually with over 80 000 deaths from dissemination of infection outside of the lungs. The insidious onset and association with skin test anergy suggests failure of the immune system to both recognise and respond to infection. To understand the immune mechanisms, we studied genome-wide whole blood RNA expression in children with TB meningitis (TBM. Findings were validated in a second cohort of children with TBM and pulmonary TB (PTB, and functional T-cell responses studied in a third cohort of children with TBM, other extrapulmonary TB (EPTB and PTB. The predominant RNA transcriptional response in children with TBM was decreased abundance of multiple genes, with 140/204 (68% of all differentially regulated genes showing reduced abundance compared to healthy controls. Findings were validated in a second cohort with concordance of the direction of differential expression in both TBM (r2 = 0.78 p = 2x10-16 and PTB patients (r2 = 0.71 p = 2x10-16 when compared to a second group of healthy controls. Although the direction of expression of these significant genes was similar in the PTB patients, the magnitude of differential transcript abundance was less in PTB than in TBM. The majority of genes were involved in activation of leucocytes (p = 2.67E-11 and T-cell receptor signalling (p = 6.56E-07. Less abundant gene expression in immune cells was associated with a functional defect in T-cell proliferation that recovered after full TB treatment (p<0.0003. Multiple genes involved in T-cell activation show decreased abundance in children with acute TB, who also have impaired functional T-cell responses. Our data suggest that childhood TB is associated with an acquired immune defect, potentially resulting in failure to contain the pathogen. Elucidation of the mechanism causing the immune paresis may identify new treatment and prevention strategies.

  2. Decreased astroglial cell adhesion and proliferation on zinc oxide nanoparticle polyurethane composites

    Science.gov (United States)

    Seil, Justin T; Webster, Thomas J

    2008-01-01

    Nanomaterials offer a number of properties that are of interest to the field of neural tissue engineering. Specifically, materials that exhibit nanoscale surface dimensions have been shown to promote neuron function while simultaneously minimizing the activity of cells such as astrocytes that inhibit central nervous system regeneration. Studies demonstrating enhanced neural tissue regeneration in electrical fields through the use of conductive materials have led to interest in piezoelectric materials (or those materials which generate a transient electrical potential when mechanically deformed) such as zinc oxide (ZnO). It has been speculated that ZnO nanoparticles possess increased piezoelectric properties over ZnO micron particles. Due to this promise in neural applications, the objective of the present in vitro study was, for the first time, to assess the activity of astroglial cells on ZnO nanoparticle polymer composites. ZnO nanoparticles embedded in polyurethane were analyzed via scanning electron microscopy to evaluate nanoscale surface features of the composites. The surface chemistry was characterized via X-ray photoelectron spectroscopy. Astroglial cell response was evaluated based on cell adhesion and proliferation. Astrocyte adhesion was significantly reduced on ZnO nanoparticle/polyurethane (PU) composites with a weight ratio of 50:50 (PU:ZnO) wt.%, 75:25 (PU:ZnO) wt.%, and 90:10 (PU:ZnO) wt.% in comparison to pure PU. The successful production of ZnO nanoparticle composite scaffolds suitable for decreasing astroglial cell density demonstrates their potential as a nerve guidance channel material with greater efficiency than what may be available today. PMID:19337420

  3. Cell-autonomous decrease in proliferative competitiveness of the aged hepatocyte.

    Science.gov (United States)

    Serra, Maria Paola; Marongiu, Fabio; Marongiu, Michela; Contini, Antonella; Laconi, Ezio

    2015-06-01

    The regenerative potential of the liver declines with age, this might be dependent on a decrease in the intensity of the stimulus and/or an increased refractoriness of the target. In the present study, we compared the in vivo growth capacity of young and old hepatocytes transplanted into the same host. We utilized the retrorsine (RS)-based model for liver repopulation, which provides a specific and effective stimulus for transplanted hepatocytes. Rats of the dipeptidyl-peptidase type IV (DPP-IV)-deficient strain were given RS and were injected with a mix of hepatocytes isolated from either a 2-month old or an 18-month old donor. To follow the fate of transplanted cells, they were each identified through a specific tag: young hepatocytes expressed the green fluorescent protein (GFP(+)), while those from old donors were DPP-IV-positive. At 1 month post-transplantation, DPP-IV-positive clusters (derived from old donor) were consistently smaller than those GFP(+) (young donor); the cross sectional area of clusters was decreased by 50%, while the mean volume was reduced to 1/3. Furthermore, when 2/3 partial hepatectomy (PH) was performed, the S-phase response of old hepatocyte-derived clusters was only 30-40% compared to that observed in cluster originating from young hepatocytes. No markers of cell senescence were expressed in clusters of transplanted hepatocytes. This is the first direct evidence in vivo that hepatocytes in the aged liver express a cell-autonomous decline in their replicative capacity and in their regenerative response to PH compared to those from a young animal. Copyright © 2015 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.

  4. Acrolein decreases endothelial cell migration and insulin sensitivity through induction of let-7a.

    Science.gov (United States)

    O'Toole, Timothy E; Abplanalp, Wesley; Li, Xiaohong; Cooper, Nigel; Conklin, Daniel J; Haberzettl, Petra; Bhatnagar, Aruni

    2014-08-01

    Acrolein is a major reactive component of vehicle exhaust, and cigarette and wood smoke. It is also present in several food substances and is generated endogenously during inflammation and lipid peroxidation. Although previous studies have shown that dietary or inhalation exposure to acrolein results in endothelial activation, platelet activation, and accelerated atherogenesis, the basis for these effects is unknown. Moreover, the effects of acrolein on microRNA (miRNA) have not been studied. Using AGILENT miRNA microarray high-throughput technology, we found that treatment of cultured human umbilical vein endothelial cells with acrolein led to a significant (>1.5-fold) upregulation of 12, and downregulation of 15, miRNAs. Among the miRNAs upregulated were members of the let-7 family and this upregulation was associated with decreased expression of their protein targets, β3 integrin, Cdc34, and K-Ras. Exposure to acrolein attenuated β3 integrin-dependent migration and reduced Akt phosphorylation in response to insulin. These effects of acrolein on endothelial cell migration and insulin signaling were reversed by expression of a let-7a inhibitor. Also, inhalation exposure of mice to acrolein (1 ppm x 6 h/day x 4 days) upregulated let-7a and led to a decrease in insulin-stimulated Akt phosphorylation in the aorta. These results suggest that acrolein exposure has broad effects on endothelial miRNA repertoire and that attenuation of endothelial cell migration and insulin signaling by acrolein is mediated in part by the upregulation of let-7a. This mechanism may be a significant feature of vascular injury caused by inflammation, oxidized lipids, and exposure to environmental pollutants. © The Author 2014. Published by Oxford University Press on behalf of the Society of Toxicology. All rights reserved. For permissions, please email: journals.permissions@oup.com.

  5. Rosiglitazone and AS601245 decrease cell adhesion and migration through modulation of specific gene expression in human colon cancer cells.

    Directory of Open Access Journals (Sweden)

    Angelo Cerbone

    Full Text Available PPARs are nuclear receptors activated by ligands. Activation of PPARγ leads to a reduction of adhesion and motility in some cancer models. PPARγ transcriptional activity can be negatively regulated by JNK-mediated phosphorylation. We postulated that the use of agents able to inhibit JNK activity could increase the effectiveness of PPARγ ligands. We analysed the effects of rosiglitazone (PPARγ ligand and AS601245 (a selective JNK inhibitor alone or in association on adhesion and migration of CaCo-2, HT29, and SW480 human colon cancer cells and investigated, through microarray analysis, the genes involved in these processes. Cell adhesion and migration was strongly inhibited by rosiglitazone and AS601245. Combined treatment with the two compounds resulted in a greater reduction of the adhesion and migration capacity. Affymetrix analysis in CaCo-2 cells revealed that some genes which were highly modulated by the combined treatment could be involved in these biological responses. Rosiglitazone, AS601245 and combined treatment down-regulated the expression of fibrinogen chains in all three cell lines. Moreover, rosiglitazone, alone or in association with AS601245, caused a decrease in the fibrinogen release. ARHGEF7/β-PIX gene was highly down-regulated by combined treatment, and western blot analysis revealed that β-PIX protein is down-modulated in CaCo-2, HT29 and SW480 cells, also. Transfection of cells with β-PIX gene completely abrogated the inhibitory effect on cell migration, determined by rosiglitazone, AS601245 and combined treatment. Results demonstrated that β-PIX protein is involved in the inhibition of cell migration and sustaining the positive interaction between PPARγ ligands and anti-inflammatory agents in humans.

  6. Decreased expression of FOXA2 promotes eutopic endometrial cell proliferation and migration in patients with endometriosis.

    Science.gov (United States)

    Lin, Anping; Yin, Juan; Cheng, Chao; Yang, Zhu; Yang, Huan

    2018-02-01

    Endometriosis is characterized by eutopic endometrial cell 'metastasis' to ectopic foci. FOXA2 is a member of the forkhead transcription factor family, which may participate in transcriptional regulation in endometrial cells and contribute to the aetiology of endometriosis. This study investigated the roles played by FOXA2 in eutopic endometrium using endometriosis samples. Western blotting showed that the relative expression of FOXA2 was significantly reduced in eutopic endometrium from patients with endometriosis (n = 14) compared with endometriosis-free controls (n = 16) (0.69 ± 0.07 versus 1.24 ± 0.06, P FOXA2 siRNA. MTT assay showed that cell viability of ESC with transfected FOXA2 siRNA increased significantly, whereas the apoptosis rate decreased as indicated by flow cytometry experiments (both P FOXA2 siRNA promoted ESC migration. Moreover, real-time PCR analysis showed progesterone-induced FOXA2 expression in ESC under physiological conditions. In conclusion, these findings indicate that FOXA2 might be a progesterone-induced gene, which may participate in the 'metastatic' process of eutopic endometrium to ectopic loci in patients with endometriosis. Copyright © 2017 Reproductive Healthcare Ltd. Published by Elsevier Ltd. All rights reserved.

  7. Colon Cancer Chemoprevention by Sage Tea Drinking: Decreased DNA Damage and Cell Proliferation.

    Science.gov (United States)

    Pedro, Dalila F N; Ramos, Alice A; Lima, Cristovao F; Baltazar, Fatima; Pereira-Wilson, Cristina

    2016-02-01

    Salvia officinalis and some of its isolated compounds have been found to be preventive of DNA damage and increased proliferation in vitro in colon cells. In the present study, we used the azoxymethane model to test effects of S. officinalis on colon cancer prevention in vivo. The results showed that sage treatment reduced the number of ACF formed only if administered before azoxymethane injection, demonstrating that sage tea drinking has a chemopreventive effect on colorectal cancer. A decrease in the proliferation marker Ki67 and in H2 O2 -induced and azoxymethane-induced DNA damage to colonocytes and lymphocytes were found with sage treatment. This confirms in vivo the chemopreventive effects of S. officinalis. Taken together, our results show that sage treatment prevented initiation phases of colon carcinogenesis, an effect due, at least in part, to DNA protection, and reduced proliferation rates of colon epithelial cell that prevent mutations and their fixation through cell replication. These chemopreventive effects of S. officinalis on colon cancer add to the many health benefits attributed to sage and encourage its consumption. Copyright © 2015 John Wiley & Sons, Ltd.

  8. Hydroxyurea treatment decreases glomerular hyperfiltration in children with sickle cell anemia.

    Science.gov (United States)

    Aygun, Banu; Mortier, Nicole A; Smeltzer, Matthew P; Shulkin, Barry L; Hankins, Jane S; Ware, Russell E

    2013-02-01

    Glomerular hyperfiltration and microalbuminuria/proteinuria are early manifestations of sickle nephropathy. The effects of hydroxyurea therapy on these renal manifestations of sickle cell anemia (SCA) are not well defined. Our objective was to investigate the effects of hydroxyurea on glomerular filtration rate (GFR) measured by (99m)Tc-DTPA clearance, and on microalbuminuria/proteinuria in children with SCA. Hydroxyurea study of long-term effects (HUSTLE) is a prospective study (NCT00305175) with the goal of describing the long-term cellular, molecular, and clinical effects of hydroxyurea therapy in SCA. Glomerular filtration rate, urine microalbumin, and serum cystatin C were measured before initiating hydroxyurea therapy and then repeated after 3 years. Baseline and Year 3 values for HUSTLE subjects were compared using the Wilcoxon Signed Rank test. Associations between continuous variables were evaluated using Spearman correlation coefficient. Twenty-three children with SCA (median age 7.5 years, range, 2.5-14.0 years) received hydroxyurea at maximum tolerated dose (MTD, 24.4 ± 4.5 mg/kg/day, range, 15.3-30.6 mg/kg/day). After 3 years of treatment, GFR measured by (99m)Tc-DTPA decreased significantly from 167 ± 46 mL/min/1.73 m² to 145 ± 27 mL/min/1.73 m² (P = 0.016). This decrease in GFR was significantly associated with increase in fetal hemoglobin (P = 0.042) and decrease in lactate dehydrogenase levels (P = 0.035). Urine microalbumin and cystatin C levels did not change significantly. Hydroxyurea at MTD is associated with a decrease in hyperfiltration in young children with SCA. Copyright © 2012 Wiley Periodicals, Inc.

  9. Targeting NADPH oxidase decreases oxidative stress in the transgenic sickle cell mouse penis.

    Science.gov (United States)

    Musicki, Biljana; Liu, Tongyun; Sezen, Sena F; Burnett, Arthur L

    2012-08-01

    Sickle cell disease (SCD) is a state of chronic vasculopathy characterized by endothelial dysfunction and increased oxidative stress, but the sources and mechanisms responsible for reactive oxygen species (ROS) production in the penis are unknown. We evaluated whether SCD activates NADPH oxidase, induces endothelial nitric oxide synthase (eNOS) uncoupling, and decreases antioxidants in the SCD mouse penis. We further tested the hypothesis that targeting NADPH oxidase decreases oxidative stress in the SCD mouse penis. SCD transgenic (sickle) mice were used as an animal model of SCD. Hemizygous (hemi) mice served as controls. Mice received an NADPH oxidase inhibitor apocynin (10 mM in drinking water) or vehicle. Penes were excised at baseline for molecular studies. Markers of oxidative stress (4-hydroxy-2-nonenal [HNE]), sources of ROS (eNOS uncoupling and NADPH oxidase subunits p67(phox) , p47(phox) , and gp91(phox) ), and enzymatic antioxidants (superoxide dismutase [SOD]1, SOD2, catalase, and glutathione peroxidase-1 [GPx1]) were measured by Western blot in penes. Sources of ROS, oxidative stress, and enzymatic antioxidants in the SCD penis. Relative to hemi mice, SCD increased (Ppenis. Apocynin treatment of sickle mice reversed (P0.05) prevented eNOS uncoupling in the penis. Apocynin treatment of hemi mice did not affect any of these parameters. NADPH oxidase and eNOS uncoupling are sources of oxidative stress in the SCD penis; decreased GPx1 further contributes to oxidative stress. Inhibition of NADPH oxidase upregulation decreases oxidative stress, implying a major role for NADPH oxidase as a ROS source and a potential target for improving vascular function in the SCD mouse penis. © 2012 International Society for Sexual Medicine.

  10. Daphnetin inhibits invasion and migration of LM8 murine osteosarcoma cells by decreasing RhoA and Cdc42 expression

    International Nuclear Information System (INIS)

    Fukuda, Hiroki; Nakamura, Seikou; Chisaki, Yugo; Takada, Tetsuya; Toda, Yuki; Murata, Hiroaki; Itoh, Kazuyuki; Yano, Yoshitaka; Takata, Kazuyuki; Ashihara, Eishi

    2016-01-01

    Daphnetin, 7,8-dihydroxycoumarin, present in main constituents of Daphne odora var. marginatai, has multiple pharmacological activities including anti-proliferative effects in cancer cells. In this study, using a Transwell system, we showed that daphnetin inhibited invasion and migration of highly metastatic murine osteosarcoma LM8 cells in a dose-dependent manner. Following treatment by daphnetin, cells that penetrated the Transwell membrane were rounder than non-treated cells. Immunofluorescence analysis revealed that daphnetin decreased the numbers of intracellular stress fibers and filopodia. Moreover, daphnetin treatment dramatically decreased the expression levels of RhoA and Cdc42. In summary, the dihydroxycoumarin derivative daphnetin inhibits the invasion and migration of LM8 cells, and therefore represents a promising agent for use against metastatic cancer. - Highlights: • Daphnetin, a coumarin-derivative, inhibited invasion and migration of LM8 cells. • Stress fibers and filopodia were decreased by daphnetin treatment. • Daphnetin decreased RhoA and Cdc42 protein expression.

  11. Daphnetin inhibits invasion and migration of LM8 murine osteosarcoma cells by decreasing RhoA and Cdc42 expression

    Energy Technology Data Exchange (ETDEWEB)

    Fukuda, Hiroki [Department of Clinical and Translational Physiology, Kyoto Pharmaceutical University, Kyoto (Japan); Nakamura, Seikou [Department of Pharmacognosy, Kyoto Pharmaceutical University, Kyoto (Japan); Chisaki, Yugo [Education and Research Center for Clinical Pharmacy, Kyoto Pharmaceutical University, Kyoto (Japan); Takada, Tetsuya [Department of Clinical and Translational Physiology, Kyoto Pharmaceutical University, Kyoto (Japan); Toda, Yuki [Department of Medicinal Chemistry, Kyoto Pharmaceutical University, Kyoto (Japan); Murata, Hiroaki [Department of Orthopedics, Graduate School of Medical Science, Kyoto Prefectural University of Medicine, Kyoto (Japan); Department of Orthopedic Surgery, Matsushita Memorial Hospital, Osaka (Japan); Itoh, Kazuyuki [Department of Biology, Osaka Medical Center of Cancer and Cardiovascular Diseases, Osaka (Japan); Yano, Yoshitaka [Education and Research Center for Clinical Pharmacy, Kyoto Pharmaceutical University, Kyoto (Japan); Takata, Kazuyuki [Department of Clinical and Translational Physiology, Kyoto Pharmaceutical University, Kyoto (Japan); Ashihara, Eishi, E-mail: ash@mb.kyoto-phu.ac.jp [Department of Clinical and Translational Physiology, Kyoto Pharmaceutical University, Kyoto (Japan)

    2016-02-26

    Daphnetin, 7,8-dihydroxycoumarin, present in main constituents of Daphne odora var. marginatai, has multiple pharmacological activities including anti-proliferative effects in cancer cells. In this study, using a Transwell system, we showed that daphnetin inhibited invasion and migration of highly metastatic murine osteosarcoma LM8 cells in a dose-dependent manner. Following treatment by daphnetin, cells that penetrated the Transwell membrane were rounder than non-treated cells. Immunofluorescence analysis revealed that daphnetin decreased the numbers of intracellular stress fibers and filopodia. Moreover, daphnetin treatment dramatically decreased the expression levels of RhoA and Cdc42. In summary, the dihydroxycoumarin derivative daphnetin inhibits the invasion and migration of LM8 cells, and therefore represents a promising agent for use against metastatic cancer. - Highlights: • Daphnetin, a coumarin-derivative, inhibited invasion and migration of LM8 cells. • Stress fibers and filopodia were decreased by daphnetin treatment. • Daphnetin decreased RhoA and Cdc42 protein expression.

  12. Stable SREBP-1a knockdown decreases the cell proliferation rate in human preadipocyte cells without inducing senescence

    International Nuclear Information System (INIS)

    Alvarez, María Soledad; Fernandez-Alvarez, Ana; Cucarella, Carme; Casado, Marta

    2014-01-01

    Highlights: • SGBS cells mostly expressed SREBP-1a variant. • SREBP-1a knockdown decreased the proliferation of SGBS cells without inducing senescence. • We have identified RBBP8 and CDKN3 genes as potential SREBP-1a targets. - Abstract: Sterol regulatory element binding proteins (SREBP), encoded by the Srebf1 and Srebf2 genes, are important regulators of genes involved in cholesterol and fatty acid metabolism. Whereas SREBP-2 controls the cholesterol synthesis, SREBP-1 proteins (-1a and -1c) function as the central hubs in lipid metabolism. Despite the key function of these transcription factors to promote adipocyte differentiation, the roles of SREBP-1 proteins during the preadipocyte state remain unknown. Here, we evaluate the role of SREBP-1 in preadipocyte proliferation using RNA interference technology. Knockdown of the SREBP-1a gene decreased the proliferation rate in human SGBS preadipocyte cell strain without inducing senescence. Furthermore, our data identified retinoblastoma binding protein 8 and cyclin-dependent kinase inhibitor 3 genes as new potential SREBP-1 targets, in addition to cyclin-dependent kinase inhibitor 1A which had already been described as a gene regulated by SREBP-1a. These data suggested a new role of SREBP-1 in adipogenesis via regulation of preadipocyte proliferation

  13. Stable SREBP-1a knockdown decreases the cell proliferation rate in human preadipocyte cells without inducing senescence

    Energy Technology Data Exchange (ETDEWEB)

    Alvarez, María Soledad [Instituto de Biomedicina de Valencia (IBV-CSIC), Jaime Roig 11, E-46010 Valencia (Spain); Fernandez-Alvarez, Ana [Fundación Instituto Leloir, IIBBA-CONICET, Av. Patricias Argentinas 435, Ciudad Autónoma de Buenos Aires C1405BWE (Argentina); Cucarella, Carme [Instituto de Biomedicina de Valencia (IBV-CSIC), Jaime Roig 11, E-46010 Valencia (Spain); Casado, Marta, E-mail: mcasado@ibv.csic.es [Instituto de Biomedicina de Valencia (IBV-CSIC), Jaime Roig 11, E-46010 Valencia (Spain)

    2014-04-25

    Highlights: • SGBS cells mostly expressed SREBP-1a variant. • SREBP-1a knockdown decreased the proliferation of SGBS cells without inducing senescence. • We have identified RBBP8 and CDKN3 genes as potential SREBP-1a targets. - Abstract: Sterol regulatory element binding proteins (SREBP), encoded by the Srebf1 and Srebf2 genes, are important regulators of genes involved in cholesterol and fatty acid metabolism. Whereas SREBP-2 controls the cholesterol synthesis, SREBP-1 proteins (-1a and -1c) function as the central hubs in lipid metabolism. Despite the key function of these transcription factors to promote adipocyte differentiation, the roles of SREBP-1 proteins during the preadipocyte state remain unknown. Here, we evaluate the role of SREBP-1 in preadipocyte proliferation using RNA interference technology. Knockdown of the SREBP-1a gene decreased the proliferation rate in human SGBS preadipocyte cell strain without inducing senescence. Furthermore, our data identified retinoblastoma binding protein 8 and cyclin-dependent kinase inhibitor 3 genes as new potential SREBP-1 targets, in addition to cyclin-dependent kinase inhibitor 1A which had already been described as a gene regulated by SREBP-1a. These data suggested a new role of SREBP-1 in adipogenesis via regulation of preadipocyte proliferation.

  14. Enhanced proliferation and decreased apoptosis in lung lavage cells of sarcoidosis patients.

    Science.gov (United States)

    Petzmann, Susanna; Maercker, Christian; Markert, Eva; Kern, Izidor; Osolnik, Katharina; Pohl, Wolfgang; Popper, Helmut H

    2006-10-01

    Sarcoidosis is a systemic autoimmune disease where an inflammatory reaction involving alveolar macrophages, T-helper lymphocytes, and epitheloid cells is mounted against unknown antigens. A genetic predisposition for sarcoidosis is supposed by studies in twins, by geographical and racial distribution. In the current investigation we compared the expression patterns between slow onset and acute sarcodosis using a whole-genome cDNA array. Bronchoalveolar lavage was performed in six patients with slow onset sarcoidosis and four patients with acute sarcoidosis (Löfgren's disease) and obtained cells were used for gene expression profiling. The results were confirmed by RT- and Taqman-PCR. In addition, protein expression was examined on paraffin sections of sarcoid granulomas by immunohistochemistry. In T-helper lymphocytes and alveolar macrophages we found an upregulation of genes belonging to the phosphoinositol-3-kinase/v-akt murine thymoma viral oncogene homolog/signal transducer and activator of transcription 3 pathway, as well as a downregulation of genes of the extrinsic and intrinsic apoptotic signaling cascades. In addition an upregulation of the genes encoding fatty acid binding protein 4 and 5, as well as peroxisome proliferative activated receptor delta in Löfgren's disease was detected. Differences in gene expression between slow onset sarcoidosis and Löfgren's syndrome were found mainly within genes of the major histocompatibility complex. In sarcoidosis enhanced cell proliferation and decreased apoptosis result in accumulation and prolonged survival of antigen-primed T-helper lymphocytes and activated macrophages. This is enhanced in Löfgren's disease, probably by hyper-stimulation via the peroxisome proliferation signaling, providing a larger pool of antigen-primed immune cells.

  15. The Human Amnion Epithelial Cell Secretome Decreases Hepatic Fibrosis in Mice with Chronic Liver Fibrosis

    Directory of Open Access Journals (Sweden)

    Majid Alhomrani

    2017-10-01

    Full Text Available Background: Hepatic stellate cells (HSCs are the primary collagen-secreting cells in the liver. While HSCs are the major cell type involved in the pathogenesis of liver fibrosis, hepatic macrophages also play an important role in mediating fibrogenesis and fibrosis resolution. Previously, we observed a reduction in HSC activation, proliferation, and collagen synthesis following exposure to human amnion epithelial cells (hAEC and hAEC-conditioned media (hAEC-CM. This suggested that specific factors secreted by hAEC might be effective in ameliorating liver fibrosis. hAEC-derived extracellular vesicles (hAEC-EVs, which are nanosized (40–100 nm membrane bound vesicles, may act as novel cell–cell communicators. Accordingly, we evaluated the efficacy of hAEC-EV in modulating liver fibrosis in a mouse model of chronic liver fibrosis and in human HSC.Methods: The hAEC-EVs were isolated and characterized. C57BL/6 mice with CCl4-induced liver fibrosis were administered hAEC-EV, hAEC-CM, or hAEC-EV depleted medium (hAEC-EVDM. LX2 cells, a human HSC line, and bone marrow-derived mouse macrophages were exposed to hAEC-EV, hAEC-CM, and hAEC-EVDM. Mass spectrometry was used to examine the proteome profile of each preparation.Results: The extent of liver fibrosis and number of activated HSCs were reduced significantly in CCl4-treated mice given hAEC-EVs, hAEC-CM, and hAEC EVDM compared to untreated controls. Hepatic macrophages were significantly decreased in all treatment groups, where a predominant M2 phenotype was observed. Human HSCs cultured with hAEC-EV and hAEC-CM displayed a significant reduction in collagen synthesis and hAEC-EV, hAEC-CM, and hAEC-EVDM altered macrophage polarization in bone marrow-derived mouse macrophages. Proteome analysis showed that 164 proteins were unique to hAEC-EV in comparison to hAEC-CM and hAEC-EVDM, and 51 proteins were co-identified components with the hAEC-EV fraction.Conclusion: This study provides novel data

  16. Depletion of arginine in yeast cells decreases the resistance to hydrostatic pressure

    Science.gov (United States)

    Nomura, Kazuki; Iwahashi, Hitoshi; Iguchi, Akinori; Shigematsu, Toru

    2015-07-01

    High hydrostatic pressure (HP) inhibits growth and inactivates microorganisms by destabilizing non-covalent molecular interactions. Arginine contributes to stress resistance because it has a guanidine side chain, which assists in the refolding of aggregated proteins. We attempted to analyze the contribution of arginine to high HP stress using a pressure-sensitive mutant strain of Saccharomyces cerevisiae and a metabolomics approach. Our results showed that the content of 136 out of 250 detected metabolites differed in the mutant and parent strains. Decreased metabolites were involved in the tricarboxylic acid cycle and arginine biosynthesis. The expression of genes contributing to arginine biosynthesis was significantly lower in the mutant strain than in the parent strain. When arginine was supplemented to the medium, the mutant strain showed more tolerance to pressure. These results suggest that yeast cells survived due to the contribution of arginine to high pressure resistance. This indicates that depletion of arginine caused by decreased activity of the biosynthesis pathway confers sensitivity to HP.

  17. X-ray sensitive strains of CHO cells show decreased frequency of stable transfection

    International Nuclear Information System (INIS)

    Jeggo, P.; Smith, J.

    1987-01-01

    Six X-ray sensitive (xrs) strains of the Chinese hamster ovary cell line have previously been isolated and shown to have a defect in double strand break rejoining. In this study, these strains have been investigated for their ability to take up and integrate foreign DNA. All the xrs strains investigated so far have shown a decreased frequency of stable transfectants compared to their parent line, in experiments using the plasmid pSV2gpt, which contains the selectable bacterial gene, guanine phosphoribosyl transferase. This decreased frequency is observed over a wide range of DNA concentrations (0.1 to 20 μg DNA) but is more pronounced at higher DNA concentrations. In contrast, these xrs strains show the same level of transfection proficiency as the wild type parent using a transient transfection system with a plasmid containing the bacterial CAT (chloramphenicol acetyl transferase) gene. Since the level of CAT activity does not depend on integration of foreign DNA, this suggests that the xrs strains are able to take up the same amount of DNA as the parent strains, but have a defect in the integration of foreign DNA. Since this integration of foreign DNA probably occurs by non-homologous recombination, this may indicate a role of the xrs gene product in this process

  18. Decreased fluidity of cell membranes causes a metal ion deficiency in recombinant Saccharomyces cerevisiae producing carotenoids.

    Science.gov (United States)

    Liu, Peitong; Sun, Liang; Sun, Yuxia; Shang, Fei; Yan, Guoliang

    2016-04-01

    The genome-wide transcriptional responses of S. cerevisiae to heterologous carotenoid biosynthesis were investigated using DNA microarray analysis. The results show that the genes involved in metal ion transport were specifically up-regulated in the recombinant strain, and metal ions, including Cu(2+), Fe(2+), Mn(2+), and Mg(2+), were deficient in the recombinant strain compared to the ion content of the parent strain. The decrease in metal ions was ascribed to a decrease in cell membrane (CM) fluidity caused by lower levels of unsaturated fatty acids and ergosterol. This was confirmed by the observation that metal ion levels were restored when CM fluidity was increased by supplying linoleic acid. In addition, a 24.3 % increase in the β-carotene concentration was observed. Collectively, our results suggest that heterologous production of carotenoids in S. cerevisiae can induce cellular stress by rigidifying the CM, which can lead to a deficiency in metal ions. Due to the importance of CM fluidity in cellular physiology, maintaining normal CM fluidity might be a potential approach to improving carotenoid production in genetically engineered S. cerevisiae.

  19. Extracellular magnesium decreases the secretory response of rat peritoneal mast cells to compound 48/80 in vitro

    DEFF Research Database (Denmark)

    Bertelsen, Niels Haldor; Johansen, Torben

    1991-01-01

    Exposure of rat peritoneal mast cells to magnesium in the absence of extracellular calcium resulted in a time- and dose-dependent decrease in the secretory response induced by compound 48/80. The decrease was prevented by a low extracellular concentration of calcium. Furthermore, the decreased se...... that magnesium may decrease the secretory response by displacing the cellular calcium which is utilized in stimulus-secretion coupling....

  20. Dihydropyridines decrease X-ray-induced DNA base damage in mammalian cells

    Energy Technology Data Exchange (ETDEWEB)

    Wojewodzka, M., E-mail: marylaw@ichtj.waw.pl [Center of Radiobiology and Biological Dosimetry, Institute of Nuclear Chemistry and Technology, Warszawa (Poland); Gradzka, I.; Buraczewska, I.; Brzoska, K.; Sochanowicz, B. [Center of Radiobiology and Biological Dosimetry, Institute of Nuclear Chemistry and Technology, Warszawa (Poland); Goncharova, R.; Kuzhir, T. [Institute of Genetics and Cytology, Belarussian National Academy of Sciences, Minsk (Belarus); Szumiel, I. [Center of Radiobiology and Biological Dosimetry, Institute of Nuclear Chemistry and Technology, Warszawa (Poland)

    2009-12-01

    Compounds with the structural motif of 1,4-dihydropyridine display a broad spectrum of biological activities, often defined as bioprotective. Among them are L-type calcium channel blockers, however, also derivatives which do not block calcium channels exert various effects at the cellular and organismal levels. We examined the effect of sodium 3,5-bis-ethoxycarbonyl-2,6-dimethyl-1,4-dihydropyridine-4-carboxylate (denoted here as DHP and previously also as AV-153) on X-ray-induced DNA damage and mutation frequency at the HGPRT (hypoxanthine-guanine phosphoribosyl transferase) locus in Chinese hamster ovary CHO-K1 cells. Using formamido-pyrimidine glycosylase (FPG) comet assay, we found that 1-h DHP (10 nM) treatment before X-irradiation considerably reduced the initial level of FPG-recognized DNA base damage, which was consistent with decreased 8-oxo-7,8-dihydro-2'-deoxyguanosine content and mutation frequency lowered by about 40%. No effect on single strand break rejoining or on cell survival was observed. Similar base damage-protective effect was observed for two calcium channel blockers: nifedipine (structurally similar to DHP) or verapamil (structurally unrelated). So far, the specificity of the DHP-caused reduction in DNA damage - practically limited to base damage - has no satisfactory explanation.

  1. Prophylactic Plasma Transfusion Is Not Associated With Decreased Red Blood Cell Requirements in Critically Ill Patients.

    Science.gov (United States)

    Warner, Matthew A; Chandran, Arun; Jenkins, Gregory; Kor, Daryl J

    2017-05-01

    Critically ill patients frequently receive plasma transfusion under the assumptions that abnormal coagulation test results confer increased risk of bleeding and that plasma transfusion will decrease this risk. However, the effect of prophylactic plasma transfusion remains poorly understood. The objective of this study was to determine the relationship between prophylactic plasma transfusion and bleeding complications in critically ill patients. This is a retrospective cohort study of adults admitted to the intensive care unit (ICU) at a single academic institution between January 1, 2009 and December 31, 2013. Inclusion criteria included age ≥18 years and an international normalized ratio measured during ICU admission. Multivariable propensity-matched analyses were used to evaluate associations between prophylactic plasma transfusion and outcomes of interest with a primary outcome of red blood cell transfusion in the ensuing 24 hours and secondary outcomes of hospital- and ICU-free days and mortality within 30 days of ICU discharge. A total of 27,561 patients were included in the investigation with 2472 (9.0%) receiving plasma therapy and 1105 (44.7%) for which plasma transfusion was prophylactic in nature. In multivariable propensity-matched analyses, patients receiving plasma had higher rates of red blood cell transfusion (odds ratio: 4.3 [95% confidence interval: 3.3-5.7], P plasma in the critically ill was not associated with improved clinical outcomes. Further investigation examining the utility of plasma transfusion in this population is warranted.

  2. Ketamine Metabolites Enantioselectively Decrease Intracellular D-Serine Concentrations in PC-12 Cells.

    Directory of Open Access Journals (Sweden)

    Nagendra S Singh

    Full Text Available D-Serine is an endogenous NMDA receptor co-agonist that activates synaptic NMDA receptors modulating neuronal networks in the cerebral cortex and plays a key role in long-term potentiation of synaptic transmission. D-serine is associated with NMDA receptor neurotoxicity and neurodegeneration and elevated D-serine concentrations have been associated with Alzheimer's and Parkinsons' diseases and amyotrophic lateral sclerosis. Previous studies have demonstrated that the ketamine metabolites (rac-dehydronorketamine and (2S,6S-hydroxynorketamine decrease intracellular D-serine concentrations in a concentration dependent manner in PC-12 cells. In the current study, PC-12 cells were incubated with a series of ketamine metabolites and the IC50 values associated with attenuated intracellular D-serine concentrations were determined. The results demonstrate that structural and stereochemical features of the studied compounds contribute to the magnitude of the inhibitory effect with (2S,6S-hydroxynorketamine and (2R,6R-hydroxynorketamine displaying the most potent inhibition with IC50 values of 0.18 ± 0.04 nM and 0.68 ± 0.09 nM. The data was utilized to construct a preliminary 3D-QSAR/pharmacophore model for use in the design of new and more efficient modulators of D-serine.

  3. Potassium-chloride cotransporter 3 interacts with Vav2 to synchronize the cell volume decrease response with cell protrusion dynamics.

    Directory of Open Access Journals (Sweden)

    Adèle Salin-Cantegrel

    Full Text Available Loss-of-function of the potassium-chloride cotransporter 3 (KCC3 causes hereditary motor and sensory neuropathy with agenesis of the corpus callosum (HMSN/ACC, a severe neurodegenerative disease associated with defective midline crossing of commissural axons in the brain. Conversely, KCC3 over-expression in breast, ovarian and cervical cancer is associated with enhanced tumor cell malignancy and invasiveness. We identified a highly conserved proline-rich sequence within the C-terminus of the cotransporter which when mutated leads to loss of the KCC3-dependent regulatory volume decrease (RVD response in Xenopus Laevis oocytes. Using SH3 domain arrays, we found that this poly-proline motif is a binding site for SH3-domain containing proteins in vitro. This approach identified the guanine nucleotide exchange factor (GEF Vav2 as a candidate partner for KCC3. KCC3/Vav2 physical interaction was confirmed using GST-pull down assays and immuno-based experiments. In cultured cervical cancer cells, KCC3 co-localized with the active form of Vav2 in swelling-induced actin-rich protruding sites and within lamellipodia of spreading and migrating cells. These data provide evidence of a molecular and functional link between the potassium-chloride co-transporters and the Rho GTPase-dependent actin remodeling machinery in RVD, cell spreading and cell protrusion dynamics, thus providing new insights into KCC3's involvement in cancer cell malignancy and in corpus callosum agenesis in HMSN/ACC.

  4. MK615 decreases RAGE expression and inhibits TAGE-induced proliferation in hepatocellular carcinoma cells.

    Science.gov (United States)

    Sakuraoka, Yuhki; Sawada, Tokihiko; Okada, Toshie; Shiraki, Takayuki; Miura, Yoshikazu; Hiraishi, Katsuya; Ohsawa, Tatsushi; Adachi, Masakazu; Takino, Jun-ichi; Takeuchi, Masayoshi; Kubota, Keiichi

    2010-11-14

    To investigate the proliferative effect of advanced glycation end-products (AGEs) and the role of their cellular receptor (RAGE) on hepatocellular carcinoma (HCC) cells, and the inhibitory effects of MK615, an extract from Japanese apricot, against AGEs were also evaluated. Two HCC cell lines, HuH7 and HepG2, were used. Expression of RAGE was investigated by polymerase chain reaction, Western blotting, and flow cytometry (FACS). The effect of MK615 on RAGE expression was also evaluated by FACS. The proliferative effects of a control (unglycated bovine serum albumin), glucose-derived AGEs (Glc-AGE), and glyceraldehyde-derived AGEs (Glycer-AGE), and the anti-proliferative effect of MK615 against AGEs, were evaluated using MTT assays. Expression of RAGE was confirmed at both the mRNA and protein levels in both HuH7 and HepG2. FACS revealed that the level of RAGE expression was higher in HuH7 than in HepG2. Treatment with 0.1 μg/mL MK615 decreased the expression level of RAGE from 24.3% to 3.7% in HuH7 and from 6.2% to 4.8% in HepG2. The growth indices for the control, Glc-AGE, and Glycer-AGE were 1.06 ± 0.08, 0.99 ± 0.04, and 1.38 ± 0.05, respectively, in HuH7 (P = 0.037), and were 1.03 ± 0.04, 1.04 ± 0.03, and 1.07 ± 0.05, respectively, in HepG2 (P > 0.05). When the cells were cultured simultaneously with Glycer-AGE and MK615, MK615 abrogated the proliferative effect of Glycer-AGE in HuH7. Only Glycer-AGE has a proliferative effect on HuH7, which expresses a higher level of RAGE. MK615 suppresses the proliferative effect of Glycer-AGE on HuH7 by decreasing the expression of RAGE.

  5. Bone stroma-derived cells change coregulators recruitment to androgen receptor and decrease cell proliferation in androgen-sensitive and castration-resistant prostate cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Villagran, Marcelo A.; Gutierrez-Castro, Francisco A.; Pantoja, Diego F.; Alarcon, Jose C.; Fariña, Macarena A.; Amigo, Romina F.; Muñoz-Godoy, Natalia A. [Molecular Endocrinology and Oncology Laboratory, University of Concepcion, Concepcion (Chile); Pinilla, Mabel G. [Department of Medical Specialties, School of Medicine, University of Concepcion, Concepcion (Chile); Peña, Eduardo A.; Gonzalez-Chavarria, Ivan; Toledo, Jorge R.; Rivas, Coralia I.; Vera, Juan C. [Department of Physiopathology, School of Biological Sciences, University of Concepcion, Concepcion (Chile); McNerney, Eileen M. [Molecular Endocrinology and Oncology Laboratory, University of Concepcion, Concepcion (Chile); Onate, Sergio A., E-mail: sergio.onate@udec.cl [Molecular Endocrinology and Oncology Laboratory, University of Concepcion, Concepcion (Chile); Department of Medical Specialties, School of Medicine, University of Concepcion, Concepcion (Chile); Department of Urology, State University of New York at Buffalo, NY (United States)

    2015-11-27

    Prostate cancer (CaP) bone metastasis is an early event that remains inactive until later-stage progression. Reduced levels of circulating androgens, due to andropause or androgen deprivation therapies, alter androgen receptor (AR) coactivator expression. Coactivators shift the balance towards enhanced AR-mediated gene transcription that promotes progression to androgen-resistance. Disruptions in coregulators may represent a molecular switch that reactivates latent bone metastasis. Changes in AR-mediated transcription in androgen-sensitive LNCaP and androgen-resistant C4-2 cells were analyzed for AR coregulator recruitment in co-culture with Saos-2 and THP-1. The Saos-2 cell line derived from human osteosarcoma and THP-1 cell line representing human monocytes were used to display osteoblast and osteoclast activity. Increased AR activity in androgen-resistant C4-2 was due to increased AR expression and SRC1/TIF2 recruitment and decreased SMRT/NCoR expression. AR activity in both cell types was decreased over 90% when co-cultured with Saos-2 or THP-1 due to dissociation of AR from the SRC1/TIF2 and SMRT/NCoR coregulators complex, in a ligand-dependent and cell-type specific manner. In the absence of androgens, Saos-2 decreased while THP-1 increased proliferation of LNCaP cells. In contrast, both Saos-2 and THP-1 decreased proliferation of C4-2 in absence and presence of androgens. Global changes in gene expression from both CaP cell lines identified potential cell cycle and androgen regulated genes as mechanisms for changes in cell proliferation and AR-mediated transactivation in the context of bone marrow stroma cells. - Highlights: • Decreased corepressor expression change AR in androgen-resistance prostate cancer. • Bone stroma-derived cells change AR coregulator recruitment in prostate cancer. • Bone stroma cells change cell proliferation in androgen-resistant cancer cells. • Global gene expression in CaP cells is modified by bone stroma cells in co

  6. Bone stroma-derived cells change coregulators recruitment to androgen receptor and decrease cell proliferation in androgen-sensitive and castration-resistant prostate cancer cells

    International Nuclear Information System (INIS)

    Villagran, Marcelo A.; Gutierrez-Castro, Francisco A.; Pantoja, Diego F.; Alarcon, Jose C.; Fariña, Macarena A.; Amigo, Romina F.; Muñoz-Godoy, Natalia A.; Pinilla, Mabel G.; Peña, Eduardo A.; Gonzalez-Chavarria, Ivan; Toledo, Jorge R.; Rivas, Coralia I.; Vera, Juan C.; McNerney, Eileen M.; Onate, Sergio A.

    2015-01-01

    Prostate cancer (CaP) bone metastasis is an early event that remains inactive until later-stage progression. Reduced levels of circulating androgens, due to andropause or androgen deprivation therapies, alter androgen receptor (AR) coactivator expression. Coactivators shift the balance towards enhanced AR-mediated gene transcription that promotes progression to androgen-resistance. Disruptions in coregulators may represent a molecular switch that reactivates latent bone metastasis. Changes in AR-mediated transcription in androgen-sensitive LNCaP and androgen-resistant C4-2 cells were analyzed for AR coregulator recruitment in co-culture with Saos-2 and THP-1. The Saos-2 cell line derived from human osteosarcoma and THP-1 cell line representing human monocytes were used to display osteoblast and osteoclast activity. Increased AR activity in androgen-resistant C4-2 was due to increased AR expression and SRC1/TIF2 recruitment and decreased SMRT/NCoR expression. AR activity in both cell types was decreased over 90% when co-cultured with Saos-2 or THP-1 due to dissociation of AR from the SRC1/TIF2 and SMRT/NCoR coregulators complex, in a ligand-dependent and cell-type specific manner. In the absence of androgens, Saos-2 decreased while THP-1 increased proliferation of LNCaP cells. In contrast, both Saos-2 and THP-1 decreased proliferation of C4-2 in absence and presence of androgens. Global changes in gene expression from both CaP cell lines identified potential cell cycle and androgen regulated genes as mechanisms for changes in cell proliferation and AR-mediated transactivation in the context of bone marrow stroma cells. - Highlights: • Decreased corepressor expression change AR in androgen-resistance prostate cancer. • Bone stroma-derived cells change AR coregulator recruitment in prostate cancer. • Bone stroma cells change cell proliferation in androgen-resistant cancer cells. • Global gene expression in CaP cells is modified by bone stroma cells in co

  7. Decreased level of phosphatidylcholine (16:0/20:4) in multiple myeloma cells compared to plasma cells: a single-cell MALDI-IMS approach.

    Science.gov (United States)

    Hossen, Md Amir; Nagata, Yasuyuki; Waki, Michihiko; Ide, Yoshimi; Takei, Shiro; Fukano, Hana; Romero-Perez, Gustavo A; Tajima, Shogo; Yao, Ikuko; Ohnishi, Kazunori; Setou, Mitsutoshi

    2015-07-01

    Lipid metabolic changes under diseased conditions, particularly in solid tumors, are attracting increased attention. However, in non-solid tumors, including most hematopoietic tumors, lipid analyses are scarce. Multiple myeloma (MM) is a plasma cell disorder arising from bone marrow, and the lipid status of MM cells has not been reported yet. In this study, we analyzed flow cytometry-sorted single MM cells and normal plasma cells (NPCs) using matrix-assisted laser desorption/ionization-imaging mass spectrometry (MALDI-IMS), a two-dimensional label-free mass spectrometry technique for biomolecular analysis, to obtain specific lipid information. We isolated 1.31-5.77% of MM cells and 0.03-0.24% of NPCs using fluorescence-activated cell sorting (FACS). Analysis of purified cells using MALDI-IMS at the single-cell level revealed that the peak intensity and ion signals of phosphatidylcholine [PC (16:0/20:4) + H](+) at m/z 782.5 were significantly decreased in MM cells compared to NPCs. By examining particular cell populations rather than cell mixtures, our method can become a suitable tool for the analysis of rare cell populations at the single-cell level and advance the understanding of MM progression.

  8. Decreased proinflammatory cytokine production by peripheral blood mononuclear cells from vitiligo patients following aspirin treatment

    International Nuclear Information System (INIS)

    Zailaie, Mohammad Z.

    2005-01-01

    Limited studies have shown that treatment of cells with aspirin modulates their cytokine production. Consequently, the aim of the present study is to investigate the pattern of important proinflammatory cytokines production by stimulated peripheral blood mononuclear cells (PBMC) from patients with active vitiligo following long-term treatment with low-dose oral aspirin. The study was conducted at the Vitiligo Unit, King Abdul-Aziz University Medical Center, Jeddah, Kingdom of Saudi Arabia between March and October 2003. Thirty-two patients (18 females and 14 males) with non-segmental vitiligo were divided into 2 equal groups, one group received a daily single dose of oral aspirin (300 mg) and the other group received placebo for a period of 12 weeks. The concentrations of interleukin (IL)-1beta, IL-6, IL-8 and tumor necrosis factor-alpha (TNF-alpha) were determined in the supernatant of isolated cultured PMBC after being stimulated with bacterial lipopolysaccharide (LPS), before the start of aspirin treatment and at end of treatment period. Cytokine levels were measured using the quantitative sandwich enzyme-linked immunosorbent assay (ELISA) technique, utilizing commercially available kits. The proinflammatory cytokine production by the PBMC of patients with active vitiligo was significantly increased compared to normal controls. Thus, the relative percentage increase in the production of IL-1beta, IL-6, IL-8 and TNF-alpha was: 39.4%, 110.5% (p<0.05), 91.5% (p<0.01), and 37% (p<0.05). At the end of treatment, proinflammatory cytokine production in the aspirin-treated group of active vitiligo patients was significantly decreased compared to the placebo group. Thus, the relative percentage decrease in the production of IL-1beta IL-6, IL-8 and TNF-alpha was: 42.5%, 45.2% (p<0.05), 30.8% (p<0.01), and 50.6% (p<0.05). The vitiligo activity was arrested in all aspirin-treated patients, while 2 patients demonstrated significant repigmentation.Chronic administration of

  9. Decreased Inflammatory Responses of Human Lung Epithelial Cells after Ethanol Exposure Are Mimicked by Ethyl Pyruvate

    Directory of Open Access Journals (Sweden)

    B. Relja

    2014-01-01

    Full Text Available Background and Purpose. Leukocyte migration into alveolar space plays a critical role in pulmonary inflammation resulting in lung injury. Acute ethanol (EtOH exposure exerts anti-inflammatory effects. The clinical use of EtOH is critical due to its side effects. Here, we compared effects of EtOH and ethyl pyruvate (EtP on neutrophil adhesion and activation of cultured alveolar epithelial cells (A549. Experimental Approach. Time course and dose-dependent release of interleukin- (IL- 6 and IL-8 from A549 were measured after pretreatment of A549 with EtP (2.5–10 mM, sodium pyruvate (NaP, 10 mM, or EtOH (85–170 mM, and subsequent lipopolysaccharide or IL-1beta stimulation. Neutrophil adhesion to pretreated and stimulated A549 monolayers and CD54 surface expression were determined. Key Results. Treating A549 with EtOH or EtP reduced substantially the cytokine-induced release of IL-8 and IL-6. EtOH and EtP (but not NaP reduced the adhesion of neutrophils to monolayers in a dose- and time-dependent fashion. CD54 expression on A549 decreased after EtOH or EtP treatment before IL-1beta stimulation. Conclusions and Implications. EtP reduces secretory and adhesive potential of lung epithelial cells under inflammatory conditions. These findings suggest EtP as a potential treatment alternative that mimics the anti-inflammatory effects of EtOH in early inflammatory response in lungs.

  10. Piperlongumine decreases cell proliferation and the expression of cell cycle-associated proteins by inhibiting Akt pathway in human lung cancer cells.

    Science.gov (United States)

    Seok, Jin Sil; Jeong, Chang Hee; Petriello, Michael C; Seo, Han Geuk; Yoo, Hyunjin; Hong, Kwonho; Han, Sung Gu

    2018-01-01

    Piperlongumine (PL) is an alkaloid of a pepper plant found in Southeast Asia. PL is known to induce selective toxicity towards a variety of cancer cell types. To explore the possible anti-lung cancer effects of PL, A549 cells were treated with PL (0-40 μM) for 24 h. Alterations in the expression of cell cycle-associated proteins (cyclin D1, cyclin-dependent kinase 4 (CDK4), CDK6 and retinoblastoma (Rb)) and intracellular signaling molecules (extracellular signal receptor-activated kinase 1/2 (ERK1/2), Akt, p38 and nuclear factor-κB (NF-κB)) were examined in cells following treatment of PL using Western blot analysis. Results showed that proliferation of cells were significantly decreased by PL in a dose-dependent manner. Flow cytometry results demonstrated increased number of cells in G1 phase in PL (40 μM)-treated group. Reactive oxygen species was significantly increased in cells treated with PL at 20-40 μM. The expression of cyclin D1, CDK4, CDK6 and p-Rb were markedly decreased in cells treated with PL at 40 μM. Treatment of cells with PL suppressed phosphorylation of Akt but increased ERK1/2 phosphorylation. Treatment of PL significantly decreased nuclear translocation of NF-κB p65 in cells. These results suggest that PL possesses antiproliferative properties in A549 cells. Copyright © 2017 Elsevier Ltd. All rights reserved.

  11. Autistic Children Exhibit Decreased Levels of Essential Fatty Acids in Red Blood Cells

    Directory of Open Access Journals (Sweden)

    Sarah A. Brigandi

    2015-05-01

    Full Text Available Omega-6 (n-6 and omega-3 (n-3 polyunsaturated fatty acids (PUFA are essential nutrients for brain development and function. However, whether or not the levels of these fatty acids are altered in individuals with autism remains debatable. In this study, we compared the fatty acid contents between 121 autistic patients and 110 non-autistic, non-developmentally delayed controls, aged 3–17. Analysis of the fatty acid composition of red blood cell (RBC membrane phospholipids showed that the percentage of total PUFA was lower in autistic patients than in controls; levels of n-6 arachidonic acid (AA and n-3 docosahexaenoic acid (DHA were particularly decreased (p < 0.001. In addition, plasma levels of the pro-inflammatory AA metabolite prostaglandin E2 (PGE2 were higher in a subset of the autistic participants (n = 20 compared to controls. Our study demonstrates an alteration in the PUFA profile and increased production of a PUFA-derived metabolite in autistic patients, supporting the hypothesis that abnormal lipid metabolism is implicated in autism.

  12. Pantoprazole Decreases Cell Viability and Function of Human Osteoclasts In Vitro

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    Markus Prause

    2015-01-01

    Full Text Available Proton pump inhibitors (PPIs are commonly prescribed drugs that decrease stomach acidity and are thus often used to treat gastroesophageal reflux disease and as a preventative agent for the adverse effects of nonsteroidal anti-inflammatory drugs on the stomach mucosa. In recently published literature, an association between proton pump inhibitor administration and increased fracture risk has been stated. In order to reveal the underlying pathomechanisms of these observations, the effects of pantoprazole, a representative of the proton pump inhibitors, on human osteoclasts in vitro were evaluated in this study. Osteoclasts were stimulated with increasing concentrations of pantoprazole ranging from 0 μg/mL to 10 μg/mL over a period of seven days. Cell viability and tartrate-resistant acid phosphatase (TRAP activity assays were performed after 1 day, 3 days, and 7 days, respectively. Here, stimulated osteoclasts presented a significantly lower viability and TRAP activity than the negative controls. Osteoclast-specific gene expression was evaluated after seven days and revealed no significant differences between all samples. Overall, the bone degrading and resorptive function of osteoclasts is inhibited by the administration of proton pump inhibitors. While PPI-related fractures through “basic multicellular unit” dysfunction are unlikely, the underlying pathomechanism remains unknown.

  13. Histone deacetylase 3 depletion in osteo/chondroprogenitor cells decreases bone density and increases marrow fat.

    Directory of Open Access Journals (Sweden)

    David F Razidlo

    2010-07-01

    Full Text Available Histone deacetylase (Hdac3 is a nuclear enzyme that contributes to epigenetic programming and is required for embryonic development. To determine the role of Hdac3 in bone formation, we crossed mice harboring loxP sites around exon 7 of Hdac3 with mice expressing Cre recombinase under the control of the osterix promoter. The resulting Hdac3 conditional knockout (CKO mice were runted and had severe deficits in intramembranous and endochondral bone formation. Calvarial bones were significantly thinner and trabecular bone volume in the distal femur was decreased 75% in the Hdac3 CKO mice due to a substantial reduction in trabecular number. Hdac3-CKO mice had fewer osteoblasts and more bone marrow adipocytes as a proportion of tissue area than their wildtype or heterozygous littermates. Bone formation rates were depressed in both the cortical and trabecular regions of Hdac3 CKO femurs. Microarray analyses revealed that numerous developmental signaling pathways were affected by Hdac3-deficiency. Thus, Hdac3 depletion in osterix-expressing progenitor cells interferes with bone formation and promotes bone marrow adipocyte differentiation. These results demonstrate that Hdac3 inhibition is detrimental to skeletal health.

  14. Nickel decreases cellular iron level and converts cytosolic aconitase to iron-regulatory protein 1 in A549 cells

    International Nuclear Information System (INIS)

    Chen Haobin; Davidson, Todd; Singleton, Steven; Garrick, Michael D.; Costa, Max

    2005-01-01

    Nickel (Ni) compounds are well-established carcinogens and are known to initiate a hypoxic response in cells via the stabilization and transactivation of hypoxia-inducible factor-1 alpha (HIF-1α). This change may be the consequence of nickel's interference with the function of several Fe(II)-dependent enzymes. In this study, the effects of soluble nickel exposure on cellular iron homeostasis were investigated. Nickel treatment decreased both mitochondrial and cytosolic aconitase (c-aconitase) activity in A549 cells. Cytosolic aconitase was converted to iron-regulatory protein 1, a form critical for the regulation of cellular iron homeostasis. The increased activity of iron-regulatory protein 1 after nickel exposure stabilized and increased transferrin receptor (Tfr) mRNA and antagonized the iron-induced ferritin light chain protein synthesis. The decrease of aconitase activity after nickel treatment reflected neither direct interference with aconitase function nor obstruction of [4Fe-4S] cluster reconstitution by nickel. Exposure of A549 cells to soluble nickel decreased total cellular iron by about 40%, a decrease that likely caused the observed decrease in aconitase activity and the increase of iron-regulatory protein 1 activity. Iron treatment reversed the effect of nickel on cytosolic aconitase and iron-regulatory protein 1. To assess the mechanism for the observed effects, human embryonic kidney (HEK) cells over expressing divalent metal transporter-1 (DMT1) were compared to A549 cells expressing only endogenous transporters for inhibition of iron uptake by nickel. The inhibition data suggest that nickel can enter via DMT1 and compete with iron for entry into the cell. This disturbance of cellular iron homeostasis by nickel may have a great impact on the ability of the cell to regulate a variety of cell functions, as well as create a state of hypoxia in cells under normal oxygen tension. These effects may be very important in how nickel exerts phenotypic

  15. Increased circulating follicular helper T cells with decreased programmed death-1 in chronic renal allograft rejection

    OpenAIRE

    Shi, Jian; Luo, Fengbao; Shi, Qianqian; Xu, Xianlin; He, Xiaozhou; Xia, Ying

    2015-01-01

    Background Chronic antibody-mediated rejection is a major issue that affects long-term renal allograft survival. Since follicular helper T (Tfh) cells promote the development of antigen-specific B cells in alloimmune responses, we investigated the potential roles of Tfh cells, B cells and their alloimmune-regulating molecules in the pathogenesis of chronic renal allograft rejection in this study. Methods The frequency of Tfh, B cells and the levels of their alloimmune-regulating molecules inc...

  16. HIV-1 transgenic rat CD4+ T cells develop decreased CD28 responsiveness and suboptimal Lck tyrosine dephosphorylation following activation

    International Nuclear Information System (INIS)

    Yadav, Anjana; Pati, Shibani; Nyugen, Anhthu; Barabitskaja, Oxana; Mondal, Prosanta; Anderson, Michael; Gallo, Robert C.; Huso, David L.; Reid, William

    2006-01-01

    Impaired CD4+ T cell responses, resulting in dysregulated T-helper 1 (Th1) effector and memory responses, are a common result of HIV-1 infection. These defects are often preceded by decreased expression and function of the α/β T cell receptor (TCR)-CD3 complex and of co-stimulatory molecules including CD28, resulting in altered T cell proliferation, cytokine secretion and cell survival. We have previously shown that HIV Tg rats have defective development of T cell effector function and generation of specific effector/memory T cell subsets. Here we identify abnormalities in activated HIV-1 Tg rat CD4+ T cells that include decreased pY505 dephosphorylation of Lck (required for Lck activation), decreased CD28 function, reduced expression of the anti-apoptotic molecule Bcl-xL, decreased secretion of the mitogenic lympokine interleukin-2 (IL-2) and increased activation induced apoptosis. These events likely lead to defects in antigen-specific signaling and may help explain the disruption of Th1 responses and the generation of specific effector/memory subsets in transgenic CD4+ T cells

  17. Function of Treg Cells Decreased in Patients With Systemic Lupus Erythematosus Due To the Effect of Prolactin.

    Science.gov (United States)

    Legorreta-Haquet, María Victoria; Chávez-Rueda, Karina; Chávez-Sánchez, Luis; Cervera-Castillo, Hernando; Zenteno-Galindo, Edgar; Barile-Fabris, Leonor; Burgos-Vargas, Rubén; Álvarez-Hernández, Everardo; Blanco-Favela, Francisco

    2016-02-01

    Prolactin has different functions, including cytokine secretion and inhibition of the suppressor effect of regulatory T (Treg) cells in healthy individuals. Systemic lupus erythematosus (SLE) is an autoimmune disease characterized by defects in the functions of B, T, and Treg cells. Prolactin plays an important role in the physiopathology of SLE. Our objective was to establish the participation of prolactin in the regulation of the immune response mediated by Treg cells from patients with SLE. CD4CD25CD127 cells were purified using magnetic beads and the relative expression of prolactin receptor was measured. The functional activity was evaluated by proliferation assay and cytokine secretion in activated cells, in the presence and absence of prolactin. We found that both percentage and function of Treg cells decrease in SLE patients compared to healthy individuals with statistical significance. The prolactin receptor is constitutively expressed on Treg and effector T (Teff) cells in SLE patients, and this expression is higher than in healthy individuals. The expression of this receptor differs in inactive and active patients: in the former, the expression is higher in Treg cells than in Teff cells, similar to healthy individuals, whereas there is no difference in the expression between Treg and Teff cells from active patients. In Treg:Teff cell cocultures, addition of prolactin decreases the suppressor effect exerted by Treg cells and increases IFNγ secretion. Our results suggest that prolactin plays an important role in the activation of the disease in inactive patients by decreasing the suppressor function exerted by Treg cells over Teff cells, thereby favoring an inflammatory microenvironment.

  18. Mast cells infiltration and decreased E-cadherin expression in ketamine-induced cystitis

    Directory of Open Access Journals (Sweden)

    Mengqiang Li

    2015-01-01

    Conclusions: Increased mast cells in bladder wall and downregulated expression of E-cadherin junction protein in epithelial cells were probably associated with interstitial inflammation and fissures in mucosa. It implied that ketamine induced an interstitial cystitis.

  19. Individual Amino Acid Supplementation Can Improve Energy Metabolism and Decrease ROS Production in Neuronal Cells Overexpressing Alpha-Synuclein.

    Science.gov (United States)

    Delic, Vedad; Griffin, Jeddidiah W D; Zivkovic, Sandra; Zhang, Yumeng; Phan, Tam-Anh; Gong, Henry; Chaput, Dale; Reynes, Christian; Dinh, Vinh B; Cruz, Josean; Cvitkovic, Eni; Placides, Devon; Frederic, Ernide; Mirzaei, Hamed; Stevens, Stanley M; Jinwal, Umesh; Lee, Daniel C; Bradshaw, Patrick C

    2017-09-01

    Parkinson's disease (PD) is a neurodegenerative disorder characterized by alpha-synuclein accumulation and loss of dopaminergic neurons in the substantia nigra (SN) region of the brain. Increased levels of alpha-synuclein have been shown to result in loss of mitochondrial electron transport chain complex I activity leading to increased reactive oxygen species (ROS) production. WT alpha-synuclein was stably overexpressed in human BE(2)-M17 neuroblastoma cells resulting in increased levels of an alpha-synuclein multimer, but no increase in alpha-synuclein monomer levels. Oxygen consumption was decreased by alpha-synuclein overexpression, but ATP levels did not decrease and ROS levels did not increase. Treatment with ferrous sulfate, a ROS generator, resulted in decreased oxygen consumption in both control and alpha-synuclein overexpressing cells. However, this treatment only decreased ATP levels and increased ROS production in the cells overexpressing alpha-synuclein. Similarly, paraquat, another ROS generator, decreased ATP levels in the alpha-synuclein overexpressing cells, but not in the control cells, further demonstrating how alpha-synuclein sensitized the cells to oxidative insult. Proteomic analysis yielded molecular insights into the cellular adaptations to alpha-synuclein overexpression, such as the increased abundance of many mitochondrial proteins. Many amino acids and citric acid cycle intermediates and their ester forms were individually supplemented to the cells with L-serine, L-proline, L-aspartate, or L-glutamine decreasing ROS production in oxidatively stressed alpha-synuclein overexpressing cells, while diethyl oxaloacetate or L-valine supplementation increased ATP levels. These results suggest that dietary supplementation with individual metabolites could yield bioenergetic improvements in PD patients to delay loss of dopaminergic neurons.

  20. Dendritic cells decreased the concomitant expanded Tregs and Tregs related IL-35 in cytokine-induced killer cells and increased their cytotoxicity against leukemia cells.

    Directory of Open Access Journals (Sweden)

    Ying Pan

    Full Text Available Regulatory T cells (Tregs are potent immunosuppressive cells and essential for inducing immune tolerance. Recent studies have reported that Tregs and Tregs related cytokines can inhibit the antitumor activity of cytokine-induced killer (CIK cells, but dendritic cells co-cultured CIK (DC-CIK cells can be used for induction of a specific immune response by blocking of Tregs and TGF-β, IL-10. As a novel identified cytokine, IL-35 is specially produced by Tregs and plays an essential role in immune regulation. However, it remains unknown whether IL-35 roles in tumor immunotherapy mediated by CIK and DC-CIK cells. In this study, we cultured CIK and DC-CIK cells from the same healthy adult samples, and investigated their phenotype, proliferation, cytotoxic activity against leukemia cell lines K562 and NB4 by FCM and CCK-8, measured IL-35, TGF-β and IL-10 protein by ELISA, detected Foxp3, IL-35 and IL-35 receptor mRNA by Real-time PCR, respectively. We found Tregs and IL-35 concomitantly expanded by a time-dependent way during the generation of CIK cells, but DC significantly down-regulated the expression of them and simultaneously up-regulated the proliferation ability as well as cytotoxic activity of CIK cells against leukemia cell lines. Therefore, our data suggested that DC decreased concomitant expanded Tregs and Tregs related IL-35 in CIK cells and might contribute to improve their cytotoxicity against leukemia cells in vitro.

  1. Prion protein-deficient mice exhibit decreased CD4 T and LTi cell numbers and impaired spleen structure.

    Science.gov (United States)

    Kim, Soochan; Han, Sinsuk; Lee, Ye Eun; Jung, Woong-Jae; Lee, Hyung Soo; Kim, Yong-Sun; Choi, Eun-Kyoung; Kim, Mi-Yeon

    2016-01-01

    The cellular prion protein is expressed in almost all tissues, including the central nervous system and lymphoid tissues. To investigate the effects of the prion protein in lymphoid cells and spleen structure formation, we used prion protein-deficient (Prnp(0/0)) Zürich I mice generated by inactivation of the Prnp gene. Prnp(0/0) mice had decreased lymphocytes, in particular, CD4 T cells and lymphoid tissue inducer (LTi) cells. Decreased CD4 T cells resulted from impaired expression of CCL19 and CCL21 in the spleen rather than altered chemokine receptor CCR7 expression. Importantly, some of the white pulp regions in spleens from Prnp(0/0) mice displayed impaired T zone structure as a result of decreased LTi cell numbers and altered expression of the lymphoid tissue-organizing genes lymphotoxin-α and CXCR5, although expression of the lymphatic marker podoplanin and CXCL13 by stromal cells was not affected. In addition, CD3(-)CD4(+)IL-7Rα(+) LTi cells were rarely detected in impaired white pulp in spleens of these mice. These data suggest that the prion protein is required to form the splenic white pulp structure and for development of normal levels of CD4 T and LTi cells. Copyright © 2015. Published by Elsevier GmbH.

  2. Smoking decreases the level of circulating CD34+ progenitor cells in young healthy women - a pilot study

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    Baumann Gert

    2010-05-01

    Full Text Available Abstract Background Decreased levels of circulating bone marrow-derived progenitor cells have been associated with risk factors and cardiovascular diseases. Smoking is the most important modifiable risk factor for atherosclerosis in young women. The aim of this pilot study was to assess in healthy premenopausal women without other risk factors for cardiovascular disease the influence of nicotine abuse on the number of circulating progenitor cells in relation to endothelial function. Methods The number of endothelial progenitor cells, measured as colony-forming units in a cell-culture assay (EPC-CFU and the number of circulating CD34 + and CD34 + /CD133 + cells, measured by flow cytometry, was estimated in 32 women at the menstrual phase of the menstrual cycle. In addition, flow-mediated dilation (FMD was assessed as a marker for vascular function. In a subgroup of these women (n = 20, progenitor cells were also investigated at the mid-follicular and luteal phases of the menstrual cycle. Results Compared to non-smokers, the abundance of circulating CD34 + cells was significantly lower in smoking women in the menstrual, mid-luteal, and mid-follicular phases of the menstrual cycle. The number of CD34 + progenitor cells was revealed to have significant positive correlation with FMD in young healthy women, whereas CD34 + /CD133 + progenitor cells and EPC-CFU showed no significant correlation. Conclusion The number of CD34 + progenitor cells positively correlates with FMD in young healthy women and is decreased by smoking.

  3. B cell dysfunction after bone marrow transplantation is associated with decreased Ca2+ flux upon membrane Ig crosslinking.

    Science.gov (United States)

    Storek, J; Hultin, L E; Ferrara, S; Ku, N; Giorgi, J V; Champlin, R E; Saxon, A

    1994-08-01

    Patients undergoing bone marrow transplantation have a long-lasting defect of B cell-mediated immunity. Both quantitative (decreased blood B cell counts) and qualitative (decreased Ig production) abnormalities of B cells have been described. To better understand the mechanism of the qualitative defect and its potential relation to B cell immaturity, we studied the in vitro responsiveness of B cells to polyclonal stimuli in patients at 2-12 months post-transplant and in normal neonates. Several key steps of the B cell program were deficient in the patients while they were relatively normal in the neonates. These included (i) early activation as assessed by Ca2+ flux; (ii) late activation as assessed by the increase in cell size and upregulation of the activation antigens CD25 and CD71; and (iii) proliferation as assessed by the number of cycling cells after stimulation. We conclude that the functional B cell defect during the early (< 1 year) post-transplant period extends back to the level of early activation and cannot be simply attributed to the relative immaturity of post-transplant B cells.

  4. High levels of SOX5 decrease proliferative capacity of human B cells, but permit plasmablast differentiation.

    Directory of Open Access Journals (Sweden)

    Mirzokhid Rakhmanov

    Full Text Available Currently very little is known about the differential expression and function of the transcription factor SOX5 during B cell maturation. We identified two new splice variants of SOX5 in human B cells, encoding the known L-SOX5B isoform and a new shorter isoform L-SOX5F. The SOX5 transcripts are highly expressed during late stages of B-cell differentiation, including atypical memory B cells, activated CD21low B cells and germinal center B cells of tonsils. In tonsillar sections SOX5 expression was predominantly polarized to centrocytes within the light zone. After in vitro stimulation, SOX5 expression was down-regulated during proliferation while high expression levels were permissible for plasmablast differentiation. Overexpression of L-SOX5F in human primary B lymphocytes resulted in reduced proliferation, less survival of CD138neg B cells, but comparable numbers of CD138+CD38hi plasmablasts compared to control cells. Thus, our findings describe for the first time a functional role of SOX5 during late B cell development reducing the proliferative capacity and thus potentially affecting the differentiation of B cells during the germinal center response.

  5. Increased intracellular Ca2+decreases cisplatin resistance by regulating iNOS expression in human ovarian cancer cells.

    Science.gov (United States)

    Yu, Yang; Xie, Qi; Liu, Weimin; Guo, Yuting; Xu, Na; Xu, Lu; Liu, Shibing; Li, Songyan; Xu, Ye; Sun, Liankun

    2017-02-01

    Previous studies have reported that intracellular Ca 2+ signals and inducible nitric oxide synthase (iNOS) are involved in cell apoptosis. However, the role of iNOS in cisplatin resistance in ovarian cancer remains unclear. Here, we demonstrate that SKOV3/DDP ovarian cancer cells were more resistant to cisplatin than were SKOV3 ovarian cancer cells. The expression of intracellular Ca 2+ and iNOS was more strongly induced by cisplatin in SKOV3 cells than in SKOV3/DDP cells. TAT-conjugated IP3R-derived peptide (TAT-IDP S ) increased cisplatin-induced iNOS expression and apoptosis in SKOV3/DDP cells. 2-Aminoethoxydiphenyl borate (2-APB) decreased cisplatin-induced iNOS expression and apoptosis in SKOV3 cells. Thus, iNOS induction may be a valuable strategy for improving the anti-tumor efficacy of cisplatin in ovarian cancer. Copyright © 2016 Elsevier Masson SAS. All rights reserved.

  6. NKT cell activation by local α-galactosylceramide administration decreases susceptibility to HSV-2 infection

    DEFF Research Database (Denmark)

    Iversen, Marie Beck; Jensen, Simon Kok; Hansen, Anne Louise

    2015-01-01

    NKT cells are a subgroup of T cells, which express a restricted TCR repertoire and are critical for the innate immune responses to viral infections. Activation of NKT cells depends on the major histocompatibility complex-related molecule CD1d, which presents bioactive lipids to NKT cells....... The marine sponge derived lipid αGalCer has recently been demonstrated as a specific agonist for activation of human and murine NKT cells. In the present study we investigated the applicability of αGalCer pre-treatment for immune protection against intra-vaginal HSV-2 infection. We found that C57BL/6 WT mice...... in vaginal tissue and elevated levels of IFN-γ in the vaginal tissue and in vaginal fluids 24h after αGalCer pre-treatment. Collectively our data demonstrate a protective effect of αGalCer induced activation of NKT cells in the innate immune protection against viral infection....

  7. Aging is associated with decreased maximal life span and accelerated senescence of bone marrow stromal cells

    DEFF Research Database (Denmark)

    Dokkedahl, Karin Stenderup; Justesen, Jeannette; Clausen, Christian

    2003-01-01

    +/- 11 population doublings [PD] vs 41 +/- 10 PD, P beta-galactosidase positive (SA beta......-gal+) cells and mean telomere length in early-passage cells obtained from young and old donors. However, MSC from old donors exhibited accelerated senescence evidenced by increased number of SA beta-gal+ cells per PD as compared with young (4% per PD vs 0.4% per PD, respectively). MSC from young and old...

  8. Shading Contributes to the Reduction of Stem Mechanical Strength by Decreasing Cell Wall Synthesis in Japonica Rice (Oryza sativa L.

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    Longmei Wu

    2017-05-01

    Full Text Available Low solar radiation caused by industrial development and solar dimming has become a limitation in crop production in China. It is widely accepted that low solar radiation influences many aspects of plant development, including slender, weak stems and susceptibility to lodging. However, the underlying mechanisms are not well understood. To clarify how low solar radiation affects stem mechanical strength formation and lodging resistance, the japonica rice cultivars Wuyunjing23 (lodging-resistant and W3668 (lodging-susceptible were grown under field conditions with normal light (Control and shading (the incident light was reduced by 60% with a black nylon net. The yield and yield components, plant morphological characteristics, the stem mechanical strength, cell wall components, culm microstructure, gene expression correlated with cellulose and lignin biosynthesis were measured. The results showed that shading significantly reduced grain yield attributed to reduction of spikelets per panicles and grain weight. The stem-breaking strength decreased significantly under shading treatment; consequently, resulting in higher lodging index in rice plant in both varieties, as revealed by decreased by culm diameter, culm wall thickness and increased plant height, gravity center height. Compared with control, cell wall components including non-structural carbohydrate, sucrose, cellulose, and lignin reduced quite higher. With histochemical straining, shading largely reduced lignin deposition in the sclerenchyma cells and vascular bundle cells compared with control, and decreased cellulose deposition in the parenchyma cells of culm tissue in both Wuyunjing23 and W3668. And under shading condition, gene expression involved in secondary cell wall synthesis, OsPAL, OsCOMT, OsCCoAOMT, OsCCR, and OsCAD2, and primary cell wall synthesis, OsCesA1, OsCesA3, and OsCesA8 were decreased significantly. These results suggest that gene expression involved in the reduction of

  9. 17β-Estradiol treatment inhibits breast cell proliferation, migration and invasion by decreasing MALAT-1 RNA level

    Energy Technology Data Exchange (ETDEWEB)

    Zhao, Ziyi [Key Laboratory of Bioresources and Ecoenvironment (Ministry of Education), College of Life Sciences, Sichuan University, Chengdu 610041 (China); Chen, Changjin [Department of Pediatrics, West China Second University Hospital, Sichuan University, Chengdu 610041 (China); Liu, Yu [Key Laboratory of Bioresources and Ecoenvironment (Ministry of Education), College of Life Sciences, Sichuan University, Chengdu 610041 (China); Wu, Chuanfang, E-mail: 879413966@qq.com [Key Laboratory of Bioresources and Ecoenvironment (Ministry of Education), College of Life Sciences, Sichuan University, Chengdu 610041 (China)

    2014-03-07

    Highlights: • E2 affects not only estrogen-receptor α positive breast cells but also negative ones. • 100 nM E2 treatment affects breast cells proliferation, migration. • 100 nM E2 treatment functions in an estrogen-receptor α-independent way. • E2 treatment decreases MALAT-1 RNA level by post-transcriptional regulation. - Abstract: Breast cancer cells, which express estrogen receptor α (ERα), respond to estrogen in a concentration dependent fashion, resulting in proliferation or apoptosis. But breast cancer cells without ERα show no effect on low concentration of estrogen treatment. Proliferation, migration and invasion of MCF10a, MCF7 and MB231 cells treated with low (1 nM) or high (100 nM) dose of 17β-Estradiol (E2) was performed. We identified the effects of E2 on these breast cell lines, and looked for the difference in the presence and absence of ERα. Specifically, we looked for the changes of long non-coding RNA metastasis associated lung adenocarcinoma transcript 1 (MALAT-1), which is found extensively and highly expressed in several kinds of tumor cells, including breast carcinoma. It was observed that proliferation, migration and invasion of breast cells were greatly affected by high concentration E2 treatment and were not affected by low concentration E2 treatment in an ERα independent way. We found that the high concentration E2 treatment largely decreased MALAT-1 RNA level. Interestingly, MALAT-1 decreasing by knocking down showed similar effects on proliferation, migration and invasion. E2 treatment affects breast tumor or non-tumor cells proliferation, migration and invasion in an ERα -independent, but a dose-dependent way by decreasing the MALAT-1 RNA level.

  10. Decreased Intracellular pH Induced by Cariporide Differentially Contributes to Human Umbilical Cord-Derived Mesenchymal Stem Cells Differentiation

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    Wei Gao

    2014-01-01

    Full Text Available Background/Aims: Na+/H+ exchanger 1 (NHE1 is an important regulator of intracellular pH (pHi. High pHi is required for cell proliferation and differentiation. Our previous study has proven that the pHi of mesenchymal stem cells is higher than that of normal differentiated cells and similar to tumor cells. NHE1 is highly expressed in both mesenchymal stem cells and tumor cells. Targeted inhibition of NHE1 could induce differentiation of K562 leukemia cells. In the present paper we explored whether inhibition of NHE1 could induce differentiation of mesenchymal stem cells. Methods: MSCs were obtained from human umbilical cord and both the surface phenotype and functional characteristics were analyzed. Selective NHE1 inhibitor cariporide was used to treat human umbilical cord-derived mesenchymal stem cells (hUC-MSCs. The pHi and the differentiation of hUC-MSCs were compared upon cariporide treatment. The putative signaling pathway involved was also explored. Results: The pHi of hUC-MSCs was decreased upon cariporide treatment. Cariporide up-regulated the osteogenic differentiation of hUC-MSCs while the adipogenic differentiation was not affected. For osteogenic differentiation, β-catenin expression was up-regulated upon cariporide treatment. Conclusion: Decreased pHi induced by cariporide differentially contributes to hUC-MSCs differentiation.

  11. Lovastatin Decreases the Expression of CD133 and Influences the Differentiation Potential of Human Embryonic Stem Cells

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    Ade Kallas-Kivi

    2016-01-01

    Full Text Available The lipophilic statin lovastatin decreases cholesterol synthesis and is a safe and effective treatment for the prevention of cardiovascular diseases. Growing evidence points at antitumor potential of lovastatin. Therefore, understanding the molecular mechanism of lovastatin function in different cell types is critical to effective therapy design. In this study, we investigated the effects of lovastatin on the differentiation potential of human embryonic stem (hES cells (H9 cell line. Multiparameter flow cytometric assay was used to detect changes in the expression of transcription factors characteristic of hES cells. We found that lovastatin treatment delayed NANOG downregulation during ectodermal and endodermal differentiation. Likewise, expression of ectodermal (SOX1 and OTX2 and endodermal (GATA4 and FOXA2 markers was higher in treated cells. Exposure of hES cells to lovastatin led to a minor decrease in the expression of SSEA-3 and a significant reduction in CD133 expression. Treated cells also formed fewer embryoid bodies than control cells. By analyzing hES with and without CD133, we discovered that CD133 expression is required for proper formation of embryoid bodies. In conclusion, lovastatin reduced the heterogeneity of hES cells and impaired their differentiation potential.

  12. Strenuous exercise decreases the percentage of type 1 T cells in the circulation

    DEFF Research Database (Denmark)

    Steensberg, A; Toft, A D; Bruunsgaard, H

    2001-01-01

    Prolonged strenuous exercise is followed by a temporary functional immune impairment. Low numbers of CD4+ T helper (Th) and CD8+ T cytotoxic (Tc) cells are found in the circulation. These cells can be divided according to their cytokine profile into type 1 (Th1 and Tc1), which produce interferon...

  13. Active Prompting to Decrease Cell Phone Use and Increase Seat Belt Use while Driving

    Science.gov (United States)

    Clayton, Michael; Helms, Bridgett; Simpson, Cathy

    2006-01-01

    Automobile crashes are the leading cause of death for those aged 3 to 33, with 43,005 (118 per day) Americans killed in 2002 alone. Seat belt use reduces the risk of serious injury in an accident, and refraining from using a cell phone while driving reduces the risk of an accident. Cell phone use while driving increases accident rates, and leads…

  14. Ubiquitin-specific protease 2 decreases p53-dependent apoptosis in cutaneous T-cell lymphoma

    DEFF Research Database (Denmark)

    Wei, Tianling; Biskup, Edyta; Gjerdrum, Lise Mette Rahbek

    2016-01-01

    Treatment of advanced cutaneous T-cell lymphomas (CTCL) is challenging because they are resistant to conventional chemotherapy. USP2 has been shown to promote resistance to chemotherapeutic agents in several cancer models.We show here USP2 is expressed in quiescent and activated T-cells and its e...

  15. Early posthatch starvation decreases satellite cell proliferation and skeletal muscle growth in chicks.

    Science.gov (United States)

    Halevy, O; Geyra, A; Barak, M; Uni, Z; Sklan, D

    2000-04-01

    The effect of posthatch starvation on skeletal muscle growth and satellite cell proliferation was examined in chicks. Chicks were either fed or starved for 48 h posthatch (d 0-d 2, d 2-d 4 or d 4-d 6) and then refed for 41 d. Body and breast muscle weights were significantly lower in starved chicks than in fed controls throughout the experiment. Histochemical staining revealed that skeletal muscle fiber development in the starved group lagged behind that of the fed group. Starvation from d 2 to 4 and d 4 to 6 posthatch had a progressively lesser effect than did immediate posthatch starvation (P < 0.05). In vitro culturing of breast muscle satellite cells revealed that DNA synthesis and number of cells per gram of muscle in the fed chicks peaked on d 2 and d 3, and then declined. In contrast, DNA synthesis in the cells of starved chicks declined on d 2 and increased on d 3 when chicks were refed. A similar pattern was seen for the number of cells per gram muscle; however, in general cell numbers tended to be higher in the starved group than in controls (P < 0.1). The results obtained with cultured cells were parallel with in situ immunostaining with 5-bromo-2'-deoxyuridine and proliferating cell nuclear antigen in breast muscle from experimental chicks, and with growth hormone receptor expression. These results suggest that satellite cell cultures are a reliable tool for evaluating muscle growth in postnatal chickens. We conclude that sufficient feed in the immediate postnatal period is critical for satellite cell proliferation and skeletal muscle development and is thus important for optimal muscle growth.

  16. Diamond, graphite, and graphene oxide nanoparticles decrease migration and invasiveness in glioblastoma cell lines by impairing extracellular adhesion.

    Science.gov (United States)

    Wierzbicki, Mateusz; Jaworski, Sławomir; Kutwin, Marta; Grodzik, Marta; Strojny, Barbara; Kurantowicz, Natalia; Zdunek, Krzysztof; Chodun, Rafał; Chwalibog, André; Sawosz, Ewa

    2017-01-01

    The highly invasive nature of glioblastoma is one of the most significant problems regarding the treatment of this tumor. Diamond nanoparticles (ND), graphite nanoparticles (NG), and graphene oxide nanoplatelets (nGO) have been explored for their biomedical applications, especially for drug delivery. The objective of this research was to assess changes in the adhesion, migration, and invasiveness of two glioblastoma cell lines, U87 and U118, after ND, NG, and nGO treatment. All treatments affected the cell surface structure, adhesion-dependent EGFR/AKT/mTOR, and β-catenin signaling pathways, decreasing the migration and invasiveness of both glioblastoma cell lines. The examined nanoparticles did not show strong toxicity but effectively deregulated cell migration. ND was effectively taken up by cells, whereas nGO and NG strongly interacted with the cell surface. These results indicate that nanoparticles could be used in biomedical applications as a low toxicity active compound for glioblastoma treatment.

  17. Anisomycin-induced GATA-6 degradation accompanying a decrease of proliferation of colorectal cancer cell

    Energy Technology Data Exchange (ETDEWEB)

    Ushijima, Hironori; Horyozaki, Akiko; Maeda, Masatomo, E-mail: mmaeda@nupals.ac.jp

    2016-09-09

    Transcription factor GATA-6 plays a key role in normal cell differentiation of the mesoderm and endoderm. On the other hand, GATA-6 is abnormally overexpressed in many clinical gastrointestinal cancer tissue samples, and accelerates cell proliferation or an anti-apoptotic response in cancerous tissues. We previously showed that activation of the JNK signaling cascade causes proteolysis of GATA-6. In this study, we demonstrated that anisomycin, a JNK activator, stimulates nuclear export of GATA-6 in a colorectal cancer cell line, DLD-1. Concomitantly, anisomycin remarkably inhibits the proliferation of DLD-1 cells via G2/M arrest in a plate culture. However, it did not induce apoptosis under growth arrest conditions. Furthermore, the growth of DLD-1 cells in a spheroid culture was suppressed by anisomycin. Although 5-FU showed only a slight inhibitory effect on 3D spheroid cultures, the same concentration of 5-FU together with a low concentration of anisomycin exhibited strong growth inhibition. These results suggest that the induction of GATA-6 dysfunction may be more effective for chemotherapy for colorectal cancer, although the mechanism underlying the synergistic effect of 5-FU and anisomycin remains unknown. - Highlights: • Anisomycin induces proteolysis of GATA-6 in DLD-1 cells. • Anisomycin remarkably inhibits the proliferation of DLD-1 cells via G2/M arrest. • Anisomycin suppresses the growth of spheroids of DLD-1, and enhances the effect of 5-FU.

  18. Decreased Cytotoxicity of Peripheral and Peritoneal Natural Killer Cell in Endometriosis.

    Science.gov (United States)

    Jeung, InCheul; Cheon, Keunyoung; Kim, Mee-Ran

    2016-01-01

    Endometriosis causes significant chronic pelvic pain, dysmenorrhea, and infertility and affects 10% of all women. In endometriosis, ectopic endometrium surviving after retrograde menstruation exhibits an abnormal immune response characterized by increased levels of activated macrophages and inflammatory cytokines. Particularly, dysfunctional natural killer (NK) cells play an important role in the pathogenesis of the disease by either facilitating or inhibiting the survival, implantation, and proliferation of endometrial cells. NK cells in the peritoneum and peritoneal fluid exhibit reduced levels of cytotoxicity in women with endometriosis. Several cytokines and inhibitory factors in the serum and peritoneal fluid also dysregulate NK cell cytotoxicity. Additionally, increased numbers of immature peripheral NK cells and induction of NK cell apoptosis are evident in the peritoneal fluid of women with endometriosis. The high rate of endometriosis recurrence after pharmaceutical or surgical treatment, which is associated with dysfunctional NK cells, indicates that new immunomodulatory management strategies are required. A good understanding of immune dysfunction would enable improvement of current treatments for endometriosis.

  19. Decreased Cytotoxicity of Peripheral and Peritoneal Natural Killer Cell in Endometriosis

    Directory of Open Access Journals (Sweden)

    InCheul Jeung

    2016-01-01

    Full Text Available Endometriosis causes significant chronic pelvic pain, dysmenorrhea, and infertility and affects 10% of all women. In endometriosis, ectopic endometrium surviving after retrograde menstruation exhibits an abnormal immune response characterized by increased levels of activated macrophages and inflammatory cytokines. Particularly, dysfunctional natural killer (NK cells play an important role in the pathogenesis of the disease by either facilitating or inhibiting the survival, implantation, and proliferation of endometrial cells. NK cells in the peritoneum and peritoneal fluid exhibit reduced levels of cytotoxicity in women with endometriosis. Several cytokines and inhibitory factors in the serum and peritoneal fluid also dysregulate NK cell cytotoxicity. Additionally, increased numbers of immature peripheral NK cells and induction of NK cell apoptosis are evident in the peritoneal fluid of women with endometriosis. The high rate of endometriosis recurrence after pharmaceutical or surgical treatment, which is associated with dysfunctional NK cells, indicates that new immunomodulatory management strategies are required. A good understanding of immune dysfunction would enable improvement of current treatments for endometriosis.

  20. Expression of HSF2 decreases in mitosis to enable stress-inducible transcription and cell survival

    Science.gov (United States)

    Elsing, Alexandra N.; Aspelin, Camilla; Björk, Johanna K.; Bergman, Heidi A.; Himanen, Samu V.; Kallio, Marko J.; Roos-Mattjus, Pia

    2014-01-01

    Unless mitigated, external and physiological stresses are detrimental for cells, especially in mitosis, resulting in chromosomal missegregation, aneuploidy, or apoptosis. Heat shock proteins (Hsps) maintain protein homeostasis and promote cell survival. Hsps are transcriptionally regulated by heat shock factors (HSFs). Of these, HSF1 is the master regulator and HSF2 modulates Hsp expression by interacting with HSF1. Due to global inhibition of transcription in mitosis, including HSF1-mediated expression of Hsps, mitotic cells are highly vulnerable to stress. Here, we show that cells can counteract transcriptional silencing and protect themselves against proteotoxicity in mitosis. We found that the condensed chromatin of HSF2-deficient cells is accessible for HSF1 and RNA polymerase II, allowing stress-inducible Hsp expression. Consequently, HSF2-deficient cells exposed to acute stress display diminished mitotic errors and have a survival advantage. We also show that HSF2 expression declines during mitosis in several but not all human cell lines, which corresponds to the Hsp70 induction and protection against stress-induced mitotic abnormalities and apoptosis. PMID:25202032

  1. Decreased SAP expression in T cells from patients with SLE contributes to early signaling abnormalities and reduced IL-2 production

    Science.gov (United States)

    Karampetsou, Maria P.; Comte, Denis; Kis-Toth, Katalin; Terhorst, Cox; Kyttaris, Vasileios C.; Tsokos, George C.

    2016-01-01

    T cells from patients with systemic lupus erythematosus (SLE) display a number of functions including increased early signaling events following engagement of the T cell receptor (TCR). Signaling lymphocytic activation molecule family (SLAMF) cell surface receptors and the X-chromosome-defined signaling lymphocytic activation molecule-associated protein (SAP) adaptor are important in the development of several immunocyte lineages and modulating immune response. Here we present evidence that SAP protein levels are decreased in T cells and in their main subsets isolated from 32 women and 3 men with SLE independently of disease activity. In SLE T cells the SAP protein is also subject to increased degradation by a caspase-3. Forced expression of SAP in SLE T cells simultaneously heightened IL-2 production, calcium (Ca2+) responses and tyrosine phosphorylation of a number of proteins. Exposure of normal T cells to SLE serum IgG, known to contain anti-CD3/TCR antibodies, resulted in SAP downregulation. We conclude that SLE T cells display reduced levels of the adaptor protein SAP probably as a result of continuous T cell activation and degradation by caspase-3. Restoration of SAP levels in SLE T cells corrects the overexcitable lupus T cell phenotype. PMID:27183584

  2. Keratin23 (KRT23 knockdown decreases proliferation and affects the DNA damage response of colon cancer cells.

    Directory of Open Access Journals (Sweden)

    Karin Birkenkamp-Demtröder

    Full Text Available Keratin 23 (KRT23 is strongly expressed in colon adenocarcinomas but absent in normal colon mucosa. Array based methylation profiling of 40 colon samples showed that the promoter of KRT23 was methylated in normal colon mucosa, while hypomethylated in most adenocarcinomas. Promoter methylation correlated with absent expression, while increased KRT23 expression in tumor samples correlated with promoter hypomethylation, as confirmed by bisulfite sequencing. Demethylation induced KRT23 expression in vitro. Expression profiling of shRNA mediated stable KRT23 knockdown in colon cancer cell lines showed that KRT23 depletion affected molecules of the cell cycle and DNA replication, recombination and repair. In vitro analyses confirmed that KRT23 depletion significantly decreased the cellular proliferation of SW948 and LS1034 cells and markedly decreased the expression of genes involved in DNA damage response, mainly molecules of the double strand break repair homologous recombination pathway. KRT23 knockdown decreased the transcript and protein expression of key molecules as e.g. MRE11A, E2F1, RAD51 and BRCA1. Knockdown of KRT23 rendered colon cancer cells more sensitive to irradiation and reduced proliferation of the KRT23 depleted cells compared to irradiated control cells.

  3. Valproic Acid Decreases the Reparation Capacity of Irradiated MOLT-4 Cells

    Czech Academy of Sciences Publication Activity Database

    Muthna, D.; Vávrová, J.; Lukášová, Emilie; Tichý, Adam; Knížek, J.; Köhlerová, R.; Mazánková, N.; Řezáčová, M.

    2012-01-01

    Roč. 46, č. 1 (2012), s. 110-116 ISSN 0026-8933 Institutional support: RVO:68081707 Keywords : HISTONE DEACETYLASE INHIBITOR * ENHANCES RADIATION SENSITIVITY * TRANSFORMED-CELLS Subject RIV: BO - Biophysics Impact factor: 0.637, year: 2012

  4. Blocking Wnt5a signaling decreases CD36 expression and foam cell formation in atherosclerosis.

    Science.gov (United States)

    Ackers, Ian; Szymanski, Candice; Duckett, K Jordan; Consitt, Leslie A; Silver, Mitchell J; Malgor, Ramiro

    2018-02-20

    Wnt5a is a highly studied member of the Wnt family and recently has been implicated in the pathogenesis of atherosclerosis, but its precise role is unknown. Foam cell development is a critical process to atherosclerotic plaque formation. In the present study, we investigated the role of noncanonical Wnt5a signaling in the development of foam cells. Human carotid atherosclerotic tissue and THP-1-derived macrophages were used to investigate the contribution of Wnt5a signaling in the formation of foam cells. Immunohistochemistry was used to evaluate protein expression of scavenger receptors and noncanonical Wnt5a receptors [frizzled 5 (Fz5) and receptor tyrosine kinase-like orphan receptor 2 (Ror2)] in human atherosclerotic macrophages/foam cells. Changes in protein expression in response to Wnt5a stimulation/inhibition were determined by Western blot, and lipid accumulation was evaluated by fluorescent lipid droplet staining. Wnt5a (Pfoam cells within the plaque. In vitro studies revealed that Wnt5a significantly increased the expression of the lipid uptake receptor CD36 (P.05). rWnt5a also significantly increased lipid accumulation in THP-1 macrophages (Pfoam cells. Copyright © 2018. Published by Elsevier Inc.

  5. Bisphenol A Disrupts Transcription and Decreases Viability in Aging Vascular Endothelial Cells

    Directory of Open Access Journals (Sweden)

    Edna Ribeiro-Varandas

    2014-09-01

    Full Text Available Bisphenol A (BPA is a widely utilized endocrine disruptor capable of mimicking endogenous hormones, employed in the manufacture of numerous consumer products, thereby interfering with physiological cellular functions. Recent research has shown that BPA alters epigenetic cellular mechanisms in mammals and may be correlated to enhanced cellular senescence. Here, the effects of BPA at 10 ng/mL and 1 µg/mL, concentrations found in human samples, were analyzed on HT29 human colon adenocarcinona cell line and Human Umbilical Vein Endothelial Cells (HUVEC. Quantitative Real-Time Polymerase Chain Reaction (qRT-PCR transcriptional analysis of the Long Interspersed Element-1 (LINE-1 retroelement showed that BPA induces global transcription deregulation in both cell lines, although with more pronounced effects in HUVEC cells. Whereas there was an increase in global transcription in HT29 exclusively after 24 h of exposure, this chemical had prolonged effects on HUVEC. Immunoblotting revealed that this was not accompanied by alterations in the overall content of H3K9me2 and H3K4me3 epigenetic marks. Importantly, cell viability assays and transcriptional analysis indicated that prolonged BPA exposure affects aging processes in senescent HUVEC. To our knowledge this is the first report that BPA interferes with senescence in primary vascular endothelial cells, therefore, suggesting its association to the etiology of age-related human pathologies, such as atherosclerosis.

  6. Bisphenol A Disrupts Transcription and Decreases Viability in Aging Vascular Endothelial Cells

    Science.gov (United States)

    Ribeiro-Varandas, Edna; Pereira, H. Sofia; Monteiro, Sara; Neves, Elsa; Brito, Luísa; Boavida Ferreira, Ricardo; Viegas, Wanda; Delgado, Margarida

    2014-01-01

    Bisphenol A (BPA) is a widely utilized endocrine disruptor capable of mimicking endogenous hormones, employed in the manufacture of numerous consumer products, thereby interfering with physiological cellular functions. Recent research has shown that BPA alters epigenetic cellular mechanisms in mammals and may be correlated to enhanced cellular senescence. Here, the effects of BPA at 10 ng/mL and 1 µg/mL, concentrations found in human samples, were analyzed on HT29 human colon adenocarcinona cell line and Human Umbilical Vein Endothelial Cells (HUVEC). Quantitative Real-Time Polymerase Chain Reaction (qRT-PCR) transcriptional analysis of the Long Interspersed Element-1 (LINE-1) retroelement showed that BPA induces global transcription deregulation in both cell lines, although with more pronounced effects in HUVEC cells. Whereas there was an increase in global transcription in HT29 exclusively after 24 h of exposure, this chemical had prolonged effects on HUVEC. Immunoblotting revealed that this was not accompanied by alterations in the overall content of H3K9me2 and H3K4me3 epigenetic marks. Importantly, cell viability assays and transcriptional analysis indicated that prolonged BPA exposure affects aging processes in senescent HUVEC. To our knowledge this is the first report that BPA interferes with senescence in primary vascular endothelial cells, therefore, suggesting its association to the etiology of age-related human pathologies, such as atherosclerosis. PMID:25207595

  7. Decreased nuclear matrix DNA topoisomerase II in human leukemia cells resistant to VM-26 and m-AMSA

    International Nuclear Information System (INIS)

    Fernandes, D.J.; Danks, M.K.; Beck, W.T.

    1990-01-01

    CEM leukemia cells selected for resistance to VM-26 (CEM/VM-1) are cross-resistant to various other DNA topoisomerase II inhibitors but not to Vinca alkaloids. Since DNA topoisomerase II is a major protein of the nuclear matrix, the authors asked if alterations in nuclear matrix topoisomerase II might be important in this form of multidrug resistance. Pretreatment of drug-sensitive CEM cells for 2 h with either 5 μM VM-26 or 3 μM m-AMSA reduced the specific activity of newly replicated DNA on the nuclear matrix by 75 and 50%, respectively, relative to that of the bulk DNA. The decatenating and unknotting activities of DNA topoisomerase II were 6- and 7-fold lower, respectively, in the nuclear matrix preparations from the CEM/VM-1 cells compared to parental CEM cells. Western blot analysis revealed that the amount of immunoreactive topoisomerase II in the nuclear matrices of the CEM/VM-1 cells decreased 3.2-fold relative to that in CEM cells. Increasing the NaCl concentration used in the matrix isolation procedure from 0.2 to 1.8 M resulted in a progressive decrease in the specific activity of topoisomerase II in matrices of CEM/VM-1 but not CEM cells, which suggested that the association of the enzyme with the matrix is altered in the resistant cells. These data support the hypothesis that resistance to VM-26 and m-AMSA is directly related to the decreased activity of nuclear matrix topoisomerase II. In CEM/VM-1 cells the interaction of either VM-26 or m-AMSA with nuclear matrix topoisomerase II is specifically diminished

  8. Up-regulation of miR-21 decreases chemotherapeutic effect of dendrosomal curcumin in breast cancer cells

    Directory of Open Access Journals (Sweden)

    Mohammad Javad Dehghan Esmatabadi

    2017-04-01

    Full Text Available Objective(s: Despite the good results of anticancer activities by curcumin, there are some hurdles that limit the use of curcumin as an anticancer agent. Many methods were examined to overcome this defect like the use of the dendrosomal curcumin (DNC. There is increasing evidence that miRNAs play important roles in biological processes. In this study, we focus on the roles of microRNA-21 in the anti-cancer effects of DNC in breast cancer. Materials and Methods: Also, we have used different methods such as MTT, apoptosis, cell cycle analysis, transwell migration assay and RT-PCR to find out more. Results: We observed that miR-21 decreased apoptotic cells in both cells (from 6.35% to 0.34 % and from 7.72% to 1.32% orderly and DNC increased it. As well as, our findings indicated that cell migration capacity was increased by miR-21 over expression and was decreased by DNC. The combination of miR-21 vector transfection and DNC treatment showed lower percentage of apoptotic cells or a higher level of penetration through the membrane compared with DNC treatment alone. Furthermore, DNC induced a marked increase in the number of cells in sub G1/G1 phase and a decrease in G2/M phase of the cell cycle in both; but, we observed reverse results compared it, after transfection with miR-21 vector. Conclusion: We observed that miR-21 suppress many aspects of anti-cancer effects of DNC in breast cancer cells, it seems that co-treatment with DNC and mir-21 down-regulation may provide a clinically useful tool for drug-resistance breast cancer cells.

  9. Up-regulation of miR-21 decreases chemotherapeutic effect of dendrosomal curcumin in breast cancer cells.

    Science.gov (United States)

    Esmatabadi, Mohammad Javad Dehghan; Farhangi, Baharak; Montazeri, Maryam; Monfared, Hamideh; Sistani, Roohollah Nakhaei; Sadeghizadeh, Majid

    2017-04-01

    Despite the good results of anticancer activities by curcumin, there are some hurdles that limit the use of curcumin as an anticancer agent. Many methods were examined to overcome this defect like the use of the dendrosomal curcumin (DNC). There is increasing evidence that miRNAs play important roles in biological processes. In this study, we focus on the roles of microRNA-21 in the anti-cancer effects of DNC in breast cancer. Also, we have used different methods such as MTT, apoptosis, cell cycle analysis, transwell migration assay and RT-PCR to find out more. We observed that miR-21 decreased apoptotic cells in both cells (from 6.35% to 0.34 % and from 7.72% to 1.32% orderly) and DNC increased it. As well as, our findings indicated that cell migration capacity was increased by miR-21 over expression and was decreased by DNC. The combination of miR-21 vector transfection and DNC treatment showed lower percentage of apoptotic cells or a higher level of penetration through the membrane compared with DNC treatment alone. Furthermore, DNC induced a marked increase in the number of cells in sub G1/G1 phase and a decrease in G2/M phase of the cell cycle in both; but, we observed reverse results compared it, after transfection with miR-21 vector. We observed that miR-21 suppress many aspects of anti-cancer effects of DNC in breast cancer cells, it seems that co-treatment with DNC and mir-21 down-regulation may provide a clinically useful tool for drug-resistance breast cancer cells.

  10. The volume of Purkinje cells decreases in the cerebellum of acrylamide-intoxicated rats, but no cells are lost

    DEFF Research Database (Denmark)

    Larsen, Jytte Overgaard; Tandrup, T; Braendgaard, H

    1994-01-01

    The effects of acrylamide intoxication on the numbers of granule and Purkinje cells and the volume of Purkinje cell perikarya have been evaluated with stereological methods. The analysis was carried out in the cerebella of rats that had received a dose of 33.3 mg/kg acrylamide, twice a week, for 7.......5 weeks. The total numbers of cerebellar granule and Purkinje cells were estimated using the optical fractionator and the mean volume of the Purkinje cell perikarya was estimated with the vertical rotator technique. The volumes of the molecular layer, the granular cell layer and the white matter were...... estimated using the Cavalieri principle. The mean weight of the cerebellum of the intoxicated rats was 7% lower than that of the control rats (2P = 0.001). The numbers of the Purkinje cells and granule cells were the same in both groups, but the mean volume of the perikarya of the Purkinje cells...

  11. Viewing a humorous film decreases IgE production by seminal B cells from patients with atopic eczema.

    Science.gov (United States)

    Kimata, Hajime

    2009-02-01

    Sperms induced IgE production by seminal B cells from patients with atopic eczema via interaction of B cells with galectin-3 on sperms. We studied the effect of viewing a humorous film on IgE production by seminal B cells cultured with sperms. Twenty-four male patients with atopic eczema viewed a humorous film (Modern Times, featuring Charlie Chaplin). Just before and immediately after viewing, semen was collected, and seminal B cells and sperms were purified. Seminal B cells were cultured with sperms and IgE production was measured, while expression of galectin-3 on sperms was assessed. After viewing the humorous film, IgE production by B cells cultured with sperms was significantly decreased. Moreover, expression of galectin-3 on sperms was reduced. Viewing a humorous film reduced galectin-3 expression on sperms, which in turn decreased IgE production by seminal B cells cultured with sperms. These results indicate that viewing a humorous film may be helpful for the study and treatment of local IgE production and allergy in the reproductive tract.

  12. Effect of decreasing electrical resistance in Characeae cell membranes caused by the flow of alternating current

    Directory of Open Access Journals (Sweden)

    Edward Śpiewla

    2014-01-01

    Full Text Available By means of the techniques of external electrodes and microelectrodes, it was found that evanescent flow of an alternating current through plasmalemma of Characeae cells neutralises oscillatory change in their electrical resistance and reversibly diminishes its value. This effect is particularly significant in the case of "high resistance cells", but it weakens with increasing temperature. The value of the estimated activation energy indicates that, after flow of the alternating current through the membrane, a rapid increase in the conductivity may be caused by an increase in conductivity of potassium channels. This result seems to support the hypothesis of electroconformational feedback.

  13. Decreased VEGF-A and sustained PEDF expression in a human retinal pigment epithelium cell line cultured under hypothermia

    Directory of Open Access Journals (Sweden)

    Masayuki Takeyama

    2015-01-01

    Full Text Available BACKGROUND: Previous reports have described a decrease in retinal temperature and clinical improvement of wet age-related macular degeneration (AMD after vitrectomy. We hypothesized that the retinal temperature decrease after vitrectomy plays a part in the suppression of wet AMD development. To test this hypothesis, we evaluated the temperature dependence of the expression of vascular endothelial growth factor-A (VEGF-A and in vitro angiogen-esis in retinal pigment epithelium (RPE. RESULTS: We cultured ARPE-19 cells at 37, 35, 33 and 31°C and measured the expression of VEGF-A, VEGF-A splicing variants, and pigment epithelium-derived factor (PEDF. We performed an in vitro tube formation assay. The dehydrogenase activity was also evaluated at each temperature. Expression of VEGF-A significantly decreased with decreased temperature while PEDF expression did not. VEGF165 expression and in vitro angiogenesis also were temperature dependent. The dehydrogenase activity significantly decreased as the culture temperature decreased. CONCLUSIONS: RPE cultured under hypothermia that decreased cellular metabolism also had decreased VEGF-A and sustained PEDF expression, creating an anti-angiogenic environment. This mechanism may be associated with a beneficial effect after vitrectomy in patients with wet AMD.

  14. Deregulation of apoptotic volume decrease and ionic movements in multidrug-resistant tumor cells: role of chloride channels

    DEFF Research Database (Denmark)

    Poulsen, Kristian Arild; Andersen, E C; Hansen, C F

    2010-01-01

    Changes in cell volume and ion gradients across the plasma membrane play a pivotal role in the initiation of apoptosis. Here we explore the kinetics of apoptotic volume decrease (AVD) and ion content dynamics in wild-type (WT) and multidrug-resistant (MDR) Ehrlich ascites tumor cells (EATC). In WT...... EATC, induction of apoptosis with cisplatin (5 muM) leads to three distinctive AVD stages: an early AVD(1) (4-12 h), associated with a 30% cell water loss; a transition stage AVD(T) ( approximately 12 to 32 h), where cell volume is partly recovered; and a secondary AVD(2) (past 32 h), where cell volume...... was further reduced. AVD(1) and AVD(2) were coupled to net loss of Cl(-), K(+), Na(+), and amino acids (ninhydrin-positive substances), whereas during AVD(T), Na(+) and Cl(-) were accumulated. MDR EATC was resistant to cisplatin, showing increased viability and less caspase 3 activation. Compared with WT EATC...

  15. Cytochalasin E alters the cytoskeleton and decreases ENaC activity in Xenopus 2F3 cells

    Science.gov (United States)

    Reifenberger, Matthew S.; Yu, Ling; Bao, Hui-Fang; Duke, Billie Jeanne; Liu, Bing-Chen; Ma, He-Ping; Eaton, Douglas C.; Alli, Abdel A.

    2014-01-01

    Numerous reports have linked cytoskeleton-associated proteins with the regulation of epithelial Na+ channel (ENaC) activity. The purpose of the present study was to determine the effect of actin cytoskeleton disruption by cytochalasin E on ENaC activity in Xenopus 2F3 cells. Here, we show that cytochalasin E treatment for 60 min can disrupt the integrity of the actin cytoskeleton in cultured Xenopus 2F3 cells. We show using single channel patch-clamp experiments and measurements of short-circuit current that ENaC activity, but not its density, is altered by cytochalasin E-induced disruption of the cytoskeleton. In nontreated cells, 8 of 33 patches (24%) had no measurable ENaC activity, whereas in cytochalasin E-treated cells, 17 of 32 patches (53%) had no activity. Analysis of those patches that did contain ENaC activity showed channel open probability significantly decreased from 0.081 ± 0.01 in nontreated cells to 0.043 ± 0.01 in cells treated with cytochalasin E. Transepithelial current from mpkCCD cells treated with cytochalasin E, cytochalasin D, or latrunculin B for 60 min was decreased compared with vehicle-treated cells. The subcellular expression of fodrin changed significantly, and several protein elements of the cytoskeleton decreased at least twofold after 60 min of cytochalasin E treatment. Cytochalasin E treatment disrupted the association between ENaC and myristoylated alanine-rich C-kinase substrate. The results presented here suggest disruption of the actin cytoskeleton by different compounds can attenuate ENaC activity through a mechanism involving changes in the subcellular expression of fodrin, several elements of the cytoskeleton, and destabilization of the ENaC-myristoylated alanine-rich C-kinase substrate complex. PMID:24829507

  16. Cytochalasin E alters the cytoskeleton and decreases ENaC activity in Xenopus 2F3 cells.

    Science.gov (United States)

    Reifenberger, Matthew S; Yu, Ling; Bao, Hui-Fang; Duke, Billie Jeanne; Liu, Bing-Chen; Ma, He-Ping; Alli, Ahmed A; Eaton, Douglas C; Alli, Abdel A

    2014-07-01

    Numerous reports have linked cytoskeleton-associated proteins with the regulation of epithelial Na(+) channel (ENaC) activity. The purpose of the present study was to determine the effect of actin cytoskeleton disruption by cytochalasin E on ENaC activity in Xenopus 2F3 cells. Here, we show that cytochalasin E treatment for 60 min can disrupt the integrity of the actin cytoskeleton in cultured Xenopus 2F3 cells. We show using single channel patch-clamp experiments and measurements of short-circuit current that ENaC activity, but not its density, is altered by cytochalasin E-induced disruption of the cytoskeleton. In nontreated cells, 8 of 33 patches (24%) had no measurable ENaC activity, whereas in cytochalasin E-treated cells, 17 of 32 patches (53%) had no activity. Analysis of those patches that did contain ENaC activity showed channel open probability significantly decreased from 0.081 ± 0.01 in nontreated cells to 0.043 ± 0.01 in cells treated with cytochalasin E. Transepithelial current from mpkCCD cells treated with cytochalasin E, cytochalasin D, or latrunculin B for 60 min was decreased compared with vehicle-treated cells. The subcellular expression of fodrin changed significantly, and several protein elements of the cytoskeleton decreased at least twofold after 60 min of cytochalasin E treatment. Cytochalasin E treatment disrupted the association between ENaC and myristoylated alanine-rich C-kinase substrate. The results presented here suggest disruption of the actin cytoskeleton by different compounds can attenuate ENaC activity through a mechanism involving changes in the subcellular expression of fodrin, several elements of the cytoskeleton, and destabilization of the ENaC-myristoylated alanine-rich C-kinase substrate complex. Copyright © 2014 the American Physiological Society.

  17. Decreased drug accumulation and increased tolerance to DNA damage in tumor cells with a low level of cisplatin resistance.

    Science.gov (United States)

    Lanzi, C; Perego, P; Supino, R; Romanelli, S; Pensa, T; Carenini, N; Viano, I; Colangelo, D; Leone, R; Apostoli, P; Cassinelli, G; Gambetta, R A; Zunino, F

    1998-04-15

    In an attempt to examine the cellular changes associated with cisplatin resistance, we selected a cisplatin-resistant (A43 1/Pt) human cervix squamous cell carcinoma cell line following continuous in vitro drug exposure. The resistant subline was characterized by a 2.5-fold degree of resistance. In particular, we investigated the expression of cellular defence systems and other cellular factors probably involved in dealing with cisplatin-induced DNA damage. Resistant cells exhibited decreased platinum accumulation and reduced levels of DNA-bound platinum and interstrand cross-link frequency after short-term drug exposure. Analysis of the effect of cisplatin on cell cycle progression revealed a cisplatin-induced G2M arrest in sensitive and resistant cells. Interestingly, a slowdown in S-phase transit was found in A431/Pt cells. A comparison of the ability of sensitive and resistant cells to repair drug-induced DNA damage suggested that resistant cells were able to tolerate higher levels of cisplatin-induced DNA damage than their parental counterparts. Analysis of the expression of proteins involved in DNA mismatch repair showed a decreased level of MSH2 in resistant cells. Since MSH2 seems to be involved in recognition of drug-induced DNA damage, this change may account for the increased tolerance to DNA damage observed in the resistant subline. In conclusion, the involvement of accumulation defects and the increased tolerance to cisplatin-induced DNA damage in these cisplatin-resistant cells support the notion that multiple changes contribute to confer a low level of cisplatin resistance.

  18. Nucleotropic doxorubicin nanoparticles decrease cancer cell viability, destroy mitochondria, induce autophagy and enhance tumour necrosis.

    Science.gov (United States)

    Friedhuber, Anna M; Chandolu, Vijay; Manchun, Somkamon; Donkor, Osaana; Sriamornsak, Pornsak; Dass, Crispin R

    2015-01-01

    Doxorubicin (Dox) is used clinically against various neoplasias, but suffers from serious side effects, and for the past three decades, this shortcoming has spurred research towards finding better drug delivery systems (DDSs) for this frontline drug. A non-targeted nucleotropic Dox-loaded nanoparticle (DNP) DDS is described, which has a simple chemical design, is easy to formulate and administer, is inexpensive, non-biohazardous and may prove to be useful clinically. The DNP formulated via vortex-assisted complex coarcevation enhanced (300-fold) cell-inhibitory activity of the drug in a panel of human cancer cells (osteosarcoma, breast, prostate and colorectal cancer) and enhanced (10-fold) efficacy against osteosarcoma (OS) in vivo. The slow-release DNPs localised to the endoplasmic reticulum disrupted the mitochondria and entered the nucleus. Prominent cytosolic vacuolisation, budding off of portions of the cytoplasm, both suggestive of autophagy, were observed. Mice that were administered with DNPs intratumorally had the smallest tumours at the end of the study, with more necrotic hotspots. This promising nucleotropic DDS enhances the cell delivery and activity of Dox against a variety of human cancer cell lines and in OS tumours in mice. © 2014 Royal Pharmaceutical Society.

  19. Poly-I:C Decreases Dendritic Cell Viability Independent of PKR Activation

    DEFF Research Database (Denmark)

    Larsen, Hjalte List; Pedersen, Anders Elm

    2012-01-01

    Vaccination with tumor-antigen pulsed, monocyte-derived dendritic cells (DCs) has emerged as a promising strategy in cancer immunotherapy. The standard DC maturation cocktail consists of a combination of tumor necrosis factor-α (TNF-α)/interleukin (IL)-1β/IL-6 and prostaglandin E2 (PGE2...

  20. Nanostructured substrate conformation can decrease osteoblast-like cell dysfunction in simulated microgravity conditions

    NARCIS (Netherlands)

    Prodanov, L.; van Loon, J.J.W.A.; te Riet, J.; Jansen, J.A.; Walboomers, X.F.

    2014-01-01

    Cells in situ are surrounded with defined structural elements formed by the nanomolecular extracellular matrix (ECM), and at the same time subjected to different mechanical stimuli arising from variety of physiological processes. In this study, using a nanotextured substrate mimicking the structural

  1. Decreased radioiodine uptake of FRTL-5 cells after {sup 131}I incubation in vitro: molecular biological investigations indicate a cell cycle-dependent pathway

    Energy Technology Data Exchange (ETDEWEB)

    Meller, Birgit; Deisting, Wibke; Baehre, Manfred [University of Luebeck, Clinic of Radiology and Nuclear Medicine, Luebeck (Germany); Gaspar, Erzsebet; Wenzel, Bjoern E. [University of Luebeck, Clinic of Internal Medicine I, Luebeck (Germany); Czarnocka, Barbara [Medical Centre of Postgraduate Education, Department of Clinical Biochemistry and Molecular Biology, Warsaw (Poland)

    2008-06-15

    In radioiodine therapy the 'stunning phenomenon' is defined as a reduction of radioiodine uptake after diagnostic application of {sup 131}I. In the current study, we established an in vitro model based on the 'Fisher rat thyrocyte cell line no. 5' (FRTL-5) to investigate the stunning. TSH-stimulated FRTL-5 cells were incubated with {sup 131}I. Time-dependent {sup 131}I uptake and the viability of FRTL-5 cells were evaluated at 4-144 h after radioiodine application. All data was corrected for number of viable cells, half life and {sup 131}I concentration. Sodium iodide symporter (NIS) and the housekeeping gene ({beta}-actin, GAPDH) levels were quantified by quantitative polymerase chain reaction (qPCR). Additionally, immunohistochemical staining (IHC) of NIS on the cell membrane was carried out. FRTL-5 monolayer cell cultures showed a specific maximum uptake of {sup 131}I 24-48 h after application. Significantly decreased {sup 131}I uptake values were observed after 72-144 h. The decrease in radioiodine uptake was correlated with decreasing mRNA levels of NIS and housekeeping genes. In parallel, unlike in controls, IHC staining of NIS on FRTL-5 cells declined significantly after {sup 131}I long-term incubation. It could be demonstrated that during {sup 131}I incubation of FRTL-5 cells, radioiodine uptake decreased significantly. Simultaneously decreasing levels of NIS mRNA and protein expression suggest a NIS-associated mechanism. Since mRNA levels of housekeeping genes decreased, too, the reduced NIS expression might be provoked by a cell cycle arrest. Our investigations recommend the FRTL-5 model as a valuable tool for further molecular biological investigations of the stunning phenomenon. (orig.)

  2. Bile acid receptor TGR5 overexpression is associated with decreased intestinal mucosal injury and epithelial cell proliferation in obstructive jaundice.

    Science.gov (United States)

    Ji, Chen-Guang; Xie, Xiao-Li; Yin, Jie; Qi, Wei; Chen, Lei; Bai, Yun; Wang, Na; Zhao, Dong-Qiang; Jiang, Xiao-Yu; Jiang, Hui-Qing

    2017-04-01

    Bile acids stimulate intestinal epithelial proliferation in vitro. We sought to investigate the role of the bile acid receptor TGR5 in the protection of intestinal epithelial proliferation in obstructive jaundice. Intestinal tissues and serum samples were obtained from patients with malignant obstructive jaundice and from bile duct ligation (BDL) rats. Intestinal permeability and morphological changes in the intestinal mucosa were observed. The functions of TGR5 in cell proliferation in intestinal epithelial injury were determined by overexpression or knockdown studies in Caco-2 and FHs 74 Int cells pretreated with lipopolysaccharide (LPS). Internal biliary drainage was superior to external biliary drainage in recovering intestinal permeability and mucosal histology in patients with obstructive jaundice. In BDL rats, feeding of chenodeoxycholic acid (CDCA) decreased intestinal mucosa injury. The levels of PCNA, a marker of proliferation, increased in response to CDCA feeding and were paralleled by elevated TGR5 expression. CDCA upregulated TGR5 expression and promoted proliferation in Caco-2 and FHs 74 Int cells pretreated with LPS. Overexpression of TGR5 resulted in increased PCNA, cell viability, EdU incorporation, and the proportion of cells in S phase, whereas knockdown of TGR5 had the opposite effect. Our data indicate that bile acids promote intestinal epithelial cell proliferation and decrease mucosal injury by upregulating TGR5 expression in obstructive jaundice. Copyright © 2016 Elsevier Inc. All rights reserved.

  3. Progesterone Increases Apoptosis and Inversely Decreases Autophagy in Human Hepatoma HA22T/VGH Cells Treated with Epirubicin

    Directory of Open Access Journals (Sweden)

    Wen-Tsan Chang

    2014-01-01

    Full Text Available Hepatocellular carcinoma (HCC is the leading cause of cancer-related deaths worldwide. Epirubicin can induce intracellular reactive oxygen species and is widely used to treat unresectable HCC. Progesterone has been found to inhibit the proliferation of hepatoma cells. This study was designed to test the combined effects of epirubicin and progesterone on human hepatoma cell line, HA22T/VGH. These cells were treated with different concentrations of epirubicin with or without the coaddition of 30 μM progesterone and then analyzed for apoptosis, autophagy, and expressions of apoptotic-related proteins and multidrug-resistant gene. Epirubicin treatment dose-dependently inhibited the growth of HA22T/VGH cells. Addition of 30 μM progesterone, which was inactive alone, augmented the effect of epirubicin on the inhibition of growth of HA22T/VGH cells. Cotreatment with progesterone enhanced epirubicin-induced apoptosis, as evidenced by greater increase in caspase-3 activity and in the ratio of the apoptosis-regulating protein, Bax/Bcl-XL. The combination also caused a decrease in autophagy and in the expression of multidrug resistance-related protein 1 mRNA compared to epirubicin alone. This study shows the epirubicin/progesterone combination was more effective in increasing apoptosis and inversely decreasing autophagy on HA22T/VGH cells treated with epirubicin alone, suggesting that this combination can potentially be used to treat HCC.

  4. Reduced Number of Transitional and Naive B Cells in Addition to Decreased BAFF Levels in Response to the T Cell Independent Immunogen Pneumovax®23.

    Directory of Open Access Journals (Sweden)

    Alena Roth

    Full Text Available Protective immunity against T cell independent (TI antigens such as Streptococcus pneumoniae is characterized by antibody production of B cells induced by the combined activation of T cell independent type 1 and type 2 antigens in the absence of direct T cell help. In mice, the main players in TI immune responses have been well defined as marginal zone (MZ B cells and B-1 cells. However, the existence of human equivalents to these B cell subsets and the nature of the human B cell compartment involved in the immune reaction remain elusive. We therefore analyzed the effect of a TI antigen on the B cell compartment through immunization of healthy individuals with the pneumococcal polysaccharide (PnPS-based vaccine Pneumovax®23, and subsequent characterization of B cell subpopulations. Our data demonstrates a transient decrease of transitional and naïve B cells, with a concomitant increase of IgA+ but not IgM+ or IgG+ memory B cells and a predominant generation of PnPS-specific IgA+ producing plasma cells. No alterations could be detected in T cells, or proposed human B-1 and MZ B cell equivalents. Consistent with the idea of a TI immune response, antigen-specific memory responses could not be observed. Finally, BAFF, which is supposed to drive class switching to IgA, was unexpectedly found to be decreased in serum in response to Pneumovax®23. Our results demonstrate that a characteristic TI response induced by Pneumovax®23 is associated with distinct phenotypical and functional changes within the B cell compartment. Those modulations occur in the absence of any modulations of T cells and without the development of a specific memory response.

  5. Preferential antitumor effect of the Src inhibitor dasatinib associated with a decreased proportion of aldehyde dehydrogenase 1-positive cells in breast cancer cells of the basal B subtype

    Directory of Open Access Journals (Sweden)

    Watanabe Mika

    2010-10-01

    Full Text Available Abstract Background Recent studies have suggested that the Src inhibitor dasatinib preferentially inhibits the growth of breast cancer cells of the basal-like subtype. To clarify this finding and further investigate combined antitumor effects of dasatinib with cytotoxic agents, a panel of breast cancer cell lines of various subtypes was treated with dasatinib and/or chemotherapeutic agents. Methods Seven human breast cancer cell lines were treated with dasatinib and/or seven chemotherapeutic agents. Effects of the treatments on c-Src activation, cell growth, cell cycle, apoptosis and the proportion of aldehyde dehydrogenase (ALDH 1-positive cells were examined. Results The 50%-growth inhibitory concentrations (IC50s of dasatinib were much lower in two basal B cell lines than those in the other cell lines. The IC50s of chemotherapeutic agents were not substantially different among the cell lines. Dasatinib enhanced antitumor activity of etoposide in the basal B cell lines. Dasatinib induced a G1-S blockade with a slight apoptosis, and a combined treatment of dasatinib with etoposide also induced a G1-S blockade in the basal B cell lines. Dasatinib decreased the expression levels of phosphorylated Src in all cell lines. Interestingly, dasatinib significantly decreased the proportion of ALDH1-positive cells in the basal B cell lines but not in the other cell lines. Conclusions The present study indicates that dasatinib preferentially inhibits the growth of breast cancer cells of the basal B subtype associated with a significant loss of putative cancer stem cell population. A combined use of dasatinib with etoposide additively inhibits their growth. Further studies targeting breast cancers of the basal B subtype using dasatinib with cytotoxic agents are warranted.

  6. Nitric oxide decreases intestinal haemorrhagic lesions in rat anaphylaxis independently of mast cell activation

    Directory of Open Access Journals (Sweden)

    J. Carvalho Tavares

    1997-01-01

    Full Text Available The purpose of this study is to assess the role of nitric oxide (NO in the intestinal lesions of passive anaphylaxis, since this experimental model resembles necrotizing enterocolitis. Sprague-Dawley rats were sensitized with IgE anti-dinitrophenol monoclonal antibody. Extravasation of protein-rich plasma and haemorrhagia were measured in the small intestine. Plasma histamine was measured to assess mast cell activation. The effect of exogenous NO on the lesions was assessed by using two structurally unrelated NO-donors: sodium nitroprusside and S-nitroso-Nacetyl-penicillamine (SNAP. An increased basal production of NO was observed in cells taken after anaphylaxis, associated with a reduced response to platelet-activating factor, interleukin 1beta, and IgE/DNP-bovine serum albumin complexes. The response to bacterial lipopolysaccharide and dibutyryl cyclic adenosine monophosphate (AMP was enhanced 24 h after challenge, but at earlier times was not significantly different from that observed in controls. Treatment with either sodium nitroprusside or SNAP produced a significant reduction of the haemorrhagic lesions, which are a hallmark of rat anaphylaxis. The extravasation of protein-rich plasma was not influenced by NO-donors. The increase of plasma histamine elicited by the anaphylactic challenge was not influenced by SNAP treatment. NO-donors protect intestinal haemorrhagic lesions of rat anaphylaxis by a mechanism apparently independent of mast cell histamine release.

  7. Luteolin decreases the attachment, invasion and cytotoxicity of UPEC in bladder epithelial cells and inhibits UPEC biofilm formation.

    Science.gov (United States)

    Shen, Xiao-fei; Ren, Lai-bin; Teng, Yan; Zheng, Shuang; Yang, Xiao-long; Guo, Xiao-juan; Wang, Xin-yuan; Sha, Kai-hui; Li, Na; Xu, Guang-ya; Tian, Han-wen; Wang, Xiao-ying; Liu, Xiao-kang; Li, Jingyu; Huang, Ning

    2014-10-01

    Urinary tract infection (UTI), primarily caused by uropathogenic Escherichia coli (UPEC), is one of the most common infectious diseases worldwide. Emerging antibiotic resistance requires novel treatment strategies. Luteolin, a dietary polyphenolic flavonoid, has been confirmed as a potential antimicrobial agent. Here, we evaluated the sub-MICs of luteolin for potential properties to modulate the UPEC infection. We found that luteolin significantly decreased the attachment and invasion of UPEC J96 or CFT073 in human bladder epithelial cell lines T24. Meanwhile, obvious decreased expression of type 1 fimbriae adhesin fimH gene, lower bacterial surface hydrophobicity and swimming motility, were observed in luteolin-pretreated UPEC. Furthermore, luteolin could attenuate UPEC-induced cytotoxicity in T24 cells, which manifested as decreased activity of lactate dehydrogenase (LDH). Simultaneously, the inhibition of luteolin on UPEC-induced cytotoxicity was confirmed by ethidium bromide/acridine orange staining. Finally, the luteolin-pretreated UPEC showed a lower ability of biofilm formation. Collectively, these results indicated that luteolin decreased the attachment and invasion of UPEC in bladder epithelial cells, attenuated UPEC-induced cytotoxicity and biofilm formation via down-regulating the expression of adhesin fimH gene, reducing the bacterial surface hydrophobicity and motility. Copyright © 2014 Elsevier Ltd. All rights reserved.

  8. The volume of Purkinje cells decreases in the cerebellum of acrylamide-intoxicated rats, but no cells are lost

    DEFF Research Database (Denmark)

    Larsen, Jytte Overgaard; Tandrup, T; Braendgaard, H

    1994-01-01

    The effects of acrylamide intoxication on the numbers of granule and Purkinje cells and the volume of Purkinje cell perikarya have been evaluated with stereological methods. The analysis was carried out in the cerebella of rats that had received a dose of 33.3 mg/kg acrylamide, twice a week, for 7...

  9. Prolonged sleep deprivation decreases cell proliferation and immature newborn neurons in both dorsal and ventral hippocampus of male rats.

    Science.gov (United States)

    Murata, Yusuke; Oka, Ayana; Iseki, Ayaka; Mori, Masayoshi; Ohe, Kenji; Mine, Kazunori; Enjoji, Munechika

    2017-09-01

    Previous studies have indicated that sleep deprivation negatively affects hippocampal neurogenesis, which may explain the reason for the relation between sleep loss and depression. Increasing evidence indicates that the hippocampus is anatomically and functionally segregated along a dorsolateral (cognitive function)/ventromedial (control for mood and stress response) axis. Thus, the present study was conducted to elucidate regional differences in the adverse effects of sleep deprivation on hippocampal neurogenesis. Male Sprague-Dawley rats were subjected to sleep deprivation using the "platform on the water" method for 24- or 72-h. Quantification of hippocampal cell proliferation and immature newborn neurons was stereologically estimated using immunostaining with Ki-67 and doublecortin (DCX), respectively, by optical fractionator method. A consecutive three days of sleep deprivation significantly reduced the density of Ki-67- and DCX-immunopositive cells both in the dorsal and ventral hippocampal subgranular zone and the decrease in DCX-labeled cells was more pronounced in the ventral hippocampus than in dorsal region. Our results indicate that prolonged sleep deprivation decreases hippocampal cell proliferation and neurogenesis in both the dorsal and ventral dentate gyrus. Future studies will be needed to clarify the impact of sleep deprivation-induced decreases in hippocampal neurogenesis on the development of depression. Copyright © 2017 Elsevier Ireland Ltd and Japan Neuroscience Society. All rights reserved.

  10. Glucocorticoids decrease the production of glucagon-like peptide-1 at the transcriptional level in intestinal L-cells.

    Science.gov (United States)

    Sato, Taiki; Hayashi, Hiroto; Hiratsuka, Masahiro; Hirasawa, Noriyasu

    2015-05-05

    Glucocorticoids are widely used as anti-inflammatory or immunosuppressive drugs, but often induce hyperglycemia as a side effect. Glucagon-like peptide-1 (GLP-1) is secreted from intestinal L cells and plays crucial roles in maintaining glucose homeostasis. However, the direct effects of glucocorticoids on the GLP-1 production pathway in L cells remain unclear. We investigated the effects of glucocorticoids on GLP-1 production in vitro and in vivo. In L cell lines, glucocorticoids decreased GLP-1 release and expression of the precursor, proglucagon, at protein and mRNA levels, which were inhibited by mifepristone. The administration of dexamethasone or budesonide to mice significantly decreased the mRNA expression of proglucagon in the ileum and partially decreased glucose-stimulated GLP-1 secretion. Compound A, a dissociated glucocorticoid receptor modulator, did not affect the expression of proglucagon in vitro. These results suggested that glucocorticoids directly reduced GLP-1 production at the transcriptional level in L cells through a glucocorticoid receptor dimerization-dependent mechanism. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

  11. Age-dependent decrease in the activity of succinic dehydrogenase in rat CA1 pyramidal cells: a quantitative cytochemical study.

    Science.gov (United States)

    Bertoni-Freddari, C; Fattoretti, P; Caselli, U; Paoloni, R; Meier-Ruge, W

    1996-09-09

    A computer-assisted morphometric study has been carried out on the ultrastructure of perikaryal CA1 pyramidal cell mitochondria positive to the copper ferricyanide cytochemical reaction for succinic dehydrogenase (SDH) in rats of 3, 12 and 23 months of age. The cytoplasmic volume fraction occupied by the positive mitochondria (Volume density: Vv), the number of organelles/micron 3 of CA1 pyramidal cell cytoplasm (Numerical density: Nv) and the average mitochondrial volume (V) were automatically calculated by means of computer-assisted morphometry. Vv was significantly decreased in 23-month-old animals versus the other age groups. Nv was unchanged between 3 and 12 months of age, but was decreased to a significant extent in old animals. V did not undergo significant changes in the three age groups taken into account. In the old animals the percent of organelles smaller than 0.16 micron 3 is above 20%, while in the young and adult groups the same size of mitochondria accounts for 7 and 3%, respectively. Thus, a reduction in the number of medium sized organelles appears to be responsible for the decrease in Vv due to age. Since SDH activity is known to support maximum rates of respiration, quantitative estimation of the active mitochondria provides information on the metabolic competence of the cells investigated when energy demand is high. In this context, our present findings document that a significant impairment in the efficiency to match actual energy provisions occurs in old CA1 pyramidal cells.

  12. Decreased viability and proliferation of CHANG conjunctival epithelial cells after contact with ultraviolet light-irradiated pollen.

    Science.gov (United States)

    Heidinger, Andrea; Rabensteiner, Dieter Franz; Rabensteiner, Jasmin; Kieslinger, Petra; Horwath-Winter, Jutta; Stabentheiner, Edith; Riedl, Regina; Wedrich, Andreas; Schmut, Otto

    2017-12-27

    Contact with pollen is the major reason for the development of allergic symptoms on the ocular surface leading to a significant increase of allergic diseases worldwide. Environmental changes such as increased ultraviolet (UV) radiation and air pollution are discussed as contributory causes for this increase. We investigated the effect of UV light on the histamine content of pollen and examined if an irradiation of pollen affects the viability and proliferation of conjunctival cells. Alder (Alnus glutinosa) and hazel (Corylus avellana) pollen were irradiated for different time periods with sunlight, UV-A or UV-B light and the histamine content was analysed and compared with non-irradiated pollen. Conjunctival epithelial cells (CHANG cells) were exposed to irradiated and non-irradiated pollen followed by an assessment of cell viability with the colorimetric MTS test and the impedance-based measurement of cell proliferation using the xCELLigence real-time analysis system. UV light irradiation increased the histamine level of alder and hazel pollen in a dose-dependent manner. CHANG cells treated with irradiated pollen induced a statistically significant higher decrease of cell viability than treatment with non-irradiated pollen. Our results indicate that UV light is able to alter pollen thus making them more harmful for conjunctival cells.

  13. Inhibition of myeloperoxidase decreases vascular oxidative stress and increases vasodilatation in sickle cell disease mice1[S

    Science.gov (United States)

    Zhang, Hao; Xu, Hao; Weihrauch, Dorothee; Jones, Deron W.; Jing, Xigang; Shi, Yang; Gourlay, David; Oldham, Keith T.; Hillery, Cheryl A.; Pritchard, Kirkwood A.

    2013-01-01

    Activated leukocytes and polymorphonuclear neutrophils (PMN) release myeloperoxidase (MPO), which binds to endothelial cells (EC), is translocated, and generates oxidants that scavenge nitric oxide (NO) and impair EC function. To determine whether MPO impairs EC function in sickle cell disease (SCD), control (AA) and SCD mice were treated with N-acetyl-lysyltyrosylcysteine-amide (KYC). SCD humans and mice have high plasma MPO and soluble L-selectin (sL-selectin). KYC had no effect on MPO but decreased plasma sL-selectin and malondialdehyde in SCD mice. MPO and 3-chlorotyrosine (3-ClTyr) were increased in SCD aortas. KYC decreased MPO and 3-ClTyr in SCD aortas to the levels in AA aortas. Vasodilatation in SCD mice was impaired. KYC increased vasodilatation in SCD mice more than 2-fold, to ∼60% of levels in AA mice. KYC inhibited MPO-dependent 3-ClTyr formation in EC proteins. SCD mice had high plasma alanine transaminase (ALT), which tended to decrease in KYC-treated SCD mice (P = 0.07). KYC increased MPO and XO/XDH and decreased 3-ClTyr and 3-nitrotyrosine (3-NO2Tyr) in SCD livers. These data support the hypothesis that SCD increases release of MPO, which generates oxidants that impair EC function and injure livers. Inhibiting MPO is an effective strategy for decreasing oxidative stress and liver injury and restoring EC function in SCD. PMID:23956444

  14. Arsenic alters transcriptional responses to Pseudomonas aeruginosa infection and decreases antimicrobial defense of human airway epithelial cells.

    Science.gov (United States)

    Goodale, Britton C; Rayack, Erica J; Stanton, Bruce A

    2017-09-15

    Arsenic contamination of drinking water and food threatens the health of hundreds of millions of people worldwide by increasing the risk of numerous diseases. Arsenic exposure has been associated with infectious lung disease in epidemiological studies, but it is not yet understood how ingestion of low levels of arsenic increases susceptibility to bacterial infection. Accordingly, the goal of this study was to examine the effect of arsenic on gene expression in primary human bronchial epithelial (HBE) cells and to determine if arsenic altered epithelial cell responses to Pseudomonas aeruginosa, an opportunistic pathogen. Bronchial epithelial cells line the airway surface, providing a physical barrier and serving critical roles in antimicrobial defense and signaling to professional immune cells. We used RNA-seq to define the transcriptional response of HBE cells to Pseudomonas aeruginosa, and investigated how arsenic affected HBE gene networks in the presence and absence of the bacterial challenge. Environmentally relevant levels of arsenic significantly changed the expression of genes involved in cellular redox homeostasis and host defense to bacterial infection, and decreased genes that code for secreted antimicrobial factors such as lysozyme. Using pathway analysis, we identified Sox4 and Nrf2-regulated gene networks that are predicted to mediate the arsenic-induced decrease in lysozyme secretion. In addition, we demonstrated that arsenic decreased lysozyme in the airway surface liquid, resulting in reduced lysis of Microccocus luteus. Thus, arsenic alters the expression of genes and proteins in innate host defense pathways, thereby decreasing the ability of the lung epithelium to fight bacterial infection. Copyright © 2017 Elsevier Inc. All rights reserved.

  15. Omega-3 Eicosapentaenoic Acid Decreases CD133 Colon Cancer Stem-Like Cell Marker Expression While Increasing Sensitivity to Chemotherapy

    Science.gov (United States)

    De Carlo, Flavia; Witte, Theodore R.; Hardman, W. Elaine; Claudio, Pier Paolo

    2013-01-01

    Colorectal cancer is the third leading cause of cancer-related death in the western world. In vitro and in vivo experiments showed that omega-3 polyunsaturated fatty acids (n-3 PUFAs) can attenuate the proliferation of cancer cells, including colon cancer, and increase the efficacy of various anticancer drugs. However, these studies address the effects of n-3 PUFAs on the bulk of the tumor cells and not on the undifferentiated colon cancer stem-like cells (CSLCs) that are responsible for tumor formation and maintenance. CSLCs have also been linked to the acquisition of chemotherapy resistance and to tumor relapse. Colon CSLCs have been immunophenotyped using several antibodies against cellular markers including CD133, CD44, EpCAM, and ALDH. Anti-CD133 has been used to isolate a population of colon cancer cells that retains stem cells properties (CSLCs) from both established cell lines and primary cell cultures. We demonstrated that the n-3 PUFA, eicosapentaenoic acid (EPA), was actively incorporated into the membrane lipids of COLO 320 DM cells. 25 uM EPA decreased the cell number of the overall population of cancer cells, but not of the CD133 (+) CSLCs. Also, we observed that EPA induced down-regulation of CD133 expression and up-regulation of colonic epithelium differentiation markers, Cytokeratin 20 (CK20) and Mucin 2 (MUC2). Finally, we demonstrated that EPA increased the sensitivity of COLO 320 DM cells (total population) to both standard-of-care chemotherapies (5-Fluorouracil and oxaliplatin), whereas EPA increased the sensitivity of the CD133 (+) CSLCs to only 5-Fluorouracil. PMID:23874993

  16. Decreased tumor cell proliferation as an indicator of the effect of preoperative radiotherapy of rectal cancer

    International Nuclear Information System (INIS)

    Adell, Gunnar; Zhang Hong; Jansson, Agneta; Sun Xiaofeng; Staal, Olle; Nordenskjoeld, Bo

    2001-01-01

    Background: Rectal cancer is a common malignancy, with significant local recurrence and death rates. Preoperative radiotherapy and refined surgical technique can improve local control rates and disease-free survival. Purpose: To investigate the relationship between the tumor growth fraction in rectal cancer measured with Ki-67 and the outcome, with and without short-term preoperative radiotherapy. Method: Ki-67 (MIB-1) immunohistochemistry was used to measure tumor cell proliferation in the preoperative biopsy and the surgical specimen. Materials: Specimens from 152 patients from the Southeast Swedish Health Care region were included in the Swedish rectal cancer trial 1987-1990. Results: Tumors with low proliferation treated with preoperative radiotherapy had a significantly reduced recurrence rate. The influence on death from rectal cancer was shown only in the univariate analysis. Preoperative radiotherapy of tumors with high proliferation did not significantly improve local control and disease-free survival. The interaction between Ki-67 status and the benefit of radiotherapy was significant for the reduced recurrence rate (p=0.03), with a trend toward improved disease-free survival (p=0.08). In the surgery-alone group, Ki-67 staining did not significantly correlate with local recurrence or survival rates. Conclusion: Many Ki-67 stained tumor cells in the preoperative biopsy predicts an increased treatment failure rate after preoperative radiotherapy of rectal cancer

  17. DECREASED LEVEL OF CORD BLOOD CIRCULATING ENDOTHELIAL COLONY-FORMING CELLS IN PREECLAMPSIA

    Science.gov (United States)

    Muñoz-Hernandez, Rocio; Miranda, Maria L.; Stiefel, Pablo; Lin, Ruei-Zeng; Praena-Fernández, Juan M.; Dominguez-Simeon, Maria J.; Villar, Jose; Moreno-Luna, Rafael; Melero-Martin, Juan M.

    2014-01-01

    Preeclampsia is a pregnancy-related disorder associated with increased cardiovascular risk for the offspring. Endothelial colony-forming cells (ECFCs) are a subset of circulating endothelial progenitor cells that participate in the formation of vasculature during development. However, the effect of preeclampsia on fetal levels of ECFCs is largely unknown. In this study, we sought to determine whether cord blood ECFC abundance and function are altered in preeclampsia. We conducted a prospective cohort study that included women with normal (n=35) and preeclamptic (n=15) pregnancies. We measured ECFC levels in the umbilical cord blood of neonates and characterized ECFC phenotype, cloning-forming ability, proliferation and migration towards VEGF-A and FGF-2, in vitro formation of capillary-like structures, and in vivo vasculogenic ability in immunodeficient mice. We found that the level of cord blood ECFCs was statistically lower in preeclampsia than in control pregnancies (P = .04), a reduction that was independent of other obstetric factors. In addition, cord blood ECFCs from preeclamptic pregnancies required more time to emerge in culture than control ECFCs. However, once derived in culture, ECFC function was deemed normal and highly similar between preeclampsia and control, including the ability to form vascular networks in vivo. This study demonstrates that preeclampsia affects ECFC abundance in neonates. A reduced level of ECFCs during preeclamptic pregnancies may contribute to an increased risk of developing future cardiovascular events. PMID:24752434

  18. Intracellular Calcium Decreases Upon Hyper Gravity-Treatment of Arabidopsis Thaliana Cell Cultures

    Science.gov (United States)

    Neef, Maren; Denn, Tamara; Ecke, Margret; Hampp, Rüdiger

    2016-06-01

    Cell cultures of Arabidopsis thaliana ( A. t.) respond to changes in the gravitational field strength with fluctuations of the amount of cytosolic calcium (Ca2+). In parabolic flight experiments, where hyper- and μg phases follow each other, μg clearly increased Ca2+, while hyper-g caused a slight reduction. Since the latter observation had not been reported before, we studied this effect in more detail. Using a special centrifuge for heavy items (ZARM, Bremen, Germany), we determined the hyper-g-dependent intracellular Ca2+ level with transgenic cell lines expressing the Ca2+ sensor, cameleon. This sensor exhibits a shift in fluorescence from 480 to 530 nm in response to Ca2+ binding. The data show a drop in the intracellular Ca2+ concentration with a threshold gravity of around 3 g. This is above hypergravity levels achieved during parabolic flights (1.8 g). The use of mutants with different sub-cellular targets of cameleon expression (nucleus, tonoplast, plasma membrane) gave the same results, i.e. Ca2+ is obviously exported from several intracellular compartments.

  19. Inhibition of MerTK increases chemosensitivity and decreases oncogenic potential in T-cell acute lymphoblastic leukemia

    International Nuclear Information System (INIS)

    Brandao, L N; Winges, A; Christoph, S; Sather, S; Migdall-Wilson, J; Schlegel, J; McGranahan, A; Gao, D; Liang, X; DeRyckere, D; Graham, D K

    2013-01-01

    Pediatric leukemia survival rates have improved dramatically over the past decades. However, current treatment protocols are still largely ineffective in cases of relapsed leukemia and are associated with a significant rate of chronic health conditions. Thus, there is a continued need for new therapeutic options. Here, we show that mer receptor tyrosine kinase (MerTK) was abnormally expressed in approximately one half of pediatric T-cell leukemia patient samples and T-cell acute lymphoblastic leukemia (T-ALL) cell lines. Stimulation of MerTK by the ligand Gas6 led to activation of the prosurvival proteins Erk 1/2 and Stat5, and MerTK-dependent activation of the STAT pathway in leukemia represents a novel finding. Furthermore, inhibition of MerTK expression increased the sensitivity of T-ALL cells to treatment with chemotherapeutic agents and decreased the oncogenic potential of the Jurkat T-ALL cell line in a methylcellulose colony-forming assay. Lastly, inhibition of MerTK expression significantly increased median survival in a xenograft mouse model of leukemia (30.5 days vs 60 days, P<0.0001). These results suggest that inhibition of MerTK is a promising therapeutic strategy for the treatment of leukemia and may allow for dose reduction of currently used chemotherapeutics resulting in decreased rates of therapy-associated toxicities

  20. Mouse retinal adaptive response to proton irradiation: correlation with increase in DNA repair and decrease in photoreceptor cell death

    International Nuclear Information System (INIS)

    Tronov, V.A.; Vinogradova, Yu.V.; Poplinskaya, V.A.; Nekrasova, E.I.; Ostrovskij, M.A.

    2015-01-01

    Retina, the visual organ, that is permanently undergone aggressive environmental influence. This implies the existence in retina of intrinsic protective and recovery mechanisms. The presence of retinal genotoxic threshold, nonlinear dose response and adaptive response of retina to genotoxicants suggest this assumption. In this article, using the model of MNU-induced degeneration of retina in mice, we investigated the adaptive response of the retina in vivo after proton irradiation and a participation of Muller glial cells in it. It was showed that the retinal recovery after the genotoxic isults correlated with increase in efficacy of DNA-damage repair and decrease in death of retinal photoreceptor cells.

  1. Decreased memory B cells and increased CD8 memory T cells in blood of breastfed children : The generation R study

    NARCIS (Netherlands)

    M.A.E. Jansen (Michelle); D. van den Heuvel (Diana); M.C. van Zelm (Menno); V.W.V. Jaddoe (Vincent); A. Hofman (Albert); J.C. de Jongste (Johan); H. Hooijkaas (Herbert); H.A. Moll (Henriëtte)

    2015-01-01

    textabstractBackground: Breastfeeding provides a protective effect against infectious diseases in infancy. Still, immunological evidence for enhanced adaptive immunity in breastfed children remains inconclusive. Objective: To determine whether breastfeeding affects B- and T-cell memory in the first

  2. MiR-371-5p facilitates pancreatic cancer cell proliferation and decreases patient survival.

    Directory of Open Access Journals (Sweden)

    De He

    Full Text Available microRNAs (miRNAs play a critical role in tumorigenesis, either as a tumor suppressor or as an oncogenic miRNA, depending on different tumor types. To date, scientists have obtained a substantial amount of knowledge with regard to miRNAs in pancreatic cancer. However, the expression and function of miR-371-5p in pancreatic cancer has not been clearly elucidated. The aim of this study was to investigate the roles of miR-371-5p in pancreatic cancer and its association with the survival of patients with pancreatic cancer.The expression of miR-371-5p was examined in pancreatic duct adenocarcinoma (PDAC and their adjacent normal pancreatic tissues (ANPT or in pancreatic cancer cell lines by qRT-PCR. The association of miR-371-5p expression with overall survival was determined. The proliferation and apoptosis of SW-1990 and Panc-1 cells, transfected with miR-371-5p mimics or inhibitor, were assessed using MTT assay and flow cytometry, respectively. The tumorigenicity was evaluated via mice xenograft experiments. miR-371-5p promoter interactions were analyzed by chromatin immunoprecipitation assays (ChIP. Protein expression was analyzed by Western blot.The expression level of miR-371-5p was dramatically upregulated in clinical PDAC tissues compared with ANPT. Patients with high miR-371-5p expression had a significantly shorter survival than those with low miR-371-5p expression. The in vitro and in vivo assays showed that overexpression of miR-371-5p resulted in cell proliferation and increased tumor growth, which was associated with inhibitor of growth 1 (ING1 downregulation. Interestingly, we also found that ING1, in turn, inhibited expression of miR-371-5p in the promoter region.our study demonstrates a novel ING1-miR-371-5p regulatory feedback loop, which may have a critical role in PDAC. Thus miR-371-5p can prove to be a novel prognostic factor and therapeutic target for pancreatic cancer treatment.

  3. Aire knockdown in medullary thymic epithelial cells affects Aire protein, deregulates cell adhesion genes and decreases thymocyte interaction.

    Science.gov (United States)

    Pezzi, Nicole; Assis, Amanda Freire; Cotrim-Sousa, Larissa Cotrim; Lopes, Gabriel Sarti; Mosella, Maritza Salas; Lima, Djalma Sousa; Bombonato-Prado, Karina F; Passos, Geraldo Aleixo

    2016-09-01

    We demonstrate that even a partial reduction of Aire mRNA levels by siRNA-induced Aire knockdown (Aire KD) has important consequences to medullary thymic epithelial cells (mTECs). Aire knockdown is sufficient to reduce Aire protein levels, impair its nuclear location, and cause an imbalance in large-scale gene expression, including genes that encode cell adhesion molecules. These genes drew our attention because adhesion molecules are implicated in the process of mTEC-thymocyte adhesion, which is critical for T cell development and the establishment of central self-tolerance. Accordingly, we consider the following: 1) mTECs contribute to the elimination of self-reactive thymocytes through adhesion; 2) Adhesion molecules play a crucial role during physical contact between these cells; and 3) Aire is an important transcriptional regulator in mTECs. However, its role in controlling mTEC-thymocyte adhesion remains unclear. Because Aire controls adhesion molecule genes, we hypothesized that the disruption of its expression could influence mTEC-thymocyte interaction. To test this hypothesis, we used a murine Aire(+) mTEC cell line as a model system to reproduce mTEC-thymocyte adhesion in vitro. Transcriptome analysis of the mTEC cell line revealed that Aire KD led to the down-modulation of more than 800 genes, including those encoding for proteins involved in cell adhesion, i.e., the extracellular matrix constituent Lama1, the CAM family adhesion molecules Vcam1 and Icam4, and those that encode peripheral tissue antigens. Thymocytes co-cultured with Aire KD mTECs had a significantly reduced capacity to adhere to these cells. This finding is the first direct evidence that Aire also plays a role in controlling mTEC-thymocyte adhesion. Copyright © 2016 Elsevier Ltd. All rights reserved.

  4. Plasmacytoid dendritic cell interferon-α production to R-848 stimulation is decreased in male infants

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    Wang Jennifer P

    2012-07-01

    Full Text Available Abstract Background Sex differences in response to microbial infections, especially viral ones, may be associated with Toll-like receptor (TLR-mediated responses by plasmacytoid dendritic cells (pDCs. Results In this study, we identified sex differences in human infant pDC interferon-α production following challenge with the TLR7/8 agonist R-848. Male pDC responses were significantly lower than those of females during early infancy. This difference may be attributed to the androgen surge experienced by males during the early infancy period. Pretreatment of human pDCs with dihydrotestosterone produced a significant reduction in interferon-α production following R-848 challenge. Conclusions Androgen-mediated regulation of pDC TLR7-driven innate immune responses may contribute to the observed sex differences in response to infections during early infancy.

  5. CXCR4 blockade decreases CD4+ T cell exhaustion and improves survival in a murine model of polymicrobial sepsis.

    Science.gov (United States)

    Ramonell, Kimberly M; Zhang, Wenxiao; Hadley, Annette; Chen, Ching-Wen; Fay, Katherine T; Lyons, John D; Klingensmith, Nathan J; McConnell, Kevin W; Coopersmith, Craig M; Ford, Mandy L

    2017-01-01

    Sepsis is a dysregulated systemic response to infection involving many inflammatory pathways and the induction of counter-regulatory anti-inflammatory processes that results in a state of immune incompetence and can lead to multi-organ failure. CXCR4 is a chemokine receptor that, following ligation by CXCL12, directs cells to bone marrow niches and also plays an important role in T cell cosignaling and formation of the immunological synapse. Here, we investigated the expression and function of CXCR4 in a murine model of polymicrobial sepsis. Results indicate that CXCR4 is selectively upregulated on naïve CD4+ and CD8+ T cells and CD4+ central memory T cells following the induction of sepsis, and that CXCR4 antagonism resulted in a significant decrease in sepsis-induced mortality. We probed the mechanistic basis for these findings and found that CXCR4 antagonism significantly increased the number of peripheral CD4+ and CD8+ T cells following sepsis. Moreover, mice treated with the CXCR4 antagonist contained fewer PD-1+ LAG-3+ 2B4+ cells, suggesting that blockade of CXCR4 mitigates CD4+ T cell exhaustion during sepsis. Taken together, these results characterize CXCR4 as an important pathway that modulates immune dysfunction and mortality following sepsis, which may hold promise as a target for future therapeutic intervention in septic patients.

  6. Human progenitor cells isolated from the developing cortex undergo decreased neurogenesis and eventual senescence following expansion in vitro

    International Nuclear Information System (INIS)

    Wright, Lynda S.; Prowse, Karen R.; Wallace, Kyle; Linskens, Maarten H.K.; Svendsen, Clive N.

    2006-01-01

    Isolation of a true self-renewing stem cell from the human brain would be of great interest as a reliable source of neural tissue. Here, we report that human fetal cortical cells grown in epidermal growth factor expressed low levels of telomerase and telomeres in these cultures shortened over time leading to growth arrest after 30 weeks. Following leukemia inhibitory factor (LIF) supplementation, growth rates and telomerase expression increased. This was best demonstrated following cell cycle synchronization and staining for telomerase using immunocytochemistry. This increase in activity resulted in the maintenance of telomeres at approximately 7 kb for more than 60 weeks in vitro. However, all cultures displayed a lack of oligodendrotye production, decreases in neurogenesis over time and underwent replicative senescence associated with increased expression of p21 before 70 weeks in vitro. Thus, under our culture conditions, these cells are not stable, multipotent, telomerase expressing self-renewing stem cells. They may be more accurately described as human neural progenitor cells (hNPC) with limited lifespan and bi-potent potential (neurons/astrocytes). Interestingly, hNPC follow a course of proliferation, neuronal production and growth arrest similar to that seen during expansion and development of the human cortex, thus providing a possible model neural system. Furthermore, due to their high expansion potential and lack of tumorogenicity, these cells remain a unique and safe source of tissue for clinical transplantation

  7. IL-21 dependent Granzyme B production of B-cells is decreased in patients with lupus nephritis.

    Science.gov (United States)

    Rabani, Mariam; Wilde, Benjamin; Hübbers, Katharina; Xu, Shilei; Kribben, Andreas; Witzke, Oliver; Dolff, Sebastian

    2018-03-01

    B-cells play a crucial role in the pathogenesis of lupus nephritis. Recently, a separate subset has been discovered characterized by expression of Granzyme B. The aim of this study is to investigate this subset in patients with systemic lupus erythematosus (SLE). Isolated PBMCs of SLE-patients (n=30) and healthy controls (n=21) were in vitro stimulated with CPG, IgG+IgM and IL-21. Patients were sub-grouped in patients with and without biopsy proven lupus nephritis. B-cells were analyzed for intracellular Granzyme B expression by flow cytometry. The strongest stimulus for Granzyme B secretion of B-cells was IgG+IgM in presence of IL-21. SLE-patients had a significant decreased percentage of Granzyme B + B-cells in particular SLE-patients with active disease and with lupus nephritis. The frequency of GrB+ producing B-cells is reduced in SLE patients. This may contribute to an imbalanced B-cell regulation towards effector B-cells which might promote the development of lupus nephritis. Copyright © 2017 Elsevier Inc. All rights reserved.

  8. Overexpression of human sperm protein 17 increases migration and decreases the chemosensitivity of human epithelial ovarian cancer cells

    International Nuclear Information System (INIS)

    Li, Fang-qiu; Han, Yan-ling; Liu, Qun; Wu, Bo; Huang, Wen-bin; Zeng, Su-yun

    2009-01-01

    Most deaths from ovarian cancer are due to metastases that are resistant to conventional therapies. But the factors that regulate the metastatic process and chemoresistance of ovarian cancer are poorly understood. In the current study, we investigated the aberrant expression of human sperm protein 17 (HSp17) in human epithelial ovarian cancer cells and tried to analyze its influences on the cell behaviors like migration and chemoresistance. Immunohistochemistry and immunocytochemistry were used to identify HSp17 in paraffin embedded ovarian malignant tumor specimens and peritoneal metastatic malignant cells. Then we examined the effect of HSp17 overexpression on the proliferation, migration, and chemoresistance of ovarian cancer cells to carboplatin and cisplatin in a human ovarian carcinoma cell line, HO8910. We found that HSp17 was aberrantly expressed in 43% (30/70) of the patients with primary epithelial ovarian carcinomas, and in all of the metastatic cancer cells of ascites from 8 patients. The Sp17 expression was also detected in the metastatic lesions the same as in ovarian lesions. None of the 7 non-epithelial tumors primarily developed in the ovaries was immunopositive for HSp17. Overexpression of HSp17 increased the migration but decreased the chemosensitivity of ovarian carcinoma cells to carboplatin and cisplatin. HSp17 is aberrantly expressed in a significant proportion of epithelial ovarian carcinomas. Our results strongly suggest that HSp17 plays a role in metastatic disease and resistance of epithelial ovarian carcinoma to chemotherapy

  9. CXCR4 blockade decreases CD4+ T cell exhaustion and improves survival in a murine model of polymicrobial sepsis.

    Directory of Open Access Journals (Sweden)

    Kimberly M Ramonell

    Full Text Available Sepsis is a dysregulated systemic response to infection involving many inflammatory pathways and the induction of counter-regulatory anti-inflammatory processes that results in a state of immune incompetence and can lead to multi-organ failure. CXCR4 is a chemokine receptor that, following ligation by CXCL12, directs cells to bone marrow niches and also plays an important role in T cell cosignaling and formation of the immunological synapse. Here, we investigated the expression and function of CXCR4 in a murine model of polymicrobial sepsis. Results indicate that CXCR4 is selectively upregulated on naïve CD4+ and CD8+ T cells and CD4+ central memory T cells following the induction of sepsis, and that CXCR4 antagonism resulted in a significant decrease in sepsis-induced mortality. We probed the mechanistic basis for these findings and found that CXCR4 antagonism significantly increased the number of peripheral CD4+ and CD8+ T cells following sepsis. Moreover, mice treated with the CXCR4 antagonist contained fewer PD-1+ LAG-3+ 2B4+ cells, suggesting that blockade of CXCR4 mitigates CD4+ T cell exhaustion during sepsis. Taken together, these results characterize CXCR4 as an important pathway that modulates immune dysfunction and mortality following sepsis, which may hold promise as a target for future therapeutic intervention in septic patients.

  10. Monosodium Glutamate Dietary Consumption Decreases Pancreatic β-Cell Mass in Adult Wistar Rats.

    Directory of Open Access Journals (Sweden)

    Piyanard Boonnate

    Full Text Available The amount of dietary monosodium glutamate (MSG is increasing worldwide, in parallel with the epidemics of metabolic syndrome. Parenteral administration of MSG to rodents induces obesity, hyperglycemia, hyperlipidemia, insulin resistance, and type 2 diabetes. However, the impact of dietary MSG is still being debated. We investigated the morphological and functional effects of prolonged MSG consumption on rat glucose metabolism and on pancreatic islet histology.Eighty adult male Wistar rats were randomly subdivided into 4 groups, and test rats in each group were supplemented with MSG for a different duration (1, 3, 6, or 9 months, n=20 for each group. All rats were fed ad libitum with a standard rat chow and water. Ten test rats in each group were provided MSG 2 mg/g body weight/day in drinking water and the 10 remaining rats in each group served as non-MSG treated controls. Oral glucose tolerance tests (OGTT were performed and serum insulin measured at 9 months. Animals were sacrificed at 1, 3, 6, or 9 months to examine the histopathology of pancreatic islets.MSG-treated rats had significantly lower pancreatic β-cell mass at 1, 6 and 9 months of study. Islet hemorrhages increased with age in all groups and fibrosis was significantly more frequent in MSG-treated rats at 1 and 3 months. Serum insulin levels and glucose tolerance in MSG-treated and untreated rats were similar at all time points we investigated.Daily MSG dietary consumption was associated with reduced pancreatic β-cell mass and enhanced hemorrhages and fibrosis, but did not affect glucose homeostasis. We speculate that high dietary MSG intake may exert a negative effect on the pancreas and such effect might become functionally significant in the presence or susceptibility to diabetes or NaCl; future experiments will take these crucial cofactors into account.

  11. Antiproliferative factor decreases Akt phosphorylation and alters gene expression via CKAP4 in T24 bladder carcinoma cells

    Directory of Open Access Journals (Sweden)

    Zhang Chen-Ou

    2010-12-01

    Full Text Available Abstract Background Urinary bladder cancer is a common malignancy worldwide, and outcomes for patients with advanced bladder cancer remain poor. Antiproliferative factor (APF is a potent glycopeptide inhibitor of epithelial cell proliferation that was discovered in the urine of patients with interstitial cystitis, a disorder with bladder epithelial thinning and ulceration. APF mediates its antiproliferative activity in primary normal bladder epithelial cells via cytoskeletal associated protein 4 (CKAP4. Because synthetic asialo-APF (as-APF has also been shown to inhibit T24 bladder cancer cell proliferation at nanomolar concentrations in vitro, and because the peptide segment of APF is 100% homologous to part of frizzled 8, we determined whether CKAP4 mediates as-APF inhibition of proliferation and/or downstream Wnt/frizzled signaling events in T24 cells. Methods T24 cells were transfected with double-stranded siRNAs against CKAP4 and treated with synthetic as-APF or inactive control peptide; cells that did not undergo electroporation and cells transfected with non-target (scrambled double-stranded siRNA served as negative controls. Cell proliferation was determined by 3H-thymidine incorporation. Expression of Akt, glycogen synthase kinase 3β (GSK3β, β-catenin, p53, and matrix metalloproteinase 2 (MMP2 mRNA was determined by quantitative reverse transcriptase polymerase chain reaction (qRT-PCR. Akt, GSK-3β, MMP2, β-catenin, and p53 protein expression, plus Akt, GSK-3β, and β-catenin phosphorylation, were determined by Western blot. Results T24 cell proliferation, MMP2 expression, Akt ser473 and thr308 phosphorylation, GSK3β tyr216 phosphorylation, and β-catenin ser45/thr41 phosphorylation were all decreased by APF, whereas p53 expression, and β-catenin ser33,37/thr41 phosphorylation, were increased by APF treatment in non-electroporated and non-target siRNA-transfected cells. Neither mRNA nor total protein expression of Akt, GSK3β, or

  12. The enlarged lysosomes in beigej cells result from decreased lysosome fission and not increased lysosome fusion

    Science.gov (United States)

    Durchfort, Nina; Verhoef, Shane; Vaughn, Michael B; Shrestha, Rishna; Adam, Dieter; Kaplan, Jerry; Ward, Diane McVey

    2011-01-01

    Chediak-Higashi Syndrome is an autosomal recessive disorder that affects vesicle morphology. The Chs1/Lyst protein is a member of the BEACH family of proteins. The absence of Chs1/Lyst gives rise to enlarged lysosomes. Lysosome size is regulated by a balance between vesicle fusion and fission and can be reversibly altered by acidifying the cytoplasm using Acetate Ringer’s or by incubating with the drug vacuolin-1. We took advantage of these procedures to determine rates of lysosome fusion and fission in the presence or absence of Chs1/Lyst. Here we show by microscopy, flow cytometry and in vitro fusion that the absence of the Chs1/Lyst protein does not increase the rate of lysosome fusion. Rather, our data indicate that loss of this protein decreases the rate of lysosome fission. We further show that overexpression of the Chs1/Lyst protein gives rise to a faster rate of lysosome fission. These results indicate that Chs1/Lyst regulates lysosome size by affecting fission. PMID:21985295

  13. Weight Loss and Decrease of Body Mass Index during Allogeneic Stem Cell Transplantation Are Common Events with Limited Clinical Impact.

    Directory of Open Access Journals (Sweden)

    Christina T Rieger

    Full Text Available Weight loss in cancer patients has been attributed with significant morbidity and mortality. During allogeneic stem cell transplantation (SCT, oral nutrition is often hampered and hence total parenteral nutrition (TPN is necessary. We therefore investigated the course of weight during stem cell transplantation and the clinical consequences of weight change.180 consecutive patients who received allogeneic SCT between January 2010 and December 2011 at our center were analyzed for weight loss, laboratory and clinical parameters.During SCT, a median decrease of 6.6% of body mass index (BMI was observed for the whole population (from 25.3 at admission to 23.6 at discharge, and a 1.6fold increase of malnutrition despite use of TPN (28.3% to 45.0%. 55.6% of patients experienced a significant weight loss of ≥5% with a median decrease of 9.2% in BMI. Serum levels of albumin, total protein and cholesterol rapidly decreased during conditioning therapy. After a median of 2.4 years, the median BMI was still only 23.4 (not different from discharge. However, we did not observe a meaningful difference in side effects and survival between patients that did or did not lose weight.Weight loss is commonly observed during allogeneic SCT despite TPN, but the clinical consequences thereof seem limited: we observed no significant impact on patients with a decrease ≥ 5% in BMI on transplant outcome, side effects or survival.

  14. Decreased HIV diversity after allogeneic stem cell transplantation of an HIV-1 infected patient: a case report

    Directory of Open Access Journals (Sweden)

    Thielen Alexander

    2010-03-01

    Full Text Available Abstract The human immunodeficiency virus type 1 (HIV-1 coreceptor use and viral evolution were analyzed in blood samples from an HIV-1 infected patient undergoing allogeneic stem cell transplantation (SCT. Coreceptor use was predicted in silico from sequence data obtained from the third variable loop region of the viral envelope gene with two software tools. Viral diversity and evolution was evaluated on the same samples by Bayesian inference and maximum likelihood methods. In addition, phenotypic analysis was done by comparison of viral growth in peripheral blood mononuclear cells and in a CCR5 (R5-deficient T-cell line which was controlled by a reporter assay confirming viral tropism. In silico coreceptor predictions did not match experimental determinations that showed a consistent R5 tropism. Anti-HIV directed antibodies could be detected before and after the SCT. These preexisting antibodies did not prevent viral rebound after the interruption of antiretroviral therapy during the SCT. Eventually, transplantation and readministration of anti-retroviral drugs lead to sustained increase in CD4 counts and decreased viral load to undetectable levels. Unexpectedly, viral diversity decreased after successful SCT. Our data evidence that only R5-tropic virus was found in the patient before and after transplantation. Therefore, blocking CCR5 receptor during stem cell transplantation might have had beneficial effects and this might apply to more patients undergoing allogeneic stem cell transplantation. Furthermore, we revealed a scenario of HIV-1 dynamic different from the commonly described ones. Analysis of viral evolution shows the decrease of viral diversity even during episodes with bursts in viral load.

  15. Inhibition of neutral sphingomyelinase decreases elevated levels of inducible nitric oxide synthase and apoptotic cell death in ocular hypertensive rats

    Energy Technology Data Exchange (ETDEWEB)

    Aslan, Mutay, E-mail: mutayaslan@akdeniz.edu.tr [Department of Medical Biochemistry, Akdeniz University Faculty of Medicine, Antalya (Turkey); Basaranlar, Goksun [Department of Biophysics, Akdeniz University Faculty of Medicine, Antalya (Turkey); Unal, Mustafa [Department of Ophthalmology, Akdeniz University Faculty of Medicine, Antalya (Turkey); Ciftcioglu, Akif [Department of Pathology, Akdeniz University Faculty of Medicine, Antalya (Turkey); Derin, Narin [Department of Biophysics, Akdeniz University Faculty of Medicine, Antalya (Turkey); Mutus, Bulent [Department of Chemistry and Biochemistry, University of Windsor, Windsor, Ontario (Canada)

    2014-11-01

    Endoplasmic reticulum (ER) stress and excessive nitric oxide production via induction of inducible nitric oxide synthase (NOS2) have been implicated in the pathogenesis of neuronal retinal cell death in ocular hypertension. Neutral sphingomyelinase (N-SMase)/ceramide pathway can regulate NOS2 expression, hence this study determined the role of selective neutral sphingomyelinase (N-SMase) inhibition on retinal NOS2 levels, ER stress, apoptosis and visual evoked potentials (VEPs) in a rat model of elevated intraocular pressure (EIOP). NOS2 expression and retinal protein nitration were significantly greater in EIOP and significantly decreased with N-SMase inhibition. A significant increase was observed in retinal ER stress markers pPERK, CHOP and GRP78 in EIOP, which were not significantly altered by N-SMase inhibition. Retinal TUNEL staining showed increased apoptosis in all EIOP groups; however N-SMase inhibition significantly decreased the percent of apoptotic cells in EIOP. Caspase-3, -8 and -9 activities were significantly increased in EIOP and returned to baseline levels following N-SMase inhibition. Latencies of all VEP components were significantly prolonged in EIOP and shortened following N-SMase inhibition. Data confirm the role of nitrative injury in EIOP and highlight the protective effect of N-SMase inhibition in EIOP via down-regulation of NOS2 levels and nitrative stress. - Highlights: • Inhibition of N-SMase decreases NOS2 levels in ocular hypertension. • Inhibition of N-SMase decreases protein nitration in ocular hypertension. • Inhibition of N-SMase decreases caspase activation in ocular hypertension. • Inhibition of N-SMase decreases apoptosis in ocular hypertension.

  16. Phagocytosis of mast cell granules results in decreased macrophage superoxide production

    Directory of Open Access Journals (Sweden)

    Bobby A. Shah

    1995-01-01

    Full Text Available The mechanism by which phagocytosed mast cell granules (MCGs inhibit macrophage superoxide production has not been defined. In this study, rat peritoneal macrophages were co-incubated with either isolated intact MCGs or MCG-sonicate, and their respiratory burst capacity and morphology were studied. Co-incubation of macrophages with either intact MCGs or MCG-sonicate resulted in a dose-dependent inhibition of superoxide- mediated cytochrome c reduction. This inhibitory effect was evident within 5 min of incubation and with MCG-sonicate was completely reversed when macrophages were washed prior to activation with PMA. In the case of intact MCGs, the inhibitory effect was only partially reversed by washing after a prolonged co-incubation time. Electron microscopic analyses revealed that MCGs were rapidly phagocytosed by macrophages and were subsequently disintegrated within the phagolysosomes. Assay of MCGs for superoxide dismutase (SOD revealed the presence of significant activity of this enzyme. A comparison of normal macrophages and those containing phagocytosed MCGs did not reveal a significant difference in total SOD activity. It is speculated that, although there was no significant increase in total SOD activity in macrophages containing phagocytosed MCGs, the phagocytosed MCGs might cause a transient increase in SOD activity within the phagolysosomes. This transient rise in SOD results in scavenging of the newly generated superoxide. Alternatively, MCG inhibition of NADPH oxidase would explain the reported observations.

  17. Recombinant nematode anticoagulant protein c2 inhibits cell invasion by decreasing uPA expression in NSCLC cells.

    Science.gov (United States)

    Tong, Yu; Yue, Jun; Mao, Meng; Liu, Qingqing; Zhou, Jing; Yang, Jiyun

    2015-04-01

    Nematode anticoagulant protein c2 (NAPc2) is an 85-residue polypeptide originally isolated from the hematophagous hookworm, Ancylostoma caninum. Several studies have shown that rNAPc2 inhibits the growth of primary and metastatic tumors in mice independently of its ability to initiate coagulation. We obtained bioactive recombinant rNAPc2 by splicing of the rNAPc2-intein-CBD fusion proteins expressed in E. coli ER2566. In the in vitro assay, rNAPc2 obviously inhibited the invasive ability of non-small cell lung cancer (NSCLC) cells in a dose-dependent manner. Furthermore, rNAPc2 suppressed tumor growth in vivo by daily intraperitoneal injection of rNAPc2 in an NSCLC cell xenograft model of nude mice. Respectively, rNAPc2 downregulated the production of urokinase plasminogen activator (uPA) (P<0.05) and suppressed nuclear factor-κB (NF-κB) activity. We also identified that inhibition of NF-κB activity impaired cell invasion and reduced the uPA production in NSCLC cells. Meanwhile, NF-κB was found to directly bind to the uPA promoter in vitro. These results demonstrated that rNAPc2 inhibits cell invasion at least in part through the downregulation of the NF-κB-dependent metastasis-related gene expression in NSCLC. Our results also suggest that uPA, a known metastasis-promoting gene, is indirectly regulated by rNAPc2 through NF-κB activation. These results indicate that rNAPc2 may be a potent agent for the prevention of NSCLC progression.

  18. Human neural progenitor cells decrease photoreceptor degeneration, normalize opsin distribution and support synapse structure in cultured porcine retina.

    Science.gov (United States)

    Mollick, Tanzina; Mohlin, Camilla; Johansson, Kjell

    2016-09-01

    Retinal neurodegenerative disorders like retinitis pigmentosa, age-related macular degeneration, diabetic retinopathy and retinal detachment decrease retinal functionality leading to visual impairment. The pathological events are characterized by photoreceptor degeneration, synaptic disassembly, remodeling of postsynaptic neurons and activation of glial cells. Despite intense research, no effective treatment has been found for these disorders. The current study explores the potential of human neural progenitor cell (hNPC) derived factors to slow the degenerative processes in adult porcine retinal explants. Retinas were cultured for 3 days with or without hNPCs as a feeder layer and investigated by terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL), immunohistochemical, western blot and quantitative real time-polymerase chain reaction (qRT-PCR) techniques. TUNEL showed that hNPCs had the capacity to limit photoreceptor cell death. Among cone photoreceptors, hNPC coculture resulted in better maintenance of cone outer segments and reduced opsin mislocalization. Additionally, maintained synaptic structural integrity and preservation of second order calbindin positive horizontal cells was also observed. However, Müller cell gliosis only seemed to be alleviated in terms of reduced Müller cell density. Our observations indicate that at 3 days of coculture, hNPC derived factors had the capacity to protect photoreceptors, maintain synaptic integrity and support horizontal cell survival. Human neural progenitor cell applied treatment modalities may be an effective strategy to help maintain retinal functionality in neurodegenerative pathologies. Whether hNPCs can independently hinder Müller cell gliosis by utilizing higher concentrations or by combination with other pharmacological agents still needs to be determined. Copyright © 2016 Elsevier B.V. All rights reserved.

  19. Decreased expression of FBXW7 is correlated with poor prognosis in patients with esophageal squamous cell carcinoma

    Science.gov (United States)

    NAGANAWA, YASUHIRO; ISHIGURO, HIDEYUKI; KUWABARA, YOSHIYUKI; KIMURA, MASAHIRO; MITSUI, AKIRA; KATADA, TAKEYASU; TANAKA, TATSUYA; SHIOZAKI, MIDORI; FUJII, YOSHITAKA; TAKEYAMA, HIROMITSU

    2010-01-01

    FBXW7 is a tumor suppressor gene that induces the degradation of positive cell-cycle regulators such as c-Myc, cyclin E, c-Jun and Notch. The loss of FBXW7 promotes cell-cycle progression and cell proliferation. In the present study, we investigated the relationship between FBXW7 expression and the clinicopathological characteristics of patients with esophageal squamous cell carcinoma (ESCC). The expression of FBXW7 was quantified by real-time reverse transcription polymerase chain reaction in 43 primary ESCCs and their paired normal esophageal mucosa in patients who had not received preoperative therapy. FBXW7 expression levels were significantly correlated with the progression of the cancer and with local invasiveness. In muscle-invasive tumor cases (T2–4), lymphatic invasive tumor cases and stage II–IV cases, FBXW7 expression levels were significantly decreased (P=0.0315, P=0.0336 and P=0.0289, respectively). Decreased expression of FBXW7 was correlated with poor prognosis (P=0.0255). In conclusion, this study examined the relationship between FBXW7 expression and tumor progression in ESCC. We suggest that FBXW7 is a molecular prognostic marker and can be used to elucidate the mechanism of carcinogenesis. PMID:22993608

  20. Cholesterol pathways affected by small molecules that decrease sterol levels in Niemann-Pick type C mutant cells.

    Directory of Open Access Journals (Sweden)

    Madalina Rujoi

    2010-09-01

    Full Text Available Niemann-Pick type C (NPC disease is a genetically inherited multi-lipid storage disorder with impaired efflux of cholesterol from lysosomal storage organelles.The effect of screen-selected cholesterol lowering compounds on the major sterol pathways was studied in CT60 mutant CHO cells lacking NPC1 protein. Each of the selected chemicals decreases cholesterol in the lysosomal storage organelles of NPC1 mutant cells through one or more of the following mechanisms: increased cholesterol efflux from the cell, decreased uptake of low-density lipoproteins, and/or increased levels of cholesteryl esters. Several chemicals promote efflux of cholesterol to extracellular acceptors in both non-NPC and NPC1 mutant cells. The uptake of low-density lipoprotein-derived cholesterol is inhibited by some of the studied compounds.Results herein provide the information for prioritized further studies in identifying molecular targets of the chemicals. This approach proved successful in the identification of seven chemicals as novel inhibitors of lysosomal acid lipase (Rosenbaum et al, Biochim. Biophys. Acta. 2009, 1791:1155-1165.

  1. Cystatin C deficiency suppresses tumor growth in a breast cancer model through decreased proliferation of tumor cells.

    Science.gov (United States)

    Završnik, Janja; Butinar, Miha; Prebanda, Mojca Trstenjak; Krajnc, Aleksander; Vidmar, Robert; Fonović, Marko; Grubb, Anders; Turk, Vito; Turk, Boris; Vasiljeva, Olga

    2017-09-26

    Cysteine cathepsins are proteases that, in addition to their important physiological functions, have been associated with multiple pathologies, including cancer. Cystatin C (CstC) is a major endogenous inhibitor that regulates the extracellular activity of cysteine cathepsins. We investigated the role of cystatin C in mammary cancer using CstC knockout mice and a mouse model of breast cancer induced by expression of the polyoma middle T oncoprotein (PyMT) in the mammary epithelium. We showed that the ablation of CstC reduced the rate of mammary tumor growth. Notably, a decrease in the proliferation of CstC knockout PyMT tumor cells was demonstrated ex vivo and in vitro , indicating a role for this protease inhibitor in signaling pathways that control cell proliferation. An increase in phosphorylated p-38 was observed in CstC knockout tumors, suggesting a novel function for cystatin C in cancer development, independent of the TGF-β pathway. Moreover, proteomic analysis of the CstC wild-type and knockout PyMT primary cell secretomes revealed a decrease in the levels of 14-3-3 proteins in the secretome of knock-out cells, suggesting a novel link between cysteine cathepsins, cystatin C and 14-3-3 proteins in tumorigenesis, calling for further investigations.

  2. Decreased STAT3 Phosphorylation Mediates Cell Swelling in Ammonia-Treated Astrocyte Cultures

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    Arumugam R. Jayakumar

    2016-12-01

    Full Text Available Brain edema, due largely to astrocyte swelling, and the subsequent increase in intracranial pressure and brain herniation, are major complications of acute liver failure (ALF. Elevated level of brain ammonia has been strongly implicated in the development of astrocyte swelling associated with ALF. The means by which ammonia brings about astrocyte swelling, however, is incompletely understood. Recently, oxidative/nitrosative stress and associated signaling events, including activation of mitogen-activated protein kinases (MAPKs, as well as activation of the transcription factor, nuclear factor-kappaB (NF-κB, have been implicated in the mechanism of ammonia-induced astrocyte swelling. Since these signaling events are known to be regulated by the transcription factor, signal transducer and activator of transcription 3 (STAT3, we examined the state of STAT3 activation in ammonia-treated cultured astrocytes, and determined whether altered STAT3 activation and/or protein expression contribute to the ammonia-induced astrocyte swelling. STAT3 was found to be dephosphorylated (inactivated at Tyrosine705 in ammonia-treated cultured astrocytes. Total STAT3 protein level was also reduced in ammonia-treated astrocytes. We also found a significant increase in protein tyrosine phosphatase receptor type-1 (PTPRT-1 protein expression in ammonia-treated cultured astrocytes, and that inhibition of PTPRT-1 enhanced the phosphorylation of STAT3 after ammonia treatment. Additionally, exposure of cultured astrocytes to inhibitors of protein tyrosine phosphatases diminished the ammonia-induced cell swelling, while cultured astrocytes over-expressing STAT3 showed a reduction in the astrocyte swelling induced by ammonia. Collectively, these studies strongly suggest that inactivation of STAT3 represents a critical event in the mechanism of the astrocyte swelling associated with acute liver failure.

  3. Fasting serum levels of ferritin are associated with impaired pancreatic beta cell function and decreased insulin sensitivity

    DEFF Research Database (Denmark)

    Bonfils, L.; Ellervik, C.; Friedrich, N.

    2014-01-01

    diabetes due to its association with impaired beta cell function and decreased insulin sensitivity. Methods: We investigated 6,392 individuals from the Danish general population. Surrogate measures of beta cell function and insulin sensitivity were calculated for approximately 6,100 individuals based...... on OGTT examinations. Results: The ORs for type 2 diabetes were 4.2 (95% CI 2.4, 7.2) for the highest vs the lowest quintile of serum ferritin, and 17 (95% CI 8.9, 33) for serum ferritin levels ≥97.5th percentile vs ... glucose levels at 0, 30 and 120 min (p beta cell function estimated as the insulinogenic index and corrected insulin response (p 

  4. Decreased CXCL12 is associated with impaired alveolar epithelial cell migration and poor lung healing after lung resection.

    Science.gov (United States)

    Kanter, Jacob A; Sun, Haiying; Chiu, Stephen; DeCamp, Malcolm M; Sporn, Peter H S; Sznajder, Jacob I; Bharat, Ankit

    2015-10-01

    Prolonged air leak (PAL) is an important cause of morbidity and mortality after lung resection, but its pathogenesis has not been elucidated. Migration of alveolar type II epithelial cells is essential for lung wound repair. Here we determined the role of C-X-C motif chemokine 12 (CXCL12) on alveolar epithelial cell migration and lung wound healing. CXCL12 in the pleural fluid of patients was analyzed using enzyme-linked immunosorbent assay. Human A549 and murine MLE12 alveolar epithelial cell lines were used for wound closure, cell migration, and proliferation assays. Western blot was used to analyze Rac1 and cofilin. Pleural CXCL12 was decreased in patients with PAL (1,389 ± 192 vs 3,270 ± 247 pg/mL; P alveolar epithelial cell migration by binding to its receptor CXCR4 and may have a role in lung healing. CXCL12-mediated alveolar epithelial cell migration is associated with Rac1 and cofilin activation. Copyright © 2015 Elsevier Inc. All rights reserved.

  5. Differentiation of Human Induced Pluripotent or Embryonic Stem Cells Decreases the DNA Damage Repair by Homologous Recombination

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    Kalpana Mujoo

    2017-11-01

    Full Text Available The nitric oxide (NO-cyclic GMP pathway contributes to human stem cell differentiation, but NO free radical production can also damage DNA, necessitating a robust DNA damage response (DDR to ensure cell survival. How the DDR is affected by differentiation is unclear. Differentiation of stem cells, either inducible pluripotent or embryonic derived, increased residual DNA damage as determined by γ-H2AX and 53BP1 foci, with increased S-phase-specific chromosomal aberration after exposure to DNA-damaging agents, suggesting reduced homologous recombination (HR repair as supported by the observation of decreased HR-related repair factor foci formation (RAD51 and BRCA1. Differentiated cells also had relatively increased fork stalling and R-loop formation after DNA replication stress. Treatment with NO donor (NOC-18, which causes stem cell differentiation has no effect on double-strand break (DSB repair by non-homologous end-joining but reduced DSB repair by HR. Present studies suggest that DNA repair by HR is impaired in differentiated cells.

  6. High Glucose-Induced PC12 Cell Death by Increasing Glutamate Production and Decreasing Methyl Group Metabolism

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    Minjiang Chen

    2016-01-01

    Full Text Available Objective. High glucose- (HG- induced neuronal cell death is responsible for the development of diabetic neuropathy. However, the effect of HG on metabolism in neuronal cells is still unclear. Materials and Methods. The neural-crest derived PC12 cells were cultured for 72 h in the HG (75 mM or control (25 mM groups. We used NMR-based metabolomics to examine both intracellular and extracellular metabolic changes in HG-treated PC12 cells. Results. We found that the reduction in intracellular lactate may be due to excreting more lactate into the extracellular medium under HG condition. HG also induced the changes of other energy-related metabolites, such as an increased succinate and creatine phosphate. Our results also reveal that the synthesis of glutamate from the branched-chain amino acids (isoleucine and valine may be enhanced under HG. Increased levels of intracellular alanine, phenylalanine, myoinositol, and choline were observed in HG-treated PC12 cells. In addition, HG-induced decreases in intracellular dimethylamine, dimethylglycine, and 3-methylhistidine may indicate a downregulation of methyl group metabolism. Conclusions. Our metabolomic results suggest that HG-induced neuronal cell death may be attributed to a series of metabolic changes, involving energy metabolism, amino acids metabolism, osmoregulation and membrane metabolism, and methyl group metabolism.

  7. Decreased intracellular granule movement and glucagon secretion in pancreatic α cells attached to superior cervical ganglion neurites.

    Science.gov (United States)

    Watabe, Kiyoto; Yokawa, Satoru; Inoh, Yoshikazu; Suzuki, Takahiro; Furuno, Tadahide

    2018-01-09

    Autonomic neurons innervate pancreatic islets of Langerhans and participate in the maintenance of blood glucose concentrations by controlling hormone levels through attachment with islet cells. We previously found that stimulated superior cervical ganglia (SCG) could induce Ca 2+ oscillation in α cells via neuropeptide substance P using an in vitro co-culture model. In this study, we studied the effect of SCG neurite adhesion on intracellular secretory granule movement and glucagon secretion in α cells stimulated by low glucose concentration. Spinning disk microscopic analysis revealed that the mean velocity of intracellular granules was significantly lower in α cells attached to SCG neurites than that in those without neurites under low (2 mM), middle (10 mM), and high (20 mM) glucose concentrations. Stimulation by a low (2 mM) glucose concentration significantly increased glucagon secretion in α cells lacking neurites but not in those bound to neurites. These results suggest that adhesion to SCG neurites decreases low glucose-induced glucagon secretion in pancreatic α cells by attenuating intracellular granule movement activity.

  8. Hypofractionated radiation induces a decrease in cell proliferation but no histological damage to organotypic multicellular spheroids of human glioblastomas

    International Nuclear Information System (INIS)

    Kaaijk, P.; Academic Medical Center, Amsterdam; Troost, D.; Leenstra, S.; Bosch, D.A.; Sminia, P.; Hulshof, M.C.C.M..; Kracht, A.H.W. van der

    1997-01-01

    The aim of this study was to examine the effect of radiation on glioblastoma, using an organotypic multicellular spheroid (OMS) model. Most glioblastoma cell lines are, in contrast to glioblastomas in vivo, relatively radiosensitive. This limits the value of using cell lines for studying the radiation effect of glioblastomas. The advantage of OMS is maintenance of the characteristics of the original tumour, which is lost in conventional cell cultures. OMS prepared from four glioblastomas were treated with hypofractionated radiation with a radiobiologically equivalent dose to standard radiation treatment for glioblastomas patients. After treatment, the histology as well as the cell proliferation of the OMS was examined. After radiation, a significant decrease in cell proliferation was found, although no histological damage to the OMS was observed. The modest effects of radiation on the OMS are in agreement with the limited therapeutic value of radiotherapy for glioblastoma patients. Therefore, OMS seems to be a good alternative for cell lines to study the radiobiological effect on glioblastomas. (author)

  9. Decreased 5-Hydroxymethylcytosine (5-hmC) predicts poor prognosis in early-stage laryngeal squamous cell carcinoma.

    Science.gov (United States)

    Zhang, Yanfang; Wu, Kexia; Shao, Yuan; Sui, Fang; Yang, Qi; Shi, Bingyin; Hou, Peng; Ji, Meiju

    2016-01-01

    Accumulating evidences suggest that large-scale loss of 5-hydroxymethylcytosine (5-hmC) is an epigenetic hallmark in different cancers. However, the levels of 5-hmC in laryngeal squamous cell carcinoma (LSCC) and its prognostic value in this cancer remain largely unknown. Using dot-blot and quantitative RT-PCR assays, we investigate 5-hmC levels and expression of TET-1, -2 and -3 in LSCCs and explore the association of 5-hmC levels with clinicopathological characteristics and clinical outcome of LSCC patients. Our data showed that 5-hmC was significantly decreased in LSCCs as compared with matched normal tissues. We also found a strong link between decreased 5-hmC and the reduction of TET-1 gene expression, but not TET-2 or -3, suggesting that decreased TET-1 expression was implicated in 5-hmC loss in LSCC. Moreover, Mann-Whitney U tests showed that 5-hmC content was significantly associated with smoking (P = 0.039) and tumor invasion (P = 0.004). Importantly, we found that decreased 5-hmC was significantly associated with poor survival of early-stage LSCC patients (P = 0.043). Altogether, our findings implicate that decreased 5-hmC probaly caused by the reduction of TET-1 is crucial to the clinical pathology of LSCC and is a poor prognostic factor in ealry-stage LSCC patients.

  10. Prognostic Significance of Decreased Expression of Six Large Common Fragile Site Genes in Oropharyngeal Squamous Cell Carcinomas

    Directory of Open Access Journals (Sweden)

    Ge Gao

    2014-12-01

    Full Text Available Common fragile sites (CFSs are large regions with profound genomic instability that often span extremely large genes a number of which have been found to be important tumor suppressors. RNA sequencing previously revealed that there was a group of six large CFS genes which frequently had decreased expression in oropharyngeal squamous cell carcinomas (OPSCCs and real-time reverse transcriptase polymerase chain reaction experiments validated that these six large CFS genes (PARK2, DLG2, NBEA, CTNNA3, DMD, and FHIT had decreased expression in most of the tumor samples. In this study, we investigated whether the decreased expression of these genes has any clinical significance in OPSCCs. We analyzed the six CFS large genes in 45 OPSCC patients and found that 27 (60% of the OPSCC tumors had decreased expression of these six genes. When we correlated the expression of these six genes to each patient’s clinical records, for 11 patients who had tumor recurrence, 10 of them had decreased expression of almost all 6 genes. When we divided the patients into two groups, one group with decreased expression of the six genes and the other group with either slight changes or increased expression of the six genes, we found that there is significant difference in the incidence of tumor recurrence between these two groups by Kaplan-Meier plot analysis (P < .05. Our results demonstrated that those OPSCC tumors with decreased expression of this select group of six large CFS genes were much more likely to be associated with tumor recurrence and these genes are potential prognostic markers for predicting tumor recurrence in OPSCC.

  11. Decreased numbers of CD4+ naive and effector memory T cells, and CD8+ naïve T cells, are associated with trichloroethylene exposure

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    H Dean eHosgood

    2012-01-01

    Full Text Available Trichloroethylene (TCE is a volatile chlorinated organic compound that is commonly used as a solvent for lipophilic compounds. Although recognized as an animal carcinogen, TCE’s carcinogenic potential in humans is still uncertain. We have carried out a cross-sectional study of 80 workers exposed to TCE and 96 unexposed controls matched on age and sex in Guangdong, China to study TCE’s early biologic effects. We previously reported that the total lymphocyte count and each of the major lymphocyte subsets (i.e., CD4+ T cells, CD8+ T cells, natural killer (NK cells, and B cells were decreased in TCE-exposed workers compared to controls, suggesting a selective effect on lymphoid progenitors and/or lymphocyte survival. To explore which T lymphocyte subsets are affected, we investigated the effect of TCE exposure on the numbers of CD4+ naïve and memory T cells, CD8+ naïve and memory T cells, and regulatory T cells by FACS analysis. Linear regression of each subset was used to test for differences between exposed workers and controls adjusting for potential confounders. We observed that CD4+ and CD8+ naïve T cell counts were about 8% (p = 0.056 and 17% (p = 0.0002 lower, respectively, among exposed workers. CD4+ effector memory T cell counts were decreased by about 20% among TCE exposed workers compared to controls (p = 0.001. The selective targeting of TCE on CD8+ naïve and possibly CD4+ naive T cells, and CD4+ effector memory T cells, provide further insights into the immunosuppression-related response of human immune cells upon TCE exposure.

  12. Heat Shock Protein 90 Inhibitor (17-AAG) Induces Apoptosis and Decreases Cell Migration/Motility of Keloid Fibroblasts.

    Science.gov (United States)

    Yun, In Sik; Lee, Mi Hee; Rah, Dong Kyun; Lew, Dae Hyun; Park, Jong-Chul; Lee, Won Jai

    2015-07-01

    The regulation of apoptosis, proliferation, and migration of fibroblasts is altered in keloids. The 90-kDa heat shock protein (heat shock protein 90) is known to play a key role in such regulation. Therefore, the authors investigated whether the inhibition of heat shock protein 90 in keloid fibroblasts could induce apoptosis and attenuate keloid fibroblast proliferation and migration. The authors evaluated heat shock protein 90 expression in keloid tissues with immunohistochemistry. The authors used cell viability [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] assays and annexin V/propidium iodide staining for apoptosis, a wound healing model and cell tracking system to assess cell migration, and Akt Western blotting analysis in keloid fibroblasts after inhibition of heat shock protein 90 with 17-allylaminodemethoxygeldanamycin (17-AAG). The expression of heat shock protein 90 in keloid tissues was significantly increased compared with normal tissues. The 17-AAG-treated keloid fibroblasts showed significantly decreased proliferation, promotion of apoptosis, and decreased expression of Akt. Furthermore, a dose-dependent decrease in cell migration was noted after 17-AAG treatment of keloid fibroblasts. The 17-AAG-treated keloid fibroblasts had less directionality to the wound center and migrated a shorter distance. The authors confirmed that the inhibition of heat shock protein 90 in keloid fibroblasts could promote apoptosis and attenuate proliferation and migration of keloid fibroblasts. Therefore, the authors think that the inhibition of heat shock protein 90 is a key factor in the regulation of biological processes in keloids. With further preclinical study, the authors will be able to apply these results clinically for keloid treatment.

  13. Male bovine GH transgenic mice have decreased adiposity with an adipose depot-specific increase in immune cell populations.

    Science.gov (United States)

    Benencia, Fabian; Harshman, Stephanie; Duran-Ortiz, Silvana; Lubbers, Ellen R; List, Edward O; Householder, Lara; Al-Naeeli, Mawadda; Liang, Xiaoyu; Welch, Lonnie; Kopchick, John J; Berryman, Darlene E

    2015-05-01

    White adipose tissue (WAT) is composed of mature adipocytes and a stromal vascular fraction (SVF), which contains a variety of cells, including immune cells that vary among the different WAT depots. Growth hormone (GH) impacts immune function and adiposity in an adipose depot-specific manner. However, its effects on WAT immune cell populations remain unstudied. Bovine GH transgenic (bGH) mice are commonly used to study the in vivo effects of GH. These giant mice have an excess of GH action, impaired glucose metabolism, decreased adiposity, increased lean mass, and a shortened lifespan. Therefore, the purpose of this study was to characterize the WAT depot-specific differences in immune cell populations in the presence of excess GH in vivo. Three WAT depots were assessed: inguinal (sc), epididymal (EPI), and mesenteric (MES). Subcutaneous and MES bGH WAT depots showed a significantly higher number of total SVF cells, yet only MES bGH WAT had higher leukocyte counts compared with control samples. By means of flow cytometry analysis of the SVF, we detected greater macrophage and regulatory T-cell infiltration in sc and MES bGH WAT depots compared with controls. However, no differences were observed in the EPI WAT depot. RNA-sequencing confirmed significant alterations in pathways related to T-cell infiltration and activation in the sc depot with fewer significant changes in the EPI bGH WAT depot. These findings collectively point to a previously unrecognized role for GH in influencing the distribution of WAT immune cell populations in a depot-specific manner.

  14. A novel approach to decrease sialic acid expression in cells by a C-3-modified N-acetylmannosamine.

    Science.gov (United States)

    Wratil, Paul R; Rigol, Stephan; Solecka, Barbara; Kohla, Guido; Kannicht, Christoph; Reutter, Werner; Giannis, Athanassios; Nguyen, Long D

    2014-11-14

    Due to its position at the outermost of glycans, sialic acid is involved in a myriad of physiological and pathophysiological cell functions such as host-pathogen interactions, immune regulation, and tumor evasion. Inhibitors of cell surface sialylation could be a useful tool in cancer, immune, antibiotic, or antiviral therapy. In this work, four different C-3 modified N-acetylmannosamine analogs were tested as potential inhibitors of cell surface sialylation. Peracetylated 2-acetylamino-2-deoxy-3-O-methyl-D-mannose decreases cell surface sialylation in Jurkat cells in a dose-dependent manner up to 80%, quantified by flow cytometry and enzyme-linked lectin assays. High-performance liquid chromatography experiments revealed that not only the concentration of membrane bound but also of cytosolic sialic acid is reduced in treated cells. We have strong evidence that the observed reduction of sialic acid expression in cells is caused by the inhibition of the bifunctional enzyme UDP-GlcNAc-2-epimerase/ManNAc kinase. 2-Acetylamino-2-deoxy-3-O-methyl-D-mannose inhibits the human ManNAc kinase domain of the UDP-GlcNAc-2-epimerase/ManNAc kinase. Binding kinetics of the inhibitor and human N-acetylmannosamine kinase were evaluated using surface plasmon resonance. Specificity studies with human N-acetylglucosamine kinase and hexokinase IV indicated a high specificity of 2-acetylamino-2-deoxy-3-O-methyl-D-mannose for MNK. This substance represents a novel class of inhibitors of sialic acid expression in cells, targeting the key enzyme of sialic acid de novo biosynthesis. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

  15. A Novel Approach to Decrease Sialic Acid Expression in Cells by a C-3-modified N-Acetylmannosamine*

    Science.gov (United States)

    Wratil, Paul R.; Rigol, Stephan; Solecka, Barbara; Kohla, Guido; Kannicht, Christoph; Reutter, Werner; Giannis, Athanassios; Nguyen, Long D.

    2014-01-01

    Due to its position at the outermost of glycans, sialic acid is involved in a myriad of physiological and pathophysiological cell functions such as host-pathogen interactions, immune regulation, and tumor evasion. Inhibitors of cell surface sialylation could be a useful tool in cancer, immune, antibiotic, or antiviral therapy. In this work, four different C-3 modified N-acetylmannosamine analogs were tested as potential inhibitors of cell surface sialylation. Peracetylated 2-acetylamino-2-deoxy-3-O-methyl-d-mannose decreases cell surface sialylation in Jurkat cells in a dose-dependent manner up to 80%, quantified by flow cytometry and enzyme-linked lectin assays. High-performance liquid chromatography experiments revealed that not only the concentration of membrane bound but also of cytosolic sialic acid is reduced in treated cells. We have strong evidence that the observed reduction of sialic acid expression in cells is caused by the inhibition of the bifunctional enzyme UDP-GlcNAc-2-epimerase/ManNAc kinase. 2-Acetylamino-2-deoxy-3-O-methyl-d-mannose inhibits the human ManNAc kinase domain of the UDP-GlcNAc-2-epimerase/ManNAc kinase. Binding kinetics of the inhibitor and human N-acetylmannosamine kinase were evaluated using surface plasmon resonance. Specificity studies with human N-acetylglucosamine kinase and hexokinase IV indicated a high specificity of 2-acetylamino-2-deoxy-3-O-methyl-d-mannose for MNK. This substance represents a novel class of inhibitors of sialic acid expression in cells, targeting the key enzyme of sialic acid de novo biosynthesis. PMID:25278018

  16. Decreased tonic inhibition in cerebellar granule cells causes motor dysfunction in a mouse model of Angelman syndrome.

    Science.gov (United States)

    Egawa, Kiyoshi; Kitagawa, Kyoko; Inoue, Koichi; Takayama, Masakazu; Takayama, Chitoshi; Saitoh, Shinji; Kishino, Tatsuya; Kitagawa, Masatoshi; Fukuda, Atsuo

    2012-12-05

    Angelman syndrome is a neurodevelopmental disorder caused by loss of function of the UBE3A gene encoding a ubiquitin E3 ligase. Motor dysfunction is a characteristic feature of Angelman syndrome, but neither the mechanisms of action nor effective therapeutic strategies have yet been elucidated. We report that tonic inhibition is specifically decreased in cerebellar granule cells of Ube3a-deficient mice, a model of Angelman syndrome. As a mechanism underlying this decrease in tonic inhibition, we show that Ube3a controls degradation of γ-aminobutyric acid (GABA) transporter 1 (GAT1) and that deficiency of Ube3a induces a surplus of GAT1 that results in a decrease in GABA concentrations in the extrasynaptic space. Administering low doses of 4,5,6,7-tetrahydroisothiazolo-[5,4-c]pyridin-3-ol (THIP), a selective extrasynaptic GABA(A) receptor agonist, improves the abnormal firing properties of a population of Purkinje cells in cerebellar brain slices and reduces cerebellar ataxia in Ube3a-deficient mice in vivo. These results suggest that pharmacologically increasing tonic inhibition may be a useful strategy for alleviating motor dysfunction in Angelman syndrome.

  17. Dietary polyherbal supplementation decreases CD3+ cell infiltration into pancreatic islets and prevents hyperglycemia in nonobese diabetic mice.

    Science.gov (United States)

    Burke, Susan J; Karlstad, Michael D; Conley, Caroline P; Reel, Danielle; Whelan, Jay; Collier, J Jason

    2015-04-01

    Type 1 diabetes mellitus results from autoimmune-mediated destruction of pancreatic islet β-cells, a process associated with inflammatory signals. We hypothesized that dietary supplementation with botanicals known to contain anti-inflammatory properties would prevent losses in functional β-cell mass in nonobese diabetic (NOD) mice, a rodent model of autoimmune-mediated islet inflammation that spontaneously develops diabetes. Female NOD mice, a model of spontaneous autoimmune diabetes, were fed a diet supplemented with herbal extracts (1.916 g total botanical extracts per 1 kg of diet) over a 12-week period. The mice consumed isocaloric matched diets without (controls) and with polyherbal supplementation (PHS) ad libitum starting at a prediabetic stage (age 6 weeks) for 12 weeks. Control mice developed hyperglycemia (>180 mg/dL) within 16 weeks (n = 9). By contrast, mice receiving the PHS diet did not develop hyperglycemia by 18 weeks (n = 8). Insulin-positive cell mass within pancreatic islets was 31.9% greater in PHS mice relative to controls. We also detected a 26% decrease in CD3(+) lymphocytic infiltration in PHS mice relative to mice consuming a control diet. In vitro assays revealed reduced β-cell expression of the chemokines CCL2 and CXCL10 after overnight PHS addition to the culture media. We conclude that dietary PHS delays initiation of autoimmune-mediated β-cell destruction and subsequent onset of diabetes mellitus by diminishing islet inflammatory responses. Copyright © 2015 Elsevier Inc. All rights reserved.

  18. siRNAs with decreased off-target effect facilitate the identification of essential genes in cancer cells.

    Science.gov (United States)

    Li, Chunyan; Liu, Zhenzhen; Yang, Fang; Liu, Wensheng; Wang, Di; Dong, Encheng; Wang, Yu; Wu, Chung-I; Lu, Xuemei

    2015-08-28

    Since the essential genes are crucial to the proliferation and survival of cancer cells, the interference of these genes is promising to be an option for cancer therapy to overcome heterogeneity. However, the essential genes are highly overestimated by RNA interference (RNAi) screenings, which is mainly caused by the pervasive off-target effect of small interference RNA (siRNA) and short hairpin RNA (shRNA). In the present study, we designed Match-Mismatch paired siRNAs to discriminate the on-target effect from off-target effect of siRNAs on cell viability. Only one of the 7 potential essential genes was validated as essential to cell viability, which demonstrates the high false positive rate in RNAi screenings. We modified the siRNA by introducing random nucleotides (N) into the guide strand to mitigate the off-target effect, without significantly compromising the on-target effect. The whole transcriptome profile analysis of cells transfected with siRNAs with or without Nindicates that siRNA-dN (with Ns on both the 2nd and the 18th bases of the guide strand) weakens the off-target effect by decreasing the unintended targets. The optimized siRNAs can be applied in the characterization of essential genes in cancer cells.

  19. Cerebral ischemic injury decreases α-synuclein expression in brain tissue and glutamate-exposed HT22 cells.

    Science.gov (United States)

    Koh, Phil-Ok

    2017-09-01

    α-Synuclein is abundantly expressed in neuronal tissue, plays an essential role in the pathogenesis of neurodegenerative disorders, and exerts a neuroprotective effect against oxidative stress. Cerebral ischemia causes severe neurological disorders and neuronal dysfunction. In this study, we examined α-synuclein expression in middle cerebral artery occlusion (MCAO)-induced cerebral ischemic injury and neuronal cells damaged by glutamate treatment. MCAO surgical operation was performed on male Sprague-Dawley rats, and brain samples were isolated 24 hours after MCAO. We confirmed neurological behavior deficit, infarction area, and histopathological changes following MCAO injury. A proteomic approach and Western blot analysis demonstrated a decrease in α-synuclein in the cerebral cortices after MCAO injury. Moreover, glutamate treatment induced neuronal cell death and decreased α-synuclein expression in a hippocampal-derived cell line in a dose-dependent manner. It is known that α-synuclein regulates neuronal survival, and low levels of α-synuclein expression result in cytotoxicity. Thus, these results suggest that cerebral ischemic injury leads to a reduction in α-synuclein and consequently causes serious brain damage.

  20. Decreased MiR-17 in glioma cells increased cell viability and migration by increasing the expression of Cyclin D1, p-Akt and Akt.

    Directory of Open Access Journals (Sweden)

    Guangwei Sun

    Full Text Available The activating mutations of micro RNA (miR-17 have been revealed in tumors such as human non-Hodgkin's lymphoma and T cell leukemia. However, it is unclear about the role of miR-17 in glioma cells. The current study aimed to investigate effects of miR-17 mimics or inhibitor on the viability and migration of rat glioma C6 cells, and explore possible mechanisms.The expression of miR-17 in rat glioma C6 cells and normal brain tissue was detected by quantitative PCR. Protein expression of Cyclin D1 in rat glioma C6 cells and normal brain tissue was measured by Western Blot. Glioma C6 cells were transfected with MiR-17 mimics or inhibitor. Cells that were not transfected (Lipofectamine only and cells that were transfected with nonsense RNA negative control served as control. MTT assay was utilized to detect cell viability, and cell wound scratch assay was utilized to examine the migration index. In addition, protein expression of Cyclin D1, p-Akt and Akt in MiR-17 mimics or inhibitor-transfected glioma C6 cells was detected by Western Blot. This study had been approved by the Medical Ethics Committee of the First Affiliated Hospital of Soochow University. All applicable international, national, and/or institutional guidelines for the care and use of animals were followed.The expression of miR-17 was significantly lower, whereas the expression of Cyclin D1 was significantly higher in glioma C6 cells compared to normal brain tissue. MiR-17 mimics decreased the viability and migration of glioma C6 cells markedly at 48 h. In addition, MiR-17 inhibitor increased the viability and migration of glioma C6 cells at 24 and 48 h. The protein expression of Cyclin D1, p-Akt and Akt in glioma C6 cells decreased after transfection with miR-17 mimics for 72 h, and increased after transfection with miR-17 inhibitor for 72 h.The reduced miR-17 levels in glioma cells increased cell viability and migration, which correlates with increased expression of Cyclin D1, p

  1. Increase of radiation damage to potassium-ion permeability in E. coli cells with decrease in membrane fluidity

    International Nuclear Information System (INIS)

    Suzuki, S.

    1980-01-01

    Membrane lipids of an auxotroph of E. coli requiring unsaturated fatty acid were manipulated by supplementing the growth medium with unsaturated fatty acids of different chain lengths and/or configurations, and the radiation damage to K + -permeability of the resulting modified cells was investigated in relation with factors influencing membrane fluidity, such as temperature and procaine. Radiation had greater effects on membranes supplemented with unsaturated fatty acids of the trans configuration with a longer chain than on those of the cis configuration with a shorter chain. Radiation damage also increased with decrease in temperature. Furthermore, procaine-treated membranes showed increased resistance to radiation. All these results indicate that the damage was affected by the physical character of membrane lipids and that it was greater in membranes with decreased fluidity. (author)

  2. Regulatory T cells generated during cytomegalovirus in vitro stimulation of mononuclear cells from HIV-infected individuals on HAART correlate with decreased lymphocyte proliferation

    International Nuclear Information System (INIS)

    Jesser, Renee D.; Li, Shaobing; Weinberg, Adriana

    2006-01-01

    HIV-infected patients fail to fully recover cell-mediated immunity despite HAART. To identify regulatory factors, we studied the phenotype and function of in vitro cytomegalovirus (CMV)-stimulated T cells from HAART recipients. CFSE-measured proliferation showed CD4 + and CD8 + cells dividing in CMV-stimulated cultures. Compared with healthy controls, CMV-stimulated lymphocytes from HAART recipients had lower 3 H-thymidine incorporation; lower IFNγ and TNFα production; higher CD4 + CD27 - CD28 - and CD8 + CD27 - CD28 - frequencies; lower CD4 + CD25 hi ; and higher FoxP3 expression in CD8 + CD25 hi cells. CMV-specific proliferation correlated with higher IFNγ, TNFα and IL10 levels and higher CD4 + perforin + and CD8 + perforin + frequencies. Decreased proliferation correlated with higher CD4 + CD27 - CD28 - frequencies and TGFβ1 production, which also correlated with each other. Anti-TGFβ1 neutralizing antibodies restored CMV-specific proliferation in a dose-dependent fashion. In HIV-infected subjects, decreased proliferation correlated with higher CMV-stimulated CD8 + CD25 hi frequencies and their FoxP3 expression. These data indicate that FoxP3- and TGFβ1-expressing regulatory T cells contribute to decreased immunity in HAART recipients

  3. PTEN-induction in U251 glioma cells decreases the expression of insulin-like growth factor binding protein-2

    International Nuclear Information System (INIS)

    Levitt, Randy J.; Georgescu, Maria-Magdalena; Pollak, Michael

    2005-01-01

    PTEN is a tumor suppressor gene whose loss of function is observed in ∼40-50% of human cancers. Although insulin-like growth factor binding protein-2 (IGFBP-2) was classically described as a growth inhibitor, multiple recent reports have shown an association of overexpression and/or high serum levels of IGFBP-2 with poor prognosis of several malignancies, including gliomas. Using an inducible PTEN expression system in the PTEN-null glioma cell line U251, we demonstrate that PTEN-induction is associated with reduced proliferation, increased apoptosis, and a substantial reduction of the high levels of IGFBP-2 expression. The PTEN-induced decrease in IGFBP-2 expression could be mimicked with the PI3-kinase inhibitor LY294002, indicating that the lipid phosphatase activity of PTEN is responsible for the observed effect. However, the rapamycin analog CCI-779 did not affect IGFBP-2 expression, suggesting that the PTEN-induced decrease in IGFBP-2 expression is not attributable to decreased mTOR signalling. Recombinant human IGFBP-2 was unable to rescue U251-PTEN cells from the antiproliferative effects of PTEN, and IGFBP-2 siRNA did not affect the IGF-dependent or -independent growth of this cell line. These results suggest that the clinical data linking IGFBP-2 expression to poor prognosis may arise, at least in part, because high levels of IGFBP-2 expression correlate with loss of function of PTEN, which is well known to lead to aggressive behavior of gliomas. Our results motivate translational research regarding the relationship between IGFBP-2 expression and loss of function of PTEN

  4. β-(1,3)-Glucan Unmasking in Some Candida albicans Mutants Correlates with Increases in Cell Wall Surface Roughness and Decreases in Cell Wall Elasticity.

    Science.gov (United States)

    Hasim, Sahar; Allison, David P; Retterer, Scott T; Hopke, Alex; Wheeler, Robert T; Doktycz, Mitchel J; Reynolds, Todd B

    2017-01-01

    Candida albicans is among the most common human fungal pathogens, causing a broad range of infections, including life-threatening systemic infections. The cell wall of C. albicans is the interface between the fungus and the innate immune system. The cell wall is composed of an outer layer enriched in mannosylated glycoproteins (mannan) and an inner layer enriched in β-(1,3)-glucan and chitin. Detection of C. albicans by Dectin-1, a C-type signaling lectin specific for β-(1,3)-glucan, is important for the innate immune system to recognize systemic fungal infections. Increased exposure of β-(1,3)-glucan to the immune system occurs when the mannan layer is altered or removed in a process called unmasking. Nanoscale changes to the cell wall during unmasking were explored in live cells with atomic force microscopy (AFM). Two mutants, the cho1Δ/Δ and kre5Δ/Δ mutants, were selected as representatives that exhibit modest and strong unmasking, respectively. Comparisons of the cho1Δ/Δ and kre5Δ/Δ mutants to the wild type reveal morphological changes in their cell walls that correlate with decreases in cell wall elasticity. In addition, AFM tips functionalized with Dectin-1 revealed that the forces of binding of Dectin-1 to all of the strains were similar, but the frequency of binding was highest for the kre5Δ/Δ mutant, decreased for the cho1Δ/Δ mutant, and rare for the wild type. These data show that nanoscale changes in surface topology are correlated with increased Dectin-1 adhesion and decreased cell wall elasticity. AFM, using tips functionalized with immunologically relevant molecules, can map epitopes of the cell wall and increase our understanding of pathogen recognition by the immune system. Copyright © 2016 American Society for Microbiology.

  5. Phenotypic Modulation of Mesenteric Vascular Smooth Muscle Cells from Type 2 Diabetic Rats is Associated with Decreased Caveolin-1 Expression

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    Maria Alicia Carrillo-Sepulveda

    2014-10-01

    Full Text Available Aims: Diabetes-induced vascular complications are associated with vascular smooth muscle cell (VSMC phenotypic modulation, switching from a contractile to a synthetic-proliferative phenotype. Loss of caveolin-1 is involved with proliferation of VSMCs. We tested the hypothesis that mesenteric VSMCs from type 2 diabetic Goto-Kakizaki (GK rat undergo phenotypic modulation and it is linked to decreased caveolin-1 expression. Methods: VSMCs were isolated from mesenteric arteries from GK rats and age-matched control Wistar rats. Western blotting was used to determine expression of target proteins such as caveolin-1, calponin (marker of differentiation, and proliferating cell nuclear antigen (PCNA, marker of proliferation. In addition, we measured intracellular reactive oxygen species (ROS production using H2DCF-DA and activation of extracellular signal-regulated kinase (ERK1/2 by western blotting in VSMCs from GK stimulated with lipopolysaccharide (LPS, an endotoxin upregulated in diabetes. Results: Mesenteric VSMCs from diabetic GK rats exhibited decreased caveolin-1 and calponin expression and increased PCNA expression compared to control. Increased levels of ROS and phospho-ERK1/2 expression were also found in GK VSMCs. LPS augmented ROS and phosphorylated ERK1/2 levels to a greater extent in GK VSMCs than in control. Likewise, high glucose decreased caveolin-1 and calponin expression, increased PCNA expression and augmented ROS production in control mesenteric VSMCs. Conclusion: These results suggest that mesenteric VSMCs from diabetic GK rats undergo phenotypic modulation and it is associated with decreased caveolin-1 expression. These alterations may be due to enhanced inflammatory stimuli and glucose levels present in diabetic milieu.

  6. TOX3 (TNRC9) Over Expression in Bladder Cancer Cells Decreases Cellular Proliferation and Triggers an Interferon-Like Response

    DEFF Research Database (Denmark)

    Birkenkamp-Demtroder, Karin; Mansilla Castaño, Francisco; Dyrskjøt, Lars

    2013-01-01

    Background: Human TOX3 (TOX high mobility group box family member 3) regulates Ca2+ dependent transcription in neurons and has been associated with breast cancer susceptibility. Aim of the study was to investigate the expression of TOX3 in bladder cancer tissue samples and to identify genes...... urothelium. Microarray expression profiling of human bladder cancer cells over expressing TOX3 followed by Pathway analysis showed that TOX3 Overexpression mainly affected the Interferon Signaling Pathway. TOX3 up regulation induced the expression of several genes with a gamma interferon activation site (GAS......), e.g. STAT1. In vitro functional studies showed that TOX3 was able to bind to the GAS-sequence located at the STAT1 promoter. siRNA mediated knockdown of TOX3 in RT4 bladder cancer cells decreased STAT1 expression suggesting a direct impact of TOX3 on STAT1. Immunoprecipitation of TOX3 over...

  7. TOX3 (TNRC9) overexpression in bladder cancer cells decreases cellular proliferation and triggers an interferon-like response

    DEFF Research Database (Denmark)

    Birkenkamp-Demtröder, Karin; Mansilla, Francisco; Andersen, Lars Dyrskjøt

    2013-01-01

    Background Human TOX3 (TOX high mobility group box family member 3) regulates Ca2+-dependent transcription in neurons and has been associated with breast cancer susceptibility. Aim of the study was to investigate the expression of TOX3 in bladder cancer tissue samples and to identify genes...... urothelium. Microarray expression profiling of human bladder cancer cells overexpressing TOX3 followed by Pathway analysis showed that TOX3 overexpression mainly affected the Interferon Signaling Pathway. TOX3 upregulation induced the expression of several genes with a gamma interferon activation site (GAS......), e.g. STAT1. In vitro functional studies showed that TOX3 was able to bind to the GAS-sequence located at the STAT1 promoter. siRNA mediated knockdown of TOX3 in RT4 bladder cancer cells decreased STAT1 expression suggesting a direct impact of TOX3 on STAT1. Immunoprecipitation of TOX3 overexpressing...

  8. Keratin23 (KRT23) knockdown decreases proliferation and affects the DNA damage response of colon cancer cells

    DEFF Research Database (Denmark)

    Birkenkamp-Demtröder, Karin; Hahn, Stephan; Mansilla, Francisco

    2013-01-01

    correlated with absent expression, while increased KRT23 expression in tumor samples correlated with promoter hypomethylation, as confirmed by bisulfite sequencing. Demethylation induced KRT23 expression in vitro. Expression profiling of shRNA mediated stable KRT23 knockdown in colon cancer cell lines showed...... response, mainly molecules of the double strand break repair homologous recombination pathway. KRT23 knockdown decreased the transcript and protein expression of key molecules as e.g. MRE11A, E2F1, RAD51 and BRCA1. Knockdown of KRT23 rendered colon cancer cells more sensitive to irradiation and reduced......Keratin 23 (KRT23) is strongly expressed in colon adenocarcinomas but absent in normal colon mucosa. Array based methylation profiling of 40 colon samples showed that the promoter of KRT23 was methylated in normal colon mucosa, while hypomethylated in most adenocarcinomas. Promoter methylation...

  9. Honokiol Increases CD4+ T Cell Activation and Decreases TNF but Fails to Improve Survival Following Sepsis.

    Science.gov (United States)

    Klingensmith, Nathan J; Chen, Ching-Wen; Liang, Zhe; Burd, Eileen M; Farris, Alton B; Arbiser, Jack L; Ford, Mandy L; Coopersmith, Craig M

    2017-10-11

    Honokiol is a biphenolic isolate extracted from the bark of the magnolia tree that has been used in traditional Chinese and Japanese medicine, and has more recently been investigated for its anti-inflammatory and anti-bacterial properties. Honokiol has previously been demonstrated to improve survival in sepsis models that have rapid 100% lethality. The purpose of this study was to determine the impact of Honokiol on the host response in a model of sepsis that more closely approximates human disease. Male and female C57BL/6 mice underwent cecal ligation and puncture (CLP) to induce polymicrobial intraabdominal sepsis. Mice were then randomized to receive an injection of either Honokiol (120 mg/kg/day) or vehicle and were sacrificed after 24 hours for functional studies or followed 7 days for survival. Honokiol treatment after sepsis increased the frequency of CD4 T cells and increased activation of CD4 T cells as measured by the activation marker CD69. Honokiol also increased splenic dendritic cells. Honokiol simultaneously decreased frequency and number of CD8 T cells. Honokiol decreased systemic TNF without impacting other systemic cytokines. Honokiol did not have a detectable effect on kidney function, lung physiology, liver function or intestinal integrity. In contrast to prior studies of Honokiol in a lethal model of sepsis, Honokiol did not alter survival at seven days (70% mortality for Honokiol vs. 60% mortality for vehicle). Honokiol is thus effective in modulating the host immune response and inflammation following a clinically relevant model of sepsis but is not sufficient to alter survival.

  10. Complex cells decrease errors for the Müller-Lyer illusion in a model of the visual ventral stream

    Directory of Open Access Journals (Sweden)

    Astrid eZeman

    2014-09-01

    Full Text Available To improve robustness in object recognition, many artificial visual systems imitate the way in which the human visual cortex encodes object information as a hierarchical set of features. These systems are usually evaluated in terms of their ability to accurately categorize well-defined, unambiguous objects and scenes. In the real world, however, not all objects and scenes are presented clearly, with well-defined labels and interpretations. Visual illusions demonstrate a disparity between perception and objective reality, allowing psychophysicists to methodically manipulate stimuli and study our interpretation of the environment. One prominent effect, the Müller-Lyer illusion, is demonstrated when the perceived length of a line is contracted (or expanded by the addition of arrowheads (or arrow-tails to its ends. HMAX, a benchmark object recognition system, consistently produces a bias when classifying Müller-Lyer images. HMAX is a hierarchical, artificial neural network that imitates the ‘simple’ and ‘complex’ cell layers found in the visual ventral stream. In this study, we perform two experiments to explore the Müller-Lyer illusion in HMAX, asking: 1 How do simple versus complex cell operations within HMAX affect illusory bias and precision? 2 How does varying the position of the figures in the input image affect classification using HMAX? In our first experiment, we assessed classification after traversing each layer of HMAX and found that in general, kernel operations performed by simple cells increase bias and uncertainty while max-pooling operations executed by complex cells decrease bias and uncertainty. In our second experiment, we increased variation in the positions of figures in the input that reduced bias and uncertainty in HMAX. Our findings suggest that the Müller-Lyer illusion is exacerbated by the vulnerability of simple cell operations to positional fluctuations, but ameliorated by the robustness of complex cell

  11. Complex cells decrease errors for the Müller-Lyer illusion in a model of the visual ventral stream.

    Science.gov (United States)

    Zeman, Astrid; Obst, Oliver; Brooks, Kevin R

    2014-01-01

    To improve robustness in object recognition, many artificial visual systems imitate the way in which the human visual cortex encodes object information as a hierarchical set of features. These systems are usually evaluated in terms of their ability to accurately categorize well-defined, unambiguous objects and scenes. In the real world, however, not all objects and scenes are presented clearly, with well-defined labels and interpretations. Visual illusions demonstrate a disparity between perception and objective reality, allowing psychophysicists to methodically manipulate stimuli and study our interpretation of the environment. One prominent effect, the Müller-Lyer illusion, is demonstrated when the perceived length of a line is contracted (or expanded) by the addition of arrowheads (or arrow-tails) to its ends. HMAX, a benchmark object recognition system, consistently produces a bias when classifying Müller-Lyer images. HMAX is a hierarchical, artificial neural network that imitates the "simple" and "complex" cell layers found in the visual ventral stream. In this study, we perform two experiments to explore the Müller-Lyer illusion in HMAX, asking: (1) How do simple vs. complex cell operations within HMAX affect illusory bias and precision? (2) How does varying the position of the figures in the input image affect classification using HMAX? In our first experiment, we assessed classification after traversing each layer of HMAX and found that in general, kernel operations performed by simple cells increase bias and uncertainty while max-pooling operations executed by complex cells decrease bias and uncertainty. In our second experiment, we increased variation in the positions of figures in the input images that reduced bias and uncertainty in HMAX. Our findings suggest that the Müller-Lyer illusion is exacerbated by the vulnerability of simple cell operations to positional fluctuations, but ameliorated by the robustness of complex cell responses to such

  12. Targeting PBK/TOPK decreases growth and survival of glioma initiating cells in vitro and attenuates tumor growth in vivo.

    Science.gov (United States)

    Joel, Mrinal; Mughal, Awais A; Grieg, Zanina; Murrell, Wayne; Palmero, Sheryl; Mikkelsen, Birthe; Fjerdingstad, Hege B; Sandberg, Cecilie J; Behnan, Jinan; Glover, Joel C; Langmoen, Iver A; Stangeland, Biljana

    2015-06-17

    Glioblastomas are invasive therapy resistant brain tumors with extremely poor prognosis. The Glioma initiating cell (GIC) population contributes to therapeutic resistance and tumor recurrence. Targeting GIC-associated gene candidates could significantly impact GBM tumorigenicity. Here, we investigate a protein kinase, PBK/TOPK as a candidate for regulating growth, survival and in vivo tumorigenicity of GICs. PBK is highly upregulated in GICs and GBM tissues as shown by RNA and protein analyses. We knocked down PBK using shRNA vectors and inhibited the function of PBK protein with a pharmacological PBK inhibitor, HITOPK-032. We assessed viability, tumorsphere formation and apoptosis in three patient derived GIC cultures. Gene knockdown of PBK led to decreased viability and sphere formation and in one culture an increase in apoptosis. Treatment of cells with inhibitor HITOPK-032 (5 μM and 10 μM) almost completely abolished growth and elicited a large increase in apoptosis in all three cultures. HI-TOPK-032 treatment (5 mg/kg and 10 mg/kg bodyweight) in vivo resulted in diminished growth of experimentally induced subcutaneous GBM tumors in mice. We also carried out multi-culture assays of cell survival to investigate the relative effects on GICs compared with the normal neural stem cells (NSCs) and their differentiated counterparts. Normal NSCs seemed to withstand treatment slightly better than the GICs. Our study of identification and functional validation of PBK suggests that this candidate can be a promising molecular target for GBM treatment.

  13. Production of cloned dogs by decreasing the interval between fusion and activation during somatic cell nuclear transfer.

    Science.gov (United States)

    Kim, Sue; Park, Sun Woo; Hossein, Mohammad Shamim; Jeong, Yeon Woo; Kim, Joung Joo; Lee, Eugine; Kim, Yeun Wook; Hyun, Sang Hwan; Shin, Taeyoung; Hwang, Woo Suk

    2009-05-01

    To improve the efficiency of somatic cell nuclear transfer (SCNT) in dogs, we evaluated whether or not the interval between fusion and activation affects the success rate of SCNT. Oocytes retrieved from outbred dogs were reconstructed with adult somatic cells from a male or female Golden Retriever. In total, 151 and 225 reconstructed oocytes were transferred to 9 and 14 naturally synchronized surrogates for male and female donor cells, respectively. Chromosomal morphology was evaluated in 12 oocytes held for an interval of 2 hr between fusion and activation and 14 oocytes held for an interval of 4 hr. Three hundred seventy-six and 288 embryos were transferred to 23 and 16 surrogates for the 2 and 4 hr interval groups, respectively. Both the male (two pregnant surrogates gave birth to three puppies) and female (one pregnant surrogate gave birth to one puppy) donor cells gave birth to live puppies (P > 0.05). In the 2 hr group, significantly more reconstructed oocytes showed condensed, metaphase-like chromosomes compared to the 4 hr group (P dogs carried pregnancies to term and four puppies were born. These results demonstrate that decreasing the interval between fusion and activation increases the success rate of clone production and pregnancy. These results may increase the overall efficiency of SCNT in the canine family. (c) 2008 Wiley-Liss, Inc.

  14. Oleic acid increases mitochondrial reactive oxygen species production and decreases endothelial nitric oxide synthase activity in cultured endothelial cells.

    Science.gov (United States)

    Gremmels, Hendrik; Bevers, Lonneke M; Fledderus, Joost O; Braam, Branko; van Zonneveld, Anton Jan; Verhaar, Marianne C; Joles, Jaap A

    2015-03-15

    Elevated plasma levels of free fatty acids (FFA) are associated with increased cardiovascular risk. This may be related to FFA-induced elevation of oxidative stress in endothelial cells. We hypothesized that, in addition to mitochondrial production of reactive oxygen species, endothelial nitric oxide synthase (eNOS)-mediated reactive oxygen species production contributes to oleic acid (OA)-induced oxidative stress in endothelial cells, due to eNOS uncoupling. We measured reactive oxygen species production and eNOS activity in cultured endothelial cells (bEnd.3) in the presence of OA bound to bovine serum albumin, using the CM-H2DCFDA assay and the L-arginine/citrulline conversion assay, respectively. OA induced a concentration-dependent increase in reactive oxygen species production, which was inhibited by the mitochondrial complex II inhibitor thenoyltrifluoroacetone (TTFA). OA had little effect on eNOS activity when stimulated by a calcium-ionophore, but decreased both basal and insulin-induced eNOS activity, which was restored by TTFA. Pretreatment of bEnd.3 cells with tetrahydrobiopterin (BH4) prevented OA-induced reactive oxygen species production and restored inhibition of eNOS activity by OA. Elevation of OA levels leads to both impairment in receptor-mediated stimulation of eNOS and to production of mitochondrial-derived reactive oxygen species and hence endothelial dysfunction. Copyright © 2015 Elsevier B.V. All rights reserved.

  15. Differentiation-associated genes regulated by c-Jun and decreased in the progression of esophageal squamous cell carcinoma.

    Directory of Open Access Journals (Sweden)

    Aiping Luo

    Full Text Available Transcription factor c-Jun plays a key role in controlling epithelium cell proliferation, apoptosis and differentiation. However, molecular mechanism and biological functions of c-Jun in squamous differentiation and the progression of esophageal squamous cell carcinoma (ESCC remain elusive. In this study, we found that c-Jun bound directly to the promoter region, and activated the transcription of differentiation-associated genes including cystatin A, involucrin and SPRR3 in vivo. Ectopic expression of c-Jun enhanced SPRR3 transactivation in KYSE450 cells. Conversely, TAM67, a dominant negative mutant of c-Jun, inhibited SPRR3 transactivation. c-Jun increased expression of SPPR3 mainly via a PKC/JNK pathway in response to TPA in KYSE450 cells. Furthermore, c-Jun was remarkably reduced in esophageal cancer. Interestingly, cystatin A, involucrin and SPRR3 were significantly downregulated as well, and associated with differentiation grade. Expression of c-Jun was correlated with the expression of these genes in normal epithelium and ESCC. Importantly, the expression of these genes was remarkably decreased during the malignant transformation from normal epithelium to low-grade intraepithelial neoplasia (LGIN or high-grade intraepithelial neoplasia (HGIN. The expression of cystatin A and involucrin was significantly reduced from LGIN to HGIN. These results suggest c-Jun was involved in the regulation of differentiation-associated genes in ESCC. These genes might serve as the potential markers in distinguishing normal epithelium from esophageal squamous intraepithelial neoplasia.

  16. Decreased Libido

    Science.gov (United States)

    ... causes decreased libido? Decreased libido often accompanies other sexual disorders. Although most men with erectile dysfunction do not complain of decreased libido, after time, persistent failure with erections and sexual performance can lead to reduced sex drive in ...

  17. Pentoxifylline sensitizes human cervical tumor cells to cisplatin-induced apoptosis by suppressing NF-kappa B and decreased cell senescence

    International Nuclear Information System (INIS)

    Hernandez-Flores, Georgina; Bravo-Cuellar, Alejandro; Ortiz-Lazareno, Pablo C; Lerma-Diaz, Jose Manuel; Dominguez-Rodriguez, Jorge R; Jave-Suarez, Luis F; Aguilar-Lemarroy, Adriana del C; Celis-Carrillo, Ruth de; Toro-Arreola, Susana del; Castellanos-Esparza, Yessica C

    2011-01-01

    Worldwide, cervical cancer is the second most common causes of cancer in women and represents an important mortality rate. Cisplatin (CIS) is a very important antitumoral agent and can lead tumor cells toward two important cellular states: apoptosis and senescence. In some types of cancers pentoxifylline (PTX) sensitizes these cells to the toxic action of chemotherapeutics drugs such as adriamycin, inducing apoptosis. In the present work, we studied in vitro whether PTX alone or in combination with CIS induces apoptosis and/or senescence in cervix cancer HeLa and SiHa cell lines infected with HPV types 16 and 18, respectively, as well as in immortalized keratinocytyes HaCaT cells. HeLa (HPV 18+), SiHa (HPV 16+) cervix cancer cells and non-tumorigenic immortalized HaCaT cells (control) were treated with PTX, CIS or both. The cellular toxicity and survival fraction of PTX and CIS were determinate by WST-1 and clonogenic assays respectively. Apoptosis, caspase activation and phosphorylation of ERK1/2, p38, p65 (NF-κB), Bcl-2 and Bcl-XL anti-apoptotic proteins were determinated by flow cytometry. Senescence by microscopy. Phosphorylation of IκBα and IκB total were measured by ELISA. Pro-apoptotic, anti-apoptotic and senescence genes, as well as HPV-E6/7 mRNA expression, were detected by RT-PCR. Our results show that after 24 hours of incubation PTX per se is toxic for cancer cells affecting cell viability and inducing apoptosis. The toxicity in HaCaT cells was minimal. CIS induces apoptosis in HeLa and SiHa cells and its effect was significantly increases when the cells were treated with PTX + CIS. In all studies there was a direct correlation with levels of caspases (-3, -6, -7, -9 and -8) activity and apoptosis. CIS induces important levels of senescence and phosphorylation of ERK1/2, p38, p65/RELA, and IκBα, and decreased the expression of anti-apoptotic protein Bcl-XL. Surprisingly these levels were significantly reduced by PTX in tumor cells, and at the same

  18. Analyzing the effect of decreasing cytosolic triosephosphate isomerase on Solanum tuberosum hairy root cells using a kinetic-metabolic model.

    Science.gov (United States)

    Valancin, Alexandre; Srinivasan, Balasubrahmanyan; Rivoal, Jean; Jolicoeur, Mario

    2013-03-01

    A kinetic-metabolic model of Solanum tuberosum hairy roots is presented in the interest of understanding the effect on the plant cell metabolism of a 90% decrease in cytosolic triosephosphate isomerase (cTPI, EC 5.3.1.1) expression by antisense RNA. The model considers major metabolic pathways including glycolysis, pentose phosphate pathway, and TCA cycle, as well as anabolic reactions leading to lipids, nucleic acids, amino acids, and structural hexoses synthesis. Measurements were taken from shake flask cultures for six extracellular nutrients (sucrose, fructose, glucose, ammonia, nitrate, and inorganic phosphate) and 15 intracellular compounds including sugar phosphates (G6P, F6P, R5P, E4P) and organic acids (PYR, aKG, SUCC, FUM, MAL) and the six nutrients. From model simulations and experimental data it can be noted that plant cell metabolism redistributes metabolic fluxes to compensate for the cTPI decrease, leading to modifications in metabolites levels. Antisense roots showed increased exchanges between the pentose phosphate pathway and the glycolysis, an increased oxygen uptake and growth rate. Copyright © 2012 Wiley Periodicals, Inc.

  19. Decreased neutrophil-associated miRNA and increased B-cell associated miRNA expression during tuberculosis.

    Science.gov (United States)

    van Rensburg, I C; du Toit, L; Walzl, G; du Plessis, N; Loxton, A G

    2018-05-20

    MicroRNAs are short non-coding RNAs that regulate gene expression by binding to, and suppressing the expression of genes. Research show that microRNAs have potential to be used as biomarkers for diagnosis, treatment response and can be used for therapeutic interventions. Furthermore, microRNA expression has effects on immune cell functions, which may lead to disease. Considering the important protective role of neutrophils and B-cells during M.tb infection, we evaluated the expression of microRNAs, known to alter function of these cells, in the context of human TB. We utilised real-time PCR to evaluate the levels of microRNA transcripts in the peripheral blood of TB cases and healthy controls. We found that neutrophil-associated miR-197-3p, miR-99b-5p and miR-191-5p transcript levels were significantly lower in TB cases. Additionally, B-cell-associated miR-320a, miR-204-5p, miR331-3p and other transcript levels were higher in TB cases. The miRNAs differentially expressed in neutrophils are predominantly implicated in signalling pathways leading to cytokine productions. Here, the decreased expression in TB cases may imply a lack of suppression on signalling pathways, which may lead to increased production of pro-inflammatory cytokines such as interferon-gamma. Furthermore, the miRNAs differentially expressed in B-cells are mostly involved in the induction/suppression of apoptosis. Further functional studies are however required to elucidate the significance and functional effects of changes in the expression of these microRNAs. Copyright © 2018 Elsevier B.V. All rights reserved.

  20. The flavanone homoeriodictyol increases SGLT-1-mediated glucose uptake but decreases serotonin release in differentiated Caco-2 cells.

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    Barbara Lieder

    Full Text Available Flavanoids and related polyphenols, among them hesperitin, have been shown to modulate cellular glucose transport by targeting SGLT-1 and GLUT-2 transport proteins. We aimed to investigate whether homoeriodictyol, which is structurally related to hesperitin, affects glucose uptake in differentiated Caco-2 cells as a model for the intestinal barrier. The results revealed that, in contrast to other polyphenols, the flavanon homoeriodictyol promotes glucose uptake by 29.0 ± 3.83% at a concentration of 100 μM. The glucose uptake stimulating effect was sensitive to phloridzin, but not to phloretin, indicating an involvement of the sodium-coupled glucose transporter SGLT-1, but not of sodium-independent glucose transporters (GLUT. In addition, in contrast to the increased extracellular serotonin levels by stimulation with 500 mM D-(+-glucose, treatment with 100 μM homoeriodictyol decreased serotonin release by -48.8 ± 7.57% in Caco-2 cells via a phloridzin-sensitive signaling pathway. Extracellular serotonin levels were also reduced by -57.1 ± 5.43% after application of 0.01 μM homoeriodictyol to human neural SH-SY5Y cells. In conclusion, we demonstrate that homoeriodictyol affects both the glucose metabolism and the serotonin system in Caco-2 cells via a SGLT-1-meditated pathway. Furthermore, the results presented here support the usage of Caco-2 cells as a model for peripheral serotonin release. Further investigations may address the value of homoeriodictyol in the treatment of anorexia and malnutrition through the targeting of SGLT-1.

  1. Overexpression of CD97 confers an invasive phenotype in glioblastoma cells and is associated with decreased survival of glioblastoma patients.

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    Michael Safaee

    Full Text Available Mechanisms of invasion in glioblastoma (GBM relate to differential expression of proteins conferring increased motility and penetration of the extracellular matrix. CD97 is a member of the epidermal growth factor seven-span transmembrane family of adhesion G-protein coupled receptors. These proteins facilitate mobility of leukocytes into tissue. In this study we show that CD97 is expressed in glioma, has functional effects on invasion, and is associated with poor overall survival. Glioma cell lines and low passage primary cultures were analyzed. Functional significance was assessed by transient knockdown using siRNA targeting CD97 or a non-target control sequence. Invasion was assessed 48 hours after siRNA-mediated knockdown using a Matrigel-coated invasion chamber. Migration was quantified using a scratch assay over 12 hours. Proliferation was measured 24 and 48 hours after confirmed protein knockdown. GBM cell lines and primary cultures were found to express CD97. Knockdown of CD97 decreased invasion and migration in GBM cell lines, with no difference in proliferation. Gene-expression based Kaplan-Meier analysis was performed using The Cancer Genome Atlas, demonstrating an inverse relationship between CD97 expression and survival. GBMs expressing high levels of CD97 were associated with decreased survival compared to those with low CD97 (p = 0.007. CD97 promotes invasion and migration in GBM, but has no effect on tumor proliferation. This phenotype may explain the discrepancy in survival between high and low CD97-expressing tumors. This data provides impetus for further studies to determine its viability as a therapeutic target in the treatment of GBM.

  2. Inhibition of deubiquitinating activity of USP14 decreases tyrosine hydroxylase phosphorylated at Ser19 in PC12D cells.

    Science.gov (United States)

    Nakashima, Akira; Ohnuma, Syuhei; Kodani, Yu; Kaneko, Yoko S; Nagasaki, Hiroshi; Nagatsu, Toshiharu; Ota, Akira

    2016-04-15

    Tyrosine hydroxylase (TH) is the rate-limiting enzyme in catecholamine biosynthesis, and its stability is a fundamental factor to maintain the level of the catecholamines in cells. However, the intracellular stability determined by the degradation pathway remains unknown. In this study, we investigated the mechanism by which phosphorylation of TH affected the proteasome pathway. The inhibition of proteasomes by MG-132 increased the percentage of TH molecules phosphorylated at their Ser19, Ser31 and/or Ser40 among the total TH proteins to about 70% in PC12D cells over a 24-hr period; although the percentage of phosphorylated TH molecules was about 20% under basal conditions. Moreover, the inhibition of proteasomes by epoxomicin with high specificity increased primarily the quantity of TH molecules phosphorylated at their Ser19. The phosphorylation of Ser19 potentiated Ser40 phosphorylation in cells by a process known as hierarchical phosphorylation. Therefore, the proteasome inhibition might result in an increase in the levels of all 3 phosphorylated TH forms, thus complicating interpretation of data. Conversely, activation of proteasome degradation by IU-1, which is an inhibitor for the deubiquitinating activity of USP14, decreased only the quantity of TH molecules phosphorylated at their Ser19, although it did not decrease that of TH phosphorylated at its Ser31 and Ser40 or that of TH molecules. These results suggest that the phosphorylation of Ser19 in the N-terminal portion of TH is critical as a trigger for the degradation of this enzyme by the ubiquitin-proteasome pathway. Copyright © 2016 Elsevier Inc. All rights reserved.

  3. Non-steroidal anti-inflammatory drugs decrease E2F1 expression and inhibit cell growth in ovarian cancer cells.

    Directory of Open Access Journals (Sweden)

    Blanca L Valle

    Full Text Available Epidemiological studies have shown that the regular use of non-steroidal anti-inflammatory (NSAIDs drugs is associated with a reduced risk of various cancers. In addition, in vitro and experiments in mouse models have demonstrated that NSAIDs decrease tumor initiation and/or progression of several cancers. However, there are limited preclinical studies investigating the effects of NSAIDs in ovarian cancer. Here, we have studied the effects of two NSAIDs, diclofenac and indomethacin, in ovarian cancer cell lines and in a xenograft mouse model. Diclofenac and indomethacin treatment decreased cell growth by inducing cell cycle arrest and apoptosis. In addition, diclofenac and indomethacin reduced tumor volume in a xenograft model of ovarian cancer. To identify possible molecular pathways mediating the effects of NSAID treatment in ovarian cancer, we performed microarray analysis of ovarian cancer cells treated with indomethacin or diclofenac. Interestingly, several of the genes found downregulated following diclofenac or indomethacin treatment are transcriptional target genes of E2F1. E2F1 was downregulated at the mRNA and protein level upon treatment with diclofenac and indomethacin, and overexpression of E2F1 rescued cells from the growth inhibitory effects of diclofenac and indomethacin. In conclusion, NSAIDs diclofenac and indomethacin exert an anti-proliferative effect in ovarian cancer in vitro and in vivo and the effects of NSAIDs may be mediated, in part, by downregulation of E2F1.

  4. Blood outgrowth endothelial cells reduce hypoxia-mediated fibroblast to myofibroblast conversion by decreasing pro-angiogenic cytokines

    Science.gov (United States)

    Torres, Evelyn Nieves; Yang, Binxia; Brahmbhatt, Akshaar; Mukhopadhyay, Debabrata; Misra, Sanjay

    2015-01-01

    Purpose Hypoxic conditions cause fibroblasts to differentiate to a smooth muscle cell actin (α -SMA) positive cells (myofibroblasts), which is a hallmark of venous neointimal hyperplasia (VNH) associated with hemodialysis vascular access. The purpose of the present study was to determine if BOEC may reduce fibroblast to myofibroblast conversion under hypoxic conditions and what are the potential mechanisms. Methods An experimental model was used in which fibroblasts and BOEC were subjected to hypoxia under contact and transwell conditions to determine if BOEC reduce the conversion of fibroblasts to myofibroblasts under hypoxic conditions. Gene expression under different conditions was performed. In addition, functional assays including cell proliferation and migration were determined. Results This study demonstrates that contact needs to occur between BOEC and fibroblast for the reduction in hypoxia driven conversion of fibroblast to α-SMA. This is associated with a decrease in several pro-angiogenic genes including vascular endothelial growth factor (VEGF-A), platelet derived growth factor (PDGF), fibroblast growth factor (FGF-1), and matrix metalloproteinase-2 (MMP-2) in fibroblasts in contact with BOEC when compared to fibroblasts alone. In addition, migration is significantly reduced while proliferation remains unchanged. Conclusion This study helps provide rationale for using BOEC delivered to the adventitia of the outflow vein of hemodialysis vascular access to reduce VNH. PMID:25677750

  5. Decreased sensitivity of the automatic white precursor cell channel (WPC) for blast flagging in patients with leukopenia.

    Science.gov (United States)

    Lee, Nuri; Jun, Ju Hyun; Lee, Dong Soon

    2016-06-01

    The advantage of Sysmex XN system is its performance of an automatic reflex test in white cell precursor (WPC) channel, which gives an accurate differential count. We performed a real-time evaluation of the automatic differential count according to WBC number with Sysmex XN series and demonstrated a significant differential impact on blast distinction depending on the number of WBC. We categorized the 49,699 specimen according to WBC number and compared the results of blast flagging in the white cell differential (WDF) channel and WPC channel on the basis of the results of manual differential count. Additionally, clinical impact of missed blasts after running WPC reflex test was analyzed. For patients with WBC under 1.5×10(9)/L, blast flagging for WPC channel showed markedly decreased sensitivity (56%) compared to that of WDF mode (100%). Most of the patients with missed blasts in WPC mode were under chemotherapy or hematopoietic stem cell transplantation. The specificity and efficiency for WPC channel were much higher than that of WDF mode in all range of WBC. Considering the additional reagent cost required for the WPC channel, WPC reflex test is not suitable for patients with leukopenia if it is operated alone. Instead, manual blood film review or double-check with other supplemental equipment should be accompanied. Copyright © 2016 The Canadian Society of Clinical Chemists. Published by Elsevier Inc. All rights reserved.

  6. miR-143 decreases COX-2 mRNA stability and expression in pancreatic cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Pham, Hung [Department of Surgery, UCLA Center of Excellence in Pancreatic Diseases, UCLA David Geffen School of Medicine, University of California – Los Angeles, Los Angeles, CA 90095 (United States); Department of Medicine, Veterans Affair Greater Los Angeles Healthcare System, Los Angeles, CA 90073 (United States); Ekaterina Rodriguez, C.; Donald, Graham W.; Hertzer, Kathleen M.; Jung, Xiaoman S.; Chang, Hui-Hua; Moro, Aune; Reber, Howard A.; Hines, O. Joe [Department of Surgery, UCLA Center of Excellence in Pancreatic Diseases, UCLA David Geffen School of Medicine, University of California – Los Angeles, Los Angeles, CA 90095 (United States); Eibl, Guido, E-mail: geibl@mednet.ucla.edu [Department of Surgery, UCLA Center of Excellence in Pancreatic Diseases, UCLA David Geffen School of Medicine, University of California – Los Angeles, Los Angeles, CA 90095 (United States)

    2013-09-13

    Highlights: •Pancreatic cancer cells express low miR-143 levels and elevated p-MEK, p-MAPK and RREB1. •MEK inhibitors U0126 and PD98059 increase miR-143 expression. •miR-143 decreases COX-2 mRNA stability and expression and PGE{sub 2}. •miR-143 decreases p-p38MAPK, p-MEK, p-MAPK and RREB1 expression. -- Abstract: Small non-coding RNAs, microRNAs (miRNA), inhibit the translation or accelerate the degradation of message RNA (mRNA) by targeting the 3′-untranslated region (3′-UTR) in regulating growth and survival through gene suppression. Deregulated miRNA expression contributes to disease progression in several cancers types, including pancreatic cancers (PaCa). PaCa tissues and cells exhibit decreased miRNA, elevated cyclooxygenase (COX)-2 and increased prostaglandin E{sub 2} (PGE{sub 2}) resulting in increased cancer growth and metastases. Human PaCa cell lines were used to demonstrate that restoration of miRNA-143 (miR-143) regulates COX-2 and inhibits cell proliferation. miR-143 were detected at fold levels of 0.41 ± 0.06 in AsPC-1, 0.20 ± 0.05 in Capan-2 and 0.10 ± 0.02 in MIA PaCa-2. miR-143 was not detected in BxPC-3, HPAF-II and Panc-1 which correlated with elevated mitogen-activated kinase (MAPK) and MAPK kinase (MEK) activation. Treatment with 10 μM of MEK inhibitor U0126 or PD98059 increased miR-143, respectively, by 187 ± 18 and 152 ± 26-fold in BxPC-3 and 182 ± 7 and 136 ± 9-fold in HPAF-II. miR-143 transfection diminished COX-2 mRNA stability at 60 min by 2.6 ± 0.3-fold in BxPC-3 and 2.5 ± 0.2-fold in HPAF-II. COX-2 expression and cellular proliferation in BxPC-3 and HPAF-II inversely correlated with increasing miR-143. PGE{sub 2} levels decreased by 39.3 ± 5.0% in BxPC-3 and 48.0 ± 3.0% in HPAF-II transfected with miR-143. Restoration of miR-143 in PaCa cells suppressed of COX-2, PGE{sub 2}, cellular proliferation and MEK/MAPK activation, implicating this pathway in regulating miR-143 expression.

  7. Progesterone receptor blockade in human breast cancer cells decreases cell cycle progression through G2/M by repressing G2/M genes

    International Nuclear Information System (INIS)

    Clare, Susan E.; Gupta, Akash; Choi, MiRan; Ranjan, Manish; Lee, Oukseub; Wang, Jun; Ivancic, David Z.; Kim, J. Julie; Khan, Seema A.

    2016-01-01

    The synthesis of specific, potent progesterone antagonists adds potential agents to the breast cancer prevention and treatment armamentarium. The identification of individuals who will benefit from these agents will be a critical factor for their clinical success. We utilized telapristone acetate (TPA; CDB-4124) to understand the effects of progesterone receptor (PR) blockade on proliferation, apoptosis, promoter binding, cell cycle progression, and gene expression. We then identified a set of genes that overlap with human breast luteal-phase expressed genes and signify progesterone activity in both normal breast cells and breast cancer cell lines. TPA administration to T47D cells results in a 30 % decrease in cell number at 24 h, which is maintained over 72 h only in the presence of estradiol. Blockade of progesterone signaling by TPA for 24 h results in fewer cells in G2/M, attributable to decreased expression of genes that facilitate the G2/M transition. Gene expression data suggest that TPA affects several mechanisms that progesterone utilizes to control gene expression, including specific post-translational modifications, and nucleosomal organization and higher order chromatin structure, which regulate access of PR to its DNA binding sites. By comparing genes induced by the progestin R5020 in T47D cells with those increased in the luteal-phase normal breast, we have identified a set of genes that predict functional progesterone signaling in tissue. These data will facilitate an understanding of the ways in which drugs such as TPA may be utilized for the prevention, and possibly the therapy, of human breast cancer. The online version of this article (doi:10.1186/s12885-016-2355-5) contains supplementary material, which is available to authorized users

  8. RNA interference-mediated c-MYC inhibition prevents cell growth and decreases sensitivity to radio- and chemotherapy in childhood medulloblastoma cells

    International Nuclear Information System (INIS)

    Bueren, André O von; Shalaby, Tarek; Oehler-Jänne, Christoph; Arnold, Lucia; Stearns, Duncan; Eberhart, Charles G; Arcaro, Alexandre; Pruschy, Martin; Grotzer, Michael A

    2009-01-01

    With current treatment strategies, nearly half of all medulloblastoma (MB) patients die from progressive tumors. Accordingly, the identification of novel therapeutic strategies remains a major goal. Deregulation of c-MYC is evident in numerous human cancers. In MB, over-expression of c-MYC has been shown to cause anaplasia and correlate with unfavorable prognosis. To study the role of c-MYC in MB biology, we down-regulated c-MYC expression by using small interfering RNA (siRNA) and investigated changes in cellular proliferation, cell cycle analysis, apoptosis, telomere maintenance, and response to ionizing radiation (IR) and chemotherapeutics in a representative panel of human MB cell lines expressing different levels of c-MYC (DAOY wild-type, DAOY transfected with the empty vector, DAOY transfected with c-MYC, D341, and D425). siRNA-mediated c-MYC down-regulation resulted in an inhibition of cellular proliferation and clonogenic growth, inhibition of G1-S phase cell cycle progression, and a decrease in human telomerase reverse transcriptase (hTERT) expression and telomerase activity. On the other hand, down-regulation of c-MYC reduced apoptosis and decreased the sensitivity of human MB cells to IR, cisplatin, and etoposide. This effect was more pronounced in DAOY cells expressing high levels of c-MYC when compared with DAOY wild-type or DAOY cells transfected with the empty vector. In human MB cells, in addition to its roles in growth and proliferation, c-MYC is also a potent inducer of apoptosis. Therefore, targeting c-MYC might be of therapeutic benefit when used sequentially with chemo- and radiotherapy rather than concomitantly

  9. Increased extracellular matrix density decreases MCF10A breast cell acinus formation in 3D culture conditions.

    Science.gov (United States)

    Lance, Amanda; Yang, Chih-Chao; Swamydas, Muthulekha; Dean, Delphine; Deitch, Sandy; Burg, Karen J L; Dréau, Didier

    2016-01-01

    The extracellular matrix (ECM) contributes to the generation and dynamic of normal breast tissue, in particular to the generation of polarized acinar and ductal structures. In vitro 3D culture conditions, including variations in the composition of the ECM, have been shown to directly influence the formation and organization of acinus-like and duct-like structures. Furthermore, the density of the ECM appears to also play a role in the normal mammary tissue and tumour formation. Here we show that the density of the ECM directly influences the number, organization and function of breast acini. Briefly, non-malignant human breast MCF10A cells were incubated in increasing densities of a Matrigel®-collagen I matrix. Elastic moduli near and distant to the acinus structures were measured by atomic force microscopy, and the number of acinus structures was determined. Immunochemistry was used to investigate the expression levels of E-cadherin, laminin, matrix metalloproteinase-14 and ß-casein in MCF10A cells. The modulus of the ECM was significantly increased near the acinus structures and the number of acinus structures decreased with the increase in Matrigel-collagen I density. As evaluated by the expression of laminin, the organization of the acinus structures present was altered as the density of the ECM increased. Increases in both E-cadherin and MMP14 expression by MCF10A cells as ECM density increased were also observed. In contrast, MCF10A cells expressed lower ß-casein levels as the ECM density increased. Taken together, these observations highlight the key role of ECM density in modulating the number, organization and function of breast acini. Copyright © 2013 John Wiley & Sons, Ltd.

  10. Decreased MiR-200a/141 Suppress Cell Migration and Proliferation by Targeting PTEN in Hirschsprung's Disease

    Directory of Open Access Journals (Sweden)

    Hongxing Li

    2014-08-01

    Full Text Available Background/Aims: Hirschsprung's disease (HSCR is a genetic disorder of neural crest development. In this study, we investigated whether and how miR-200a and miR-141, belonging to miR-200 family, were involved in the pathogenesis of HSCR. Methods: Quantitative real time PCR and Western blot were used to detect the levels of miRNA, mRNAs, and proteins in colon tissues from 88 HSCR patients and 75 controls. The direct regulation of specific mRNA by miRNAs was validated by dual-luciferase reporter assay and RNA interference in cell lines. Transwell assays, CCK8 assay, and flow cytometry were inplemented to measure viability and activities of human 293T and SH-SY5Y cells, respectively. Results: Aberrant suppression of miR-200a was observed in colon tissues of HSCR patients. A decreased level of miR-200a and miR-141 correlated with increased levels of PTEN mRNA and protein. The Dual-Luciferase reporter gene assay demonstrated that miR-200a and miR-141 binded directly to 3'UTR of PTEN and resulting in the inhibition of PTEN. The reductions in miR-200a and miR-141 inhibited migration and proliferation of 293T and SH-SY5Y cells through up-regulating the expression of PTEN. Moreover, knocking-down of PTEN rescued the extent of suppressed cell migration and proliferation induced by miR-200a and miR-141. Conclusions: The miR-200 family may play a crucial role in the pathogenesis of HSCR by co-regulating PTEN.

  11. PA-X protein decreases replication and pathogenicity of swine influenza virus in cultured cells and mouse models.

    Science.gov (United States)

    Gong, Xiao-Qian; Sun, Ying-Feng; Ruan, Bao-Yang; Liu, Xiao-Min; Wang, Qi; Yang, Hai-Ming; Wang, Shuai-Yong; Zhang, Peng; Wang, Xiu-Hui; Shan, Tong-Ling; Tong, Wu; Zhou, Yan-Jun; Li, Guo-Xin; Zheng, Hao; Tong, Guang-Zhi; Yu, Hai

    2017-06-01

    Swine influenza viruses have been circulating in pigs throughout world and might be potential threats to human health. PA-X protein is a newly discovered protein produced from the PA gene by ribosomal frameshifting and the effects of PA-X on the 1918 H1N1, the pandemic 2009 H1N1, the highly pathogenic avian H5N1 and the avian H9N2 influenza viruses have been reported. However, the role of PA-X in the pathogenesis of swine influenza virus is still unknown. In this study, we rescued the H1N1 wild-type (WT) classical swine influenza virus (A/Swine/Guangdong/1/2011 (H1N1)) and H1N1 PA-X deficient virus containing mutations at the frameshift motif, and compared their replication properties and pathogenicity of swine influenza virus in vitro and in vivo. Our results show that the expression of PA-X inhibits virus replication and polymerase activity in cultured cells and decreases virulence in mouse models. Therefore, our study demonstrates that PA-X protein acts as a negative virulence regulator for classical H1N1 swine influenza virus and decreases virulence by inhibiting viral replication and polymerase activity, deepening our understanding of the pathogenesis of swine influenza virus. Copyright © 2017 Elsevier B.V. All rights reserved.

  12. Leydig cell number and sperm production decrease induced by chronic ametryn exposure: a negative impact on animal reproductive health.

    Science.gov (United States)

    Dantas, T A; Cancian, G; Neodini, D N R; Mano, D R S; Capucho, C; Predes, F S; Pulz, R Barbieri; Pigoso, A A; Dolder, H; Severi-Aguiar, G D C

    2015-06-01

    Ametryn is an herbicide used to control broadleaf and grass weeds and its acute and chronic toxicity is expected to be low. Since toxicological data on ametryn is scarce, the aim of this study was to evaluate rat reproductive toxicity. Thirty-six adult male Wistar rats (90 days) were divided into three groups: Co (control) and T1 and T2 exposed to 15 and 30 mg/kg/day of ametryn, respectively, for 56 days. Testicular analysis demonstrated that ametryn decreased sperm number per testis, daily sperm production, and Leydig cell number in both treated groups, although little perceptible morphological change has been observed in seminiferous tubule structure. Lipid peroxidation was higher in group T2, catalase activity decreased in T1 group, superoxide dismutase activity diminished, and a smaller number of sulphydryl groups of total proteins were verified in both exposed groups, suggesting oxidative stress. These results showed negative ametryn influence on the testes and can compromise animal reproductive performance and survival.

  13. Early transcutaneous electrical nerve stimulation reduces hyperalgesia and decreases activation of spinal glial cells in mice with neuropathic pain.

    Science.gov (United States)

    Matsuo, Hideaki; Uchida, Kenzo; Nakajima, Hideaki; Guerrero, Alexander Rodriguez; Watanabe, Shuji; Takeura, Naoto; Sugita, Daisuke; Shimada, Seiichiro; Nakatsuka, Terumasa; Baba, Hisatoshi

    2014-09-01

    Although transcutaneous electrical nerve stimulation (TENS) is widely used for the treatment of neuropathic pain, its effectiveness and mechanism of action in reducing neuropathic pain remain uncertain. We investigated the effects of early TENS (starting from the day after surgery) in mice with neuropathic pain, on hyperalgesia, glial cell activation, pain transmission neuron sensitization, expression of proinflammatory cytokines, and opioid receptors in the spinal dorsal horn. Following nerve injury, TENS and behavioral tests were performed every day. Immunohistochemical, immunoblot, and flow cytometric analysis of the lumbar spinal cord were performed after 8 days. Early TENS reduced mechanical and thermal hyperalgesia and decreased the activation of microglia and astrocytes (PEarly TENS decreased p-p38 within microglia (Pearly TENS relieved hyperalgesia in our mouse model of neuropathic pain by inhibiting glial activation, MAP kinase activation, PKC-γ, and p-CREB expression, and proinflammatory cytokines expression, as well as maintenance of spinal opioid receptors. The findings indicate that TENS treatment is more effective when applied as early after nerve injury as possible. Copyright © 2014 International Association for the Study of Pain. Published by Elsevier B.V. All rights reserved.

  14. Arsenic trioxide decreases the amount and inhibits the function of regulatory T cells, which may contribute to its efficacy in the treatment of acute promyelocytic leukemia.

    Science.gov (United States)

    Xu, Wen; Li, Xiaoxia; Quan, Lina; Yao, Jiying; Mu, Guannan; Guo, Jingjie; Wang, Yitong

    2018-03-01

    Arsenic trioxide (ATO) exhibits substantial clinical efficacy in the treatment of acute promyelocytic leukemia (APL). Here, we investigated whether ATO exerts its efficacy by affecting regulatory T (Treg) cells. We determined whether ATO treatment influenced the amount and function of purified Treg cells. We also examined the effect of ATO treatment on Treg cells from APL patients. ATO treatment induced apoptosis in purified Treg cells and dampened the inhibition of effector T (Teff) cells proliferation and the secretion of cytokine by Treg cells. Treg cell levels in the peripheral blood and serum IL-10 levels were dramatically decreased in APL patients after single ATO treatment. In summary, our results show that ATO decreases the amount and inhibits the function of Treg cells, thereby enhancing Teff cell function and overall anti-tumor immunity.

  15. An earthworm protease cleaving serum fibronectin and decreasing HBeAg in HepG2.2.15 cells

    Directory of Open Access Journals (Sweden)

    Zhao Jing

    2008-11-01

    Full Text Available Abstract Background Virus-binding activity is one of the important functions of fibronectin (FN. It has been reported that a high concentration of FN in blood improves the transmission frequency of hepatitis viruses. Therefore, to investigate a protease that hydrolyzes FN rapidly is useful to decrease the FN concentration in blood and HBV infection. So far, however, no specific protease digesting FN in serum has been reported. Methods We employed a purified earthworm protease to digest serum proteins. The rapidly cleaved protein (FN was identified by MALDI-TOF MS and western blotting. The cleavage sites were determined by N-terminus amino acid residues sequencing. The protease was orally administrated to rats to investigate whether serum FN in vivo became decreased. The serum FN was determined by western blotting and ELISA. In cytological studies, the protease was added to the medium in the culture of HepG2.2.15 cells and then HBsAg and HBeAg were determined by ELISA. Results The protease purified from earthworm Eisenia fetida was found to function as a fibronectinase (FNase. The cleavage sites on FN by the FNase were at R and K, exhibiting a trypsin alkaline serine-like function. The earthworm fibronectinase (EFNase cleaved FN at four sites, R259, R1005, K1557 and R2039, among which the digested fragments at R259, K1557 and R2039 were related to the virus-binding activity as reported. The serum FN was significantly decreased when the earthworm fibronectinase was orally administrated to rats. The ELISA results showed that the secretion of HBeAg from HepG2.2.15 cells was significantly inhibited in the presence of the FNase. Conclusion The earthworm fibronectinase (EFNase cleaves FN much faster than the other proteins in serum, showing a potential to inhibit HBV infection through its suppressing the level of HBeAg. This suggests that EFNase is probably used as one of the candidates for the therapeutic agents to treat hepatitis virus infection.

  16. miR-122 targets NOD2 to decrease intestinal epithelial cell injury in Crohn’s disease

    Energy Technology Data Exchange (ETDEWEB)

    Chen, Yu; Wang, Chengxiao; Liu, Ying; Tang, Liwei; Zheng, Mingxia [Department of Pediatrics, Jiangwan Hospital of Shanghai, Shanghai 200434 (China); Xu, Chundi [Department of Pediatrics, Ruijin affiliated to Shanghai Jiaotong University School of Medicine, Shanghai, 200025 (China); Song, Jian, E-mail: jiansongkxy@126.com [Department of Gastroenterology, Jiangwan Hospital of Shanghai, Shanghai 200434 (China); Meng, Xiaochun [Department of Pediatrics, Jiangwan Hospital of Shanghai, Shanghai 200434 (China)

    2013-08-16

    Highlights: •NOD2 is a target gene of miR-122. •miR-122 inhibits LPS-induced apoptosis by suppressing NOD2 in HT-29 cells. •miR-122 reduces the expression of pro-inflammatory cytokines (TNF-α and IFN-γ). •miR-122 promotes the release of anti-inflammatory cytokines (IL-4 and IL-10). •NF-κB signaling pathway is involved in inflammatory response induced by LPS. -- Abstract: Crohn’s disease (CD) is one of the two major types of inflammatory bowel disease (IBD) thought to be caused by genetic and environmental factors. Recently, miR-122 was found to be deregulated in association with CD progression. However, the underlying molecular mechanisms remain unclear. In the present study, the gene nucleotide-binding oligomerization domain 2 (NOD2/CARD15), which is strongly associated with susceptibility to CD, was identified as a functional target of miR-122. MiR-122 inhibited LPS-induced apoptosis by suppressing NOD2 in HT-29 cells. NOD2 interaction with LPS initiates signal transduction mechanisms resulting in the activation of nuclear factor κB (NF-κB) and the stimulation of downstream pro-inflammatory events. The activation of NF-κB was inhibited in LPS-stimulated HT-29 cells pretreated with miR-122 precursor or NOD2 shRNA. The expression of the pro-inflammatory cytokines TNF-α and IFN-γ was significantly decreased, whereas therelease of the anti-inflammatory cytokines IL-4 and IL-10 was increased in LPS-stimulated HT-29 cells pretreated with miR-122 precursor, NOD2 shRNA or the NF-κB inhibitor QNZ. Taken together, these results indicate that miR-122 and its target gene NOD2 may play an important role in the injury of intestinal epithelial cells induced by LPS.

  17. Aspirin rectifies calcium homeostasis, decreases reactive oxygen species, and increases NO production in high glucose-exposed human endothelial cells.

    Science.gov (United States)

    Dragomir, Elena; Manduteanu, Ileana; Voinea, Manuela; Costache, Gabi; Manea, Adrian; Simionescu, Maya

    2004-01-01

    Aspirin's pharmacological action is mainly related to its property to inhibit prostaglandin synthesis; apart from this, aspirin has some beneficial side effects that are not completely understood, yet. Since aspirin possesses antioxidant properties and antioxidants prevent high d-glucose enhanced endothelial [Ca(2+)](i), we questioned whether aspirin also has an effect on this process as well as on high-glucose-impaired nitric oxide (NO) production. For these purposes, human endothelial cells (HECs) were cultured in normal concentration (5 mM) glucose (NG) or high concentration (33 mM) glucose (HG) and after confluence, exposed for 48 h to HG in the absence or presence of 1 mM aspirin. Then, the [Ca(2+)](i) was measured fluorimetrically using fura-2, NO production was determined by Griess reaction, superoxide anions (O(2)) was evaluated by ferricytochrome c reduction, the intracellular reactive oxygen species (ROS) were evaluated by fluorimetry, and the levels of protein kinase C (PKC) by Western blot. The results showed that HECs exposed to HG displayed: (i) increased [Ca(2+)](i); (ii) enhanced O(2) release; (iii) augmented level of intracellular ROS; and (iv) PKC translocation to the membrane fraction. By comparison, exposure to cells grown in HG to 1 mM aspirin resulted in: (i) a reduction of histamine stimulated [Ca(2+)](i) release to control level and of [Ca(2+)](i) entry by 30%; (ii) a twofold increase in NO production; (iii) a decrease of O(2)(-) accumulation in both culture medium and cell homogenate (by 60.4% and 70%, respectively); (iv) a decline of ROS to the control levels; and (v) a reduction of PKC translocation to the control levels. These data indicate that aspirin corrects the high-glucose-induced changes in cellular Ca(2+) homeostasis and NO production, via a mechanism involving the reduction of the O(2)(-) levels possible by acting on PKC-induced NADPH activity.

  18. HU-446 and HU-465, Derivatives of the Non-psychoactive Cannabinoid Cannabidiol, Decrease the Activation of Encephalitogenic T Cells.

    Science.gov (United States)

    Kozela, Ewa; Haj, Christeene; Hanuš, Lumir; Chourasia, Mukesh; Shurki, Avital; Juknat, Ana; Kaushansky, Nathali; Mechoulam, Raphael; Vogel, Zvi

    2016-01-01

    Cannabidiol (CBD), the non-psychoactive cannabinoid, has been previously shown by us to decrease peripheral inflammation and neuroinflammation in mouse experimental autoimmune encephalomyelitis (EAE) model of multiple sclerosis (MS). Here we have studied the anti-inflammatory effects of newly synthesized derivatives of natural (-)-CBD ((-)-8,9-dihydro-7-hydroxy-CBD; HU-446) and of synthetic (+)-CBD ((+)-8,9-dihydro-7-hydroxy-CBD; HU-465) on activated myelin oligodendrocyte glycoprotein (MOG)35-55-specific mouse encephalitogenic T cells (T(MOG) ) driving EAE/MS-like pathologies. Binding assays followed by molecular modeling revealed that HU-446 has negligible affinity toward the cannabinoid CB1 and CB2 receptors while HU-465 binds to both CB1 and CB2 receptors at the high nanomolar concentrations (Ki = 76.7 ± 5.8 nm and 12.1 ± 2.3 nm, respectively). Both, HU-446 and HU-465, at 5 and 10 μm (but not at 0.1 and 1 μm), inhibited the MOG35-55-induced proliferation of autoreactive T(MOG) cells via CB1/CB2 receptor independent mechanisms. Moreover, both HU-446 and HU-465, at 5 and 10 μm, inhibited the release of IL-17, a key autoimmune cytokine, from MOG35-55-stimulated T(MOG) cells. These results suggest that HU-446 and HU-465 have anti-inflammatory potential in inflammatory and autoimmune diseases. © 2015 John Wiley & Sons A/S.

  19. Increased IL-4- and IL-17-producing CD8+ cells are related to decreased CD39+CD4+Foxp3+ cells in allergic asthma.

    Science.gov (United States)

    Li, Ping; Yang, Qun-Zhen; Wang, Wei; Zhang, Gu-Qin; Yang, Jiong

    2018-01-01

    In allergic asthma, regulatory T cell (Treg) number and function are decreased. Antigen-primed CD8 + T cells play an indispensable role in the full development of airway inflammation and airway hyper-responsiveness (AHR) occurring in asthma. In this study, we investigated the relationship between subpopulations of CD8 + T cells and CD39 + Tregs. Female C57BL/6 mice were used to develop the model of allergic asthma. Experimental mice were immunized with ovalbumin (OVA) by intra-peritoneal (i.p) injection and then challenged with OVA by intra-tracheal administration. Control mice were immunized with vehicle by i.p injection and challenged with OVA. Airway inflammation was determined by histology and AHR was measured by an invasive method. Levels of interferon (IFN)-γ, IL-4, and IL-17 in bronchoalveolar lavage fluid (BALF) were determined by enzyme-linked immunosorbent assay. The frequencies of CD8 + IFN-γ + cells (Tc1), CD8 + IL-4 + cells (Tc2), CD8 + IL-17 + cells (Tc17), and CD39 + Tregs were measured by flow cytometry. The correlation between CD39 + Tregs and Tc subsets was analyzed by Pearson's test. Experimental mice displayed phenotypes of allergic asthma, including inflammatory cell infiltration into the lungs, goblet cell hyperplasia, increased airway resistance, and increased IL-4 and IL-17 in BALF. Compared to control mice, experimental mice displayed lower CD39 + Tregs and Tc1 but higher Tc2 and Tc17. There was a negative correlation between CD39 + Tregs and Tc2 or Tc17. In allergic asthma, increased Tc2 and Tc17 are possibly related to insufficient CD39 + Tregs.

  20. Nitrogen deficiency inhibits leaf blade growth in Lolium perenne by increasing cell cycle duration and decreasing mitotic and post-mitotic growth rates.

    Science.gov (United States)

    Kavanová, Monika; Lattanzi, Fernando Alfredo; Schnyder, Hans

    2008-06-01

    Nitrogen deficiency severely inhibits leaf growth. This response was analysed at the cellular level by growing Lolium perenne L. under 7.5 mM (high) or 1 mM (low) nitrate supply, and performing a kinematic analysis to assess the effect of nitrogen status on cell proliferation and cell growth in the leaf blade epidermis. Low nitrogen supply reduced leaf elongation rate (LER) by 43% through a similar decrease in the cell production rate and final cell length. The former was entirely because of a decreased average cell division rate (0.023 versus 0.032 h(-1)) and thus longer cell cycle duration (30 versus 22 h). Nitrogen status did not affect the number of division cycles of the initial cell's progeny (5.7), and accordingly the meristematic cell number (53). Meristematic cell length was unaffected by nitrogen deficiency, implying that the division and mitotic growth rates were equally impaired. The shorter mature cell length arose from a considerably reduced post-mitotic growth rate (0.033 versus 0.049 h(-1)). But, nitrogen stress did not affect the position where elongation stopped, and increased cell elongation duration. In conclusion, nitrogen deficiency limited leaf growth by increasing the cell cycle duration and decreasing mitotic and post-mitotic elongation rates, delaying cell maturation.

  1. When larger brains do not have more neurons: increased numbers of cells are compensated by decreased average cell size across mouse individuals

    Science.gov (United States)

    Herculano-Houzel, Suzana; Messeder, Débora J.; Fonseca-Azevedo, Karina; Pantoja, Nilma A.

    2015-01-01

    There is a strong trend toward increased brain size in mammalian evolution, with larger brains composed of more and larger neurons than smaller brains across species within each mammalian order. Does the evolution of increased numbers of brain neurons, and thus larger brain size, occur simply through the selection of individuals with more and larger neurons, and thus larger brains, within a population? That is, do individuals with larger brains also have more, and larger, neurons than individuals with smaller brains, such that allometric relationships across species are simply an extension of intraspecific scaling? Here we show that this is not the case across adult male mice of a similar age. Rather, increased numbers of neurons across individuals are accompanied by increased numbers of other cells and smaller average cell size of both types, in a trade-off that explains how increased brain mass does not necessarily ensue. Fundamental regulatory mechanisms thus must exist that tie numbers of neurons to numbers of other cells and to average cell size within individual brains. Finally, our results indicate that changes in brain size in evolution are not an extension of individual variation in numbers of neurons, but rather occur through step changes that must simultaneously increase numbers of neurons and cause cell size to increase, rather than decrease. PMID:26082686

  2. Growth and maturation of heart valves leads to changes in endothelial cell distribution, impaired function, decreased metabolism and reduced cell proliferation.

    Science.gov (United States)

    Anstine, Lindsey J; Bobba, Chris; Ghadiali, Samir; Lincoln, Joy

    2016-11-01

    Risk factors of heart valve disease are well defined and prolonged exposure throughout life leads to degeneration and dysfunction in up to 33% of the population. While aortic valve replacement remains the most common need for cardiovascular surgery particularly in those aged over 65, the underlying mechanisms of progressive deterioration are unknown. In other cardiovascular systems, a decline in endothelial cell integrity and function play a major role in promoting pathological changes, and while similar mechanisms have been speculated in the valves, studies to support this are lacking. The goal of this study was to examine age-related changes in valve endothelial cell (VEC) distribution, morphology, function and transcriptomes during critical stages of valve development (embryonic), growth (postnatal (PN)), maintenance (young adult) and aging (aging adult). Using a combination of in vivo mouse, and in vitro porcine assays we show that VEC function including, nitric oxide bioavailability, metabolism, endothelial-to-mesenchymal potential, membrane self-repair and proliferation decline with age. In addition, density of VEC distribution along the endothelium decreases and this is associated with changes in morphology, decreased cell-cell interactions, and increased permeability. These changes are supported by RNA-seq analysis showing that focal adhesion-, cell cycle-, and oxidative phosphorylation-associated biological processes are negatively impacted by aging. Furthermore, by performing high-throughput analysis we are able to report the differential and common transcriptomes of VECs at each time point that can provide insights into the mechanisms underlying age-related dysfunction. These studies suggest that maturation of heart valves over time is a multifactorial process and this study has identified several key parameters that may contribute to impairment of the valve to maintain critical structure-function relationships; leading to degeneration and disease

  3. Anti-IL-2 receptor antibody decreases cytokine-induced apoptosis of human renal tubular epithelial cells (TEC).

    Science.gov (United States)

    Wang, Shuang; Zhang, Zhu-Xu; Yin, Ziqin; Liu, Weihua; Garcia, Bertha; Huang, Xuyan; Acott, Philip; Jevnikar, Anthony M

    2011-07-01

    Transplant rejection is mediated by T-cell activation which is modulated by interleukin-2 (IL-2) binding to IL-2R (CD25). Monoclonal anti-IL-2 receptor antibody is used in renal transplantation to reduce rejection. Interestingly, proximal tubular epithelial cells (TEC) express CD25, similar to T cells. We have demonstrated that IL-2 induces murine TEC apoptosis through down-regulation of the caspase-8 inhibitor protein c-FLIP. Anti-CD25 antibody may be useful clinically to limit renal injury, but this has not been tested in human TEC. Human PT-2 TEC were isolated and cloned from the urine of transplant patients. Apoptosis was determined by FACS with Annexin-V FITC. Protein expression was studied using western blot, and mRNA levels by quantitative real-time (PR-PCR). We demonstrated that the morphology of a human kidney cell line (PT-2) cloned from urine was consistent with proximal TEC and expresses alkaline phosphatase, cytokeratin, vimentin, CD13, CD26, and low levels of E-cadherin. Basal IL-2 receptor (CD25) was up-regulated by IL-2/IFN-γ stimulation, and cytokine exposure induced apoptosis in a dose-dependent manner. Apoptosis with IL-2/IFN-γ was associated with increased caspase-8 activity and decreased endogenous caspase-8 inhibitor c-FLIP mRNA and protein expression. IL-2/IFN-γ-induced apoptosis could be blocked by pre-treatment of PT-2 with anti IL-2R antibody (basiliximab) but not control IgG antibody. These data demonstrate for the first time in human TEC that IL-2 and IFN-γ can induce TEC apoptosis which can be blocked by CD25 blockade antibody. These data suggest that anti-CD25 mAb might similarly attenuate inflammation-induced TEC injury in vivo. Kidney-expressed CD25 may represent a clinically important new target for attenuating early inflammatory injury in donor kidneys and preserving renal function during anti-rejection therapy.

  4. GroEL1, a heat shock protein 60 of Chlamydia pneumoniae, impairs neovascularization by decreasing endothelial progenitor cell function.

    Directory of Open Access Journals (Sweden)

    Yi-Wen Lin

    Full Text Available The number and function of endothelial progenitor cells (EPCs are sensitive to hyperglycemia, hypertension, and smoking in humans, which are also associated with the development of atherosclerosis. GroEL1 from Chlamydia pneumoniae has been found in atherosclerotic lesions and is related to atherosclerotic pathogenesis. However, the actual effects of GroEL1 on EPC function are unclear. In this study, we investigate the EPC function in GroEL1-administered hind limb-ischemic C57BL/B6 and C57BL/10ScNJ (a toll-like receptor 4 (TLR4 mutation mice and human EPCs. In mice, laser Doppler imaging, flow cytometry, and immunohistochemistry were used to evaluate the degree of neo-vasculogenesis, circulating level of EPCs, and expression of CD34, vWF, and endothelial nitric oxide synthase (eNOS in vessels. Blood flow in the ischemic limb was significantly impaired in C57BL/B6 but not C57BL/10ScNJ mice treated with GroEL1. Circulating EPCs were also decreased after GroEL1 administration in C57BL/B6 mice. Additionally, GroEL1 inhibited the expression of CD34 and eNOS in C57BL/B6 ischemic muscle. In vitro, GroEL1 impaired the capacity of differentiation, mobilization, tube formation, and migration of EPCs. GroEL1 increased senescence, which was mediated by caspases, p38 MAPK, and ERK1/2 signaling in EPCs. Furthermore, GroEL1 decreased integrin and E-selectin expression and induced inflammatory responses in EPCs. In conclusion, these findings suggest that TLR4 and impaired NO-related mechanisms could contribute to the reduced number and functional activity of EPCs in the presence of GroEL1 from C. pneumoniae.

  5. Surgical Management of Advanced Non-Small Cell Lung Cancer Is Decreasing But Is Associated With Improved Survival.

    Science.gov (United States)

    David, Elizabeth A; Canter, Robert J; Chen, Yingjia; Cooke, David T; Cress, Rosemary D

    2016-10-01

    For patients with advanced stage non-small cell lung cancer (NSCLC), chemotherapy and chemoradiation are the principal treatment modalities, and the role of surgical resection remains unclear. Our objective was to evaluate current trends and oncologic outcomes for advanced stage NSCLC. We hypothesized that surgery is associated with increased survival and may be an underutilized treatment modality. The California Cancer Registry was queried from 2004 to 2012 for cases of stage IIIA, IIIB, and IV NSCLC, and we identified 34,016 cases. Patients were categorized by treatment group, and linear regression was used to calculate trends in treatment and predictors of treatment group. Kaplan-Meier and Cox regression modeling were used to determine the influence of treatment group on overall survival. Twenty-seven percent of patients (9,223 of 34,016) received no treatment. For the entire cohort, treatment with chemotherapy alone increased (p < 0.001), but treatment with radiation alone, surgery alone, or in any combination decreased (p = 0.011, p < 0.001, p = 0.021, p = 0.007, and p = 0.094). Treatment group, age, sex, race, socioeconomic status, stage, histology, and tumor size were all significant predictors of overall survival. Overall survival was significantly longer for patients who had surgery as part of their treatment regimen (p < 0.001). For patients with advanced stage NSCLC, the use of multimodality regimens that include surgery are decreasing despite longer overall survival. Future studies are needed to identify the demographics and clinical characteristics of patients with advanced stage NSCLC who may benefit from surgery. Copyright © 2016 The Society of Thoracic Surgeons. Published by Elsevier Inc. All rights reserved.

  6. Increase vs. decrease of calcium uptake by isolated heart cells induced by H2O2 vs. HOCl

    International Nuclear Information System (INIS)

    Kaminishi, T.; Matsuoka, T.; Yanagishita, T.; Kako, K.J.

    1989-01-01

    Adult rat heart myocytes were labeled rapidly with exogenous [45Ca2+]. Addition of 2.5 mM H2O2 to the heart cell suspension raised the content of rapidly exchangeable intracellular Ca2+ twofold, whereas addition of 1-30 mM HOCl decreased the Ca2+ content. The H2O2-induced increase in Ca2+ content was dependent on the medium Na+, pH, and temperature but was not significantly affected by addition of verapamil, diltiazem, amiloride, or 3-aminobenzamide. The [3H]ouabain binding to myocytes was suppressed by H2O2, whereas the Ca2+ efflux from myocytes was not influenced. An uncoupler, carbonyl cyanide m-chlorophenylhydrazone, reduced Ca2+ content, implying that the H2O2-induced change in Ca2+ content was not directly related to ATP depletion. On the other hand, the H2O2-induced Ca2+ accumulation in myocytes was prevented by deferoxamine or o-phenanthroline. These results suggest that H2O2 inhibited Na+-K+-ATPase, resulting in an increase in intracellular Na+ concentration and stimulation of sarcolemmal Na+-Ca2+ exchange activity, which caused a transient net Ca2+ influx into myocytes. By contrast, HOCl decreased the Ca2+ content of the rapidly exchangeable pool below control levels and this action of HOCl was antagonized by 1,4-dithiothreitol. HOCl accelerated Ca2+ efflux from myocytes. Ca2+ uptake and Ca2+-ATPase of the isolated sarcoplasmic reticular (SR) fraction were highly sensitive to the action of HOCl. Ca2+ uptake by intracellular sites, studied with myocytes permeabilized with digitonin, was inhibited by both H2O2 and HOCl. Thus these results suggest that HOCl inhibits the SR Ca2+ pump, resulting in the observed acceleration of Ca2+ efflux from and decline in Ca2+ content of myocytes

  7. Quercus infectoria and Terminalia chebula decrease melanin content and tyrosinase activity in B16/F10 cell lines

    Directory of Open Access Journals (Sweden)

    Akram Jamshidzadeh

    2017-10-01

    Full Text Available Context: One of the most complained skin cares in ethnic skin like Asian women is hyperpigmentation, and lightening preparations have been long-standing desired. Due to the side effects of current drugs, medicinal plants have attracted more attentions as a source of novel drugs. Mazo (Quercus infectoria galls and Terminalia chebula fruits have been suggested in Persian Traditional Medicine as a safe treatment for hyperpigmentation. Aims: To evaluate the cytotoxicity and the effect on melanin synthesis in B16/F10 melanoma of Q. infectoria and T. chebula extracts. Methods: After collection and scientific authentication, plants were extracted by maceration method with methanol and were standardized based on total phenolic content. MTT assay and colorimetric method were used for cytotoxicity and determination of melanin content and tyrosinase activity in B16/F10 cells, respectively. Kojic acid was used as a reference compound. Results: Total phenolic content of Q. infectoria and T. chebula was determined as 287.34 ± 4.21 and 172.61 ± 8.67 mg gallic acid equivalent/g dried extract, respectively. Both plants decreased cell viability at 100 µg/mL and significantly reduced intercellular melanin to 66.25% and 71.1%, respectively in comparison to kojic acid (56.63% at 50 µg/mL. In the same concentration, 65.7% and 71.2% tyrosinase activity was inhibited by Q. infectoria and T. chebula, which significantly were different from control (p<0.001. Conclusions: These findings suggest that both plants especially Q. infectoria could inhibit melanogenesis in non-toxic concentrations and would be a good candidate for further studies.

  8. High COX-2 expression in canine mast cell tumours is associated with proliferation, angiogenesis and decreased overall survival.

    Science.gov (United States)

    Gregório, H; Raposo, T; Queiroga, F L; Pires, I; Pena, L; Prada, J

    2017-12-01

    COX-2 overexpression is associated with several hallmarks of carcinogenesis such as proliferation, angiogenesis, invasion and metastasis. Fifty cases of canine mast cell tumours (MCT) were retrospectively evaluated and submitted to immunohistochemistry for COX-2, CD31, Ki-67, MAC-387 and CD3. Furthermore its relationship with clinicopathological variables and overall survival (OS) was analysed. COX-2 intensity (P = 0.016), but not COX-2 extension nor score was associated with decreased OS and higher grades of malignancy according to Patnaik (P = 0.002) and Kiupel (P Cox-2 intensity was also associated with higher Ki-67 scores (P = 0.009), higher mitotic index (P = 0.022) and higher microvascularization density (P = 0.045). No association was observed for COX-2 intensity and CD3-T lymphocyte (P = 0.377) and macrophage infiltration (P = 0.261) by MAC-387 immunollabelling, suggesting an active role of COX-2 in MCT oncogenesis mainly through proliferation and angiogenesis stimulation making it a potentially clinical relevant prognosis marker and therapeutic target. © 2016 John Wiley & Sons Ltd.

  9. Morphological abnormalities, impaired fetal development and decrease in myostatin expression following somatic cell nuclear transfer in dogs.

    Science.gov (United States)

    Hong, Il-Hwa; Jeong, Yeon-Woo; Shin, Taeyoung; Hyun, Sang-Hwan; Park, Jin-Kyu; Ki, Mi-Ran; Han, Seon-Young; Park, Se-Il; Lee, Ji-Hyun; Lee, Eun-Mi; Kim, Ah-Young; You, Sang-Young; Hwang, Woo-Suk; Jeong, Kyu-Shik

    2011-05-01

    Several mammals, including dogs, have been successfully cloned using somatic cell nuclear transfer (SCNT), but the efficiency of generating normal, live offspring is relatively low. Although the high failure rate has been attributed to incomplete reprogramming of the somatic nuclei during the cloning process, the exact cause is not fully known. To elucidate the cause of death in cloned offspring, 12 deceased offspring cloned by SCNT were necropsied. The clones were either stillborn just prior to delivery or died with dyspnea shortly after birth. On gross examination, defects in the anterior abdominal wall and increased heart and liver sizes were found. Notably, a significant increase in muscle mass and macroglossia lesions were observed in deceased SCNT-cloned dogs. Interestingly, the expression of myostatin, a negative regulator of muscle growth during embryogenesis, was down-regulated at the mRNA level in tongues and skeletal muscles of SCNT-cloned dogs compared with a normal dog. Results of the present study suggest that decreased expression of myostatin in SCNT-cloned dogs may be involved in morphological abnormalities such as increased muscle mass and macroglossia, which may contribute to impaired fetal development and poor survival rates. Copyright © 2011 Wiley-Liss, Inc.

  10. ∆(9)-Tetrahydrocannabinol decreases NOP receptor density and mRNA levels in human SH-SY5Y cells.

    Science.gov (United States)

    Cannarsa, Rosalia; Carretta, Donatella; Lattanzio, Francesca; Candeletti, Sanzio; Romualdi, Patrizia

    2012-02-01

    Several studies demonstrated a cross-talk between the opioid and cannabinoid system. The NOP receptor and its endogenous ligand nociceptin/orphanin FQ represent an opioid-related functional entity that mediates some non-classical opioid effects. The relationship between cannabinoid and nociceptin/NOP system is yet poorly explored. In this study, we used the neuroblastoma SH-SY5Y cell line to investigate the effect of delta-9-tetrahydrocannabinol (∆(9)-THC) on nociceptin/NOP system. Results revealed that the exposure to ∆(9)-THC (100, 150, and 200 nM) for 24 h produces a dose-dependent NOP receptor B (max) down-regulation. Moreover, ∆(9)-THC caused a dose-dependent decrease in NOP mRNA levels. The selective cannabinoid receptor CB1 antagonist AM251 (1-(2,4-dichlorophenyl)-5-(4-iodophenyl)-4-methyl-N-1-piperidinyl-1H-pyrazole-3-carboxamide) reduces both effects, suggesting that ∆(9)-THC activation of CB1 receptor is involved in the observed effects. These data show evidence of a cross-talk between NOP and CB1 receptors, thus suggesting a possible interplay between cannabinoid and nociceptin/NOP system.

  11. 17β-Estradiol and progesterone decrease MDP induced NOD2 expression in bovine mammary epithelial cells.

    Science.gov (United States)

    Xu, Dan-Dan; Wang, Gang; He, Xian-Jing; Wang, Jian-Fa; Yang, Bin; Sun, Zhi-Peng; Sun, Dong-Bo; He, Qian-Yu; Zhang, Xu; Wu, Rui

    2017-06-01

    During the periparturient period, many neuroendocrine changes develop in cows. Periparturient hormone fluxes may adversely affect mammary gland immunity and mastitis susceptibility. 17β-Estradiol (E 2 ) and progesterone (P 4 ) have been reported to function on immune regulation, and their concentration fluctuates dramatically during the perinatal period. Nucleotide-binding oligomerization domain (NOD)-like receptors (NLRs) mediate numerous aspects of innate immunity in humans and experimental animals. This study aimed to explore the effects of E 2 and P 4 on NOD2 expression in bovine mammary epithelial cells (BMECs). BMECs were isolated and purified from bovine mammary tissue and treated with E 2 /P 4 and muramyl dipeptide (MDP). After these treatments, the mRNA levels of NOD2, receptor-interacting protein kinase (RIP) 2, interleukin (IL) 1β, IL-6, IL-8 and tumor necrosis factor (TNF) α were assessed by quantitative real-time polymerase chain reaction (qRT-PCR) respectively, and the protein levels of NOD2 were analyzed by western blotting. The results showed that E 2 and P 4 decreased MDP-induced transcriptional expression of NOD2 and the downstream molecules. Moreover, E 2 reduced MDP-induced NOD2 protein expression levels. Our study suggests that down-regulation of NOD2 by E 2 and P 4 may be one of the reasons for mastitis susceptibility in periparturient dairy cows. Copyright © 2017 Elsevier B.V. All rights reserved.

  12. Splenectomy suppresses growth and metastasis of hepatocellular carcinoma through decreasing myeloid-derived suppressor cells in vivo.

    Science.gov (United States)

    Long, Xin; Wang, Jian; Zhao, Jian-Ping; Liang, Hui-Fang; Zhu, Peng; Cheng, Qi; Chen, Qian; Wu, Yan-Hui; Zhang, Zhan-Guo; Zhang, Bi-Xiang; Chen, Xiao-Ping

    2016-10-01

    The function of the spleen in tumor development has been investigated for years. The relationship of the spleen with hepatocellular carcinoma (HCC), a huge health burden worldwide, however, remains unknown. The present study aimed to examine the effect of splenectomy on the development of HCC and the possible mechanism. Mouse hepatic carcinoma lines H22 and Hepa1-6 as well as BALB/c and C57 mice were used to establish orthotopic and metastatic mouse models of liver cancer. Mice were divided into four groups, including control group, splenectomy control group (S group), tumor group (T group) and tumor plus splenectomy group (T+S group). Tumor growth, metastases and overall survival were assessed at determined time points. Meanwhile, myeloid-derived suppressor cells (MDSCs) were isolated from the peripheral blood (PB), the spleen and liver tumors, and then measured by flow cytometery. It was found that liver cancer led to splenomegaly, and increased the percentage of MDSCs in the PB and spleen in the mouse models. Splenectomy inhibited the growth and progression of liver cancer and prolonged the overall survival time of orthotopic and metastatic models, which was accompanied by decreased proportion of MDSCs in the PB and tumors of liver cancer-bearing mouse. It was suggested that splenectomy could be considered an adjuvant therapy to treat liver cancer.

  13. The influence of continuous γ-irradiation at decreasing dose-rate on the survival rote and induction of gene mutations in cultured Chinese hamster cells

    International Nuclear Information System (INIS)

    Feoktistova, T.P.; Elisova, E.V.; Stavrakova, N.M.

    1991-01-01

    Continuous γ-irradiation at decreasing dose-rate was shown to be less effective than acute exposure with regard to the lethal effect and frequency of mutations of resistance to 6-thioguanine in cultured Chinese hamster cells. The cell population subjected to continuons irradiation was d more radioresistant than the intact one. Lethal and genetic effects of continuous irradiation at decreasing dose-rate were mainly determined by the contribution of the radiation dose received during the first 24 h of exposure

  14. Defected red blood cell membranes and direct correlation with the uraemic milieu: the connection with the decreased red blood cell lifespan observed in haemodialysis patients

    International Nuclear Information System (INIS)

    Stamopoulos, D; Manios, E; Gogola, V; Grapsa, E; Bakirtzi, N

    2012-01-01

    Together with impaired production of erythropoietin and iron deficiency, the decreased lifespan of red blood cells (RBCs) is a main factor contributing to the chronic anaemia observed in haemodialysis (HD) patients. Atomic force microscopy is employed in this work to thoroughly survey the membrane of intact RBCs (iRBCs) of HD patients in comparison to those of healthy donors, aiming to obtain direct information on the structural status of RBCs that can be related to their decreased lifespan. We observed that the iRBC membrane of the HD patients is overpopulated with extended circular defects, termed ‘orifices’, that have typical dimension ranging between 0.2 and 1.0 μm. The ‘orifice’ index—that is, the mean population of ‘orifices’ per top membrane surface—exhibits a pronounced relative increase of order 54 ± 12% for the HD patients as compared to healthy donors. Interestingly, for the HD patients, the ‘orifice’ index, which relates to the structural status of the RBC membrane, correlates strongly with urea concentration, which is a basic index of the uraemic milieu. Thus, these results indicate that the uraemic milieu downgrades the structural status of the RBC membrane, possibly triggering biochemical processes that result in their premature elimination from the circulation. This process could decrease the lifespan of RBCs, as observed in HD patients. (paper)

  15. Dietary fat without body weight gain increases in vivo MCF-7 human breast cancer cell growth and decreases natural killer cell cytotoxicity.

    Science.gov (United States)

    Lamas, Bruno; Nachat-Kappes, Rachida; Goncalves-Mendes, Nicolas; Mishellany, Florence; Rossary, Adrien; Vasson, Marie-Paule; Farges, Marie-Chantal

    2015-01-01

    High-calorie (HC) diet contributes to the increased incidence of obesity, which is a risk factor for breast cancer in postmenopausal women, and in particular for estrogen receptor (ER) positive tumors. This study investigated whether an HC diet increases human ER-positive breast cancer progression and modulates natural killer (NK) cell functions. Four-week-old female BALB/c athymic nude mice were fed a HC diet (5320 kcal/kg) or standard calorie diet (SC, 2820 kcal/kg) for 6 mo. After 5 mo, the mice were randomly implanted with MCF-7 breast cancer cells (SCT and HCT) or received an isovolumic injection (SC and HC) in both inguinal fat pads. Tumor growth was greater in the HCT group than in the SC group without change in body weight. The HC diet decreased the tumor expression of genes involved in the citrate cycle and in adiponectin and lipid metabolism but increased that of genes controlling glycolysis and angiogenesis. The tumor expression level of Ki67 was increased while that of the cleaved caspase 3 and the ER-β and progesterone receptors was reduced. Tumor development in response to the HC diet was associated with smaller numbers and lower cytotoxicity of splenic NK cells. These results indicate that an HC diet without body weight gain increases ER-positive breast cancer cell proliferation and reduces tumor apoptosis. The underlying mechanisms might involve a downexpression of tumor hormonal receptor and reduced NK cell functions, and might also result in the regulation of genes involved in several cellular functions. © 2013 Wiley Periodicals, Inc.

  16. miR-518f-5p decreases tetraspanin CD9 protein levels and differentially affects non-tumourigenic prostate and prostate cancer cell migration and adhesion.

    Science.gov (United States)

    Bond, Danielle R; Naudin, Crystal; Carroll, Adam P; Goldie, Belinda J; Brzozowski, Joshua S; Jankowski, Helen M; Cairns, Murray J; Ashman, Leonie K; Scarlett, Christopher J; Weidenhofer, Judith

    2018-01-05

    Tetraspanin CD9 is generally considered to be a metastasis suppressor, with decreased levels associated with progression and metastasis in many advanced stage cancers. Little is known about the cause of CD9 dysregulation in prostate cancer, however there are several miRNA-binding sites in the 3´UTR of the transcript suggesting it could be post-transcriptionally regulated. Using microarrays and luciferase assays in tumourigenic and non-tumourigenic prostate cell lines we identified miR-518f-5p as a regulator of the CD9 3'UTR gene expression, and decreased expression of endogenous CD9 in non-tumorigenic prostate RWPE1 and prostate cancer DU145 cells. This resulted in differential functional effects, in which RWPE1 cells showed increased migration and decreased adhesion to extracellular matrix substrates, whereas DU145 cells showed decreased migration and increased adhesion. Moreover, overexpression of miR-518f-5p significantly increased proliferation between 48h and 72h in normal RWPE1 cells, with no effect on tumourigenic DU145 cell proliferation. These results show that tetraspanin CD9 is regulated by miRNAs in prostate cell lines and that due to differential functional effects in non-tumourigenic versus tumourigenic prostate cells, miR-518f-5p may be an effective biomarker and/or therapeutic target for prostate cancer progression.

  17. MicroRNA-29a is up-regulated in beta-cells by glucose and decreases glucose-stimulated insulin secretion

    DEFF Research Database (Denmark)

    Bagge, Annika; Clausen, Trine R; Larsen, Sylvester

    2012-01-01

    Chronically elevated levels of glucose impair pancreatic beta-cell function while inducing beta-cell proliferation. MicroRNA-29a (miR-29a) levels are increased in several tissues in diabetic animals and mediate decreased insulin-stimulated glucose-transport of adipocytes. The aim was to investiga...

  18. Ataxia-telangiectasia mutated (ATM) deficiency decreases reprogramming efficiency and leads to genomic instability in iPS cells

    Energy Technology Data Exchange (ETDEWEB)

    Kinoshita, Taisuke [Department of Cell Differentiation, The Sakaguchi Laboratory, School of Medicine, Keio University, Tokyo 160-8582 (Japan); Nagamatsu, Go, E-mail: gonag@sc.itc.keio.ac.jp [Department of Cell Differentiation, The Sakaguchi Laboratory, School of Medicine, Keio University, Tokyo 160-8582 (Japan); Precursory Research for Embryonic Science and Technology, Japan Science and Technology Agency, Kawaguchi, Saitama 332-0012 (Japan); Kosaka, Takeo [Department of Urology, School of Medicine, Keio University, Tokyo 160-8582 (Japan); Takubo, Keiyo [Department of Cell Differentiation, The Sakaguchi Laboratory, School of Medicine, Keio University, Tokyo 160-8582 (Japan); Hotta, Akitsu [Precursory Research for Embryonic Science and Technology, Japan Science and Technology Agency, Kawaguchi, Saitama 332-0012 (Japan); Department of Reprogramming Science, Center for iPS Cell Research and Application (CiRA), Kyoto University, Kyoto (Japan); Ellis, James [Ontario Human iPS Cell Facility, Molecular Genetics, University of Toronto, Developmental and Stem Cell Biology, SickKids, Toronto, Canada MG1L7 (Canada); Suda, Toshio, E-mail: sudato@sc.itc.keio.ac.jp [Department of Cell Differentiation, The Sakaguchi Laboratory, School of Medicine, Keio University, Tokyo 160-8582 (Japan)

    2011-04-08

    Highlights: {yields} iPS cells were induced with a fluorescence monitoring system. {yields} ATM-deficient tail-tip fibroblasts exhibited quite a low reprogramming efficiency. {yields} iPS cells obtained from ATM-deficient cells had pluripotent cell characteristics. {yields} ATM-deficient iPS cells had abnormal chromosomes, which were accumulated in culture. -- Abstract: During cell division, one of the major features of somatic cell reprogramming by defined factors, cells are potentially exposed to DNA damage. Inactivation of the tumor suppressor gene p53 raised reprogramming efficiency but resulted in an increased number of abnormal chromosomes in established iPS cells. Ataxia-telangiectasia mutated (ATM), which is critical in the cellular response to DNA double-strand breaks, may also play an important role during reprogramming. To clarify the function of ATM in somatic cell reprogramming, we investigated reprogramming in ATM-deficient (ATM-KO) tail-tip fibroblasts (TTFs). Although reprogramming efficiency was greatly reduced in ATM-KO TTFs, ATM-KO iPS cells were successfully generated and showed the same proliferation activity as WT iPS cells. ATM-KO iPS cells had a gene expression profile similar to ES cells and WT iPS cells, and had the capacity to differentiate into all three germ layers. On the other hand, ATM-KO iPS cells accumulated abnormal genome structures upon continuous passages. Even with the abnormal karyotype, ATM-KO iPS cells retained pluripotent cell characteristics for at least 20 passages. These data indicate that ATM does participate in the reprogramming process, although its role is not essential.

  19. Effect of tributyltin (TBT) on ATP levels in human natural killer (NK) cells: relationship to TBT-induced decreases in NK function.

    Science.gov (United States)

    Dudimah, Fred D; Odman-Ghazi, Sabah O; Hatcher, Frank; Whalen, Margaret M

    2007-01-01

    The purpose of this study was to investigate the role that tributyltin (TBT)-induced decreases in ATP levels may play in TBT-induced decreases in the tumor lysing (lytic) function of natural killer (NK) cells. NK cells are a subset of lymphocytes that act as an initial immune defense against tumor cells and virally infected cells. TBT is an environmental contaminant that has been detected in human blood, which has been shown to interfere with ATP synthesis. Previous studies have shown that TBT is able to decrease very significantly the lytic function of NK cells. In this study NK cells were exposed to various concentrations of TBT and to two other compounds that interfere with ATP synthesis (rotenone a complex I inhibitor and oligomycin an ATP synthase inhibitor) for various lengths of time before determining the levels of ATP and lytic function. Exposures of NK cells to 10, 25, 50 and 100 nm TBT did not significantly reduce ATP levels after 24 h. However, these same exposures caused significant decreases in cytotoxic function. Studies of brief 1 h exposures to a range of TBT, rotenone and oligomycin concentrations followed by 24 h, 48 h and 6 day periods in compound-free media prior to assaying for ATP levels or cytotoxic function showed that each of the compounds caused persistent decreases in ATP levels and lytic function of NK cells. Exposures to 0.05-5 microm rotenone or oligomycin for 1 h reduced ATP levels by 20-25% but did not have any measurable effect on the ability of NK cells to lyse tumor cells. ATP levels were also decreased by about 20-25% after 24 h or 48 h exposures to rotenone or oligomycin (0.5 microm ), and the lytic function was decreased by about 50%. The results suggest that TBT-induced decreases in ATP levels were not responsible for the loss of cytotoxic function seen at 1 h and 24 h. However, TBT-induced decreases of NK-ATP levels may be at least in part responsible for losses of NK-cytotoxic function seen after 48 h and 6 day exposures

  20. Reversible lysine-specific demethylase 1 antagonist HCI-2509 inhibits growth and decreases c-MYC in castration- and docetaxel-resistant prostate cancer cells.

    Science.gov (United States)

    Gupta, S; Weston, A; Bearrs, J; Thode, T; Neiss, A; Soldi, R; Sharma, S

    2016-12-01

    Lysine-specific demethylase 1 (LSD1 or KDM1A) overexpression correlates with poor survival and castration resistance in prostate cancer. LSD1 is a coregulator of ligand-independent androgen receptor signaling promoting c-MYC expression. We examined the antitumor efficacy of LSD1 inhibition with HCI-2509 in advanced stages of prostate cancer. Cell survival, colony formation, histone methylation, c-MYC level, c-MYC expression, cell cycle changes and in vivo efficacy were studied in castration-resistant prostate cancer cells upon treatment with HCI-2509. In vitro combination studies, using HCI-2509 and docetaxel, were performed to assess the synergy. Cell survival, colony formation, histone methylation and c-myc levels were studied in docetaxel-resistant prostate cancer cells treated with HCI-2509. HCI-2509 is cytotoxic and inhibits colony formation in castration-resistant prostate cancer cells. HCI-2509 treatment causes a dose-dependent increase in H3K9me2 (histone H3lysine 9) levels, a decrease in c-MYC protein, inhibition of c-MYC expression and accumulation in the G0/G1 phase of the cell cycle in these cells. PC3 xenografts in mice have a significant reduction in tumor burden upon treatment with HCI-2509 with no associated myelotoxicity or weight loss. More synergy is noted at sub-IC 50 (half-maximal inhibitory concentration) doses of docetaxel and HCI-2509 in PC3 cells than in DU145 cells. HCI-2509 has growth-inhibitory efficacy and decreases the c-myc level in docetaxel-resistant prostate cancer cells. LSD1 inhibition with HCI-2509 decreases the c-MYC level in poorly differentiated prostate cancer cell lines and has a therapeutic potential in castration- and docetaxel-resistant prostate cancer.

  1. Amplifying IFN-γ Signaling in Dendritic Cells by CD11c-Specific Loss of SOCS1 Increases Innate Immunity to Infection while Decreasing Adaptive Immunity.

    Science.gov (United States)

    Alice, Alejandro F; Kramer, Gwen; Bambina, Shelly; Baird, Jason R; Bahjat, Keith S; Gough, Michael J; Crittenden, Marka R

    2018-01-01

    Although prophylactic vaccines provide protective humoral immunity against infectious agents, vaccines that elicit potent CD8 T cell responses are valuable tools to shape and drive cellular immunity against cancer and intracellular infection. In particular, IFN-γ-polarized cytotoxic CD8 T cell immunity is considered optimal for protective immunity against intracellular Ags. Suppressor of cytokine signaling (SOCS)1 is a cross-functional negative regulator of TLR and cytokine receptor signaling via degradation of the receptor-signaling complex. We hypothesized that loss of SOCS1 in dendritic cells (DCs) would improve T cell responses by accentuating IFN-γ-directed immune responses. We tested this hypothesis using a recombinant Listeria monocytogenes vaccine platform that targets CD11c + DCs in mice in which SOCS1 is selectively deleted in all CD11c + cells. Unexpectedly, in mice lacking SOCS1 expression in CD11c + cells, we observed a decrease in CD8 + T cell response to the L. monocytogenes vaccine. NK cell responses were also decreased in mice lacking SOCS1 expression in CD11c + cells but did not explain the defect in CD8 + T cell immunity. We found that DCs lacking SOCS1 expression were functional in driving Ag-specific CD8 + T cell expansion in vitro but that this process was defective following infection in vivo. Instead, monocyte-derived innate TNF-α and inducible NO synthase-producing DCs dominated the antibacterial response. Thus, loss of SOCS1 in CD11c + cells skewed the balance of immune response to infection by increasing innate responses while decreasing Ag-specific adaptive responses to infectious Ags. Copyright © 2017 by The American Association of Immunologists, Inc.

  2. RNAi Knockdown of Hypoxia-Inducible Factor-1α Decreased the Proliferation, Migration, and Invasion of Hypoxic Hepatocellular Carcinoma Cells.

    Science.gov (United States)

    Chen, ChengShi; Liu, Rong; Wang, JianHua; Yan, ZhiPing; Qian, Sheng; Zhang, Wei

    2015-04-01

    The obstruction of hepatic arterial blood flow results in tumor tissue hypoxia and elevated expression of hypoxia-inducible factor-1alpha (HIF-1α). Our study evaluated whether lentivirus-mediated short interference RNA against HIF-1α inhibits proliferation, invasion, and migration of hepatocellular carcinoma (HCC) cells under hypoxia. RNA interference knockdown of HIF-1α was achieved by HIF-1α-directed lentiviral shRNA, in a rat HCC cell line cultured under hypoxia condition for varying length of times. The expression levels of HIF-1α and vascular endothelial growth factor were examined using reverse transcription polymerase chain reaction and western blot analyses. Cell proliferation, migration, and invasion were measured by cell viability, transwell migration, and invasion assays, respectively. Inhibition of HIF-1α expression by shRNA suppressed vascular endothelial growth factor mRNA and protein levels under both normoxia and hypoxia. It also suppressed cell migration and invasion, which were enhanced under hypoxic conditions. RNAi knockdown of HIF-1α further suppressed hypoxia-mediated inhibition of the cell proliferation. These data suggest that shRNA of HIF-1α could antagonize the hypoxia-mediated increase in hepatic cancer cell migration and invasion, and synergize with hypoxia to inhibit the cell proliferation in HCC cells.

  3. Ascorbic acid inhibits TPA-induced HL-60 cell differentiation by decreasing cellular H₂O₂ and ERK phosphorylation.

    Science.gov (United States)

    Yiang, Giou-Teng; Chen, Jen-Ni; Wu, Tsai-Kun; Wang, Hsueh-Fang; Hung, Yu-Ting; Chang, Wei-Jung; Chen, Chinshuh; Wei, Chyou-Wei; Yu, Yung-Luen

    2015-10-01

    Retinoic acid (RA), vitamin D and 12-O‑tetradecanoyl phorbol-13-acetate (TPA) can induce HL-60 cells to differentiate into granulocytes, monocytes and macrophages, respectively. Similar to RA and vitamin D, ascorbic acid also belongs to the vitamin family. High‑dose ascorbic acid (>100 µM) induces HL‑60 cell apoptosis and induces a small fraction of HL‑60 cells to express the granulocyte marker, CD66b. In addition, ascorbic acid exerts an anti‑oxidative stress function. Oxidative stress is required for HL‑60 cell differentiation following treatment with TPA, however, the effect of ascorbic acid on HL‑60 cell differentiation in combination with TPA treatment remains to be fully elucidated. The aim of the present study was to investigate the cellular effects of ascorbic acid treatment on TPA-differentiated HL-60 cells. TPA-differentiated HL-60 cells were used for this investigation, this study and the levels of cellular hydrogen peroxide (H2O2), caspase activity and ERK phosphorylation were determined following combined treatment with TPA and ascorbic acid. The results demonstrated that low‑dose ascorbic acid (5 µM) reduced the cellular levels of H2O2 and inhibited the differentiation of HL‑60 cells into macrophages following treatment with TPA. In addition, the results of the present study further demonstrated that low‑dose ascorbic acid inactivates the ERK phosphorylation pathway, which inhibited HL‑60 cell differentiation following treatment with TPA.

  4. Men with Sickle Cell Anemia and Priapism Exhibit Increased Hemolytic Rate, Decreased Red Blood Cell Deformability and Increased Red Blood Cell Aggregate Strength.

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    Kizzy-Clara Cita

    Full Text Available To investigate the association between priapism in men with sickle cell anemia (SCA and hemorheological and hemolytical parameters.Fifty-eight men with SCA (median age: 38 years were included; 28 who had experienced priapism at least once during their life (priapism group and 30 who never experienced this complication (control group. Twenty-two patients were treated with hydroxycarbamide, 11 in each group. All patients were at steady state at the time of inclusion. Hematological and biochemical parameters were obtained through routine procedures. The Laser-assisted Optical Rotational Cell Analyzer was used to measure red blood cell (RBC deformability at 30 Pa (ektacytometry and RBC aggregation properties (laser backscatter versus time. Blood viscosity was measured at a shear rate of 225 s-1 using a cone/plate viscometer. A principal component analysis was performed on 4 hemolytic markers (i.e., lactate dehydrogenase (LDH, aspartate aminotransferase (ASAT, total bilirubin (BIL levels and reticulocyte (RET percentage to calculate a hemolytic index.Compared to the control group, patients with priapism exhibited higher ASAT (p = 0.01, LDH (p = 0.03, RET (p = 0.03 levels and hemolytic indices (p = 0.02. Higher RBC aggregates strength (p = 0.01 and lower RBC deformability (p = 0.005 were observed in patients with priapism compared to controls. After removing the hydroxycarbamide-treated patients, RBC deformability (p = 0.01 and RBC aggregate strength (p = 0.03 were still different between the two groups, and patients with priapism exhibited significantly higher hemolytic indices (p = 0.01 than controls.Our results confirm that priapism in SCA is associated with higher hemolytic rates and show for the first time that this complication is also associated with higher RBC aggregate strength and lower RBC deformability.

  5. Blood T-cell receptor diversity decreases during the course of HIV infection, but the potential for a diverse repertoire persists

    Science.gov (United States)

    Young, Jennifer J.; Schmidt, Diane; Zhang, Qianjun; Hoh, Rebecca; Busch, Michael; Martin, Jeffrey; Deeks, Steven; McCune, Joseph M.

    2012-01-01

    HIV infection results in a decrease in circulating CD4+ T-cell and naive T-cell numbers. If such losses were associated with an erosion of T-cell receptor (TCR) repertoire diversity in the peripheral T-cell pool, this might exacerbate the state of persistent immunodeficiency. Existing methods for the analysis of the TCR repertoire have demonstrated skewed distributions of TCR genes in HIV-infected subjects but cannot directly measure TCR diversity. Here we used AmpliCot, a quantitative assay based on DNA hybridization kinetics, to measure TCR diversity in a cross-sectional comparison of 19 HIV-infected persons to 18 HIV-uninfected controls. HIV-infected persons had a 10-fold decrease in total TCR repertoire diversity in 1.5 mL of blood compared with uninfected controls, with decreased diversity correlating most closely with a lower CD4+ T-cell percentage. Nonetheless, the TCR repertoire diversity of sort-purified T-cell subpopulations in HIV-infected and HIV-uninfected subjects was comparable. These observations suggest that the TCR repertoire diversity changes in whole blood during HIV disease progression are primarily the result of changes in the number and proportion of T-cell subpopulations and that most HIV-infected persons may retain a sufficiently diverse TCR repertoire to permit immune reconstitution with antiretroviral therapy alone, without thymopoiesis. PMID:22371879

  6. Increase of Intracellular BAFF in B Cells of Sjögren’s Patients Is Not Affected by Decrease of BAFFR

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    Jan Krejsek

    2015-07-01

    Full Text Available The presence of a broad spectrum of autoantibodies in Sjögren’s syndrome (SjS patients is the result of abnormal B-cell regulation that can be at least partially explained by abnormal BAFF/BAFFR regulation. The objective of this study was to determine both membrane and intracellular expression of BAFF/BAFFR in monocytes and B-cells in peripheral blood of 19 primary Sjögren’s syndrome patients and 20 healthy controls using flow cytometry. We also measured sBAFF in serum. Compared to healthy controls, both surface and intracellular expression of BAFF was significantly increased in monocytes and B-cells of SjS patients. Also serum sBAFF level was elevated. Expression of BAFFR on B-cells of SjS patients was surprisingly decreased, but there was no clear increase or decrease within monocytes. Our results indicate that activated monocytes communicate with B-cells via BAFF and BAFFR, so that B-cells are stimulated, but BAFF is also produced to stimulate cells in autocrine way. The decrease of BAFFR expression in SjS patients suggests that there is the mechanism that attempts to take over in order to balance the high level of BAFF.

  7. SIRT1 overexpression decreases cisplatin-induced acetylation of NF-κB p65 subunit and cytotoxicity in renal proximal tubule cells

    International Nuclear Information System (INIS)

    Jung, Yu Jin; Lee, Jung Eun; Lee, Ae Sin; Kang, Kyung Pyo; Lee, Sik; Park, Sung Kwang; Lee, Sang Yong; Han, Myung Kwan; Kim, Duk Hoon; Kim, Won

    2012-01-01

    Highlights: ► Cisplatin increases acetylation of NF-κB p65 subunit in HK2 cells. ► SIRT1 overexpression decreases cisplatin-induced p65 acetylation and -cytotoxicity. ► Resveratrol decreased cisplatin-induced cell viability through deacetylation of p65. -- Abstract: As the increased acetylation of p65 is linked to nuclear factor-κB (NF-κB) activation, the regulation of p65 acetylation can be a potential target for the treatment of inflammatory injury. Cisplatin-induced nephrotoxicity is an important issue in chemotherapy of cancer patients. SIRT1, nicotinamide adenine dinucleotide (NAD + )-dependent protein deacetylase, has been implicated in a variety of cellular processes such as inflammatory injury and the control of multidrug resistance in cancer. However, there is no report on the effect of SIRT1 overexpression on cisplatin-induced acetylation of p65 subunit of NF-κB and cell injury. To investigate the effect of SIRT1 in on cisplatin-induced acetylation of p65 subunit of NF-κB and cell injury, HK2 cells were exposed with SIRT1 overexpression, LacZ adenovirus or dominant negative adenovirus after treatment with cisplatin. While protein expression of SIRT1 was decreased by cisplatin treatment compared with control buffer treatment, acetylation of NF-κB p65 subunit was significantly increased after treatment with cisplatin. Overexpression of SIRT1 ameliorated the increased acetylation of p65 of NF-κB during cisplatin treatment and cisplatin-induced cytotoxicity. Further, treatment of cisplatin-treated HK2 cells with resveratrol, a SIRT1 activator, also decreased acetylation of NF-κB p65 subunit and cisplatin-induced increase of the cell viability in HK2 cells. Our findings suggests that the regulation of acetylation of p65 of NF-κB through SIRT1 can be a possible target to attenuate cisplatin-induced renal cell damage.

  8. Curcumin decreases the expression of Pokemon by suppressing the binding activity of the Sp1 protein in human lung cancer cells.

    Science.gov (United States)

    Cui, Jiajun; Meng, Xianfeng; Gao, Xudong; Tan, Guangxuan

    2010-03-01

    Pokemon, which stands for POK erythroid myeloid ontogenic factor, can regulate expression of many genes and plays an important role in tumorigenesis. Curcumin, a natural and non-toxic yellow compound, has capacity for antioxidant, free radical scavenger, anti-inflammatory properties. Recent studies shows it is a potential inhibitor of cell proliferation in a variety of tumour cells. To investigate whether curcumin can regulate the expression of Pokemon, a series of experiments were carried out. Transient transfection experiments demonstrated that curcumin could decrease the activity of the Pokemon promoter. Western blot analysis suggested that curcumin could significantly decrease the expression of the Pokemon. Overexpression of Sp1 could enhance the activity of the Pokemon promoter, whereas knockdown of Sp1 could decrease its activity. More important, we also found that curcumin could decrease the expression of the Pokemon by suppressing the stimulation of the Sp1 protein. Therefore, curcumin is a potential reagent for tumour therapy which may target Pokemon.

  9. Loss of CD28 on Peripheral T Cells Decreases the Risk for Early Acute Rejection after Kidney Transplantation.

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    Burç Dedeoglu

    Full Text Available End-stage renal disease patients have a dysfunctional, prematurely aged peripheral T-cell system. Here we hypothesized that the degree of premature T-cell ageing before kidney transplantation predicts the risk for early acute allograft rejection (EAR.222 living donor kidney transplant recipients were prospectively analyzed. EAR was defined as biopsy proven acute allograft rejection within 3 months after kidney transplantation. The differentiation status of circulating T cells, the relative telomere length and the number of CD31+ naive T cells were determined as T-cell ageing parameters.Of the 222 patients analyzed, 30 (14% developed an EAR. The donor age and the historical panel reactive antibody score were significantly higher (p = 0.024 and p = 0.039 respectively and the number of related donor kidney transplantation was significantly lower (p = 0.018 in the EAR group. EAR-patients showed lower CD4+CD28null T-cell numbers (p<0.01 and the same trend was observed for CD8+CD28null T-cell numbers (p = 0.08. No differences regarding the other ageing parameters were found. A multivariate Cox regression analysis showed that higher CD4+CD28null T-cell numbers was associated with a lower risk for EAR (HR: 0.65, p = 0.028. In vitro, a significant lower percentage of alloreactive T cells was observed within CD28null T cells (p<0.001.Immunological ageing-related expansion of highly differentiated CD28null T cells is associated with a lower risk for EAR.

  10. Exogenous NAD+ decreases oxidative stress and protects H2O2-treated RPE cells against necrotic death through the up-regulation of autophagy

    Science.gov (United States)

    Zhu, Ying; Zhao, Ke-ke; Tong, Yao; Zhou, Ya-li; Wang, Yi-xiao; Zhao, Pei-quan; Wang, Zhao-yang

    2016-01-01

    Increased oxidative stress, which can lead to the retinal pigment epithelium (RPE) cell death by inducing ATP depletion and DNA repair, is believed to be a prominent pathology in age-related macular degeneration (AMD). In the present study, we showed that and 0.1 mM nicotinamide adenine dinucleotide (NAD+) administration significantly blocked RPE cell death induced by 300 μM H2O2. Further investigation showed that H2O2 resulted in increased intracellular ROS level, activation of PARP-1 and subsequently necrotic death of RPE cells. Exogenous NAD+ administration significantly decreased intracellular and intranuclear ROS levels in H2O2-treated RPE cells. In addition, NAD+ administration to H2O2-treated RPE cells inhibited the activation of PARP-1 and protected the RPE cells against necrotic death. Moreover, exogenous NAD+ administration up-regulated autophagy in the H2O2-treated RPE cells. Inhibition of autophagy by LY294002 blocked the decrease of intracellular and intranuclear ROS level. Besides, inhibition of autophagy by LY294002 abolished the protection of exogenous NAD+ against H2O2-induced cell necrotic death. Taken together, our findings indicate that that exogenous NAD+ administration suppresses H2O2-induced oxidative stress and protects RPE cells against PARP-1 mediated necrotic death through the up-regulation of autophagy. The results suggest that exogenous NAD+ administration might be potential value for the treatment of AMD. PMID:27240523

  11. The mast cell stabiliser ketotifen decreases visceral hypersensitivity and improves intestinal symptoms in patients with irritable bowel syndrome

    NARCIS (Netherlands)

    Klooker, Tamira K.; Braak, Breg; Koopman, Karin E.; Welting, Olaf; Wouters, Mira M.; van der Heide, Sicco; Schemann, Michael; Bischoff, Stephan C.; van den Wijngaard, Rene M.; Boeckxstaens, Guy E.

    Background Mast cell activation is thought to be involved in visceral hypersensitivity, one of the main characteristics of the irritable bowel syndrome (IBS). A study was therefore undertaken to investigate the effect of the mast cell stabiliser ketotifen on rectal sensitivity and symptoms in

  12. The mast cell stabiliser ketotifen decreases visceral hypersensitivity and improves intestinal symptoms in patients with irritable bowel syndrome

    NARCIS (Netherlands)

    Klooker, T.K.; Braak, B.; Koopman, K.E.; Welting, O.; Wouters, M.M.; van der Heide, S.; Schemann, M.; Bischoff, S.C.; van den Wijngaard, R.M.; Boeckxstaens, G.E.

    2010-01-01

    Background Mast cell activation is thought to be involved in visceral hypersensitivity, one of the main characteristics of the irritable bowel syndrome (IBS). A study was therefore undertaken to investigate the effect of the mast cell stabiliser ketotifen on rectal sensitivity and symptoms in

  13. Hypofractionated radiation induces a decrease in cell proliferation but no histological damage to organotypic multicellular spheroids of human glioblastomas

    NARCIS (Netherlands)

    Kaaijk, P.; Troost, D.; Sminia, P.; Hulshof, M. C.; van der Kracht, A. H.; Leenstra, S.; Bosch, D. A.

    1997-01-01

    The aim of this study was to examine the effect of radiation on glioblastoma, using an organotypic multicellular spheroid (OMS) model. Most glioblastoma cell lines are, in contrast to glioblastomas in vivo, relatively radiosensitive. This limits the value of using cell lines for studying the

  14. Selective decrease in circulating V alpha 24+V beta 11+ NKT cells during HIV type 1 infection

    NARCIS (Netherlands)

    van der Vliet, Hans J. J.; von Blomberg, B. Mary E.; Hazenberg, Mette D.; Nishi, Nobusuke; Otto, Sigrid A.; van Benthem, Birgit H.; Prins, Maria; Claessen, Frans A.; van den Eertwegh, Alfons J. M.; Giaccone, Giuseppe; Miedema, Frank; Scheper, Rik J.; Pinedo, Herbert M.

    2002-01-01

    CD1d-restricted NKT cells express an invariant TCR and have been demonstrated to play an important regulatory role in a variety of immune responses. Invariant NKT cells down-regulate autoimmune responses by production of type 2 cytokines and can initiate antitumor and antimicrobial immune responses

  15. Body Temperature during Hibernation Is Highly Correlated with a Decrease in Circulating Innate Immune Cells in the Brown Bear (Ursus arctos): A Common Feature among Hibernators?

    OpenAIRE

    Sahdo, Berolla; Evans, Alina L.; Arnemo, Jon Martin; Fröbert, Ole; Särndahl, Eva; Blanc, Stéphane

    2013-01-01

    International audience; Background: Hibernation involves periods of severely depressed metabolism (torpor) and decreases in body temperature (Tb). Small arctic mammals ( < 5kg), in which Tb generally drop drastically, display leukopenia during hibernation. This raised the question of whether the decreased leukocyte counts in mammalian hibernators is due to torpor per se or is secondary to low Tb. The present study examined immune cell counts in brown bears (Ursus arctos), where torpor is only...

  16. Increased miR-223 expression in T cells from patients with rheumatoid arthritis leads to decreased insulin-like growth factor-1-mediated interleukin-10 production.

    Science.gov (United States)

    Lu, M-C; Yu, C-L; Chen, H-C; Yu, H-C; Huang, H-B; Lai, N-S

    2014-09-01

    We hypothesized that the aberrant expression of microRNAs (miRNAs) in rheumatoid arthritis (RA) T cells was involved in the pathogenesis of RA. The expression profile of 270 human miRNAs in T cells from the first five RA patients and five controls were analysed by real-time polymerase chain reaction. Twelve miRNAs exhibited potentially aberrant expression in RA T cells compared to normal T cells. After validation with another 22 RA patients and 19 controls, miR-223 and miR-34b were over-expressed in RA T cells. The expression levels of miR-223 were correlated positively with the titre of rheumatoid factor (RF) in RA patients. Transfection of Jurkat cells with miR-223 mimic suppressed insulin-like growth factor-1 receptor (IGF-1R) and transfection with miR-34b mimic suppressed cAMP response element binding protein (CREB) protein expression by Western blotting. The protein expression of IGF-1R but not CREB was decreased in RA T cells. The addition of recombinant IGF-1-stimulated interleukin (IL)-10 production by activated normal T cells, but not RA T cells. The transfection of miR-223 mimic impaired IGF-1-mediated IL-10 production in activated normal T cells. The expression levels of SCD5, targeted by miR-34b, were decreased in RA T cells after microarray analysis. In conclusion, both miR-223 and miR-34b were over-expressed in RA T cells, but only the miR-223 expression levels were correlated positively with RF titre in RA patients. Functionally, the increased miR-223 expression could impair the IGF-1-mediated IL-10 production in activated RA T cells in vivo, which might contribute to the imbalance between proinflammatory and anti-inflammatory cytokines. © 2014 British Society for Immunology.

  17. Decreased UV-induced DNA repair synthesis in peripheral leukocytes from patients with the nevoid basal cell carcinoma syndrome

    International Nuclear Information System (INIS)

    Ringborg, U.; Lambert, B.; Landergen, J.; Lewensohn, R.

    1981-01-01

    The uv-induced DNA repair synthesis in peripheral leukocytes from 7 patients with the nevoid basal cell carcinoma syndrome was compared to that in peripheral leukocytes from 5 patients with basal cell carcinomas and 39 healthy subjects. A dose response curve was established for each individual, and maximum DNA repair synthesis was used as a measure of the capacity for DNA repair. The patients with the nevoid basal cell carcinoma syndrome had about 25% lower level of maximum DNA repair synthesis as compared to the patients with basal cell carcinomas and control individuals. The possibility that DNA repair mechanisms may be involved in the etiology to the nevoid basal cell carcinoma syndrome is discussed

  18. Chronic exposure of HIT cells to high glucose concentrations paradoxically decreases insulin gene transcription and alters binding of insulin gene regulatory protein.

    Science.gov (United States)

    Olson, L K; Redmon, J B; Towle, H C; Robertson, R P

    1993-07-01

    Chronically culturing HIT-T15 cells in media containing high glucose concentrations leads to decreased insulin mRNA levels, insulin content, and insulin secretion. These changes can be prevented by culturing the cells in media containing lower glucose levels (Robertson, R. P., H.-J. Zhang, K. L. Pyzdrowski, and T. F. Walseth. 1992. J. Clin. Invest. 90:320-325). The mechanism of this seemingly paradoxical phenomenon was examined by transiently transfecting HIT cells with a chloramphenicol acetyl transferase (CAT) reporter gene controlled by the 5'-regulatory domain of the human insulin gene (INSCAT). Early passages of HIT cells readily expressed INSCAT, whereas late passages of cells chronically cultured in 11.1 mM glucose expressed only 28.7 +/- 2.3% (mean +/- SEM) of the CAT activity expressed in early passages. In contrast, late passages of HIT cells chronically cultured in 0.8 mM glucose retained the ability to express the INSCAT reporter gene to 69.6 +/- 10.0% of the CAT activity observed in early passages. The decrease in INSCAT expression in late passages of cells serially cultured in 11.1 mM glucose was associated with the inability to form a specific nuclear protein-DNA complex with the CT motifs of the human insulin promoter. Formation of this specific protein-DNA complex was preserved in late passages of HIT cells when serially cultured in 0.8 mM glucose. Mutations of the CT motifs caused markedly diminished CAT activity in all passages examined. These data indicate that chronic exposure of the beta cell to high glucose concentrations can paradoxically decrease insulin gene transcription, in part, by altering the ability of a regulatory protein (GSTF) to interact with the insulin gene promoter. This provides a potential mechanism for glucotoxic effects on the beta cell at the level of the insulin gene.

  19. Notch signaling mediates the age-associated decrease in adhesion of germline stem cells to the niche.

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    Chen-Yuan Tseng

    2014-12-01

    Full Text Available Stem cells have an innate ability to occupy their stem cell niche, which in turn, is optimized to house stem cells. Organ aging is associated with reduced stem cell occupancy in the niche, but the mechanisms involved are poorly understood. Here, we report that Notch signaling is increased with age in Drosophila female germline stem cells (GSCs, and this results in their removal from the niche. Clonal analysis revealed that GSCs with low levels of Notch signaling exhibit increased adhesiveness to the niche, thereby out-competing their neighbors with higher levels of Notch; adhesiveness is altered through regulation of E-cadherin expression. Experimental enhancement of Notch signaling in GSCs hastens their age-dependent loss from the niche, and such loss is at least partially mediated by Sex lethal. However, disruption of Notch signaling in GSCs does not delay GSC loss during aging, and nor does it affect BMP signaling, which promotes self-renewal of GSCs. Finally, we show that in contrast to GSCs, Notch activation in the niche (which maintains niche integrity, and thus mediates GSC retention is reduced with age, indicating that Notch signaling regulates GSC niche occupancy both intrinsically and extrinsically. Our findings expose a novel role of Notch signaling in controlling GSC-niche adhesion in response to aging, and are also of relevance to metastatic cancer cells, in which Notch signaling suppresses cell adhesion.

  20. Melatonin Decreases Glucose Metabolism in Prostate Cancer Cells: A 13C Stable Isotope-Resolved Metabolomic Study

    Science.gov (United States)

    Hevia, David; Gonzalez-Menendez, Pedro; Fernandez-Fernandez, Mario; Cueto, Sergio; Mayo, Juan C.

    2017-01-01

    The pineal neuroindole melatonin exerts an exceptional variety of systemic functions. Some of them are exerted through its specific membrane receptors type 1 and type 2 (MT1 and MT2) while others are mediated by receptor-independent mechanisms. A potential transport of melatonin through facilitative glucose transporters (GLUT/SLC2A) was proposed in prostate cancer cells. The prostate cells have a particular metabolism that changes during tumor progression. During the first steps of carcinogenesis, oxidative phosphorylation is reactivated while the switch to the “Warburg effect” only occurs in advanced tumors and in the metastatic stage. Here, we investigated whether melatonin might change prostate cancer cell metabolism. To do so, 13C stable isotope-resolved metabolomics in androgen sensitive LNCaP and insensitive PC-3 prostate cancer cells were employed. In addition to metabolite 13C-labeling, ATP/AMP levels, and lactate dehydrogenase or pentose phosphate pathway activity were measured. Melatonin reduces lactate labeling in androgen-sensitive cells and it also lowers 13C-labeling of tricarboxylic acid cycle metabolites and ATP production. In addition, melatonin reduces lactate 13C-labeling in androgen insensitive prostate cancer cells. Results demonstrated that melatonin limits glycolysis as well as the tricarboxylic acid cycle and pentose phosphate pathway in prostate cancer cells, suggesting that the reduction of glucose uptake is a major target of the indole in this tumor type. PMID:28933733

  1. Non integrative strategy decreases chromosome instability and improves endogenous pluripotency genes reactivation in porcine induced pluripotent-like stem cells.

    Science.gov (United States)

    Congras, Annabelle; Barasc, Harmonie; Canale-Tabet, Kamila; Plisson-Petit, Florence; Delcros, Chantal; Feraud, Olivier; Oudrhiri, Noufissa; Hadadi, Eva; Griscelli, Franck; Bennaceur-Griscelli, Annelise; Turhan, Ali; Afanassieff, Marielle; Ferchaud, Stéphane; Pinton, Alain; Yerle-Bouissou, Martine; Acloque, Hervé

    2016-06-01

    The pig is an emerging animal model, complementary to rodents for basic research and for biomedical and agronomical purposes. However despite the progress made on mouse and rat models to produce genuine pluripotent cells, it remains impossible to produce porcine pluripotent cell lines with germline transmission. Reprogramming of pig somatic cells using conventional integrative strategies remains also unsatisfactory. In the present study, we compared the outcome of both integrative and non-integrative reprogramming strategies on pluripotency and chromosome stability during pig somatic cell reprogramming. The porcine cell lines produced with integrative strategies express several pluripotency genes but they do not silence the integrated exogenes and present a high genomic instability upon passaging. In contrast, pig induced pluripotent-like stem cells produced with non-integrative reprogramming system (NI-iPSLCs) exhibit a normal karyotype after more than 12 months in culture and reactivate endogenous pluripotency markers. Despite the persistent expression of exogenous OCT4 and MYC, these cells can differentiate into derivatives expressing markers of the three embryonic germ layers and we propose that these NI-iPSLCs can be used as a model to bring new insights into the molecular factors controlling and maintaining pluripotency in the pig and other non-rodent mammalians.

  2. Stathmin decreases cholangiocarcinoma cell line sensitivity to staurosporine-triggered apoptosis via the induction of ERK and Akt signaling

    Science.gov (United States)

    Bo, Xiaobo; Wang, Yaojie; Wang, Jiwen; Shen, Sheng; Liu, Han; Suo, Tao; Pan, Hongtao; Ai, Zhilong; Liu, Houbao

    2017-01-01

    Cholangiocarcinoma is a rare, but highly fatal malignancy. However, the intrinsic mechanism involved in its tumorigenesis remains obscure. An urgent need remains for a promising target for cholangiocarcinoma biological therapies. Based on comparative proteomical technologies, we found 253 and 231 different spots in gallbladder tumor cell lines and cholangiocarcinoma cell lines, respectively, relative to non-malignant cells. Using Mass Spectrometry (MS) and database searching, we chose seven differentially expressed proteins. High Stathmin expression was found in both cholangiocarcinoma and gallbladder carcinoma cells. Stathmin expression was validated using immunohistochemistry and western blot in cholangiocarcinoma tissue samples and peritumoral tissue. It was further revealed that high Stathmin expression was associated with the repression of staurosporine-induced apoptosis in the cholangiocarcinoma cell. Moreover, we found that Stathmin promoted cancer cell proliferation and inhibited its apoptosis through protein kinase B (Akt) and extracellular signal-regulated kinase (ERK) signaling. Integrin, β1 appears to serve as a partner of Stathmin induction of ERK and Akt signaling by inhibiting apoptosis in the cholangiocarcinoma cell. Understanding the regulation of anti-apoptosis effect by Stathmin might provide new insight into how to overcome therapeutic resistance in cholangiocarcinoma. PMID:28178656

  3. CD8+ lymphocytes control viral replication in SIVmac239-infected rhesus macaques without decreasing the lifespan of productively infected cells.

    Directory of Open Access Journals (Sweden)

    Nichole R Klatt

    2010-01-01

    Full Text Available While CD8+ T cells are clearly important in controlling virus replication during HIV and SIV infections, the mechanisms underlying this antiviral effect remain poorly understood. In this study, we assessed the in vivo effect of CD8+ lymphocyte depletion on the lifespan of productively infected cells during chronic SIVmac239 infection of rhesus macaques. We treated two groups of animals that were either CD8+ lymphocyte-depleted or controls with antiretroviral therapy, and used mathematical modeling to assess the lifespan of infected cells either in the presence or absence of CD8+ lymphocytes. We found that, in both early (day 57 post-SIV and late (day 177 post-SIV chronic SIV infection, depletion of CD8+ lymphocytes did not result in a measurable increase in the lifespan of either short- or long-lived productively infected cells in vivo. This result indicates that the presence of CD8+ lymphocytes does not result in a noticeably shorter lifespan of productively SIV-infected cells, and thus that direct cell killing is unlikely to be the main mechanism underlying the antiviral effect of CD8+ T cells in SIV-infected macaques with high virus replication.

  4. Decreased proliferative, migrative and neuro-differentiative potential of postnatal rat enteric neural crest-derived cells during culture in vitro

    International Nuclear Information System (INIS)

    Yu, Hui; Pan, Wei-Kang; Zheng, Bai-Jun; Wang, Huai-Jie; Chen, Xin-Lin; Liu, Yong; Gao, Ya

    2016-01-01

    A growing body of evidence supports the potential use of enteric neural crest-derived cells (ENCCs) as a cell replacement therapy for Hirschsprung's disease. Based on previous observations of robust propagation of primary ENCCs, as opposed to their progeny, it is suggested that their therapeutic potential after in vitro expansion may be restricted. We therefore examined the growth and differentiation activities and phenotypic characteristics of continuous ENCC cultures. ENCCs were isolated from the intestines of postnatal rats and were identified using an immunocytochemical approach. During continuous ENCC culture expansion, proliferation, migration, apoptosis, and differentiation potentials were monitored. The Cell Counting Kit-8 was used for assessment of ENCC vitality, Transwell inserts for cell migration, immunocytochemistry for cell counts and identification, and flow cytometry for apoptosis. Over six continuous generations, ENCC proliferation potency was reduced and with prolonged culture, the ratio of migratory ENCCs was decreased. The percentage of apoptosis showed an upward trend with prolonged intragenerational culture, but showed a downward trend with prolonged culture of combined generations. Furthermore, the percentage of peripherin + cells decreased whilst the percentage of GFAP + cells increased with age. The results demonstrated that alterations in ENCC growth characteristics occur with increased culture time, which may partially account for the poor results of proposed cell therapies. - Highlights: • Differences were identified between primary and daughter ENCCs. • Daughter ENCCs had reduced proliferation, migration and differentiation. • Daughter ENCCs also had increased apoptosis. • These altered characteristics warrant further investigation.

  5. Decrease of miR-622 expression promoted the proliferation, migration and invasion of cholangiocarcinoma cells by targeting regulation of c-Myc.

    Science.gov (United States)

    Wu, Yi-Fei; Li, Zhuo-Ri; Cheng, Zhi-Qi; Yin, Xin-Min; Wu, Jin-Shu

    2017-12-01

    To explore the mechanism of miR-622 in regulating the proliferation, migration and invasion of cholangiocarcinoma (CCA) cells. Quantitative real-time PCR was conducted to measure the expression of miR-622 and c-Myc in CCA tissues and cell lines. Protein level of c-Myc was measured by Western blot. The effect of miR-622 on cell proliferation, migration and invasion was analyzed by MTT assay and Transwell chamber migration assay. Luciferase reporter assay was performed to measure the effect of miR-622 on c-Myc. miR-622 expression was downregulated in both CCA tissues and cell lines, while c-Myc expression was uregulated. Overexpression of miR-622 in CCA cells was statistically correlated with a decrease of cell proliferation, migration and invasion, while inhibition of miR-622 made an inverse result. We also proved c-Myc was identified as a target gene of miR-622 in CCA. Moreover, we found overexpression of c-Myc can strengthen the effects of miR-622 on the proliferation, migration and invasion of CCA cells. Decrease of miR-622 promotes the proliferation, migration and invasion of CCA cells by directly targeting c-Myc. Copyright © 2017. Published by Elsevier Masson SAS.

  6. Decreased proliferative, migrative and neuro-differentiative potential of postnatal rat enteric neural crest-derived cells during culture in vitro

    Energy Technology Data Exchange (ETDEWEB)

    Yu, Hui [Department of Pediatric Surgery, the Second Affiliated Hospital, Xi’an Jiaotong University, No 157, Xi Wu Road, Xi’an 710004, Shaanxi (China); Institute of Neurobiology, Environment and Genes Related to Diseases Key Laboratory of Chinese Ministry of Education, Xi’an Jiaotong University, No 96, Yan Ta Xi Road, Xi’an 710061, Shaanxi (China); Pan, Wei-Kang; Zheng, Bai-Jun; Wang, Huai-Jie [Department of Pediatric Surgery, the Second Affiliated Hospital, Xi’an Jiaotong University, No 157, Xi Wu Road, Xi’an 710004, Shaanxi (China); Chen, Xin-Lin; Liu, Yong [Institute of Neurobiology, Environment and Genes Related to Diseases Key Laboratory of Chinese Ministry of Education, Xi’an Jiaotong University, No 96, Yan Ta Xi Road, Xi’an 710061, Shaanxi (China); Gao, Ya, E-mail: ygao@mail.xjtu.edu.cn [Department of Pediatric Surgery, the Second Affiliated Hospital, Xi’an Jiaotong University, No 157, Xi Wu Road, Xi’an 710004, Shaanxi (China)

    2016-05-01

    A growing body of evidence supports the potential use of enteric neural crest-derived cells (ENCCs) as a cell replacement therapy for Hirschsprung's disease. Based on previous observations of robust propagation of primary ENCCs, as opposed to their progeny, it is suggested that their therapeutic potential after in vitro expansion may be restricted. We therefore examined the growth and differentiation activities and phenotypic characteristics of continuous ENCC cultures. ENCCs were isolated from the intestines of postnatal rats and were identified using an immunocytochemical approach. During continuous ENCC culture expansion, proliferation, migration, apoptosis, and differentiation potentials were monitored. The Cell Counting Kit-8 was used for assessment of ENCC vitality, Transwell inserts for cell migration, immunocytochemistry for cell counts and identification, and flow cytometry for apoptosis. Over six continuous generations, ENCC proliferation potency was reduced and with prolonged culture, the ratio of migratory ENCCs was decreased. The percentage of apoptosis showed an upward trend with prolonged intragenerational culture, but showed a downward trend with prolonged culture of combined generations. Furthermore, the percentage of peripherin{sup +} cells decreased whilst the percentage of GFAP{sup +} cells increased with age. The results demonstrated that alterations in ENCC growth characteristics occur with increased culture time, which may partially account for the poor results of proposed cell therapies. - Highlights: • Differences were identified between primary and daughter ENCCs. • Daughter ENCCs had reduced proliferation, migration and differentiation. • Daughter ENCCs also had increased apoptosis. • These altered characteristics warrant further investigation.

  7. Hypomethylating drugs efficiently decrease cytosine methylation in telomeric DNA and activate telomerase without affecting telomere lengths in tobacco cells

    Czech Academy of Sciences Publication Activity Database

    Majerová, E.; Fojtová, M.; Mozgová, I.; Bittová, M.; Fajkus, Jiří

    2011-01-01

    Roč. 77, 4-5 (2011), s. 371-380 ISSN 0167-4412 Institutional support: RVO:68081707 Keywords : Nicotiana tabacum * Cell culture * Telomere Subject RIV: BO - Biophysics Impact factor: 4.150, year: 2011

  8. Lipid-induced insulin resistance in cultured hepatoma cells is associated with a decreased insulin receptor tyrosine kinase activity.

    OpenAIRE

    Hubert, P; Bruneau-Wack, C; Cremel, G; Le Marchand-Brustel, Y; Staedel, C

    1991-01-01

    We have shown previously that experimental modifications of the cellular lipid composition of an insulin-sensitive rat hepatoma cell line (Zajdela Hepatoma Culture, ZHC) affect both binding and biological actions of insulin. Discrepancies between insulin binding and actions implied a postbinding defect, responsible for the observed insulin resistance in lipid-treated cells. To elucidate the mechanism for this defect, we have studied insulin binding and insulin receptor kinase activity in part...

  9. Decreased expression of the mitochondrial solute carrier SLC25A43 in basal cell carcinoma compared with healthy skin.

    Science.gov (United States)

    Prosén, Sara; Eremo, Anna Göthlin; Tsegai, Alexander Duarte; Lindberg, Magnus; Tina, Elisabet

    2017-08-01

    Basal cell carcinoma is the most common type of cancer in fair-skinned individuals, and its incidence is rapidly increasing. The aim of the present study was to investigate the gene and protein expression of the mitochondrial solute carrier family 25 member 43 (SLC25A43) in basal cell carcinoma. SLC25A43 has previously been identified to be genetically altered and associated with cell proliferation in human epidermal growth factor receptor 2-positive breast cancer. However, the knowledge about SLC25A43 is limited, and its role in other cancers is unknown. The SLC25A43 gene and protein expression was analysed in 14 basal cell carcinomas and healthy skin samples from the same individuals by quantitative polymerase chain reaction and immunohistochemistry, respectively. The results demonstrated a significantly lower (≥50%) SLC25A43 gene expression in all carcinomas compared with that in healthy skin. In addition, SLC25A43 protein expression was absent in >90% of all visual fields in the basal cell carcinomas, and the H-score was significantly lower in tumours compared with the adjacent epidermis. These results demonstrate that SLC25A43 expression is altered at the gene and protein levels in basal cell carcinoma. The underlying mechanisms and the clinical relevance of these data must be elucidated in additional experimental and clinical studies.

  10. Antagonism of serotonin receptor 1B decreases viability and promotes apoptosis in the COS canine osteosarcoma cell line.

    Science.gov (United States)

    Viall, A K; Goodall, C P; Stang, B; Marley, K; Chappell, P E; Bracha, S

    2016-06-01

    Serotonin receptor 1B (5HTR1B) traditionally exhibits anti-proliferative activity in osteoblasts. We examined the expression and function of 5HTR1B in the COS canine osteosarcoma cell line and normal canine osteoblasts. Equal levels of 5HTR1B gene and protein expression were found between normal and malignant osteoblasts. Treatment with serotonin enhanced viability of osteosarcoma cells but not normal osteoblasts. Challenge with the 5HTR1B agonist anpirtoline caused no change in cell viability. Rather incubation with the specific receptor antagonist SB224289 caused reduction in osteoblast viability, with this effect more substantial in osteosarcoma cells. Investigation of this inhibitory activity showed 5HTR1B antagonism induces apoptosis in malignant cells. Evaluation of phosphorylated levels of CREB and ERK, transcriptional regulators associated with serotonin receptor signalling in osteoblasts, revealed aberrant 5HTR1B signalling in COS. Our results confirm the presence of 5HTR1B in a canine osteosarcoma cell line and highlight this receptor as a possible novel therapeutic target. © 2014 John Wiley & Sons Ltd.

  11. Tumor cells have decreased ability to metabolize H2O2: Implications for pharmacological ascorbate in cancer therapy

    Directory of Open Access Journals (Sweden)

    Claire M. Doskey

    2016-12-01

    Full Text Available Ascorbate (AscH− functions as a versatile reducing agent. At pharmacological doses (P-AscH−; [plasma AscH−] ≥≈20 mM, achievable through intravenous delivery, oxidation of P-AscH− can produce a high flux of H2O2 in tumors. Catalase is the major enzyme for detoxifying high concentrations of H2O2. We hypothesize that sensitivity of tumor cells to P-AscH− compared to normal cells is due to their lower capacity to metabolize H2O2. Rate constants for removal of H2O2 (kcell and catalase activities were determined for 15 tumor and 10 normal cell lines of various tissue types. A differential in the capacity of cells to remove H2O2 was revealed, with the average kcell for normal cells being twice that of tumor cells. The ED50 (50% clonogenic survival of P-AscH− correlated directly with kcell and catalase activity. Catalase activity could present a promising indicator of which tumors may respond to P-AscH−.

  12. Loss of Selenium-Binding Protein 1 Decreases Sensitivity to Clastogens and Intracellular Selenium Content in HeLa Cells.

    Science.gov (United States)

    Zhao, Changhui; Zeng, Huawei; Wu, Ryan T Y; Cheng, Wen-Hsing

    2016-01-01

    Selenium-binding protein 1 (SBP1) is not a selenoprotein but structurally binds selenium. Loss of SBP1 during carcinogenesis usually predicts poor prognosis. Because genome instability is a hallmark of cancer, we hypothesize that SBP1 sequesters cellular selenium and sensitizes cancer cells to DNA-damaging agents. To test this hypothesis, we knocked down SBP1 expression in HeLa cervical cancer cells by employing a short hairpin RNA (shRNA) approach. Reduced sensitivity to hydrogen peroxide, paraquat and camptothecin, reactive oxygen species content, and intracellular retention of selenium after selenomethionine treatment were observed in SBP1 shRNA HeLa cells. Results from Western analyses showed that treatment of HeLa cells with selenomethionine resulted in increased SBP1 protein expression in a dose-dependent manner. Knockdown of SBP1 rendered HeLa cells increased expression of glutathione peroxidase-1 but not glutathione peroxidase-4 protein levels and accelerated migration from a wound. Altogether, SBP1 retains supplemental selenium and sensitizes HeLa cancer cells to clastogens, suggesting a new cancer treatment strategy by sequestering selenium through SBP1.

  13. Decreased number of CD4+ and CD8+ T cells that express the interleukin-7 receptor in blood and tissues of SIV-infected macaques

    International Nuclear Information System (INIS)

    Moniuszko, Marcin; Edghill-Smith, Yvette; Venzon, David; Stevceva, Liljana; Nacsa, Janos; Tryniszewska, Elzbieta; Tsai, Wen-Po; Franchini, Genoveffa

    2006-01-01

    Acute HIV/SIV (human/simian immunodeficiency virus) infection results in severe CD4 + T cell depletion in lymphoid compartments. During the chronic phase of infection, CD4 + T cell numbers rebound in blood but remain low in the gut-associated lymphoid tissue (GALT), even when viral replication is suppressed by antiretroviral therapy (ART). Thus, strategies to repopulate lymphoid compartments may ameliorate the clinical outcome of HIV/SIV infection. Interleukin (IL)-7 is a key cytokine for the maintenance of homeostatic proliferation of T cells. In HIV/SIV infection, IL-7 expression is increased, likely to compensate for T cell loss, suggesting that supraphysiological administration of IL-7 could provide additional benefit. However, the ability of T cells to respond to IL-7 is dependent on the level of expression of the IL-7 receptor (IL-7R) in T cells in various body compartments. In here, we investigated the proportion of IL-7R + T cells in blood, spleen, gut, and genitourinary tract of healthy and SIV-infected macaques with various degrees of CD4 + T cell depletion. We found that the percentage of T cells expressing IL-7R was significantly lower in both CD4 + and CD8 + T cell subsets in SIV-infected macaques than in healthy animals and this decrease directly correlated with the CD4 + T cell number. Importantly, the proportion of CD4 + and CD8 + T cells expressing IL-7R in blood paralleled that found in tissues. IL-7R + T cells within the SIV-specific CD8 + T cells varied and were lowest in most tissues of viremic macaques, likely reflecting continuous antigen stimulation of effector cells

  14. SET overexpression decreases cell detoxification efficiency: ALDH2 and GSTP1 are downregulated, DDR is impaired and DNA damage accumulates.

    Science.gov (United States)

    Almeida, Luciana O; Goto, Renata N; Pestana, Cezar R; Uyemura, Sérgio A; Gutkind, Silvio; Curti, Carlos; Leopoldino, Andréia M

    2012-12-01

    Alcohol and tobacco consumption are risk factors for head and neck squamous cell carcinoma (HNSCC). Aldehyde dehydrogenase 2 (ALDH2) and glutathione S-transferase pi 1 (GSTP1) are important enzymes for cellular detoxification and low efficiencies are implicated in cancer. We assessed the potential role of SET protein overexpression, a histone acetylation modulator accumulated in HNSCC, in gene regulation and protein activity of ALDH2 and GSTP1. SET was knocked down in HN13, HN12 and Cal27, and overexpressed in HEK293 cells; ethanol and cisplatin were the chemical agents. Cells with SET overexpression (HEK293/SET, HN13 and HN12) showed lower ALDH2 and GSTP1 mRNA levels and trichostatin A increased them (real-time PCR). Ethanol upregulated GSTP1 and ALDH2 mRNAs, whereas cisplatin upregulated GSTP1 in HEK293 cells. SET-chromatin binding revealed SET interaction with ALDH2 and GSTP1 promoters, specifically via SET NAP domain; ethanol and cisplatin abolished SET binding. ALDH2 and GSTP1 efficiency was assessed by enzymatic and comet assay. A lower ALDH2 activity was associated with greater DNA damage (tail intensity) in HEK293/SET compared with HEK293 cells, whereas HN13/siSET showed ALDH2 activity higher than HN13 cells. HN13/siSET cells showed increased tail intensity. Cisplatin-induced DNA damage response showed negative relationship between SET overexpression and BRCA2 recruitment. SET downregulated repair genes ATM, BRCA1 and CHEK2 and upregulated TP53. Cisplatin-induced cell-cycle arrest occurred in G(0) /G(1) and S in HEK293 cells, whereas HEK293/SET showed G(2) /M stalling. Overall, cisplatin was more cytotoxic for HN13 than HN13/siSET cells. Our data suggest a role for SET in cellular detoxification, DNA damage response and genome integrity. © 2012 The Authors Journal compilation © 2012 FEBS.

  15. Decrease in paracellular permeability and chemosensitivity to doxorubicin by claudin-1 in spheroid culture models of human lung adenocarcinoma A549 cells.

    Science.gov (United States)

    Akizuki, Risa; Maruhashi, Ryohei; Eguchi, Hiroaki; Kitabatake, Kazuki; Tsukimoto, Mitsutoshi; Furuta, Takumi; Matsunaga, Toshiyuki; Endo, Satoshi; Ikari, Akira

    2018-03-07

    Chemotherapy resistance is a major problem in the treatment of cancer, but the underlying mechanisms are not fully understood. We found that the expression levels of claudin-1 (CLDN1) and 3, tight junctional proteins, are upregulated in cisplatin (CDDP)-resistant human lung adenocarcinoma A549 (A549R) cells. A549R cells showed cross-resistance to doxorubicin (DXR). Here, the expression mechanism and function of CLDN1 and 3 were examined. CLDN1 and 3 were mainly localized at tight junctions concomitant with zonula occludens (ZO)-1, a scaffolding protein, in A549 and A549R cells. The phosphorylation levels of Src, MEK, ERK, c-Fos, and Akt in A549R cells were higher than those in A549 cells. The expression levels of CLDN1 and 3 were decreased by LY-294002, a phosphoinositide 3-kinase (PI3K) inhibitor, and BAY 11-7082, an NF-κB inhibitor. The overexpression of CLDN1 and 3 decreased the paracellular permeability of DXR in A549 cells. Hypoxia levels in A549R and CLDN1-overexpressing cells (CLDN1/A549) were greater than those in A549, mock/A549, and CLDN3/A549 cells in a spheroid culture model. In contrast, accumulation in the region inside the spheroids and the toxicity of DXR in A549R and CLDN1/A549 cells were lower than those in other cells. Furthermore, the accumulation and toxicity of DXR were rescued by CLDN1 siRNA in A549R cells. We suggest that CLDN1 is upregulated by CDDP resistance through activation of a PI3K/Akt/NF-κB pathway, resulting in the inhibition of penetration of anticancer drugs into the inner area of spheroids. Copyright © 2018 Elsevier B.V. All rights reserved.

  16. Stroke promotes survival of nearby transplanted neural stem cells by decreasing their activation of caspase 3 while not affecting their differentiation.

    Science.gov (United States)

    Kosi, Nina; Alić, Ivan; Salamon, Iva; Mitrečić, Dinko

    2018-02-14

    Although transplantation of stem cells improves recovery of the nervous tissue, little is known about the influence of different brain regions on transplanted cells. After we confirmed that cells with uniform differentiation potential can be generated in independent experiments, one million of neural stem cells isolated from B6.Cg-Tg(Thy1-YFP)16Jrs/J mouse embryos were transplanted into the brain 24 h after induction of stroke. The lateral ventricles, the corpus callosum and the striatum were tested. Two and four weeks after the transplantation, the cells transplanted in all three regions have been attracted to the ischemic core. The largest number of attracted cells has been observed after transplantation into the striatum. Their differentiation pattern and expression of neuroligin 1, SynCAM 1, postsynaptic density protein 95 and synapsin 1 followed the same pattern observed during in vitro cultivation and it did not differ among the tested regions. Differentiation pattern of the cells transplanted in the stroke-affected and healthy animals was the same. On the other hand, neural stem cells transplanted in the striatum of the animals affected by stroke exhibited significantly increased survival rates reaching 260 ± 19%, when compared to cells transplanted in their wild type controls. Surprisingly, improved survival two and four weeks after transplantation was not due to increased proliferation of the grafted cells and it was accompanied by decreased levels of activity of Casp3 (19.56 ± 3.1% in the stroke-affected vs. 30.14 ± 2.4% in healthy animals after four weeks). We assume that the decreased levels of Casp3 in cells transplanted near the ischemic region was linked to increased vasculogenesis, synaptogenesis, astrocytosis and axonogenesis detected in the host tissue affected by ischemia. Copyright © 2017 Elsevier B.V. All rights reserved.

  17. Treatment with a JNK inhibitor increases, whereas treatment with a p38 inhibitor decreases, H2O2-induced calf pulmonary arterial endothelial cell death.

    Science.gov (United States)

    Park, Woo Hyun

    2017-08-01

    Oxidative stress induces apoptosis in endothelial cells (ECs). Reactive oxygen species (ROS) promote cell death by regulating the activity of various mitogen-activated protein kinases (MAPKs) in ECs. The present study investigated the effects of MAPK inhibitors on cell survival and glutathione (GSH) levels upon H 2 O 2 treatment in calf pulmonary artery ECs (CPAECs). H 2 O 2 treatment inhibited the growth and induced the death of CPAECs, as well as causing GSH depletion and the loss of mitochondrial membrane potential (MMP). While treatment with the MEK or JNK inhibitor impaired the growth of H 2 O 2 -treated CPAECs, treatment with the p38 inhibitor attenuated this inhibition of growth. Additionally, JNK inhibitor treatment increased the proportion of sub-G 1 phase cells in H 2 O 2 -treated CPAECs and further decreased the MMP. However, treatment with a p38 inhibitor reversed the effects of H 2 O 2 treatment on cell growth and the MMP. Similarly, JNK inhibitor treatment further increased, whereas p38 inhibitor treatment decreased, the proportion of GSH-depleted cells in H 2 O 2 -treated CPAECs. Each of the MAPK inhibitors affected cell survival, and ROS or GSH levels differently in H 2 O 2 -untreated, control CPAECs. The data suggest that the exposure of CPAECs to H 2 O 2 caused the cell growth inhibition and cell death through GSH depletion. Furthermore, JNK inhibitor treatment further enhanced, whereas p38 inhibitors attenuated, these effects. Thus, the results of the present study suggest a specific protective role for the p38 inhibitor, and not the JNK inhibitor, against H 2 O 2 -induced cell growth inhibition and cell death.

  18. Decreased SAP Expression in T Cells from Patients with Systemic Lupus Erythematosus Contributes to Early Signaling Abnormalities and Reduced IL-2 Production.

    Science.gov (United States)

    Karampetsou, Maria P; Comte, Denis; Kis-Toth, Katalin; Terhorst, Cox; Kyttaris, Vasileios C; Tsokos, George C

    2016-06-15

    T cells from patients with systemic lupus erythematosus (SLE) display a number of abnormalities, including increased early signaling events following engagement of the TCR. Signaling lymphocytic activation molecule family cell surface receptors and the X-chromosome-defined signaling lymphocytic activation molecule-associated protein (SAP) adaptor are important in the development of several immunocyte lineages and modulating the immune response. We present evidence that SAP protein levels are decreased in T cells and in their main subsets isolated from 32 women and three men with SLE, independent of disease activity. In SLE T cells, SAP protein is also subject to increased degradation by caspase-3. Forced expression of SAP in SLE T cells normalized IL-2 production, calcium (Ca(2+)) responses, and tyrosine phosphorylation of a number of proteins. Exposure of normal T cells to SLE serum IgG, known to contain anti-CD3/TCR Abs, resulted in SAP downregulation. We conclude that SLE T cells display reduced levels of the adaptor protein SAP, probably as a result of continuous T cell activation and degradation by caspase-3. Restoration of SAP levels in SLE T cells corrects the overexcitable lupus T cell phenotype. Copyright © 2016 by The American Association of Immunologists, Inc.

  19. MicroRNA-29a is up-regulated in beta-cells by glucose and decreases glucose-stimulated insulin secretion

    International Nuclear Information System (INIS)

    Bagge, Annika; Clausen, Trine R.; Larsen, Sylvester; Ladefoged, Mette; Rosenstierne, Maiken W.; Larsen, Louise; Vang, Ole; Nielsen, Jens H.; Dalgaard, Louise T.

    2012-01-01

    Highlights: ► MicroRNA-29a (miR-29a) levels are increased by glucose in human and rat islets and INS-1E cells. ► miR-29a increases proliferation of INS-1E beta-cells. ► Forced expression of miR-29a decreases glucose-stimulated insulin secretion (GSIS). ► Depletion of beta-cell miR-29a improves GSIS. ► miR-29a may be a mediator of glucose toxicity in beta-cells. -- Abstract: Chronically elevated levels of glucose impair pancreatic beta-cell function while inducing beta-cell proliferation. MicroRNA-29a (miR-29a) levels are increased in several tissues in diabetic animals and mediate decreased insulin-stimulated glucose-transport of adipocytes. The aim was to investigate the impact of glucose on miR-29a levels in INS-1E beta-cells and in human islets of Langerhans and furthermore to evaluate the impact of miR-29a on beta-cell function and proliferation. Increased glucose levels up-regulated miR-29a in beta-cells and human and rat islets of Langerhans. Glucose-stimulated insulin-secretion (GSIS) of INS-1E beta-cells was decreased by forced expression of miR-29a, while depletion of endogenous miR-29a improved GSIS. Over-expression of miR-29a increased INS-1E proliferation. Thus, miR-29a up-regulation is involved in glucose-induced proliferation of beta-cells. Furthermore, as depletion of miR-29a improves beta-cell function, miR-29a is a mediator of glucose-induced beta-cell dysfunction. Glucose-induced up-regulation of miR-29a in beta-cells could be implicated in progression from impaired glucose tolerance to type 2 diabetes.

  20. Benzyl butyl phthalate decreases myogenic differentiation of endometrial mesenchymal stem/stromal cells through miR-137-mediated regulation of PITX2.

    Science.gov (United States)

    Chen, Hung-Sheng; Hsu, Chia-Yi; Chang, Yu-Chia; Chuang, Hui-Yu; Long, Cheng-Yu; Hsieh, Tsung-Hua; Tsai, Eing-Mei

    2017-03-15

    Phthalate, an environmental toxin, has been considered as an endocrine-disrupting chemical. Growing evidence has demonstrated links between endocrine-disrupting chemicals, tissue development, and reproductive physiology, but the mechanisms of gene expression regulation by environmental factors that affect cell differentiation are unclear. Herein, we investigated the effects of butyl benzyl phthalate (BBP) on human endometrial mesenchymal stem/stromal cell (EN-MSC) differentiation and identified a novel signaling pathway. Differentiation of endometrial mesenchymal stem/stromal cells decreased after administration of BBP. We analyzed BBP regulation of gene expression in EN-MSC using cDNA microarrays and Ingenuity Pathway Analysis software to identify affected target genes and their biological functions. PITX2 emerged as a common gene hit from separate screens targeting skeletal and muscular disorders, cell morphology, and tissue development. BBP decreased transcription of PITX2 and elevated expression of the microRNA miR-137, the predicted upstream negative regulator of PITX2. These data indicated that BBP affects PITX2 expression through miR-137 targeting of the 3' untranslated region of PITX2 mRNA. PITX2 down-regulation also decreased MyoD transcript levels in EN-MSC. Our results demonstrate that BBP decreases EN-MSC myogenic differentiation through up-regulation of miR-137, contribute to our understanding of EN-MSC differentiation, and underline the hazardous potential of environmental hormones.

  1. Acidosis Decreases c-Myc Oncogene Expression in Human Lymphoma Cells: A Role for the Proton-Sensing G Protein-Coupled Receptor TDAG8

    Directory of Open Access Journals (Sweden)

    Zhigang Li

    2013-10-01

    Full Text Available Acidosis is a biochemical hallmark of the tumor microenvironment. Here, we report that acute acidosis decreases c-Myc oncogene expression in U937 human lymphoma cells. The level of c-Myc transcripts, but not mRNA or protein stability, contributes to c-Myc protein reduction under acidosis. The pH-sensing receptor TDAG8 (GPR65 is involved in acidosis-induced c-Myc downregulation. TDAG8 is expressed in U937 lymphoma cells, and the overexpression or knockdown of TDAG8 further decreases or partially rescues c-Myc expression, respectively. Acidic pH alone is insufficient to reduce c-Myc expression, as it does not decrease c-Myc in H1299 lung cancer cells expressing very low levels of pH-sensing G protein-coupled receptors (GPCRs. Instead, c-Myc is slightly increased by acidosis in H1299 cells, but this increase is completely inhibited by ectopic overexpression of TDAG8. Interestingly, TDAG8 expression is decreased by more than 50% in human lymphoma samples in comparison to non-tumorous lymph nodes and spleens, suggesting a potential tumor suppressor function of TDAG8 in lymphoma. Collectively, our results identify a novel mechanism of c-Myc regulation by acidosis in the tumor microenvironment and indicate that modulation of TDAG8 and related pH-sensing receptor pathways may be exploited as a new approach to inhibit Myc expression.

  2. Gomisin G Inhibits the Growth of Triple-Negative Breast Cancer Cells by Suppressing AKT Phosphorylation and Decreasing Cyclin D1.

    Science.gov (United States)

    Maharjan, Sony; Park, Byoung Kwon; Lee, Su In; Lim, Yoonho; Lee, Keunwook; Kwon, Hyung-Joo

    2018-03-28

    A type of breast cancer with a defect in three molecular markers such as the estrogen receptor, progesterone receptor, and human epidermal growth factor receptor is called triple-negative breast cancer (TNBC). Many patients with TNBC have a lower survival rate than patients with other types due to a poor prognosis. In this study, we confirmed the anti-cancer effect of a natural compound, Gomisin G, in TNBC cancer cells. Treatment with Gomisin G suppressed the viability of two TNBC cell lines, MDA-MB-231 and MDA-MB-468 but not non-TNBC cell lines such as MCF-7, T47D, and ZR75-1. To investigate the molecular mechanism of this activity, we examined the signal transduction pathways after treatment with Gomisin G in MDA-MB-231 cells. Gomisin G did not induce apoptosis but drastically inhibited AKT phosphorylation and reduced the amount of retinoblastoma tumor suppressor protein (Rb) and phosphorylated Rb. Gomisin G induced in a proteasome-dependent manner a decrease in Cyclin D1. Consequently, Gomisin G causes cell cycle arrest in the G1 phase. In contrast, there was no significant change in T47D cells except for a mild decrease in AKT phosphorylation. These results show that Gomisin G has an anti-cancer activity by suppressing proliferation rather than inducing apoptosis in TNBC cells. Our study suggests that Gomisin G could be used as a therapeutic agent in the treatment of TNBC patients.

  3. Downregulation of heat shock protein B8 decreases osteogenic differentiation potential of dental pulp stem cells during in vitro proliferation.

    Science.gov (United States)

    Flanagan, M; Li, C; Dietrich, M A; Richard, M; Yao, S

    2018-04-01

    Tissue-derived stem cells, such as dental pulp stem cells (DPSCs), reduce differentiation capability during in vitro culture. We found that cultured DPSCs reduce expression of heat shock protein B8 (HspB8) and GIPC PDZ domain containing family member 2 (Gipc2). Our objectives were to evaluate the changes in DPSC composition during in vitro proliferation and to determine whether HspB8 and Gipc2 have function in differentiation potential of DPSCs. Different passages of rat DPSCs were evaluated for changes in CD90+ and/or CD271+ stem cells and changes in osteogenic potential. Real-time RT-PCR and immunostaining were conducted to determine expression of HspB8 and Gipc2. Expression of the genes in DPSCs was knocked down by siRNA, followed by osteogenic induction to evaluate the function of the genes. About 90% of cells in the DPSC cultures were CD90+ and/or CD271+ cells without dramatic change during in vitro proliferation. The DPSCs at passages 3 to 5 (P3 to P5) possess strong osteogenic potential, but such potential was greatly reduced at later passages. Expression of HspB8 and Gipc2 was significantly reduced at P11 versus P3. Knock-down of HspB8 expression abolished osteogenic potential of the DPSCs, but knock-down of Gipc2 had no effect. CD90+ and CD271+ cells are the major components of DPSCs in in vitro culture. High-level expression of HspB8 was critical for maintaining differentiation potential of DPSCs. © 2017 John Wiley & Sons Ltd.

  4. Dietary n-3 polyunsaturated fatty acids (PUFA) decrease obesity-associated Th17 cell-mediated inflammation during colitis.

    Science.gov (United States)

    Monk, Jennifer M; Hou, Tim Y; Turk, Harmony F; Weeks, Brad; Wu, Chaodong; McMurray, David N; Chapkin, Robert S

    2012-01-01

    Clinical and experimental evidence suggests that obesity-associated inflammation increases disease activity during colitis, attributed in part to the effects of Th17 cells. Using a model of concurrent obesity and colitis, we monitored changes in critical immune cell subsets and inflammatory biomarker expression in three key tissues: visceral adipose tissue, colon (local inflammatory site) and spleen (systemic inflammatory site), and we hypothesized that n-3 PUFA would reduce the percentage of inflammatory immune cell subsets and suppress inflammatory gene expression, thereby improving the disease phenotype. Obesity was induced in C57BL/6 mice by feeding a high fat (HF) diet (59.2% kcal) alone or an isocaloric HF diet supplemented with fish oil (HF-FO) for 12 weeks. Colitis was induced via a 2.5% trinitrobenzene sulfonic acid (TNBS) enema. The HF-FO diet improved the obese phenotype by reducing i) serum hormone concentrations (leptin and resistin), ii) adipose tissue mRNA expression of inflammatory cytokines (MCP-1, IFNγ, IL-6, IL17F and IL-21) and iii) total (F4/80⁺ CD11b⁺) and inflammatory adipose tissue M1 (F4/80⁺ CD11c⁺) macrophage content compared to HF (Pdiet reduced both colitis-associated disease severity and colonic mRNA expression of the Th17 cell master transcription factor (RORγτ) and critical cytokines (IL-6, IL-17A, IL-17F, IL-21, IL-23 and IFNγ) versus HF (P<0.05). Compared to HF, the percentage of both splenic Th17 and Th1 cells were reduced by the HF-FO group (P<0.05). Under ex vivo polarizing conditions, the percentage of HF-FO derived CD4⁺ T cells that reached Th17 cell effector status was suppressed (P = 0.05). Collectively, these results indicate that n-3 PUFA suppress Th1/Th17 cells and inflammatory macrophage subsets and reconfigure the inflammatory gene expression profile in diverse tissue sites in obese mice following the induction of colitis.

  5. Dietary n-3 polyunsaturated fatty acids (PUFA decrease obesity-associated Th17 cell-mediated inflammation during colitis.

    Directory of Open Access Journals (Sweden)

    Jennifer M Monk

    Full Text Available Clinical and experimental evidence suggests that obesity-associated inflammation increases disease activity during colitis, attributed in part to the effects of Th17 cells. Using a model of concurrent obesity and colitis, we monitored changes in critical immune cell subsets and inflammatory biomarker expression in three key tissues: visceral adipose tissue, colon (local inflammatory site and spleen (systemic inflammatory site, and we hypothesized that n-3 PUFA would reduce the percentage of inflammatory immune cell subsets and suppress inflammatory gene expression, thereby improving the disease phenotype. Obesity was induced in C57BL/6 mice by feeding a high fat (HF diet (59.2% kcal alone or an isocaloric HF diet supplemented with fish oil (HF-FO for 12 weeks. Colitis was induced via a 2.5% trinitrobenzene sulfonic acid (TNBS enema. The HF-FO diet improved the obese phenotype by reducing i serum hormone concentrations (leptin and resistin, ii adipose tissue mRNA expression of inflammatory cytokines (MCP-1, IFNγ, IL-6, IL17F and IL-21 and iii total (F4/80⁺ CD11b⁺ and inflammatory adipose tissue M1 (F4/80⁺ CD11c⁺ macrophage content compared to HF (P<0.05. In addition, the HF-FO diet reduced both colitis-associated disease severity and colonic mRNA expression of the Th17 cell master transcription factor (RORγτ and critical cytokines (IL-6, IL-17A, IL-17F, IL-21, IL-23 and IFNγ versus HF (P<0.05. Compared to HF, the percentage of both splenic Th17 and Th1 cells were reduced by the HF-FO group (P<0.05. Under ex vivo polarizing conditions, the percentage of HF-FO derived CD4⁺ T cells that reached Th17 cell effector status was suppressed (P = 0.05. Collectively, these results indicate that n-3 PUFA suppress Th1/Th17 cells and inflammatory macrophage subsets and reconfigure the inflammatory gene expression profile in diverse tissue sites in obese mice following the induction of colitis.

  6. Retinal vessel diameters decrease with macular ganglion cell layer thickness in autosomal dominant optic atrophy and in healthy subjects

    DEFF Research Database (Denmark)

    Rönnbäck, Cecilia; Grønskov, Karen; Larsen, Michael

    2014-01-01

    diameters (central retinal artery equivalent, CRAE, and central retinal vein equivalent, CRVE). Statistical analysis was corrected for age, gender, spherical equivalent refraction, axial length and mean arterial blood pressure (MABP) in a mixed model analysis. RESULTS: Retinal arteries and veins were...... ganglion cell-inner plexiform layer (GC-IPL) thickness (p = 0.0017 and p = 0.0057, respectively). CONCLUSION: Narrow retinal arteries and veins were associated not only with the severity of ADOA but with ganglion cell volume in patients with ADOA and in healthy subjects. This suggests that narrow vessels...

  7. Decrease of Functional Activated T and B Cells and Treatment of Glomerulonephitis in Lupus-Prone Mice Using a Natural Flavonoid Astilbin.

    Directory of Open Access Journals (Sweden)

    Lele Guo

    Full Text Available Treatment of systemic lupus erythematosus (SLE, a chronic inflammatory disease, involves the long-term use of immunosuppressive agents with significant side effects. New therapeutic approaches are being explored to find better treatment possibilities. In this study, age-matched female MRL/lpr mice were treated orally with a natural flavonoid astilbin. Astilbin administration started either at week 8 or week 12 of age though week 20. In the early treatment regimen, the treatment with astilbin reduced splenomegaly/lymphomegaly, autoantibody production and ameliorated lupus nephritis. Several serum cytokines were significantly decreased upon treatment including IFN-g, IL-17A, IL-1b, TNF-a and IL-6. Both spleen CD44 hi CD62L lo activated T cells and CD138+B220- plasma cells greatly declined. Furthermore, astilbin treatment resulted in decreased mitochondrial membrane potential in activated T cells and downregulated expression of the co-stimulatory molecules CD80 and CD86 on LPS stimulated B cells. Similar but less profound effectiveness was observed in the mice with established disease in the late treatment regimen. These results indicate that the natural product astilbin can mitigate disease development in lupus-prone mice by decreasing functional activated T and B cells.

  8. Growth arrest and decrease of alpha-SMA and type I collagen expression by palmitic acid in the rat hepatic stellate cell line PAV-1.

    Science.gov (United States)

    Abergel, Armand; Sapin, Vincent; Dif, Nicolas; Chassard, Christophe; Darcha, Claude; Marcand-Sauvant, Julie; Gaillard-Martinie, Brigitte; Rock, Edmond; Dechelotte, Pierre; Sauvant, Patrick

    2006-05-01

    Liver fibrosis is characterized by an activation of hepatic stellate cells (HSC). During primary culture HSC evolve from a quiescent into an activated phenotype which is characterized by alpha-smooth muscle actin (alpha-SMA) up-regulation, increase in cell growth, and extracellular matrix secretion. HSC culture with trans-resveratrol can lead to deactivation of myofibroblast-like HSC. We used an HSC line, PAV-1, to check the role of retinol and palmitic acid in the deactivation process of HSC. Using mass and metabolic-based methods, Western blot and immunocytochemistry assays, we demonstrated that treatment with palmitic acid (75 muM) alone or in combination with retinol (2 muM) significantly decreased cell proliferation and alpha-SMA expression. We also established that the association of both compounds strongly decreased collagen type I expression. Our results suggest the potential use of palmitic acid alone or in combination with retinol to induce HSC deactivation.

  9. Pulsed electromagnetic fields decrease proinflammatory cytokine secretion (IL-1β and TNF-α) on human fibroblast-like cell culture.

    Science.gov (United States)

    Gómez-Ochoa, Ignacio; Gómez-Ochoa, Pablo; Gómez-Casal, Francisco; Cativiela, Encarna; Larrad-Mur, Luis

    2011-10-01

    The clinical use of pulsed electromagnetic fields (PEMF) in osteoarticular pathology is widely extended, although the mechanisms involved are unknown. The aim of this study was to evaluate the action of a new protocol of treatment with PEMF on liquid medium cultures of fibroblast-like cells derivates of mononuclear peripheral blood cells. Fibroblast-like cells growth was obtained in liquid medium culture from mononuclear cells (MNC) of human peripheral blood. The PEMF irradiation protocol included an intensity of 2.25 mT, a frequency of 50 Hz and an application time of 15 min on days 7, 8 and 9 of cell culture. Immunophenotype was performed with specific heterologous monoclonal antibodies for each cell receptor (Vimentin, Cytokeratin, CD34, CD41, CD61 and CD68). The cytokines' production was determined in the supernatant of the culture medium by means of the Luminex technology. The immunophenotype did not show any statistical difference on comparing treated against non-treated cell cultures on any of the days. In the treatment cell population, the proinflammatory cytokines, IL-1β and TNF-α showed a significant decrease on days 14 and 21 of the culture, whilst IL-10 increased significantly on day 21. It is concluded that PEMF irradiation does not alter the cell immunophenotype of the fibroblast-like cell population, but does provoke a decrease in the production of inflammatory-type cytokines (IL-1β, TNF-α) and an increase in cytokines of lymphocytic origin (IL-10). These facts coincide with the chronology of the clinical effect undergone by patients with osteoarticular pathology after PEMF irradiation.

  10. Cytochrome P4502E1 inhibitor, chlormethiazole, decreases lipopolysaccharide-induced inflammation in rat Kupffer cells with ethanol treatment

    Science.gov (United States)

    To investigate the role of Cytochrome P4502E1 in sensitizing Kupffer cells to lipopolysaccharide (LPS)-mediated inflammation after ethanol induction. Sprague-Dawley rats were fed a liquid ethanol diet, control diet or ethanol diet supplemented with CYP2E1 inhibitor, chlormethiazole (CMZ), for 4'week...

  11. Decreased helenalin-induced cytotoxicity by flavonoids from Arnica as studied in a human lung carcinoma cell line

    NARCIS (Netherlands)

    Woerdenbag, HJ; Merfort, [No Value; Schmidt, TJ; Passreiter, CM; Willuhn, G; vanUden, W; Pras, N; Konings, AWT

    1995-01-01

    The effect of the flavones apigenin, luteolin, hispidulin and eupafolin, and of the flavonols kaempferol, quercetin, 6-methoxykaempferol and patuletin from Amica spp, on the cytotoxicity of the sesquiterpene lactone helenalin was studied in the human lung carcinoma cell line GLC(4) using the

  12. Rapid decrease in hepatitis C viremia by direct acting antivirals improves the natural killer cell response to IFNα.

    Science.gov (United States)

    Serti, Elisavet; Park, Heiyoung; Keane, Meghan; O'Keefe, Ashley C; Rivera, Elenita; Liang, T Jake; Ghany, Marc; Rehermann, Barbara

    2017-04-01

    Chronic HCV infection is characterised by innate immune activation with increased interferon-stimulated genes (ISG) expression and by an altered phenotype of interferon-responsive natural killer (NK) cells. Here, we asked whether a rapid reduction in viremia by daclatasvir (DCV) and asunaprevir (ASV) improves the response to pegylated interferon (PegIFN) in patients who had previously failed a standard course of PegIFN/ribavirin (RBV) therapy. Twenty-two HCV-infected non-responders to previous PegIFN/RBV therapy were studied for IFN-responsiveness of NK cells during quadruple (QUAD) therapy with DCV, ASV, PegIFN and RBV. A direct comparison of early NK cell responses in PegIFN/RBV therapy and QUAD therapy was performed for seven patients using paired cryopreserved peripheral blood mononuclear cells (PBMC) from both treatment courses. As a validation cohort, nine DCV/ASV-treated patients were studied for their NK cell response to in vitro stimulation with IFNα. The 24 h virological response to QUAD therapy correlated with an increase in signal transducer and activator of transcription 1 (STAT1), phosphorylated STAT1 (pSTAT1) and tumour necrosis factor-related apoptosis-inducing ligand (TRAIL) expression in NK cells, and the STAT1/pSTAT1/TRAIL induction was greater during QUAD therapy than during previous PegIFN/RBV therapy. Successful QUAD therapy as well as successful IFN-free DCV/ASV regimen resulted in an improved functional NK cell response (degranulation and TRAIL expression) to in vitro stimulation with IFNα. IFN-responsiveness can be improved by inhibiting HCV replication and reducing the HCV-induced activation of the innate immune response. This may provide a rationale for clinical trials of a brief period of direct acting antiviral therapy followed by PegIFN/RBV therapy to reduce the overall treatment costs in low-income and middle-income countries. NCT01888900 and NCT00718172. Published by the BMJ Publishing Group Limited. For permission to use (where

  13. A microRNA-mediated decrease in eukaryotic initiation factor 2α promotes cell survival during PS-341 treatment.

    Science.gov (United States)

    Jiang, Lili; Zang, Dan; Yi, Songgang; Li, Xiaofen; Yang, Changshan; Dong, Xiaoxian; Zhao, Chong; Lan, Xiaoying; Chen, Xin; Liu, Shouting; Liu, Ningning; Huang, Hongbiao; Shi, Xianping; Wang, Xuejun; Liu, Jinbao

    2016-02-22

    MicroRNAs (miRs) play pivotal roles in carcinogenesis and endoplasmic reticulum (ER) that performs the folding, modification and trafficking of proteins targeted to the secretory pathway. Cancer cells often endure ER stress during tumor progression but use the adaptive ER stress response to gain survival advantage. Here we report: (i) A group of miRs, including miR-30b-5p and miR-30c-5p, are upregulated by proteasome inhibitor PS-341 treatment, in HepG2 and MDA-MB-453 cells. (ii) Two representative PS-341-induced miRs: miR-30b-5p and miR-30c-5p are found to promote cell proliferation and anti-apoptosis in both tumor cells. (iii) eIF2α is confirmed as the congenerous target of miR-30b-5p and miR-30c-5p, essential to the anti-apoptotic function of these miRs. (iv) Upregulation of miR-30b-5p or miR-30c-5p, which occurs latter than the increase of phosphorylated eIF2α (p-eIF2α) in the cell under ER stress, suppresses the p-eIF2α/ATF4/CHOP pro-apoptotic pathway. (v) Inhibition of the miR-30b-5p or miR-30c-5p sensitizes the cancer cells to the cytotoxicity of proteasome inhibition. In conclusion, we unravels a new miRs-based mechanism that helps maintain intracellular proteostasis and promote cell survival during ER stress through upregulation of miR-30b-5p and miR-30c-5p which target eIF2α and thereby inhibit the p-eIF2α/ATF4/CHOP pro-apoptotic pathway, identifying miR-30b-5p and miR-30c-5p as potentially new targets for anti-cancer therapies.

  14. Comparison of Monoamine Oxidase Inhibitors in Decreasing Production of the Autotoxic Dopamine Metabolite 3,4-Dihydroxyphenylacetaldehyde in PC12 Cells

    Science.gov (United States)

    Jinsmaa, Yunden; Sullivan, Patti; Holmes, Courtney; Kopin, Irwin J.; Sharabi, Yehonatan

    2016-01-01

    According to the catecholaldehyde hypothesis, the toxic dopamine metabolite 3,4-dihydroxyphenylacetaldehyde (DOPAL) contributes to the loss of nigrostriatal dopaminergic neurons in Parkinson’s disease. Monoamine oxidase-A (MAO-A) catalyzes the conversion of intraneuronal dopamine to DOPAL and may serve as a therapeutic target. The “cheese effect”—paroxysmal hypertension evoked by tyramine-containing foodstuffs—limits clinical use of irreversible MAO-A inhibitors. Combined MAO-A/B inhibition decreases DOPAL production in rat pheochromocytoma PC12 cells, but whether reversible MAO-A inhibitors or MAO-B inhibitors decrease endogenous DOPAL production is unknown. We compared the potencies of MAO inhibitors in attenuating DOPAL production and examined possible secondary effects on dopamine storage, constitutive release, synthesis, and auto-oxidation. Catechol concentrations were measured in cells and medium after incubation with the irreversible MAO-A inhibitor clorgyline, three reversible MAO-A inhibitors, or the MAO-B inhibitors selegiline or rasagiline for 180 minutes. Reversible MAO-A inhibitors were generally ineffective, whereas clorgyline (1 nM), rasagiline (500 nM), and selegiline (500 nM) decreased DOPAL levels in the cells and medium. All three drugs also increased dopamine and norepinephrine, decreased 3,4-dihydroxyphenylalanine, and increased cysteinyl-dopamine concentrations in the medium, suggesting increased vesicular uptake and constitutive release, decreased dopamine synthesis, and increased dopamine spontaneous oxidation. In conclusion, clorgyline, rasagiline, and selegiline decrease production of endogenous DOPAL. At relatively high concentrations, the latter drugs probably lose their selectivity for MAO-B. Possibly offsetting increased formation of potentially toxic oxidation products and decreased formation of DOPAL might account for the failure of large clinical trials of MAO-B inhibitors to demonstrate slowing of neurodegeneration in

  15. Methamphetamine decreases CD4 T cell frequency and alters pro-inflammatory cytokine production in a model of drug abuse

    Science.gov (United States)

    Mata, Mariana M.; Napier, T. Celeste; Graves, Steven M.; Mahmood, Fareeha; Raeisi, Shohreh; Baum, Linda L.

    2015-01-01

    The reason co-morbid methamphetamine use and HIV infection lead to more rapid progression to AIDS is unclear. We used a model of methamphetamine self-administration to measure the effect of methamphetamine on the systemic immune system to better understand the comorbidity of methamphetamine and HIV. Catheters were implanted into the jugular veins of male, Sprague Dawley rats so they could self-administer methamphetamine (n = 18) or be given saline (control; n = 16) for 14 days. One day after the last self-administration session, blood and spleens were collected. We measured serum levels of pro-inflammatory cytokines, intracellular IFN-γand TNF-α, and frequencies of CD4+, CD8+, CD200+ and CD11b/c+ lymphocytes in the spleen. Rats that self-administer methamphetamine had a lower frequency of CD4+ T cells, but more of these cells produced IFN-γ. Methamphetamine did not alter the frequency of TNF-α-producing CD4+ T cells. Methamphetamine using rats had a higher frequency of CD8+ T cells, but fewer of them produced TNF-α. CD11b/c and CD200 expression were unchanged. Serum cytokine levels of IFN-γ, TNF-α and IL-6 in methamphetamine rats were unchanged. Methamphetamine lifetime dose inversely correlated with serum TNF-α levels. Or data suggest that methamphetamine abuse may exacerbate HIV disease progression by activating CD4 T cells, making them more susceptible to HIV infection, and contributing to their premature demise. Methamphetamine may also increase susceptibility to HIV infection, explaining why African American men who have sex with men (MSM) and frequently use methamphetamine are at the highest risk of HIV infection. PMID:25678251

  16. Methamphetamine decreases CD4 T cell frequency and alters pro-inflammatory cytokine production in a model of drug abuse.

    Science.gov (United States)

    Mata, Mariana M; Napier, T Celeste; Graves, Steven M; Mahmood, Fareeha; Raeisi, Shohreh; Baum, Linda L

    2015-04-05

    The reason co-morbid methamphetamine use and HIV infection lead to more rapid progression to AIDS is unclear. We used a model of methamphetamine self-administration to measure the effect of methamphetamine on the systemic immune system to better understand the co-morbidity of methamphetamine and HIV. Catheters were implanted into the jugular veins of male, Sprague Dawley rats so they could self-administer methamphetamine (n=18) or be given saline (control; n=16) for 14 days. One day after the last operant session, blood and spleens were collected. We measured serum levels of pro-inflammatory cytokines, intracellular IFN-γ and TNF-α, and frequencies of CD4(+), CD8(+), CD200(+) and CD11b/c(+) lymphocytes in the spleen. Rats that self-administered methamphetamine had a lower frequency of CD4(+) T cells, but more of these cells produced IFN-γ. Methamphetamine did not alter the frequency of TNF-α-producing CD4(+) T cells. Methamphetamine using rats had a higher frequency of CD8(+) T cells, but fewer of them produced TNF-α. CD11b/c and CD200 expression were unchanged. Serum cytokine levels of IFN-γ, TNF-α and IL-6 in methamphetamine rats were unchanged. Methamphetamine lifetime dose inversely correlated with serum TNF-α levels. Our data suggest that methamphetamine abuse may exacerbate HIV disease progression by activating CD4 T cells, making them more susceptible to HIV infection, and contributing to their premature demise. Methamphetamine may also increase susceptibility to HIV infection, explaining why men who have sex with men (MSM) and frequently use methamphetamine are at the highest risk of HIV infection. Copyright © 2015 Elsevier B.V. All rights reserved.

  17. Cuprizone decreases intermediate and late-stage progenitor cells in hippocampal neurogenesis of rats in a framework of 28-day oral dose toxicity study

    Energy Technology Data Exchange (ETDEWEB)

    Abe, Hajime; Tanaka, Takeshi; Kimura, Masayuki; Mizukami, Sayaka [Laboratory of Veterinary Pathology, Tokyo University of Agriculture and Technology, 3-5-8 Saiwai-cho, Fuchu-shi, Tokyo 183-8509 (Japan); Pathogenetic Veterinary Science, United Graduate School of Veterinary Sciences, Gifu University, 1-1 Yanagido, Gifu-shi, Gifu 501-1193 (Japan); Saito, Fumiyo; Imatanaka, Nobuya; Akahori, Yumi [Chemicals Evaluation and Research Institute, Japan, 1-4-25 Koraku, Bunkyo-ku, Tokyo 112-0004 (Japan); Yoshida, Toshinori [Laboratory of Veterinary Pathology, Tokyo University of Agriculture and Technology, 3-5-8 Saiwai-cho, Fuchu-shi, Tokyo 183-8509 (Japan); Shibutani, Makoto, E-mail: mshibuta@cc.tuat.ac.jp [Laboratory of Veterinary Pathology, Tokyo University of Agriculture and Technology, 3-5-8 Saiwai-cho, Fuchu-shi, Tokyo 183-8509 (Japan)

    2015-09-15

    Developmental exposure to cuprizone (CPZ), a demyelinating agent, impairs intermediate-stage neurogenesis in the hippocampal dentate gyrus of rat offspring. To investigate the possibility of alterations in adult neurogenesis following postpubertal exposure to CPZ in a framework of general toxicity studies, CPZ was orally administered to 5-week-old male rats at 0, 120, or 600 mg/kg body weight/day for 28 days. In the subgranular zone (SGZ), 600 mg/kg CPZ increased the number of cleaved caspase-3{sup +} apoptotic cells. At ≥ 120 mg/kg, the number of SGZ cells immunoreactive for TBR2, doublecortin, or PCNA was decreased, while that for SOX2 was increased. In the granule cell layer, CPZ at ≥ 120 mg/kg decreased the number of postmitotic granule cells immunoreactive for NEUN, CHRNA7, ARC or FOS. In the dentate hilus, CPZ at ≥ 120 mg/kg decreased phosphorylated TRKB{sup +} interneurons, although the number of reelin{sup +} interneurons was unchanged. At 600 mg/kg, mRNA levels of Bdnf and Chrna7 were decreased, while those of Casp4, Casp12 and Trib3 were increased in the dentate gyrus. These data suggest that CPZ in a scheme of 28-day toxicity study causes endoplasmic reticulum stress-mediated apoptosis of granule cell lineages, resulting in aberrations of intermediate neurogenesis and late-stage neurogenesis and following suppression of immediate early gene-mediated neuronal plasticity. Suppression of BDNF signals to interneurons caused by decreased cholinergic signaling may play a role in these effects of CPZ. The effects of postpubertal CPZ on neurogenesis were similar to those observed with developmental exposure, except for the lack of reelin response, which may contribute to a greater decrease in SGZ cells. - Highlights: • Effect of 28-day CPZ exposure on hippocampal neurogenesis was examined in rats. • CPZ suppressed intermediate neurogenesis and late-stage neurogenesis in the dentate gyrus. • CPZ suppressed BDNF signals to interneurons by decrease of

  18. Cuprizone decreases intermediate and late-stage progenitor cells in hippocampal neurogenesis of rats in a framework of 28-day oral dose toxicity study

    International Nuclear Information System (INIS)

    Abe, Hajime; Tanaka, Takeshi; Kimura, Masayuki; Mizukami, Sayaka; Saito, Fumiyo; Imatanaka, Nobuya; Akahori, Yumi; Yoshida, Toshinori; Shibutani, Makoto

    2015-01-01

    Developmental exposure to cuprizone (CPZ), a demyelinating agent, impairs intermediate-stage neurogenesis in the hippocampal dentate gyrus of rat offspring. To investigate the possibility of alterations in adult neurogenesis following postpubertal exposure to CPZ in a framework of general toxicity studies, CPZ was orally administered to 5-week-old male rats at 0, 120, or 600 mg/kg body weight/day for 28 days. In the subgranular zone (SGZ), 600 mg/kg CPZ increased the number of cleaved caspase-3 + apoptotic cells. At ≥ 120 mg/kg, the number of SGZ cells immunoreactive for TBR2, doublecortin, or PCNA was decreased, while that for SOX2 was increased. In the granule cell layer, CPZ at ≥ 120 mg/kg decreased the number of postmitotic granule cells immunoreactive for NEUN, CHRNA7, ARC or FOS. In the dentate hilus, CPZ at ≥ 120 mg/kg decreased phosphorylated TRKB + interneurons, although the number of reelin + interneurons was unchanged. At 600 mg/kg, mRNA levels of Bdnf and Chrna7 were decreased, while those of Casp4, Casp12 and Trib3 were increased in the dentate gyrus. These data suggest that CPZ in a scheme of 28-day toxicity study causes endoplasmic reticulum stress-mediated apoptosis of granule cell lineages, resulting in aberrations of intermediate neurogenesis and late-stage neurogenesis and following suppression of immediate early gene-mediated neuronal plasticity. Suppression of BDNF signals to interneurons caused by decreased cholinergic signaling may play a role in these effects of CPZ. The effects of postpubertal CPZ on neurogenesis were similar to those observed with developmental exposure, except for the lack of reelin response, which may contribute to a greater decrease in SGZ cells. - Highlights: • Effect of 28-day CPZ exposure on hippocampal neurogenesis was examined in rats. • CPZ suppressed intermediate neurogenesis and late-stage neurogenesis in the dentate gyrus. • CPZ suppressed BDNF signals to interneurons by decrease of cholinergic

  19. Global Decrease of Histone H3K27 Acetylation in ZEB1-Induced Epithelial to Mesenchymal Transition in Lung Cancer Cells

    International Nuclear Information System (INIS)

    Roche, Joëlle; Nasarre, Patrick; Gemmill, Robert; Baldys, Aleksander; Pontis, Julien; Korch, Christopher; Guilhot, Joëlle; Ait-Si-Ali, Slimane; Drabkin, Harry

    2013-01-01

    The epithelial to mesenchymal transition (EMT) enables epithelial cells with a migratory mesenchymal phenotype. It is activated in cancer cells and is involved in invasion, metastasis and stem-like properties. ZEB1, an E-box binding transcription factor, is a major suppressor of epithelial genes in lung cancer. In the present study, we show that in H358 non-small cell lung cancer cells, ZEB1 downregulates EpCAM (coding for an epithelial cell adhesion molecule), ESRP1 (epithelial splicing regulatory protein), ST14 (a membrane associated serine protease involved in HGF processing) and RAB25 (a small G-protein) by direct binding to these genes. Following ZEB1 induction, acetylation of histone H4 and histone H3 on lysine 9 (H3K9) and 27 (H3K27) was decreased on ZEB1 binding sites on these genes as demonstrated by chromatin immunoprecipitation. Of note, decreased H3K27 acetylation could be also detected by western blot and immunocytochemistry in ZEB1 induced cells. In lung cancers, H3K27 acetylation level was higher in the tumor compartment than in the corresponding stroma where ZEB1 was more often expressed. Since HDAC and DNA methylation inhibitors increased expression of ZEB1 target genes, targeting these epigenetic modifications would be expected to reduce metastasis

  20. Global Decrease of Histone H3K27 Acetylation in ZEB1-Induced Epithelial to Mesenchymal Transition in Lung Cancer Cells

    Energy Technology Data Exchange (ETDEWEB)

    Roche, Joëlle, E-mail: joelle.roche@univ-poitiers.fr [Department of Medicine, Hematology Oncology Division, MUSC, 96 Jonathan Lucas St., Charleston, SC 29425 (United States); CNRS FRE 3511, University of Poitiers, 1 rue Georges Bonnet, F-86022 Poitiers Cédex (France); Nasarre, Patrick; Gemmill, Robert [Department of Medicine, Hematology Oncology Division, MUSC, 96 Jonathan Lucas St., Charleston, SC 29425 (United States); Baldys, Aleksander [Department of Medicine, Nephrology Division, MUSC, Ralph H. Johnson Veterans Affairs Medical Center, Charleston, SC 29425 (United States); Pontis, Julien [Epigénétique & Destin Cellulaire, CNRS UMR 7216, University of Paris Diderot, Sorbonne Paris Cité, F-75013 Paris (France); Korch, Christopher [CU DNA Sequencing and Analysis Core, University of Colorado, School of Medicine, Anschutz Medical Campus, 12801 E. 17th Ave., Aurora, CO 80045 (United States); Guilhot, Joëlle [INSERM, CIC 0802, CHU de Poitiers, F-86021 (France); Ait-Si-Ali, Slimane [Epigénétique & Destin Cellulaire, CNRS UMR 7216, University of Paris Diderot, Sorbonne Paris Cité, F-75013 Paris (France); Drabkin, Harry [Department of Medicine, Hematology Oncology Division, MUSC, 96 Jonathan Lucas St., Charleston, SC 29425 (United States)

    2013-04-03

    The epithelial to mesenchymal transition (EMT) enables epithelial cells with a migratory mesenchymal phenotype. It is activated in cancer cells and is involved in invasion, metastasis and stem-like properties. ZEB1, an E-box binding transcription factor, is a major suppressor of epithelial genes in lung cancer. In the present study, we show that in H358 non-small cell lung cancer cells, ZEB1 downregulates EpCAM (coding for an epithelial cell adhesion molecule), ESRP1 (epithelial splicing regulatory protein), ST14 (a membrane associated serine protease involved in HGF processing) and RAB25 (a small G-protein) by direct binding to these genes. Following ZEB1 induction, acetylation of histone H4 and histone H3 on lysine 9 (H3K9) and 27 (H3K27) was decreased on ZEB1 binding sites on these genes as demonstrated by chromatin immunoprecipitation. Of note, decreased H3K27 acetylation could be also detected by western blot and immunocytochemistry in ZEB1 induced cells. In lung cancers, H3K27 acetylation level was higher in the tumor compartment than in the corresponding stroma where ZEB1 was more often expressed. Since HDAC and DNA methylation inhibitors increased expression of ZEB1 target genes, targeting these epigenetic modifications would be expected to reduce metastasis.

  1. Granulocyte colony-stimulating factor decreases the Th1/Th2 ratio in peripheral blood mononuclear cells from patients with chronic immune thrombocytopenic purpura in vitro.

    Science.gov (United States)

    Ge, Fei; Zhang, Zhuo; Hou, Jinxiao; Cao, Fenglin; Zhang, Yingmei; Wang, Ping; Wei, Hong; Zhou, Jin

    2016-12-01

    Chronic immune thrombocytopenia purpura (ITP) is an autoimmune disease that exhibits an abnormally high Th1/Th2 ratio. Granulocyte colony-stimulating factor (G-CSF) has been shown to decrease the Th1/Th2 ratio in healthy donors. In this study, we investigated the effects of G-CSF treatment on the Th1/Th2 cells and the underlying mechanisms in patients with ITP in vitro. Peripheral blood mononuclear cells (PBMCs) isolated from patients with ITP and healthy controls were treated with G-CSF. Expression levels of interferon (IFN)-γ, interleukin (IL)-2, IL-4, and IL-13 in supernatants were measured by enzyme-linked immunosorbent assays. The expression of IFN-γ, IL-4, and G-CSF receptor (G-CSFR) on Th1 and Th2 cells were examined by flow cytometry and confocal microscopy. The mRNA expression of IFN-γ, IL-2, IL-4, IL-13, and T-box expressed in T cells (T-bet) and GATA-binding protein 3 (GATA-3) in PBMCs was evaluated by reverse transcription polymerase chain reaction. The results showed that G-CSF could significantly reduce the Th1/Th2 ratio in PBMCs from patients with ITP in vitro. As the concentration of G-CSF increased, Th1/Th2 ([IFN-γ+IL-2]/[IL-4+IL-13]) cytokine ratios and T-bet/GATA-3 mRNA ratios decreased in a concentration-dependent manner. Th1 cells and Th2 cells both expressed G-CSFR. These results suggest that G-CSF could decrease the Th1/Th2 ratio in the context of ITP, and elucidate the direct and indirect immunomodulatory mechanisms underlying G-CSF functions in Th1/Th2 cells, thus supporting the therapeutic potential of G-CSF in the treatment of patients with ITP. Copyright © 2016 Elsevier Ltd. All rights reserved.

  2. MiR-133b is frequently decreased in gastric cancer and its overexpression reduces the metastatic potential of gastric cancer cells

    International Nuclear Information System (INIS)

    Zhao, Yu; Zhu, Zhenggang; Huang, Jie; Zhang, Li; Qu, Ying; Li, Jianfang; Yu, Beiqin; Yan, Min; Yu, Yingyan; Liu, Bingya

    2014-01-01

    Emerging evidence has shown that microRNAs are involved in gastric cancer development and progression. Here we examine the role of miR-133b in gastric cancer. Quantitative real-time PCR analysis was performed in 140 patient gastric cancer tissues and 8 gastric cancer cell lines. The effects of miR-133b in gastric cancer cells metastasis were examined by scratch assay, transwell migration and matrigel invasion. In vivo effects of miR-133b were examined in an intraperitoneal mouse tumor model. Targets of miR-133b were predicted by bioinformatics tools and validated by luciferase reporter analyses, western blot, and quantitative real-time PCR. MiR-133b was significantly downregulated in 70% (98/140) of gastric cancer patients. Expression of miR-133b was negatively correlated with lymph node metastasis of gastric cancer in patients. Similarly, the expression of miR-133b was significantly lower in seven tested gastric cancer cell lines than in the immortalized non-cancerous GES-1 gastric epithelial cells. Overexpression of miR-133b markedly inhibited metastasis of gastric cancer cells in vitro and in vivo. Moreover, the transcriptional factor Gli1 was identified as a direct target for miR-133b. Level of Gli1 protein but not mRNA was decreased by miR-133b. Activity of luciferase with Gli1 3′-untranslated region was markedly decreased by miR-133b in gastric cancer cells. Gli1 target genes, OPN and Zeb2, were also inhibited by miR133b. MiR-133b is frequently decreased in gastric cancer. Overexpression of miR-133b inhibits cell metastasis in vitro and in vivo partly by directly suppressing expression of Gli1 protein. These results suggested that miR-133b plays an important role in gastric cancer metastasis

  3. Prolactin receptor attenuation induces zinc pool redistribution through ZnT2 and decreases invasion in MDA-MB-453 breast cancer cells

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    Bostanci, Zeynep, E-mail: zbostanci@hmc.psu.edu [The Pennsylvania State University, Department of Nutritional Sciences, 209 Chandlee Lab, University Park, PA 16802 (United States); The Pennsylvania State University Milton S. Hershey Medical Center, Department of Surgery, 500 University Dr., Hershey, PA 17033 (United States); Alam, Samina, E-mail: sra116@psu.edu [The Pennsylvania State University, Department of Nutritional Sciences, 209 Chandlee Lab, University Park, PA 16802 (United States); The Pennsylvania State University Milton S. Hershey Medical Center, Department of Surgery, 500 University Dr., Hershey, PA 17033 (United States); Soybel, David I., E-mail: dsoybel@hmc.psu.edu [The Pennsylvania State University, Department of Nutritional Sciences, 209 Chandlee Lab, University Park, PA 16802 (United States); The Pennsylvania State University Milton S. Hershey Medical Center, Department of Surgery, 500 University Dr., Hershey, PA 17033 (United States); The Pennsylvania State University College of Medicine, Department of Cell and Molecular Physiology, 500 University Dr., Hershey, PA 17033 (United States); Kelleher, Shannon L., E-mail: slk39@psu.edu [The Pennsylvania State University, Department of Nutritional Sciences, 209 Chandlee Lab, University Park, PA 16802 (United States); The Pennsylvania State University Milton S. Hershey Medical Center, Department of Surgery, 500 University Dr., Hershey, PA 17033 (United States); The Pennsylvania State University College of Medicine, Department of Cell and Molecular Physiology, 500 University Dr., Hershey, PA 17033 (United States)

    2014-02-15

    Prolactin receptor (PRL-R) activation regulates cell differentiation, proliferation, cell survival and motility of breast cells. Prolactin (PRL) and PRL-R over-expression are strongly implicated in breast cancer, particularly contributing to tumor growth and invasion in the more aggressive estrogen-receptor negative (ER−) disease. PRL-R antagonists have been suggested as potential therapeutic agents; however, mechanisms through which PRL-R antagonists exert their actions are not well-understood. Zinc (Zn) is a regulatory factor for over 10% of the proteome, regulating critical cell processes such as proliferation, cell signaling, transcription, apoptosis and autophagy. PRL-R signaling regulates Zn metabolism in breast cells. Herein we determined effects of PRL-R attenuation on cellular Zn metabolism and cell function in a model of ER-, PRL-R over-expressing breast cancer cells (MDA-MB-453). PRL-R attenuation post-transcriptionally increased ZnT2 abundance and redistributed intracellular Zn pools into lysosomes and mitochondria. ZnT2-mediated lysosomal Zn sequestration was associated with reduced matrix metalloproteinase 2 (MMP-2) activity and decreased invasion. ZnT2-mediated Zn accumulation in mitochondria was associated with increased mitochondrial oxidation. Our results suggest that PRL-R antagonism in PRL-R over-expressing breast cancer cells may reduce invasion through the redistribution of intracellular Zn pools critical for cellular function. - Highlights: • PRL-R attenuation increased ZnT2 expression. • PRL-R attenuation increased lysosomal and mitochondrial Zn accumulation. • PRL-R attenuation decreased MMP-2 and invasion. • PRL-R antagonists may modulate lysosomal and mitochondrial Zn pools.

  4. Treatment with a Small Molecule Mutant IDH1 Inhibitor Suppresses Tumorigenic Activity and Decreases Production of the Oncometabolite 2-Hydroxyglutarate in Human Chondrosarcoma Cells.

    Directory of Open Access Journals (Sweden)

    Luyuan Li

    Full Text Available Chondrosarcomas are malignant bone tumors that produce cartilaginous matrix. Mutations in isocitrate dehydrogenase enzymes (IDH1/2 were recently described in several cancers including chondrosarcomas. The IDH1 inhibitor AGI-5198 abrogates the ability of mutant IDH1 to produce the oncometabolite D-2 hydroxyglutarate (D-2HG in gliomas. We sought to determine if treatment with AGI-5198 would similarly inhibit tumorigenic activity and D-2HG production in IDH1-mutant human chondrosarcoma cells. Two human chondrosarcoma cell lines, JJ012 and HT1080 with endogenous IDH1 mutations and a human chondrocyte cell line C28 with wild type IDH1 were employed in our study. Mutation analysis of IDH was performed by PCR-based DNA sequencing, and D-2HG was detected using tandem mass spectrometry. We confirmed that JJ012 and HT1080 harbor IDH1 R132G and R132C mutation, respectively, while C28 has no mutation. D-2HG was detectable in cell pellets and media of JJ012 and HT1080 cells, as well as plasma and urine from an IDH-mutant chondrosarcoma patient, which decreased after tumor resection. AGI-5198 treatment decreased D-2HG levels in JJ012 and HT1080 cells in a dose-dependent manner, and dramatically inhibited colony formation and migration, interrupted cell cycling, and induced apoptosis. In conclusion, our study demonstrates anti-tumor activity of a mutant IDH1 inhibitor in human chondrosarcoma cell lines, and suggests that D-2HG is a potential biomarker for IDH mutations in chondrosarcoma cells. Thus, clinical trials of mutant IDH inhibitors are warranted for patients with IDH-mutant chondrosarcomas.

  5. tmRNA decreases the bactericidal activity of aminoglycosides and the susceptibility to inhibitors of cell wall synthesis.

    Science.gov (United States)

    Luidalepp, Hannes; Hallier, Marc; Felden, Brice; Tenson, Tanel

    2005-04-01

    Trans-translation is a process that recycles ribosomes stalled on problematic mRNAs. tmRNA, coded by the DeltassrA gene, is a major component of trans-translation. Bacteria lacking tmRNA are more sensitive to several inhibitors of protein synthesis when compared to a wild type strain. We measured bacterial growth of the DeltassrA and wild type strains in Escherichia coli in the presence of 14 antibiotics including some that do not target protein synthesis. Both the optical density of the bacterial cultures and the number of viable cells were monitored. For the ribosome-targeted antibiotics, sensitization was observed on erythromycin, chloramphenicol, kanamycin, puromycin and streptomycin. Minor or no effects were observed with clindamycin, tetracycline and spectinomycin. Surprisingly, the DeltassrA strain is more sensitive than wild type to inhibitors of cell wall synthesis: fosfomycin and ampicillin. No growth difference was observed on drugs with other target sites: ofloxacin, norfloxacin, rifampicin and trimethoprim. Sensitization to antibiotics having target sites other than the ribosome suggests that trans-translation could influence antibiotic-induced stress responses. In trans-translation-deficient bacteria, cell death is significantly enhanced by the two aminoglycosides that induce translational misreading, streptomycin and kanamycin.

  6. Purple carrot (Daucus carota L.) polyacetylenes decrease lipopolysaccharide-induced expression of inflammatory proteins in macrophage and endothelial cells.

    Science.gov (United States)

    Metzger, Brandon T; Barnes, David M; Reed, Jess D

    2008-05-28

    Carrots ( Daucus carota L.) contain phytochemicals including carotenoids, phenolics, polyacetylenes, isocoumarins, and sesquiterpenes. Purple carrots also contain anthocyanins. The anti-inflammatory activity of extracts and phytochemicals from purple carrots was investigated by determining attenuation of the response to lipopolysaccharide (LPS). A bioactive chromatographic fraction (Sephadex LH-20) reduced LPS inflammatory response. There was a dose-dependent reduction in nitric oxide production and mRNA of pro-inflammatory cytokines (IL-6, IL-1beta, TNF-alpha) and iNOS in macrophage cells. Protein secretions of IL-6 and TNF-alpha were reduced 77 and 66% in porcine aortic endothelial cells treated with 6.6 and 13.3 microg/mL of the LH-20 fraction, respectively. Preparative liquid chromatography resulted in a bioactive subfraction enriched in the polyacetylene compounds falcarindiol, falcarindiol 3-acetate, and falcarinol. The polyacetylenes were isolated and reduced nitric oxide production in macrophage cells by as much as 65% without cytotoxicity. These results suggest that polyacetylenes, not anthocyanins, in purple carrots are responsible for anti-inflammatory bioactivity.

  7. Inhibition of host extracellular signal-regulated kinase (ERK) activation decreases new world alphavirus multiplication in infected cells

    International Nuclear Information System (INIS)

    Voss, Kelsey; Amaya, Moushimi; Mueller, Claudius; Roberts, Brian; Kehn-Hall, Kylene; Bailey, Charles; Petricoin, Emanuel; Narayanan, Aarthi

    2014-01-01

    New World alphaviruses belonging to the family Togaviridae are classified as emerging infectious agents and Category B select agents. Our study is focused on the role of the host extracellular signal-regulated kinase (ERK) in the infectious process of New World alphaviruses. Infection of human cells by Venezuelan equine encephalitis virus (VEEV) results in the activation of the ERK-signaling cascade. Inhibition of ERK1/2 by the small molecule inhibitor Ag-126 results in inhibition of viral multiplication. Ag-126-mediated inhibition of VEEV was due to potential effects on early and late stages of the infectious process. While expression of viral proteins was down-regulated in Ag-126 treated cells, we did not observe any influence of Ag-126 on the nuclear distribution of capsid. Finally, Ag-126 exerted a broad-spectrum inhibitory effect on New World alphavirus multiplication, thus indicating that the host kinase, ERK, is a broad-spectrum candidate for development of novel therapeutics against New World alphaviruses. - Highlights: • VEEV infection activated multiple components of the ERK signaling cascade. • Inhibition of ERK activation using Ag-126 inhibited VEEV multiplication. • Activation of ERK by Ceramide C6 increased infectious titers of TC-83. • Ag-126 inhibited virulent strains of all New World alphaviruses. • Ag-126 treatment increased percent survival of infected cells

  8. Inhibition of host extracellular signal-regulated kinase (ERK) activation decreases new world alphavirus multiplication in infected cells

    Energy Technology Data Exchange (ETDEWEB)

    Voss, Kelsey; Amaya, Moushimi [National Center for Biodefense and Infectious Diseases, School of Systems Biology, George Mason University, 10650 Pyramid Place, Manassas, VA (United States); Mueller, Claudius [Center for Applied Proteomics and Personalized Medicine, George Mason University, 10900 University Boulevard, Manassas, VA (United States); Roberts, Brian [Leidos Health Life Sciences, 5202 Presidents Court, Suite 110, Frederick, MD (United States); Kehn-Hall, Kylene; Bailey, Charles [National Center for Biodefense and Infectious Diseases, School of Systems Biology, George Mason University, 10650 Pyramid Place, Manassas, VA (United States); Petricoin, Emanuel [Center for Applied Proteomics and Personalized Medicine, George Mason University, 10900 University Boulevard, Manassas, VA (United States); Narayanan, Aarthi, E-mail: anaraya1@gmu.edu [National Center for Biodefense and Infectious Diseases, School of Systems Biology, George Mason University, 10650 Pyramid Place, Manassas, VA (United States)

    2014-11-15

    New World alphaviruses belonging to the family Togaviridae are classified as emerging infectious agents and Category B select agents. Our study is focused on the role of the host extracellular signal-regulated kinase (ERK) in the infectious process of New World alphaviruses. Infection of human cells by Venezuelan equine encephalitis virus (VEEV) results in the activation of the ERK-signaling cascade. Inhibition of ERK1/2 by the small molecule inhibitor Ag-126 results in inhibition of viral multiplication. Ag-126-mediated inhibition of VEEV was due to potential effects on early and late stages of the infectious process. While expression of viral proteins was down-regulated in Ag-126 treated cells, we did not observe any influence of Ag-126 on the nuclear distribution of capsid. Finally, Ag-126 exerted a broad-spectrum inhibitory effect on New World alphavirus multiplication, thus indicating that the host kinase, ERK, is a broad-spectrum candidate for development of novel therapeutics against New World alphaviruses. - Highlights: • VEEV infection activated multiple components of the ERK signaling cascade. • Inhibition of ERK activation using Ag-126 inhibited VEEV multiplication. • Activation of ERK by Ceramide C6 increased infectious titers of TC-83. • Ag-126 inhibited virulent strains of all New World alphaviruses. • Ag-126 treatment increased percent survival of infected cells.

  9. Body temperature during hibernation is highly correlated with a decrease in circulating innate immune cells in the brown bear (Ursus arctos): a common feature among hibernators?

    Science.gov (United States)

    Sahdo, Berolla; Evans, Alina L; Arnemo, Jon M; Fröbert, Ole; Särndahl, Eva; Blanc, Stéphane

    2013-01-01

    Hibernation involves periods of severely depressed metabolism (torpor) and decreases in body temperature (Tb). Small arctic mammals (hibernation. This raised the question of whether the decreased leukocyte counts in mammalian hibernators is due to torpor per se or is secondary to low Tb. The present study examined immune cell counts in brown bears (Ursus arctos), where torpor is only associated with shallow decreases in Tb. The results were compared across hibernator species for which immune and Tb data were available. The white blood cell counts were determined by flow cytometry in 13 bears captured in the field both during summer and winter over 2 years time. Tb dropped from 39.6±0.8 to 33.5±1.1°C during hibernation. Blood neutrophils and monocytes were lower during hibernation than during the active period (47%, p= 0.001; 43%, p=0.039, respectively), whereas no change in lymphocyte counts was detected (p=0.599). Further, combining our data and those from 10 studies on 9 hibernating species suggested that the decline in Tb explained the decrease in innate immune cells (R(2)=0.83, phibernation, which may represent a suppressed innate immune system. Across species comparison suggests that, both in small and large hibernators, Tb is the main driver of immune function regulation during winter dormancy. The lack of a difference in lymphocyte counts in this context requires further investigations.

  10. Apoptosis is increased and cell proliferation is decreased in out-of-phase endometria from infertile and recurrent abortion patients

    Directory of Open Access Journals (Sweden)

    Irigoyen Marcela

    2010-10-01

    Full Text Available Abstract Background Various endometrial abnormalities have been associated with luteal phase deficiency: a significant dyssynchrony in the maturation of the glandular epithelium and the stroma and a prevalence of out-of-phase endometrial biopsy specimens. Out-of phase endometrium is a controversial disorder related to failed implantation, infertility and early pregnancy loss. Given that the regulation of the apoptotic process in endometrium of luteal phase deficiency is still unknown, the aim of this study was to evaluate cell proliferation, apoptosis and the levels of the main effector caspase, caspase-3 in the luteal in-phase and out-of-phase endometrium. Methods Thirty-seven endometrial samples from sterile or recurrent abortion patients were included in this study: 21 in-phase samples (controls and 16 samples with out-of-phase endometrium. Biopsy specimens of eutopic endometrium were obtained from all subjects during days 21-25 of the menstrual cycle. The endometrium with endometrial maturity of cycle day 25 or less at the time of menstruation was considered out-of phase. Endometrial tissues were fixed in 10% buffered formaldehyde. For apoptosis quantification, sections were processed for in situ immunohistochemical localization of nuclei exhibiting DNA fragmentation, by the terminal deoxynucleotidyl transferase (TdT-mediated dUTP digoxygenin nick-end labeling (TUNEL technique. Expressions of Proliferating Cell Nuclear Antigen (PCNA as a marker of cell proliferation, and of cleaved caspase-3 as a marker of apoptosis, were assessed by immunohistochemistry in the luteal in-phase and out-of-phase endometrium from infertile and recurrent abortion patients. Results Luteal out-of-phase endometrium had increased apoptosis levels compared to in-phase endometrium (p Conclusions this study represents the first report describing variations at the cell proliferation and cell death levels in the out-of-phase endometrium in comparison with in

  11. Inhibition of Cholesterol Synthesis in HepG2 Cells by GINST-Decreasing HMG-CoA Reductase Expression Via AMP-Activated Protein Kinase.

    Science.gov (United States)

    Han, Joon-Seung; Sung, Jong Hwan; Lee, Seung Kwon

    2017-11-01

    GINST, a hydrolyzed ginseng extract, has been reported to have antidiabetic effects and to reduce hyperglycemia and hyperlipidemia. Hypercholesterolemia is caused by diet or genetic factors and can lead to atherosclerosis and coronary heart disease. Thus, the purpose of this study is to determine whether GINST and the ginsenoside metabolite, IH-901 (compound K), reduce cholesterol synthesis in HepG2 cells and the signal transduction pathways involved. Concentrations of cholesterol were measured by using an enzymatic method. Expression levels of sterol regulatory element-binding protein 2 (SREBP2), HMG-CoA reductase (HMGCR), peroxisome proliferators-activated receptor γ (PPARγ), CCAAT/enhancer-binding proteins α (C/EBPα), GAPDH, and phosphorylation of AMP-activated protein kinase α (AMPKα), protein kinase B (PKB, also known as Akt), and mechanistic target of rapamycin complex 1 (mTORC1) were measured using western blot. Total cholesterol concentration decreased after GINST treatment for 24 and 48 h. Expression of HMGCR decreased more with GINST than with the inhibitors, U18666A and atorvastatin, after 48 h in a dose-dependent manner. Phosphorylation of AMPKα increased 2.5x by GINST after 360 min of treatment, and phosphorylation of Akt decreased after 120 and 360 min. We separated compound K from GINST extracts flash chromatography. Compound K decreased cholesterol synthesis in HepG2 cells at 24 and 48 h. Therefore, we conclude that GINST inhibits cholesterol synthesis in HepG2 cells by decreasing HMGCR expression via AMPKα activation. GINST, a hydrolyzed ginseng extract, can inhibit cholesterol synthesis in liver cells via activation of AMPKα. IH-901 (compound K), which is the main component with bioactivity in GINST, also has anticholesterol effects. Thus, we suggest that GINST can be used to reduce hypercholesterolemia. © 2017 Institute of Food Technologists®.

  12. Processing of Kansui Roots Stir-Baked with Vinegar Reduces Kansui-Induced Hepatocyte Cytotoxicity by Decreasing the Contents of Toxic Terpenoids and Regulating the Cell Apoptosis Pathway

    Directory of Open Access Journals (Sweden)

    Xiaojing Yan

    2014-06-01

    Full Text Available Euphorbia kansui is a Traditional Chinese Medicine widely used for the treatment of oedema, ascites and asthma. However, its serious hepatotoxicity hinders its safe clinical application. The process of stir-baking with vinegar is regularly used to reduce the toxicity of kansui. Up till now, the exact mechanism of the reduction in hepatotoxicity of kansui stir-baked with vinegar has been poorly defined. In this study, decreased  contents of five diterpene and one triterpene in kansui (GS-1 after stir-baking with vinegar (GS-2 was investigated by UPLC-QTOF/MS. Flow cytometry and Hoechst staining were used to show that the stir-baking with vinegar process reduces kansui-induced cell apoptosis. Furthermore, the result also indicated that kansui stir-baked with vinegar protects LO2 cells from apoptosis by increasing the cell mitochondrial membrane potential (ΔΨm, decreasing the release of cytochrome c and inhibiting the activities of caspase-9 and caspase-3 as evidenced by means of high content screening (HCS, ELISA and western blotting. These results suggested that the stir-baking vinegar could reduce the hepatotoxicity of kansui by effectively decreasing the contents of toxic terpenoids and inhibiting the intrinsic pathway of hepatocyte cell apoptosis. In conclusion, the study provided significant data for promoting safer and better clinical use of this herb.

  13. Decreased Expression of T-Cell Costimulatory Molecule CD28 on CD4 and CD8 T Cells of Mexican Patients with Pulmonary Tuberculosis

    Directory of Open Access Journals (Sweden)

    German Bernal-Fernandez

    2010-01-01

    Full Text Available Patients with tuberculosis frequently develop anergy, a state of T-cell hyporesponsiveness in which defective T-cell costimulation could be a factor. To know if the expression of T-cell costimulatory molecules was altered in tuberculosis, we analyzed the peripheral blood T-cell phenotype of 23 Mexican patients with pulmonary tuberculosis. There was severe CD4 (P<.001 and CD8 (P<.01 lymphopenia and upregulation of costimulatory molecule CD30 on CD4 and CD8 T cells (P<.05; this increase was higher in relapsing tuberculosis. The main finding was severe downregulation of the major costimulatory molecule CD28 on both CD8 and CD4 T cells (P<.001. Depletion of the CD4/CD28 subset, a hitherto undescribed finding, is relevant because CD4 T cells constitute the main arm of the cell-mediated antimycobacterial immune response.

  14. SIRT1 overexpression decreases cisplatin-induced acetylation of NF-{kappa}B p65 subunit and cytotoxicity in renal proximal tubule cells

    Energy Technology Data Exchange (ETDEWEB)

    Jung, Yu Jin; Lee, Jung Eun; Lee, Ae Sin [Department of Internal Medicine, Chonbuk National University Medical School, Jeonju (Korea, Republic of); Kang, Kyung Pyo; Lee, Sik; Park, Sung Kwang [Department of Internal Medicine, Chonbuk National University Medical School, Jeonju (Korea, Republic of); Institute for Medical Sciences, Chonbuk National University Medical School, Jeonju (Korea, Republic of); Lee, Sang Yong [Department of Diagnostic Radiology, Chonbuk National University Medical School, Jeonju (Korea, Republic of); Institute for Medical Sciences, Chonbuk National University Medical School, Jeonju (Korea, Republic of); Han, Myung Kwan [Department of Microbiology, Institute for Medical Sciences, Chonbuk National University Medical School, Jeonju (Korea, Republic of); Kim, Duk Hoon [Division of Forensic Medicine, National Forensic Service, Seoul (Korea, Republic of); Kim, Won, E-mail: kwon@jbnu.ac.kr [Department of Internal Medicine, Chonbuk National University Medical School, Jeonju (Korea, Republic of); Institute for Medical Sciences, Chonbuk National University Medical School, Jeonju (Korea, Republic of)

    2012-03-09

    Highlights: Black-Right-Pointing-Pointer Cisplatin increases acetylation of NF-{kappa}B p65 subunit in HK2 cells. Black-Right-Pointing-Pointer SIRT1 overexpression decreases cisplatin-induced p65 acetylation and -cytotoxicity. Black-Right-Pointing-Pointer Resveratrol decreased cisplatin-induced cell viability through deacetylation of p65. -- Abstract: As the increased acetylation of p65 is linked to nuclear factor-{kappa}B (NF-{kappa}B) activation, the regulation of p65 acetylation can be a potential target for the treatment of inflammatory injury. Cisplatin-induced nephrotoxicity is an important issue in chemotherapy of cancer patients. SIRT1, nicotinamide adenine dinucleotide (NAD{sup +})-dependent protein deacetylase, has been implicated in a variety of cellular processes such as inflammatory injury and the control of multidrug resistance in cancer. However, there is no report on the effect of SIRT1 overexpression on cisplatin-induced acetylation of p65 subunit of NF-{kappa}B and cell injury. To investigate the effect of SIRT1 in on cisplatin-induced acetylation of p65 subunit of NF-{kappa}B and cell injury, HK2 cells were exposed with SIRT1 overexpression, LacZ adenovirus or dominant negative adenovirus after treatment with cisplatin. While protein expression of SIRT1 was decreased by cisplatin treatment compared with control buffer treatment, acetylation of NF-{kappa}B p65 subunit was significantly increased after treatment with cisplatin. Overexpression of SIRT1 ameliorated the increased acetylation of p65 of NF-{kappa}B during cisplatin treatment and cisplatin-induced cytotoxicity. Further, treatment of cisplatin-treated HK2 cells with resveratrol, a SIRT1 activator, also decreased acetylation of NF-{kappa}B p65 subunit and cisplatin-induced increase of the cell viability in HK2 cells. Our findings suggests that the regulation of acetylation of p65 of NF-{kappa}B through SIRT1 can be a possible target to attenuate cisplatin-induced renal cell damage.

  15. Baicalein induces G1 arrest in oral cancer cells by enhancing the degradation of cyclin D1 and activating AhR to decrease Rb phosphorylation

    Energy Technology Data Exchange (ETDEWEB)

    Cheng, Ya-Hsin, E-mail: yhcheng@mail.cmu.edu.tw [Department of Physiology, School of Medicine, China Medical University, Taichung 40402, Taiwan, ROC (China); Li, Lih-Ann; Lin, Pinpin; Cheng, Li-Chuan [Division of Environmental Health and Occupational Medicine, National Health Research Institutes, Zhunan, Miaoli 35053, Taiwan, ROC (China); Hung, Chein-Hui [Graduate Institute of Clinical Medicine Sciences, Chang Gung University, Puizi City, Chiayi 613, Taiwan, ROC (China); Chang, Nai Wen [Department of Biochemistry, School of Medicine, China Medical University, Taichung, Taiwan, ROC (China); Lin, Chingju [Department of Physiology, School of Medicine, China Medical University, Taichung 40402, Taiwan, ROC (China)

    2012-09-15

    Baicalein is a flavonoid, known to have anti-inflammatory and anti-cancer effects. As an aryl hydrocarbon receptor (AhR) ligand, baicalein at high concentrations blocks AhR-mediated dioxin toxicity. Because AhR had been reported to play a role in regulating the cell cycle, we suspected that the anti-cancer effect of baicalein is associated with AhR. This study investigated the molecular mechanism involved in the anti-cancer effect of baicalein in oral cancer cells HSC-3, including whether such effect would be AhR-mediated. Results revealed that baicalein inhibited cell proliferation and increased AhR activity in a dose-dependent manner. Cell cycle was arrested at the G1 phase and the expression of CDK4, cyclin D1, and phosphorylated retinoblastoma (pRb) was decreased. When the AhR was suppressed by siRNA, the reduction of pRb was partially reversed, accompanied by a decrease of cell population at G1 phase and an increase at S phase, while the reduction of cyclin D1 and CDK4 did not change. This finding suggests that the baicalein activation of AhR is indeed associated with the reduction of pRb, but is independent of the reduction of cyclin D1 and CDK4. When cells were pre-treated with LiCl, the inhibitor of GSK-3β, the decrease of cyclin D1 was blocked and the reduction of pRb was recovered. The data indicates that in HSC-3 the reduction of pRb is both mediated by baicalein through activation of AhR and facilitation of cyclin D1 degradation, which causes cell cycle arrest at the G1 phase, and results in the inhibition of cell proliferation. -- Highlights: ► Baicalein causes the G1 phase arrest by decreasing Rb phosphorylation. ► Baicalein modulates AhR-mediated cell proliferation. ► Both AhR activation and cyclin D1 degradation results in hypophosphorylation of Rb. ► Baicalein facilitates cyclin D1 degradation by signalling the GSK-3β pathway.

  16. Bromodomain and extra-terminal protein mimic JQ1 decreases inflammation in human vascular endothelial cells: Implications for pulmonary arterial hypertension.

    Science.gov (United States)

    Mumby, Sharon; Gambaryan, Natalia; Meng, Chao; Perros, Frederic; Humbert, Marc; Wort, S John; Adcock, Ian M

    2017-01-01

    Nuclear factor kappa B (NF-kB)-mediated inflammatory gene expression and vascular endothelial cell proliferation/remodelling are implicated in the pathophysiology of the fatal disease, pulmonary arterial hypertension (PAH). Bromodomain and extra-terminal (BET) proteins are essential for the expression of a subset of NF-kB-induced inflammatory genes. BET mimics including JQ1+ prevent binding of BETs to acetylated histones and down-regulate the expression of selected genes. The effects of JQ1+ on the proliferation of primary human pulmonary microvascular endothelial cells (HPMECs) from healthy subjects were measured by bromodeoxyuridine (BrdU) incorporation. Cell cycle progression was assessed by flow cytometry; mRNA and protein levels of cyclin-dependent kinases (CDKs), inhibitors and cytokines were determined by reverse transcription-quantitative PCR (RT-qPCR), Western blotting or ELISA. Histone acetyltransferase (HAT) and deacetylase (HDAC) activities were determined in nuclear extracts from whole lung of PAH and control patients. JQ1+ significantly inhibited IL6 and IL8 (IL6 and CXCL8) mRNA and protein in HPMECs compared with its inactive enantiomer JQ1-. JQ1+ decreased NF-kB p65 recruitment to native IL6 and IL8 promoters. JQ1+ showed a concentration-dependent decrease in HPMEC proliferation compared with JQ1--treated cells. JQ1+ induced G1 cell cycle arrest by increasing the expression of the CDK inhibitors (CDKN) 1A (p21 cip ) and CDKN2D (p19 INK4D ) and decreasing that of CDK2, CDK4 and CDK6. JQ1+ also inhibited serum-stimulated migration of HPMECs. Finally, HAT activity was significantly increased in the lung of PAH patients. Inhibition of BETs in primary HPMECs decreases inflammation and remodelling. BET proteins could be a target for future therapies for PAH. © 2016 The Authors. Respirology published by John Wiley & Sons Australia, Ltd on behalf of Asian Pacific Society of Respirology.

  17. Expression of a fungal ferulic acid esterase in suspension cultures of tall fescue (Festuca arundinacea) decreases cell wall feruloylation and increases rates of cell wall digestion.

    Science.gov (United States)

    Morris, Phillip; Dalton, Sue; Langdon, Tim; Hauck, Barbara; de Buanafina, Marcia M O

    2017-01-01

    In the cell walls of grasses ferulic acid is esterified to arabinosyl residues in arabinoxylans that can then undergo oxidative coupling reactions to form ferulate dehydrodimers, trimers and oligomers which function to cross-link cell-wall polysaccharides, limiting cell wall degradability. Fungal ferulic acid esterase can release both esterified monomeric and dimeric ferulic acids from these cell wall arabinoxylans making the cell wall more susceptible to further enzymatic attack and increasing cell wall degradability. Non-embryogenic cell suspension cultures of Festuca arundinacea expressing a Aspergillus niger ferulic acid esterase ( faeA ) targeted to either the apoplast, or endoplasmic reticulum under the control of a constitutive actin promoter, or to the vacuole under the control of a soybean heat shock promoter, were established and FAE activity determined in the cells and medium during a growth cycle. Analysis of the ester-linked ferulates of the cell walls showed that all three transformed cell lines had both reduced ferulate levels and increased levels of xylanase mediated release of wall phenolics on autodigestion as well as increased rates of cell wall digestion in a simulated rumen environment, when compared to control non-transformed cells.

  18. Suppression of Glut1 and Glucose Metabolism by Decreased Akt/mTORC1 Signaling Drives T Cell Impairment in B Cell Leukemia

    NARCIS (Netherlands)

    Siska, Peter J.; van der Windt, Gerritje J. W.; Kishton, Rigel J.; Cohen, Sivan; Eisner, William; MacIver, Nancie J.; Kater, Arnon P.; Weinberg, J. Brice; Rathmell, Jeffrey C.

    2016-01-01

    Leukemia can promote T cell dysfunction and exhaustion that contributes to increased susceptibility to infection and mortality. The treatment-independent mechanisms that mediate leukemia-associated T cell impairments are poorly understood, but metabolism tightly regulates T cell function and may

  19. Glucocorticoid receptor knockdown decreases the antioxidant protection of B16 melanoma cells: an endocrine system-related mechanism that compromises metastatic cell resistance to vascular endothelium-induced tumor cytotoxicity.

    Science.gov (United States)

    Obrador, Elena; Valles, Soraya L; Benlloch, María; Sirerol, J Antoni; Pellicer, José A; Alcácer, Javier; Coronado, Javier Alcácer-F; Estrela, José M

    2014-01-01

    We previously reported an interorgan system in which stress-related hormones (corticosterone and noradrenaline), interleukin-6, and glutathione (GSH) coordinately regulate metastatic growth of highly aggressive B16-F10 melanoma cells. Corticosterone, at levels measured in tumor-bearing mice, also induces apoptotic cell death in metastatic cells with low GSH content. In the present study we explored the potential role of glucocorticoids in the regulation of metastatic cell death/survival during the early stages of organ invasion. Glucocorticoid receptor (GCR) knockdown decreased the expression and activity of γ-glutamylcysteine synthetase (γ-GCS), the rate-limiting step in GSH synthesis, in metastatic cells in vivo independent of the tumor location (liver, lung, or subcutaneous). The decrease in γ-GCS activity was associated with lower intracellular GSH levels. Nrf2- and p53-dependent down-regulation of γ-GCS was associated with a decrease in the activities of superoxide dismutase 1 and 2, catalase, glutathione peroxidase, and glutathione reductase, but not of the O2--generating NADPH oxidase. The GCR knockdown-induced decrease in antioxidant protection caused a drastic decrease in the survival of metastatic cells during their interaction with endothelial cells, both in vitro and in vivo; only 10% of cancer cells attached to the endothelium survived compared to 90% survival observed in the controls. This very low rate of metastatic cell survival was partially increased (up to 52%) in vivo by inoculating B16-F10 cells preloaded with GSH ester, which enters the cell and delivers free GSH. Taken together, our results indicate that glucocorticoid signaling influences the survival of metastatic cells during their interaction with the vascular endothelium.

  20. Decreased 5-Hydroxymethylcytosine (5-hmC) predicts poor prognosis in early-stage laryngeal squamous cell carcinoma

    OpenAIRE

    Zhang, Yanfang; Wu, Kexia; Shao, Yuan; Sui, Fang; Yang, Qi; Shi, Bingyin; Hou, Peng; Ji, Meiju

    2016-01-01

    Accumulating evidences suggest that large-scale loss of 5-hydroxymethylcytosine (5-hmC) is an epigenetic hallmark in different cancers. However, the levels of 5-hmC in laryngeal squamous cell carcinoma (LSCC) and its prognostic value in this cancer remain largely unknown. Using dot-blot and quantitative RT-PCR assays, we investigate 5-hmC levels and expression of TET-1, -2 and -3 in LSCCs and explore the association of 5-hmC levels with clinicopathological characteristics and clinical outcome...

  1. Over-expression of CD8+ T-cell activation is associated with decreased CD4+ cells in patients seeking treatment of Alcohol Use Disorder.

    Science.gov (United States)

    Zuluaga, Paola; Sanvisens, Arantza; Martínez-Cáceres, Eva; Teniente, Aina; Tor, Jordi; Muga, Robert

    2017-11-01

    Harmful alcohol consumption may have an impact on the adaptive immune system through an imbalance in T cell subpopulations and changes in cell activation. We aimed to analyze profiles of CD4 and CD8T cell activation in patients with alcohol use disorder (AUD). We used a cross-sectional study with patients seeking treatment of the disorder. Blood samples for immunophenotyping were obtained at admission. Profiles of T cell activation were defined: (I) CD38 + /HLA-DR + , (II) CD38 + /HLA-DR - , (III) CD38 - /HLA-DR + , (IV) CD38 - /HLA-DR - and compared with healthy controls. We calculated a CD8 + T cell activation indicator (AI) that was defined as the quotient of non-activated cells (CD38 - /HLA-DR - ) and activated cells (CD38 + /HLA-DR + ). 60 patients were eligible (83%M); median age was 49 years [IQR: 44-54] and alcohol consumption was 145g/day [IQR: 90-205]. Mean±SD of CD38 + /HLA-DR - was 50.3±50.6 cells/μL in patients and 33.5±24.5 cells/μL in controls (p=0.03), for the CD38 - /HLA-DR + it was 61±62.2 cells/μL in patients and 21.2±17.3 cells/μL in controls (pcells/μL in patients and 10.8±10.3 cells/μL in controls (pcells, and the percentage of CD38 + /HLA-DR + CD8 + T cells (r=0.37, p=0.003; r=0.2, p=0.086, respectively). Patients with AUD have an increased expression of immune activation with respect to healthy individuals. This excess of activated CD8 + T cells correlates with the absolute CD4 + T cells. Copyright © 2017 Elsevier B.V. All rights reserved.

  2. Decreased nuclear stiffness via FAK-ERK1/2 signaling is necessary for osteopontin-promoted migration of bone marrow-derived mesenchymal stem cells

    International Nuclear Information System (INIS)

    Liu, Lingling; Luo, Qing; Sun, Jinghui; Wang, Aoli; Shi, Yisong; Ju, Yang; Morita, Yasuyuki; Song, Guanbin

    2017-01-01

    Migration of bone marrow-derived mesenchymal stem cells (BMSCs) plays an important role in many physiological and pathological settings, including wound healing. During the migration of BMSCs through interstitial tissues, the movement of the nucleus must be coordinated with the cytoskeletal dynamics, which in turn affects the cell migration efficiency. Our previous study indicated that osteopontin (OPN) significantly promotes the migration of rat BMSCs. However, the nuclear behaviors and involved molecular mechanisms in OPN-mediated BMSC migration are largely unclear. In the present study, using an atomic force microscope (AFM), we found that OPN could decrease the nuclear stiffness of BMSCs and reduce the expression of lamin A/C, which is the main determinant of nuclear stiffness. Increased lamin A/C expression attenuates BMSC migration by increasing nuclear stiffness. Decreased lamin A/C expression promotes BMSC migration by decreasing nuclear stiffness. Furthermore, OPN promotes BMSC migration by diminishing lamin A/C expression and decreasing nuclear stiffness via the FAK-ERK1/2 signaling pathway. This study provides strong evidence for the role of nuclear mechanics in BMSC migration as well as new insight into the molecular mechanisms of OPN-promoted BMSC migration. - Highlights: • OPN promotes BMSC migration by decreasing nuclear stiffness. • Lamin A/C knockdown decreases, while its overexpression enhances, the nuclear stiffness of BMSCs. • Lamin A/C overexpression and downregulation affect the migration of BMSCs. • OPN diminishes lamin A/C expression and decreases nuclear stiffness through the activation of the FAK-ERK1/2 signaling pathway. • OPN promotes BMSC migration via the FAK-ERK1/2 signaling pathway.

  3. Decreased nuclear stiffness via FAK-ERK1/2 signaling is necessary for osteopontin-promoted migration of bone marrow-derived mesenchymal stem cells

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Lingling, E-mail: liulingling2012@163.com [Key Laboratory of Biorheological Science and Technology, Ministry of Education, College of Bioengineering, Chongqing University, Chongqing 400044 (China); Luo, Qing, E-mail: qing.luo@cqu.edu.cn [Key Laboratory of Biorheological Science and Technology, Ministry of Education, College of Bioengineering, Chongqing University, Chongqing 400044 (China); Sun, Jinghui, E-mail: sunjhemail@163.com [Key Laboratory of Biorheological Science and Technology, Ministry of Education, College of Bioengineering, Chongqing University, Chongqing 400044 (China); Wang, Aoli, E-mail: leaf13332@163.com [Key Laboratory of Biorheological Science and Technology, Ministry of Education, College of Bioengineering, Chongqing University, Chongqing 400044 (China); Shi, Yisong, E-mail: shiyis@cqu.edu.cn [Key Laboratory of Biorheological Science and Technology, Ministry of Education, College of Bioengineering, Chongqing University, Chongqing 400044 (China); Ju, Yang, E-mail: ju@mech.nagoya-u.ac.jp [Department of Mechanical Science and Engineering, Nagoya University, Nagoya 464-8603 (Japan); Morita, Yasuyuki, E-mail: morita@mech.nagoya-u.ac.jp [Department of Mechanical Science and Engineering, Nagoya University, Nagoya 464-8603 (Japan); Song, Guanbin, E-mail: song@cqu.edu.cn [Key Laboratory of Biorheological Science and Technology, Ministry of Education, College of Bioengineering, Chongqing University, Chongqing 400044 (China)

    2017-06-15

    Migration of bone marrow-derived mesenchymal stem cells (BMSCs) plays an important role in many physiological and pathological settings, including wound healing. During the migration of BMSCs through interstitial tissues, the movement of the nucleus must be coordinated with the cytoskeletal dynamics, which in turn affects the cell migration efficiency. Our previous study indicated that osteopontin (OPN) significantly promotes the migration of rat BMSCs. However, the nuclear behaviors and involved molecular mechanisms in OPN-mediated BMSC migration are largely unclear. In the present study, using an atomic force microscope (AFM), we found that OPN could decrease the nuclear stiffness of BMSCs and reduce the expression of lamin A/C, which is the main determinant of nuclear stiffness. Increased lamin A/C expression attenuates BMSC migration by increasing nuclear stiffness. Decreased lamin A/C expression promotes BMSC migration by decreasing nuclear stiffness. Furthermore, OPN promotes BMSC migration by diminishing lamin A/C expression and decreasing nuclear stiffness via the FAK-ERK1/2 signaling pathway. This study provides strong evidence for the role of nuclear mechanics in BMSC migration as well as new insight into the molecular mechanisms of OPN-promoted BMSC migration. - Highlights: • OPN promotes BMSC migration by decreasing nuclear stiffness. • Lamin A/C knockdown decreases, while its overexpression enhances, the nuclear stiffness of BMSCs. • Lamin A/C overexpression and downregulation affect the migration of BMSCs. • OPN diminishes lamin A/C expression and decreases nuclear stiffness through the activation of the FAK-ERK1/2 signaling pathway. • OPN promotes BMSC migration via the FAK-ERK1/2 signaling pathway.

  4. Decreased expression of protein tyrosine phosphatase non-receptor type 12 is involved in the proliferation and recurrence of bladder transitional cell carcinoma

    Science.gov (United States)

    PIAO, YONGRUI; LIU, XIANKUI; LIN, ZHENHUA; JIN, ZHEHU; JIN, XUANSHUN; YUAN, KUICHANG; WU, WENYUAN

    2015-01-01

    Protein tyrosine phosphatase non-receptor type 12 (PTPN12) has been shown to be involved in the development of a number of types of carcinoma. However, the effect of PTPN12 on the proliferation and recurrence of human bladder transitional cell carcinoma (TCC) is unclear. The present study aimed to investigate the expression and function of PTPN12 in human TCC. Samples from 164 patients with TCC, in addition to 146 patients undergoing bladder surgery for indications other than TCC, were examined. PTPN12 protein expression was examined using immunohistochemistry and western blotting, and PTPN12 mRNA expression was examined using reverse transcription-quantitative polymerase chain reaction. PTPN12 expression was increased following transfection with the PTPN12-expressing, pcDEF3 vector, and PTPN12 expression was decreased by RNA interference, in four TCC cell lines. The proliferation of TCC cells was analyzed by a WST-1 assay and in xenografts on BALB/C nude mice. The effect of PTPN12 on tumor recurrence was analyzed by adhesion, migration and invasion assays in TCC cell lines. PTPN12 expression was significantly decreased in TCC tissues compared with that in normal urothelium, and the level of PTPN12 expression was negatively correlated with tumor size, pathological grade, clinical stage and tumor recurrence. Furthermore, decreased expression of PTPN12 significantly enhanced the proliferation of TCC cells in vitro and in vivo. TCC cells with lower levels of PTPN12 exhibited greater adhesion, migration and invasion. In conclusion, PTPN12 expression is downregulated in human TCC. Restoring PTPN12 activity may represent a novel therapeutic strategy for this disease. PMID:26622721

  5. Endocan, a putative endothelial cell marker, is elevated in preeclampsia, decreased in acute pyelonephritis, and unchanged in other obstetrical syndromes

    Science.gov (United States)

    Adekola, Henry; Romero, Roberto; Chaemsaithong, Piya; Korzeniewski, Steven J.; Dong, Zhong; Yeo, Lami; Hassan, Sonia S.; Chaiworapongsa, Tinnakorn

    2015-01-01

    Abstract Objective: Endocan, a dermatan sulphate proteoglycan produced by endothelial cells, is considered a biomarker for endothelial cell activation/dysfunction. Preeclampsia is characterized by systemic vascular inflammation, and endothelial cell activation/dysfunction. Therefore, the objectives of this study were to determine whether: (1) plasma endocan concentrations in preeclampsia differ from those in uncomplicated pregnancies; (2) changes in plasma endocan concentration relate to the severity of preeclampsia, and whether these changes are specific or observed in other obstetrical syndromes such as small-for-gestational age (SGA), fetal death (FD), preterm labor (PTL) or preterm prelabor rupture of membranes (PROM); (3) a correlation exists between plasma concentration of endocan and angiogenic (placental growth factor or PlGF)/anti-angiogenic factors (soluble vascular endothelial growth factor receptor or sVEGFR-1, and soluble endoglin or sEng) among pregnancies complicated by preeclampsia; and (4) plasma endocan concentrations in patients with preeclampsia and acute pyelonephritis (both conditions in which there is endothelial cell activation) differ. Method: This cross-sectional study included the following groups: (1) uncomplicated pregnancy (n = 130); (2) preeclampsia (n = 102); (3) pregnant women without preeclampsia who delivered an SGA neonate (n = 51); (4) FD (n = 49); (5) acute pyelonephritis (AP; n = 35); (6) spontaneous PTL (n = 75); and (7) preterm PROM (n = 64). Plasma endocan concentrations were determined in all groups, and PIGF, sEng and VEGFR-1 plasma concentrations were measured by ELISA in the preeclampsia group. Results: (1) Women with preeclampsia had a significantly higher median plasma endocan concentration than those with uncomplicated pregnancies (p = 0.004); (2) among women with preeclampsia, the median plasma endocan concentration did not differ significantly according to disease severity (p = 0

  6. Changes in T-cell subpopulations and cytokine network during early period of ibrutinib therapy in chronic lymphocytic leukemia patients: the significant decrease in T regulatory cells number.

    Science.gov (United States)

    Podhorecka, Monika; Goracy, Aneta; Szymczyk, Agnieszka; Kowal, Malgorzata; Ibanez, Blanca; Jankowska-Lecka, Olga; Macheta, Arkadiusz; Nowaczynska, Aleksandra; Drab-Urbanek, Elzbieta; Chocholska, Sylwia; Jawniak, Dariusz; Hus, Marek

    2017-05-23

    B cell receptor (BCR) stimulation signal plays an important role in the pathogenesis of chronic lymphocytic leukemia (CLL), and kinase inhibitors directed toward the BCR pathway are now the promising anti-leukemic drugs. Ibrutinib, a Bruton tyrosine kinase inhibitor, demonstrates promising clinical activity in CLL. It is reported that ibrutinib, additionally to directly targeting leukemic cells, also inhibits the interactions of these cells with T cells, macrophages and accessory cells. Assessment of these mechanisms is important because of their non -direct anti-leukemic effects and to identify possible side effects connected with long-term drug administration.The aim of this study was to assess the in vivo effects of ibrutinib on T-cell subpopulations and cytokine network in CLL. The analysis was performed on a group of 19 patients during first month of ibrutinib therapy. The standard multicolor flow cytometry and cytometric bead array methods were used for assessment of T-cell subsets and cytokines/chemokines, respectively.The data obtained indicates that Ibrutinib treatment results in changes in T-cell subpopulations and cytokine network in CLL patients. Particularly, a significant reduction of T regulatory cells in peripheral blood was observed. By targeting these populations of T cells Ibrutinib can stimulate rejection of tumor cells by the immune system.

  7. Glutathione export by human lymphoid cells: depletion of glutathione by inhibition of its synthesis decreases export and increases sensitivity to irradiation.

    Science.gov (United States)

    Dethmers, J K; Meister, A

    1981-12-01

    Glutathione (in the form of GSH) is transported out of cultured human lymphoid cells at rates proportional to the intracellular glutathione levels. Inhibition of glutathione synthesis by buthionine sulfoximine, a potent selective inhibitor of gamma-glutamylcysteine synthetase, leads to exponential decrease in intracellular glutathione, a large fraction of which appears extracellularly, indicating that glutathione turnover is associated with its export. Although cells with 0.09 mM glutathione (4% of controls) were 85% viable, further decrease was associated with marked loss of viability. Cells with 4-5% of control glutathione levels were much more sensitive than control cells to the effects of gamma radiation and of 5,5'-dithiobis(2-nitrobenzoate). Depletion of glutathione by use of buthionine sulfoximine has advantages over other reagents (such as diamide, other oxidizing agents, and diethylmaleate, which affect other cellular components and may increase glutathione disulfide levels) and therefore has potential usefulness in sensitizing cells to the effects of radiation and to therapeutic agents that are detoxified by reactions involving glutathione.

  8. Mitigating the Effects of Xuebijing Injection on Hematopoietic Cell Injury Induced by Total Body Irradiation with γ rays by Decreasing Reactive Oxygen Species Levels

    Directory of Open Access Journals (Sweden)

    Deguan Li

    2014-06-01

    Full Text Available Hematopoietic injury is the most common side effect of radiotherapy. However, the methods available for the mitigating of radiation injury remain limited. Xuebijing injection (XBJ is a traditional Chinese medicine used to treat sepsis in the clinic. In this study, we investigated the effects of XBJ on the survival rate in mice with hematopoietic injury induced by γ ray ionizing radiation (IR. Mice were intraperitoneally injected with XBJ daily for seven days after total body irradiation (TBI. Our results showed that XBJ (0.4 mL/kg significantly increased 30-day survival rates in mice exposed to 7.5 Gy TBI. This effect may be attributable to improved preservation of white blood cells (WBCs and hematopoietic cells, given that bone marrow (BM cells from XBJ-treated mice produced more granulocyte-macrophage colony forming units (CFU-GM than that in the 2 Gy/TBI group. XBJ also decreased the levels of reactive oxygen species (ROS by increasing glutathione (GSH and superoxide dismutase (SOD levels in serum and attenuated the increased BM cell apoptosis caused by 2 Gy/TBI. In conclusion, these findings suggest that XBJ enhances the survival rate of irradiated mice and attenuates the effects of radiation on hematopoietic injury by decreasing ROS production in BM cells, indicating that XBJ may be a promising therapeutic candidate for reducing hematopoietic radiation injury.

  9. AGE-modified basement membrane cooperates with Endo180 to promote epithelial cell invasiveness and decrease prostate cancer survival

    DEFF Research Database (Denmark)

    Rodriguez-Teja, Mercedes; Gronau, Julian H; Breit, Claudia

    2015-01-01

    in non-transformed PECs via a molecular mechanism linked to cancer progression. This study provides a rationale for targeting CTLD2 in Endo180 in prostate cancer and other pathologies in which increased basal lamina thickness and tissue stiffness are driving factors. © 2014 The Authors. The Journal......Biomechanical strain imposed by age-related thickening of the basal lamina and augmented tissue stiffness in the prostate gland coincides with increased cancer risk. Here we hypothesized that the structural alterations in the basal lamina associated with age can induce mechanotransduction pathways...... in prostate epithelial cells (PECs) to promote invasiveness and cancer progression. To demonstrate this, we developed a 3D model of PEC acini in which thickening and stiffening of basal lamina matrix was induced by advanced glycation end-product (AGE)-dependent non-enzymatic crosslinking of its major...

  10. Exogenous administration of gangliosides inhibits FcepsilonRI-mediated mast cell degranulation by decreasing the activity of phospholipase Cgamma

    Czech Academy of Sciences Publication Activity Database

    Dráberová, Lubica; Dudková, L.; Boubelík, Michael; Tolarová, Helena; Šmíd, F.; Dráber, Petr

    2003-01-01

    Roč. 171, č. 7 (2003), s. 3585-3593 ISSN 0022-1767 R&D Projects: GA MŠk LN00A026; GA ČR GA204/00/0204; GA ČR GA204/03/0594; GA ČR GA310/00/0205; GA ČR GA301/03/0596; GA AV ČR IAA5052005; GA AV ČR IAA7052006; GA AV ČR IAA5052310; GA MZd NB6758 Institutional research plan: CEZ:AV0Z5052915 Keywords : mast cell * plasma membrane * IgE Subject RIV: EC - Immunology Impact factor: 6.702, year: 2003

  11. Long none coding RNA HOTTIP/HOXA13 act as synergistic role by decreasing cell migration and proliferation in Hirschsprung disease

    Energy Technology Data Exchange (ETDEWEB)

    Xie, Hua; Zhu, Dongmei; Xu, Cao; Zhu, Hairong; Chen, Pingfa; Li, Hongxing [Department of Pediatric Surgery, State Key Laboratory of Reproductive Medicine, Nanjing Children' s Hospital Affiliated Nanjing Medical University, Nanjing 210008 (China); Key Laboratory of Modern Toxicology (Nanjing Medical University), Institute of Toxicology, School of Public Health, Nanjing Medical University, Ministry of Education, Nanjing 211166 (China); Liu, Xiang [Department of Pediatric Surgery, Anhui Provincial Children' s Hospital, Anhui 230000 (China); Xia, Yankai [Department of Pediatric Surgery, State Key Laboratory of Reproductive Medicine, Nanjing Children' s Hospital Affiliated Nanjing Medical University, Nanjing 210008 (China); Key Laboratory of Modern Toxicology (Nanjing Medical University), Institute of Toxicology, School of Public Health, Nanjing Medical University, Ministry of Education, Nanjing 211166 (China); Tang, Weibing, E-mail: twbcn@njmu.com [Department of Pediatric Surgery, State Key Laboratory of Reproductive Medicine, Nanjing Children' s Hospital Affiliated Nanjing Medical University, Nanjing 210008 (China); Key Laboratory of Modern Toxicology (Nanjing Medical University), Institute of Toxicology, School of Public Health, Nanjing Medical University, Ministry of Education, Nanjing 211166 (China)

    2015-08-07

    Long noncoding RNAs (lncRNAs) have been confirmed to be associated with various human diseases. However, whether they are associated with Hirschsprung disease (HSCR) progression remains unclear. In this study, we designed the experiment to explore the relationship between lncRNA HOTTIP and HOXA13, and their pathogenicity to HSCR. Quantitative real-time PCR and Western blot were performed to detect the levels of lncRNA, mRNAs, and proteins in colon tissues from 79 patients with HSCR and 79 controls. Small RNA interference transfection was used to study the function experiments in human 293T and SK-N-BE cell lines. The cell viability and activities were detected by the transwell assays, CCK8 assay, and flow cytometry, respectively. LncRNA HOTTIP and HOXA13 were significantly down-regulated in HSCR compared to the controls. Meanwhile, the declined extent of their expression levels makes sense between two main phenotype of HSCR. SiRNA-mediated knock-down of HOTTIP or HOXA13 correlated with decreased levels of each other and both reduced the cell migration and proliferation without affecting cell apoptosis or cell cycle. Our study demonstrates that aberrant reduction of HOTTIP and HOXA13, which have a bidirectional regulatory loop, may play an important role in the pathogenesis of HSCR. - Highlights: • LncRNA HOTTIP and HOXA13 are both down-regulated in HSCR. • HOTTIP and HOXA13 can regulate each other in 293T and SK-N-BE(2) cell lines. • Both HOTTIP and HOXA13 can decrease cell migration and proliferation.

  12. Arecoline inhibits interleukin-2 secretion in Jurkat cells by decreasing the expression of alpha7-nicotinic acetylcholine receptors and prostaglandin E2.

    Science.gov (United States)

    Hwang, G S; Hu, S; Lin, Y H; Chen, S T; Tang, T K; Wang, P S; Wang, S W

    2013-10-01

    The purpose of the present study was to explore the effect of arecoline on phytohemagglutinin (PHA)-stimulated interleukin-2 (IL-2) secretion, the expression of alpha7-nicotinic acetylcholine receptors (α7-nAChRs), prostaglandin E2(PGE2) protein, and IL-2 mRNA in human lymphocyte cells (Jurkat cell line). The IL-2 and PGE2 were determined by enzyme-linked immunosorbent assay (ELISA). The expressions of phosphorylated extracellular signal-regulated kinase (ERK) and α7-nAChRs were determined by Western blotting. The level of IL-2 mRNA was determined by reverse-transcriptase polymerase chain reaction (RT-PCR). Arecoline, in a dose-dependent manner, significantly decreased IL-2 and PGE2 secretion by Jurkat cells incubated with 0 or 5 μg/ml 5 μg/ml PHA. PGE2 also significantly inhibited IL-2 secretion by Jurkat cells in a dose-dependent manner. In addition, reduced expression of PHA-induced ERK phosphorylation was observed in Jurkat cells treated with arecoline. PHA-enhanced IL-2 mRNA expression was also inhibited by arecoline. These results imply that arecoline inhibits the release of PGE2 and PHA-induced IL-2 secretion by Jurkat cells and that these effects seem to occur, at least in part, either through the attenuation of ERK in conjunction with a decrease of PHA-induced IL-2 mRNA expression. These results imply that arecoline inhibits the protein expression of α7-nAChRs , the release of PGE2 and PHA-induced IL-2 secretion by Jurkat cells.

  13. Processing of stored packed red blood cells using autotransfusion devices decreases potassium and microaggregates: a prospective, randomized, single-blinded in vitro study.

    Science.gov (United States)

    Westphal-Varghese, B; Erren, M; Westphal, M; Van Aken, H; Ertmer, C; Lange, M; Booke, M

    2007-04-01

    The aim of the study was to compare the potential of autotransfusion devices to reduce non-infectious complications related to transfusion of long-stored packed red blood cells (PRBC; n= 57), such as changes in electrolytes, blood cells and the load of free microaggregates. Following a baseline measurement, a blood pool of three PRBC was divided into three equal volumes and washed with either the Haemonetics Cell Saver (HCS) or the continuous autotransfusion system (C.A.T.S), using the quality (CATS(quality)) and emergency (CATS(emergency)) mode. After the washing procedure, measurements for electrolytes, blood cells and free microaggregates were repeated (n= 19 each). Compared with baseline, the investigated autotransfusion devices reduced the median load of potassium (baseline: 52 mEq L(-1); HCS: 4 mEq L(-1); CATS(quality): 4 mEq L(-1); CATS(emergency): 17 mEq L(-1); each P glucose and protein. Whereas the quantity of microaggregates was not reduced by HCS, CATS(emergency) decreased the load of cell fragments below 7.8 microm (P CATS(quality) decreased the load of cell fragments not only to a diameter below 7.8 microm (P < 0.001 vs. baseline) but also of microaggregates between 7.8 and 17.6 microm (P < 0.05 vs. baseline). In situations where long-stored PRBC have to be transfused, the procedure described here may be feasible to reduce clinically relevant side effects, i.e. hyperkalaemia and microvascular obstruction secondary to free cell fragments. This approach could be especially useful in patients undergoing massive transfusion and/or suffering from renal failure.

  14. Triptolide, a diterpenoid triepoxide, induces antitumor proliferation via activation of c-Jun NH2-terminal kinase 1 by decreasing phosphatidylinositol 3-kinase activity in human tumor cells

    International Nuclear Information System (INIS)

    Miyata, Yoshiki; Sato, Takashi; Ito, Akira

    2005-01-01

    Triptolide, a diterpenoid triepoxide extracted from the Chinese herb Tripterygium wilfordii Hook f., exerts antitumorigenic actions against several tumor cells, but the intracellular target signal molecule(s) for this antitumorigenesis activity of triptolide remains to be identified. In the present study, we demonstrated that triptolide, in a dose-dependent manner, inhibited the proliferation of human fibrosarcoma HT-1080, human squamous carcinoma SAS, and human uterine cervical carcinoma SKG-II cells. In addition, triptolide was found to decrease phosphatidylinositol 3-kinase (PI3K) activity. A PI3K inhibitor, LY-294002, mimicked the triptolide-induced antiproliferative activity in HT-1080, SAS, and SKG-II cells. There was no change in the activity of Akt or protein kinase C (PKC), both of which are downstream effectors in the PI3K pathway. Furthermore, the phosphorylation of Ras, Raf, and mitogen-activated protein/extracellular signal-regulated kinase 1/2 was not modified in HT-1080 cells treated with triptolide. However, the phosphorylation of c-Jun NH 2 -terminal kinase 1 (JNK1) was found to increase in both triptolide- and LY-294002-treated cells. Furthermore, the triptolide-induced inhibition of HT-1080 cell proliferation was not observed by JNK1 siRNA-treatment. These results provide novel evidence that PI3K is a crucial target molecule in the antitumorigenic action of triptolide. They further suggest a possible triptolide-induced inhibitory signal for tumor cell proliferation that is initiated by the decrease in PI3K activity, which in turn leads to the augmentation of JNK1 phosphorylation via the Akt and/or PKC-independent pathway(s). Moreover, it is likely that the activation of JNK1 is required for the triptolide-induced inhibition of tumor proliferation

  15. Light-Emitting Diode (LED) therapy improves occipital cortex damage by decreasing apoptosis and increasing BDNF-expressing cells in methanol-induced toxicity in rats.

    Science.gov (United States)

    Ghanbari, Amir; Ghareghani, Majid; Zibara, Kazem; Delaviz, Hamdallah; Ebadi, Elham; Jahantab, Mohammad Hossein

    2017-05-01

    Methanol-induced retinal toxicity, frequently associated with elevated free radicals and cell edema, is characterized by progressive retinal ganglion cell (RGC) death and vision loss. Previous studies investigated the effect of photomodulation on RGCs, but not the visual cortex. In this study, the effect of 670nm Light-Emitting Diode (LED) therapy on RGCs and visual cortex recovery was investigated in a seven-day methanol-induced retinal toxicity protocol in rats. Methanol administration showed a reduction in the number of RGCs, loss of neurons (neuronal nuclear antigen, NeuN+), activation of glial fibrillary acidic protein (GFAP+) expressing cells, suppression of brain-derived neurotrophic factor (BDNF+) positive cells, increase in apoptosis (caspase 3+) and enhancement of nitric oxide (NO) release in serum and brain. On the other hand, LED therapy significantly reduced RGC death, in comparison to the methanol group. In addition, the number of BDNF positive cells was significantly higher in the visual cortex of LED-treated group, in comparison to methanol-intoxicated and control groups. Moreover, LED therapy caused a significant decrease in cell death (caspase 3+ cells) and a significant reduction in the NO levels, both in serum and brain tissue, in comparison to methanol-intoxicated rats. Overall, LED therapy demonstrated a number of beneficial effects in decreasing oxidative stress and in functional recovery of RGCs and visual cortex. Our data suggest that LED therapy could be a potential condidate as a non-invasive approach for treatment of retinal damage, which needs further clinicl studies. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

  16. Decreasing Eukaryotic Initiation Factor 3C (EIF3C) Suppresses Proliferation and Stimulates Apoptosis in Breast Cancer Cell Lines Through Mammalian Target of Rapamycin (mTOR) Pathway.

    Science.gov (United States)

    Zhao, Weipeng; Li, Xichuan; Wang, Jun; Wang, Chen; Jia, Yongsheng; Yuan, Shunzong; Huang, Yong; Shi, Yehui; Tong, Zhongsheng

    2017-08-30

    BACKGROUND Translation initiation is the rate limiting step of protein synthesis and is highly regulated. Eukaryotic initiation factor 3C (EIF3C), an oncogene overexpressed in several human cancers, plays an important role in tumorigenesis and cell proliferation. MATERIAL AND METHODS Immunohistochemistry was used to determine the expression of EIF3C in breast cancer tissues from 42 patients. We investigated whether EIF3C silencing decreases breast cancer cell proliferation as assessed by colony formation assay, and whether EIF3C gene knockdown induces apoptosis as assessed by flow cytometry analysis. We utilized the stress and apoptosis signaling antibody array kit, while p-ERK1/2, p-Akt, p-Smad2, p-p38 MAPK, cleaved caspase-3, and cleaved caspase-7 were explored between EIF3C-siRNA and controls. Furthermore, the effects of EIF3C gene knockdown in mTOR pathway were analyzed by western blotting for different cell lines. RESULTS In EIF3C-positive tumors, 32 out of 42 showed significantly higher frequencies of high grade group by immunoreactivity (p=0.0016). BrdU incorporation after four days of cell plating was significantly suppressed in MDA-MB-231 cells by EIF3C knockdown compared with controls, with average changes of 7.8-fold (pcells by EIF3C knockdown compared with controls (pCell apoptosis was significantly increased in the EIF3C-siRNA group when compared with the cells that were transfected with scrambled siRNA (3.51±0.0842 versus 13.24±0.2307, p<0.01). The mTOR signaling pathway was involved in decreasing EIF3C translational efficiency. CONCLUSIONS Unveiling the mechanisms of EIF3 action in tumorigenesis may help identify attractive targets for cancer therapy.

  17. Long none coding RNA HOTTIP/HOXA13 act as synergistic role by decreasing cell migration and proliferation in Hirschsprung disease

    International Nuclear Information System (INIS)

    Xie, Hua; Zhu, Dongmei; Xu, Cao; Zhu, Hairong; Chen, Pingfa; Li, Hongxing; Liu, Xiang; Xia, Yankai; Tang, Weibing

    2015-01-01

    Long noncoding RNAs (lncRNAs) have been confirmed to be associated with various human diseases. However, whether they are associated with Hirschsprung disease (HSCR) progression remains unclear. In this study, we designed the experiment to explore the relationship between lncRNA HOTTIP and HOXA13, and their pathogenicity to HSCR. Quantitative real-time PCR and Western blot were performed to detect the levels of lncRNA, mRNAs, and proteins in colon tissues from 79 patients with HSCR and 79 controls. Small RNA interference transfection was used to study the function experiments in human 293T and SK-N-BE cell lines. The cell viability and activities were detected by the transwell assays, CCK8 assay, and flow cytometry, respectively. LncRNA HOTTIP and HOXA13 were significantly down-regulated in HSCR compared to the controls. Meanwhile, the declined extent of their expression levels makes sense between two main phenotype of HSCR. SiRNA-mediated knock-down of HOTTIP or HOXA13 correlated with decreased levels of each other and both reduced the cell migration and proliferation without affecting cell apoptosis or cell cycle. Our study demonstrates that aberrant reduction of HOTTIP and HOXA13, which have a bidirectional regulatory loop, may play an important role in the pathogenesis of HSCR. - Highlights: • LncRNA HOTTIP and HOXA13 are both down-regulated in HSCR. • HOTTIP and HOXA13 can regulate each other in 293T and SK-N-BE(2) cell lines. • Both HOTTIP and HOXA13 can decrease cell migration and proliferation

  18. High Fat Diet-Induced Skeletal Muscle Wasting Is Decreased by Mesenchymal Stem Cells Administration: Implications on Oxidative Stress, Ubiquitin Proteasome Pathway Activation, and Myonuclear Apoptosis

    Directory of Open Access Journals (Sweden)

    Johanna Abrigo

    2016-01-01

    Full Text Available Obesity can lead to skeletal muscle atrophy, a pathological condition characterized by the loss of strength and muscle mass. A feature of muscle atrophy is a decrease of myofibrillar proteins as a result of ubiquitin proteasome pathway overactivation, as evidenced by increased expression of the muscle-specific ubiquitin ligases atrogin-1 and MuRF-1. Additionally, other mechanisms are related to muscle wasting, including oxidative stress, myonuclear apoptosis, and autophagy. Stem cells are an emerging therapy in the treatment of chronic diseases such as high fat diet-induced obesity. Mesenchymal stem cells (MSCs are a population of self-renewable and undifferentiated cells present in the bone marrow and other mesenchymal tissues of adult individuals. The present study is the first to analyze the effects of systemic MSC administration on high fat diet-induced skeletal muscle atrophy in the tibialis anterior of mice. Treatment with MSCs reduced losses of muscle strength and mass, decreases of fiber diameter and myosin heavy chain protein levels, and fiber type transitions. Underlying these antiatrophic effects, MSC administration also decreased ubiquitin proteasome pathway activation, oxidative stress, and myonuclear apoptosis. These results are the first to indicate that systemically administered MSCs could prevent muscle wasting associated with high fat diet-induced obesity and diabetes.

  19. Dithiolethione modified valproate and diclofenac increase E-cadherin expression and decrease proliferation of non-small cell lung cancer cells

    OpenAIRE

    Moody, Terry W.; Switzer, Christopher; Santana-Flores, Wilmarie; Ridnour, Lisa A.; Berna, Marc; Thill, Michelle; Jensen, Robert T.; Sparatore, Anna; Del Soldato, Piero; Yeh, Grace C; Roberts, David D.; Giaccone, Giuseppe; Wink, David A.

    2009-01-01

    The effects of dithiolethione-modified valproate, diclofenac and sulindac on non-small cell lung cancer (NSCLC) cells were investigated. Sulfur(S)-valproate and S-diclofenac at 1 μg/ml concentrations significantly reduced prostaglandin (PG)E2 levels in NSCLC cell lines A549 and NCI-H1299 as did the COX-2 inhibitor DuP-697. In vitro, S-valproate, S-diclofenac and S-sulindac half-maximally inhibited the clonal growth of NCI-H1299 cells at 6, 6 and 15 μg/ml, respectively. Using the MTT assay, 10...

  20. Decreased camptothecin sensitivity of the stem-cell-like fraction of Caco2 cells correlates with an altered phosphorylation pattern of topoisomerase I.

    Science.gov (United States)

    Roy, Amit; Tesauro, Cinzia; Frøhlich, Rikke; Hede, Marianne S; Nielsen, Maria J; Kjeldsen, Eigil; Bonven, Bjarne; Stougaard, Magnus; Gromova, Irina; Knudsen, Birgitta R

    2014-01-01

    The CD44+ and CD44- subpopulations of the colorectal cancer cell line Caco2 were analyzed separately for their sensitivities to the antitumor drug camptothecin. CD44+ cells were less sensitive to camptothecin than CD44- cells. The relative resistance of CD44+ cells was correlated with (i) reduced activity of the nuclear enzyme topoisomerase I and (ii) insensitivity of this enzyme to camptothecin when analyzed in extracts. In contrast, topoisomerase I activity was higher in extracts from CD44- cells and the enzyme was camptothecin sensitive. Topoisomerase I from the two subpopulations were differentially phosphorylated in a manner that appeared to determine the drug sensitivity and activity of the enzyme. This finding was further supported by the fact that phosphorylation of topoisomerase I in CD44+ cell extract by protein kinase CK2 converted the enzyme to a camptothecin sensitive, more active form mimicking topoisomerase I in extracts from CD44- cells. Conversely, dephosphorylation of topoisomerase I in extracts from CD44- cells rendered the enzyme less active and camptothecin resistant. These findings add to our understanding of chemotherapy resistance in the Caco2 CD44+ cancer stem cell model.

  1. Decreased camptothecin sensitivity of the stem-cell-like fraction of Caco2 cells correlates with an altered phosphorylation pattern of topoisomerase I.

    Directory of Open Access Journals (Sweden)

    Amit Roy

    Full Text Available The CD44+ and CD44- subpopulations of the colorectal cancer cell line Caco2 were analyzed separately for their sensitivities to the antitumor drug camptothecin. CD44+ cells were less sensitive to camptothecin than CD44- cells. The relative resistance of CD44+ cells was correlated with (i reduced activity of the nuclear enzyme topoisomerase I and (ii insensitivity of this enzyme to camptothecin when analyzed in extracts. In contrast, topoisomerase I activity was higher in extracts from CD44- cells and the enzyme was camptothecin sensitive. Topoisomerase I from the two subpopulations were differentially phosphorylated in a manner that appeared to determine the drug sensitivity and activity of the enzyme. This finding was further supported by the fact that phosphorylation of topoisomerase I in CD44+ cell extract by protein kinase CK2 converted the enzyme to a camptothecin sensitive, more active form mimicking topoisomerase I in extracts from CD44- cells. Conversely, dephosphorylation of topoisomerase I in extracts from CD44- cells rendered the enzyme less active and camptothecin resistant. These findings add to our understanding of chemotherapy resistance in the Caco2 CD44+ cancer stem cell model.

  2. Decreased nuclear stiffness via FAK-ERK1/2 signaling is necessary for osteopontin-promoted migration of bone marrow-derived mesenchymal stem cells.

    Science.gov (United States)

    Liu, Lingling; Luo, Qing; Sun, Jinghui; Wang, Aoli; Shi, Yisong; Ju, Yang; Morita, Yasuyuki; Song, Guanbin

    2017-06-15

    Migration of bone marrow-derived mesenchymal stem cells (BMSCs) plays an important role in many physiological and pathological settings, including wound healing. During the migration of BMSCs through interstitial tissues, the movement of the nucleus must be coordinated with the cytoskeletal dynamics, which in turn affects the cell migration efficiency. Our previous study indicated that osteopontin (OPN) significantly promotes the migration of rat BMSCs. However, the nuclear behaviors and involved molecular mechanisms in OPN-mediated BMSC migration are largely unclear. In the present study, using an atomic force microscope (AFM), we found that OPN could decrease the nuclear stiffness of BMSCs and reduce the expression of lamin A/C, which is the main determinant of nuclear stiffness. Increased lamin A/C expression attenuates BMSC migration by increasing nuclear stiffness. Decreased lamin A/C expression promotes BMSC migration by decreasing nuclear stiffness. Furthermore, OPN promotes BMSC migration by diminishing lamin A/C expression and decreasing nuclear stiffness via the FAK-ERK1/2 signaling pathway. This study provides strong evidence for the role of nuclear mechanics in BMSC migration as well as new insight into the molecular mechanisms of OPN-promoted BMSC migration. Copyright © 2017 Elsevier Inc. All rights reserved.

  3. Decrease in circulating CD25(hi)Foxp3(+) regulatory T cells following vaccination with the candidate malaria vaccine RTS,S.

    Science.gov (United States)

    Parsons, Emily; Epstein, Judith; Sedegah, Martha; Villasante, Eileen; Stewart, Ann

    2016-08-31

    Regulatory T (Treg) cells have been shown in some cases to limit vaccine-specific immune responses and impact efficacy. Very little is known about the regulatory responses to the leading malaria vaccine candidate, RTS,S. The goal of this study was to begin to characterize the regulatory responses to the RTS,S vaccine. Using multi-parameter flow cytometry, we examined responses in 13 malaria naïve adult volunteers who received 2 doses of RTS,S given eight weeks apart. Five of these volunteers had previously received 3 doses of a candidate DNA-CSP vaccine, with the final dose given approximately one year prior to the first dose of the RTS,S vaccine. We found that the frequency of CD25(hi)Foxp3(+) Treg cells decreased following administration of RTS,S (p=0.0195), with no differences based on vaccine regimen. There was a concomitant decrease in CTLA-4 expression on CD25(hi)Foxp3(+) Treg cells (p=0.0093) and PD-1 levels on CD8(+) T cells (p=0.0002). Additionally, the frequency of anergic CTLA-4(+)CCR7(+) T cells decreased following vaccination. An inverse correlation was observed between the frequency of Plasmodium falciparum circumsporozoite protein (PfCSP)-specific IFN-γ and PfCSP-specific IL-10, as well as an inverse correlation between IL-10 induced by Hepatitis B surface antigen, the carrier of RTS,S, and PfCSP-specific IFN-γ, suggesting that immunity against the vaccine backbone could impact vaccine immunogenicity. These results have implications for future malaria vaccine design. Copyright © 2016. Published by Elsevier Ltd.

  4. 7-ketocholesterol inhibits Na,K-ATPase activity by decreasing expression of its α1-subunit and membrane fluidity in human endothelial cells.

    Science.gov (United States)

    Duran, M J; Pierre, S V; Lesnik, P; Pieroni, G; Bourdeaux, M; Dignat-Georges, F; Sampol, J; Maixent, J M

    2010-11-09

    As cholesterol, oxysterols, can insert the cell membrane and thereby modify the functions of membrane-bound proteins. The Na,K-ATPase is very sensitive to its lipid environment, seems to be involved in important endothelial functions as the regulation of nitric oxide (NO) release. The effects of 7-ketocholesterol , an oxysterol present in oxidized LDL, was investigated on Na,K-ATPase in isolated human endothelial cells. Cells were incubated 24h with lecithin-, cholesterol- or 7-ketocholesterol liposomes (6 μg/ml). K+-stimulated paranitrophenyl phosphatase activity, reflecting Na,K-ATPase activity, was evaluated as well as cell viability and lipoperoxidation. The expression of Na,K-ATPase subunits mRNAs and membrane fluidity were also investigated. As Na,K-ATPase and nitric oxide seem to be related, we determined the production of NO and the expression of endothelial NO synthase mRNAs. Na,K-ATPase activity was strongly decreased by 7-ketocholesterol. This decrease, not related to lipoperoxidation, was correlated with a decreased expression of the Na,K-ATPase α1-subunit messengers and with rigidity of plasma membranes. Cholesterol induced similar effects but was less potent than 7-ketocholesterol. Basal NO production and expression of endothelial NO synthase mRNAs were not modified by 7-ketocholesterol. Our new findings demonstrate that 7-ketocholesterol, used at non toxic doses, was very potent to disrupt the transport of ions by Na,K-ATPase and perturb membrane structure. These data demonstrate that 7-ketocholesterol induces endothelial dysfunction without cell death that may contribute to early events in atherosclerosis.

  5. Progenitor-like cells derived from mouse kidney protect against renal fibrosis in a remnant kidney model via decreased endothelial mesenchymal transition.

    Science.gov (United States)

    Chen, C L; Chou, K J; Fang, H C; Hsu, C Y; Huang, W C; Huang, C W; Huang, C K; Chen, H Y; Lee, P T

    2015-12-02

    Pathophysiological changes associated with chronic kidney disease impair angiogenic processes and increase renal fibrosis. Progenitor-like cells derived from adult kidney have been previously used to promote regeneration in acute kidney injury, even though it remained unclear whether the cells could be beneficial in chronic kidney disease (CKD). In this study, we established a CKD model by five-sixths nephrectomy and mouse kidney progenitor-like cells (MKPCs) were intravenously administered weekly for 5 weeks after establishing CKD. We examined the impact of MKPCs on the progression of renal fibrosis and the potential of MKPCs to preserve the angiogenic process and prevent endothelial mesenchymal transition in vivo and in vitro. Our results demonstrate that the MKPCs delayed interstitial fibrosis and the progression of glomerular sclerosis and ameliorated the decline of kidney function. At 17 weeks, the treated mice exhibited lower blood pressures, higher hematocrit levels, and larger kidney sizes than the control mice. In addition, the MKPC treatment prolonged the survival of the mice with chronic kidney injuries. We observed a decreased recruitment of macrophages and myofibroblasts in the interstitium and the increased tubular proliferation. Notably, MKPC both decreased the level of vascular rarefaction and prevented endothelial mesenchymal transition (EndoMT) in the remnant kidneys. Moreover, the conditioned medium from the MKPCs ameliorated endothelial cell death under hypoxic culture conditions and prevented TGF-β-induced EndoMT through downregulation of phosphorylated Smad 3 in vitro. MKPCs may be a beneficial treatment for kidney diseases characterized by progressive renal fibrosis. The enhanced preservation of angiogenic processes following MKPC injections may be associated with decreased fibrosis in the remnant kidney. These findings provide further understanding of the mechanisms involved in these processes and will help develop new cell

  6. Decrease in newly generated oligodendrocytes leads to motor dysfunctions and changed myelin structures that can be rescued by transplanted cells.

    Science.gov (United States)

    Schneider, Sarah; Gruart, Agnès; Grade, Sofia; Zhang, Yina; Kröger, Stephan; Kirchhoff, Frank; Eichele, Gregor; Delgado García, José M; Dimou, Leda

    2016-12-01

    NG2-glia in the adult brain are known to proliferate and differentiate into mature and myelinating oligodendrocytes throughout lifetime. However, the role of these newly generated oligodendrocytes in the adult brain still remains little understood. Here we took advantage of the Sox10-iCreER T2 x CAG-eGFP x Esco2 fl/fl mouse line in which we can specifically ablate proliferating NG2-glia in adult animals. Surprisingly, we observed that the generation of new oligodendrocytes in the adult brain was severely affected, although the number of NG2-glia remained stable due to the enhanced proliferation of non-recombined cells. This lack of oligodendrogenesis led to the elongation of the nodes of Ranvier as well as the associated paranodes, which could be locally rescued by myelinating oligodendrocytes differentiated from transplanted NG2-glia deriving from wildtype mice. Repetitive measurements of conduction velocity in the corpus callosum of awake animals revealed a progressive deceleration specifically in the mice lacking adult oligodendrogenesis that resulted in progressive motor deficits. In summary, here we demonstrated for the first time that axon function is not only controlled by the reliable organization of myelin, but also requires a dynamic and continuous generation of new oligodendrocytes in the adult brain. GLIA 2016;64:2201-2218. © 2016 Wiley Periodicals, Inc.

  7. Hoxa9 collaborates with E2A-PBX1 in mouse B cell leukemia in association with Flt3 activation and decrease of B cell gene expression.

    Science.gov (United States)

    Hassawi, Mona; Shestakova, Elena A; Fournier, Marilaine; Lebert-Ghali, Charles-Étienne; Vaisson, Gratianne; Frison, Héloïse; Sinnett, Daniel; Vidal, Ramon; Thompson, Alexander; Bijl, Janet J

    2014-01-01

    The fusion protein E2A-PBX1 induces pediatric B cell leukemia in human. Previously, we reported oncogenic interactions between homeobox (Hox) genes and E2A-PBX1 in murine T cell leukemia. A proviral insertional mutagenesis screen with our E2A-PBX1 B cell leukemia mouse model identified Hoxa genes as potential collaborators to E2A-PBX1. Here we studied whether Hoxa9 could enhance E2A-PBX1 leukemogenesis. We show that Hoxa9 confers a proliferative advantage to E2A-PBX1 B cells. Transplantation experiments with E2A-PBX1 transgenic B cells overexpressing Hoxa9 isolated from bone marrow chimeras showed that Hoxa9 accelerates the generation of E2A-PBX1 B cell leukemia, but Hoxa9 is unable to transform B cells alone. Quantitative-reverse transcriptase polymerase chain reaction analysis demonstrated a strong repression of B cell specific genes in these E2A-PBX1/Hoxa9 leukemias in addition to Flt3 activation, indicating inhibition of B cell differentiation in combination with enhanced proliferation. Overexpression of Hoxa9 in established E2A-PBX1 mouse leukemic B cells resulted in a growth advantage in vitro, which was also characterized by an enhanced expression of Flt3. we show for the first time that Hoxa9 collaborates with E2A-PBX1 in the oncogenic transformation of B cells in a mouse model that involves Flt3 signaling, which is potentially relevant to human disease. Copyright © 2013 Wiley Periodicals, Inc.

  8. Decreased donor-specific cytotoxic T cell precursor frequencies one year after clinical lung transplantation do not reflect transplantation tolerance : A comparison of lung transplant recipients with or without bronchiolitis obliterans syndrome

    NARCIS (Netherlands)

    de Haan, A; van der Gun, [No Value; Hepkema, BG; van der Bij, W; de Leij, LFMH; Prop, J

    2000-01-01

    Background. Decreased in vitro T cell alloreactivity, demonstrated by decreased frequencies of peripheral blood donor-specific T cell precursors, may reflect a tolerant state after transplantation and lower the risk for development of chronic graft dysfunction. It is unknown whether a decrease in

  9. Erastin-Like Anti-Warburg Agents Prevent Mitochondrial Depolarization Induced by Free Tubulin and Decrease Lactate Formation in Cancer Cells.

    Science.gov (United States)

    DeHart, David N; Lemasters, John J; Maldonado, Eduardo N

    2018-01-01

    In Warburg metabolism, suppression of mitochondrial metabolism contributes to a low cytosolic ATP/ADP ratio favoring enhanced aerobic glycolysis. Flux of metabolites across the mitochondrial outer membrane occurs through voltage-dependent anion channels (VDAC). In cancer cells, free dimeric tubulin induces VDAC closure and dynamically regulates mitochondrial membrane potential (ΔΨ). Erastin, a small molecule that binds to VDAC, antagonizes the inhibitory effect of tubulin on VDAC and hyperpolarizes mitochondria in intact cells. Here, our aim was to identify novel compounds from the ChemBridge DIVERSet library that block the inhibitory effect of tubulin on ΔΨ using cell-based screening. HCC4006 cells were treated with nocodazole (NCZ) to increase free tubulin and decrease ΔΨ in the presence or absence of library compounds. Tetramethylrhodamine methylester (TMRM) fluorescence was assessed by high-content imaging to determine changes in ΔΨ. Compounds were considered positive if ΔΨ increased in the presence of NCZ. Using confocal microscopy, we identified and validated six lead molecules that antagonized the depolarizing effect of NCZ. Lead compounds and erastin did not promote microtubule stabilization, so changes in ΔΨ were independent of tubulin dynamics. The most potent lead compound also decreased lactate formation. These novel small molecules represent a potential new class of anti-Warburg drugs.

  10. Neural Mobilization Treatment Decreases Glial Cells and Brain-Derived Neurotrophic Factor Expression in the Central Nervous System in Rats with Neuropathic Pain Induced by CCI in Rats.

    Science.gov (United States)

    Giardini, Aline Carolina; Dos Santos, Fabio Martinez; da Silva, Joyce Teixeira; de Oliveira, Mara Evany; Martins, Daniel Oliveira; Chacur, Marucia

    2017-01-01

    Background . Glial cells are implicated in the development of chronic pain and brain-derived neurotropic factor (BDNF) released from activated microglia contributes to the nociceptive transmission. Neural mobilization (NM) technique is a method clinically effective in reducing pain sensitivity. Here we examined the involvement of glial cells and BDNF expression in the thalamus and midbrain after NM treatment in rats with chronic constriction injury (CCI). CCI was induced and rats were subsequently submitted to 10 sessions of NM, every other day, beginning 14 days after CCI. Thalamus and midbrain were analyzed for glial fibrillary acidic protein (GFAP), microglial cell OX-42, and BDNF using Immunohistochemistry and Western blot assays. Results . Thalamus and midbrain of CCI group showed increases in GFAP, OX-42, and BDNF expression compared with control group and, in contrast, showed decreases in GFAP, OX-42, and BDNF after NM when compared with CCI group. The decreased immunoreactivity for GFAP, OX-42, and BDNF in ventral posterolateral nucleus in thalamus and the periaqueductal gray in midbrain was shown by immunohistochemistry. Conclusions . These findings may improve the knowledge about the involvement of astrocytes, microglia, and BDNF in the chronic pain and show that NM treatment, which alleviates neuropathic pain, affects glial cells and BDNF expression.

  11. Esculetin Downregulates the Expression of AML1-ETO and C-Kit in Kasumi-1 Cell Line by Decreasing Half-Life of mRNA

    Directory of Open Access Journals (Sweden)

    Sharad Sawney

    2015-01-01

    Full Text Available One of the most frequent genetic aberrations in acute myeloid leukemia (AML is chromosomal translocation between AML1/RUNX1 on chromosome 21 and ETO gene on chromosome 8 resulting in the expression of chimeric oncogene AML1-ETO. Although patients with t(8;21 translocation have good prognosis, 5-year survival is observed only in 50% of the cases. AML1-ETO translocation is usually accompanied by overexpression of mutant C-Kit, a tyrosine kinase, which contributes to uncontrolled proliferation of premature blood cells leading to relapse and poor prognosis. We illustrate the potential use of esculetin on leukemic cell line, Kasumi-1, bearing t(8;21 translocation and mutated C-Kit gene. Esculetin decreases the expression of AML1-ETO at both protein and transcript level within 24 hours of treatment. Half-life of AML1-ETO mRNA was reduced from 7 hours to 1.5 hours. Similarly half-life of C-Kit mRNA was reduced to 2 hours from 5 hours in esculetin treated cells. Esculetin also perturbed the expression of ectopically expressed AML1-ETO in U937 cells. The decreased expression of AML1-ETO chimeric gene was associated with increased expression of LAT1 and RUNX3 genes, targets of AML1. We envisage that discovery of a drug candidate which could target both these mutated genes would be a considerable breakthrough for future application.

  12. Elevation of arginine decarboxylase-dependent putrescine production enhances aluminum tolerance by decreasing aluminum retention in root cell walls of wheat.

    Science.gov (United States)

    Yu, Yan; Jin, Chongwei; Sun, Chengliang; Wang, Jinghong; Ye, Yiquan; Lu, Lingli; Lin, Xianyong

    2015-12-15

    Aluminum (Al) stress induces putrescine (Put) accumulation in several plants and this response is proposed to alleviate Al toxicity. However, the mechanisms underlying this alleviation remain largely unknown. Here, we show that exposure to Al clearly increases Put accumulation in the roots of wheat plants (Triticum aestivum L. 'Xi Aimai-1') and that this was accompanied by significant increase in the activity of arginine decarboxylase (ADC), a Put producing enzyme. Application of an ADC inhibitor (d-arginine) terminated the Al-induced Put accumulation, indicating that increased ADC activity may be responsible for the increase in Put accumulation in response to Al. The d-arginine treatment also increased the Al-induced accumulation of cell wall polysaccharides and the degree of pectin demethylation in wheat roots. Thus, it elevated Al retention in cell walls and exacerbated Al accumulation in roots, both of which aggravate Al toxicity in wheat plants. The opposite effects were true for exogenous Put application. These results suggest that ADC-dependent Put accumulation plays important roles in providing protection against Al toxicity in wheat plants through decreasing cell wall polysaccharides and increasing the degree of pectin methylation, thus decreasing Al retention in the cell walls. Copyright © 2015 Elsevier B.V. All rights reserved.

  13. Low Molecular Weight Protein Tyrosine Phosphatase Slow Isoform Knockdown in MDA-MB-435 Cells Decreases RAW 264.7 Osteoclastic Differentiation.

    Science.gov (United States)

    Alho, Irina; Costa, Luis; Bicho, Manuel; Coelho, Constança

    2016-05-01

    During the bone metastatic process, tumor cells and bone cells drive a vicious cycle stimulating growth and activity of each other. We here address how low molecular weight protein tyrosine phosphatase (LMW-PTP) could be involved in this process. We targeted LMW-PTP by siRNA and evaluated the effect of various soluble factors released to the culture medium by the MDA-MB-435 breast cancer cell line, in RAW 264.7 osteoclastogenesis. We showed that these soluble factors did not change RAW 264.7 osteoclastogenic potential. The knockdown of the LMW-PTP slow isoform decreased osteoclastogenesis of RAW 264.7, showing less active Src. The knockdown of LMW-PTP and its slow isoform decreased the release of IL-8 but not IL-6 in MDA-MB-435. The LMW-PTP slow isoform can be an important protein in bone metastatic disease, with a fundamental role in the interplay between tumor cells and osteoclasts, through the regulation of Src activity and IL-8 secretion. Copyright© 2016 International Institute of Anticancer Research (Dr. John G. Delinassios), All rights reserved.

  14. The novel curcumin analog FLLL32 decreases STAT3 DNA binding activity and expression, and induces apoptosis in osteosarcoma cell lines

    International Nuclear Information System (INIS)

    Fossey, Stacey L; London, Cheryl A; Bear, Misty D; Lin, Jiayuh; Li, Chenglong; Schwartz, Eric B; Li, Pui-Kai; Fuchs, James R; Fenger, Joelle; Kisseberth, William C

    2011-01-01

    Curcumin is a naturally occurring phenolic compound shown to have a wide variety of antitumor activities; however, it does not attain sufficient blood levels to do so when ingested. Using structure-based design, a novel compound, FLLL32, was generated from curcumin. FLLL32 possesses superior biochemical properties and more specifically targets STAT3, a transcription factor important in tumor cell survival, proliferation, metastasis, and chemotherapy resistance. In our previous work, we found that several canine and human osteosarcoma (OSA) cell lines, but not normal osteoblasts, exhibit constitutive phosphorylation of STAT3. Compared to curcumin, we hypothesized that FLLL32 would be more efficient at inhibiting STAT3 function in OSA cells and that this would result in enhanced downregulation of STAT3 transcriptional targets and subsequent death of OSA cells. Human and canine OSA cells were treated with vehicle, curcumin, or FLLL32 and the effects on proliferation (CyQUANT ® ), apoptosis (SensoLyte ® Homogeneous AMC Caspase- 3/7 Assay kit, western blotting), STAT3 DNA binding (EMSA), and vascular endothelial growth factor (VEGF), survivin, and matrix metalloproteinase-2 (MMP2) expression (RT-PCR, western blotting) were measured. STAT3 expression was measured by RT-PCR, qRT- PCR, and western blotting. Our data showed that FLLL32 decreased STAT3 DNA binding by EMSA. FLLL32 promoted loss of cell proliferation at lower concentrations than curcumin leading to caspase-3- dependent apoptosis, as evidenced by PARP cleavage and increased caspase 3/7 activity; this could be inhibited by treatment with the pan-caspase inhibitor Z-VAD-FMK. Treatment of OSA cells with FLLL32 decreased expression of survivin, VEGF, and MMP2 at both mRNA and protein levels with concurrent decreases in phosphorylated and total STAT3; this loss of total STAT3 occurred, in part, via the ubiquitin-proteasome pathway. These data demonstrate that the novel curcumin analog FLLL32 has biologic activity

  15. NT-020, a natural therapeutic approach to optimize spatial memory performance and increase neural progenitor cell proliferation and decrease inflammation in the aged rat.

    Science.gov (United States)

    Acosta, S; Jernberg, J; Sanberg, C D; Sanberg, P R; Small, Brent J; Gemma, Carmelina; Bickford, Paula C

    2010-10-01

    The process of aging is linked to oxidative stress, microglial activation, and proinflammatory factors, which are known to decrease cell proliferation and limit neuroplasticity. These factors may lead the transition from normal aging to more severe cognitive dysfunction associated with neurodegenerative diseases. We have shown that natural compounds such as polyphenols from blueberry and green tea and amino acids like carnosine are high in antioxidant and antiinflammatory activity that decreases the damaging effects of reactive oxygen species (ROS), in the blood, brain, and other tissues of the body. Furthermore, we have shown that the combination of these nutrients (called NT-020) creates a synergistic effect that promotes the proliferation of stem cells in vitro and in vivo. In the current study, we examined the effects of NT-020 on neurogenesis and performance on a Morris water maze (MWM). Aged (20-month-old) male Fischer 344 rats were treated with 135.0 mg/kg per day (n = 13) of NT-020. Young (3-month-old) (n = 10) and aged (20-month-old) (n = 13) control male Fischer 344 rats were treated with water by oral gavage. All groups were treated for a period of 4 weeks. Although there was no difference in performance in the MWM when comparing all aged rats, when the data for aged impaired rats were compared, there was a significant difference between groups on the last day of training with the treatment group performing better than controls. Using the cell cycle-regulating protein (Ki67), doublecortin (DCX), and OX6 antibody markers, cell proliferation, neurogenesis, and microglial activation were estimated in the dentate gyrus (DG) of young and aged animals. Cell proliferation was also examined in the subventricular zone (SVZ). A decreased number of OX6 MHC II-positive cells, increased neurogenesis, and increased number of proliferating cells were found in rats treated with NT-020 in comparison with aged control rats. In sum, NT-020 may promote health, proliferation

  16. Major components of energy drinks (caffeine, taurine, and guarana) exert cytotoxic effects on human neuronal SH-SY5Y cells by decreasing reactive oxygen species production.

    Science.gov (United States)

    Zeidán-Chuliá, Fares; Gelain, Daniel Pens; Kolling, Eduardo Antônio; Rybarczyk-Filho, José Luiz; Ambrosi, Priscilla; Terra, Silvia Resende; Pires, André Simões; da Rocha, João Batista Teixeira; Behr, Guilherme Antônio; Moreira, José Cláudio Fonseca

    2013-01-01

    To elucidate the morphological and biochemical in vitro effects exerted by caffeine, taurine, and guarana, alone or in combination, since they are major components in energy drinks (EDs). On human neuronal SH-SY5Y cells, caffeine (0.125-2 mg/mL), taurine (1-16 mg/mL), and guarana (3.125-50 mg/mL) showed concentration-dependent nonenzymatic antioxidant potential, decreased the basal levels of free radical generation, and reduced both superoxide dismutase (SOD) and catalase (CAT) activities, especially when combined together. However, guarana-treated cells developed signs of neurite degeneration in the form of swellings at various segments in a beaded or pearl chain-like appearance and fragmentation of such neurites at concentrations ranging from 12.5 to 50 mg/mL. Swellings, but not neuritic fragmentation, were detected when cells were treated with 0.5 mg/mL (or higher doses) of caffeine, concentrations that are present in EDs. Cells treated with guarana also showed qualitative signs of apoptosis, including membrane blebbing, cell shrinkage, and cleaved caspase-3 positivity. Flow cytometric analysis confirmed that cells treated with 12.5-50 mg/mL of guarana and its combinations with caffeine and/or taurine underwent apoptosis. Excessive removal of intracellular reactive oxygen species, to nonphysiological levels (or "antioxidative stress"), could be a cause of in vitro toxicity induced by these drugs.

  17. Non-cytotoxic Concentration of Cisplatin Decreases Neuroplasticity-Related Proteins and Neurite Outgrowth Without Affecting the Expression of NGF in PC12 Cells.

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    Ferreira, Rafaela Scalco; Dos Santos, Neife Aparecida Guinaim; Martins, Nádia Maria; Fernandes, Laís Silva; Dos Santos, Antonio Cardozo

    2016-11-01

    Cisplatin is the most effective and neurotoxic platinum chemotherapeutic agent. It induces a peripheral neuropathy characterized by distal axonal degeneration that might progress to degeneration of cell bodies and apoptosis. Most symptoms occur nearby distal axonal branches and axonal degeneration might induce peripheral neuropathy regardless neuronal apoptosis. The toxic mechanism of cisplatin has been mainly associated with DNA damage, but cisplatin might also affect neurite outgrowth. Nevertheless, the neurotoxic mechanism of cisplatin remains unclear. We investigated the early effects of cisplatin on axonal plasticity by using non-cytotoxic concentrations of cisplatin and PC12 cells as a model of neurite outgrowth and differentiation. PC12 cells express NGF-receptors (trkA) and respond to NGF by forming neurites, branches and synaptic vesicles. For comparison, we used a neuronal model (SH-SY5Y cells) that does not express trkA nor responds to NGF. Cisplatin did not change NGF expression in PC12 cells and decreased neurite outgrowth in both models, suggesting a NGF/trkA independent mechanism. It also reduced axonal growth (GAP-43) and synaptic (synapsin I and synaptophysin) proteins in PC12 cells, without inducing mitochondrial damage or apoptosis. Therefore, cisplatin might affect axonal plasticity before DNA damage, NGF/trkA down-regulation, mitochondrial damage or neuronal apoptosis. This is the first study to show that neuroplasticity-related proteins might be early targets of the neurotoxic action of cisplatin and their role on cisplatin-induced peripheral neuropathy should be investigated in vivo.

  18. Leptin and insulin up-regulate miR-4443 to suppress NCOA1 and TRAF4, and decrease the invasiveness of human colon cancer cells

    International Nuclear Information System (INIS)

    Meerson, Ari; Yehuda, Hila

    2016-01-01

    Obesity is a risk factor for colorectal cancer (CRC). Normal and tumor cells respond to metabolic hormones, such as leptin and insulin. Thus, obesity-associated resistance to these hormones likely leads to changes in gene expression and behavior of tumor cells. However, the mechanisms affected by leptin and insulin signaling in CRC cells remain mostly unknown. We hypothesized that microRNAs (miRNAs) are involved in the regulation of tumorigenesis-related gene expression in CRC cells by leptin and insulin. To test this hypothesis, miRNA levels in the CRC-derived cell lines HCT-116, HT-29 and DLD-1 were profiled, following leptin and insulin treatment. Candidate miRNAs were validated by RT-qPCR. Predicted miRNA targets with known roles in cancer, were validated by immunoblots and reporter assays in HCT-116 cells. Transfection of HCT-116 cells with candidate miRNA mimic was used to test in vitro effects on proliferation and invasion. Of ~800 miRNAs profiled, miR-4443 was consistently up-regulated by leptin and insulin in HCT-116 and HT-29, but not in DLD-1, which lacked normal leptin receptor expression. Dose response experiments showed that leptin at 100 ng/ml consistently up-regulated miR-4443 in HCT-116 cells, concomitantly with a significant decrease in cell invasion ability. Transfection with miR-4443 mimic decreased invasion and proliferation of HCT-116 cells. Moreover, leptin and miR-4443 transfection significantly down-regulated endogenous NCOA1 and TRAF4, both predicted targets of miR-4443 with known roles in cancer metastasis. miR-4443 was found to directly regulate TRAF4 and NCOA1, as validated by a reporter assay. The up-regulation of miR-4443 by leptin or insulin was attenuated by the inhibition of MEK1/2. Our findings suggest that miR-4443 acts in a tumor-suppressive manner by down-regulating TRAF4 and NCOA1 downstream of MEK-C/EBP-mediated leptin and insulin signaling, and that insulin and/or leptin resistance (e.g. in obesity) may suppress this pathway

  19. Decreasing NF-κB expression enhances odontoblastic differentiation and collagen expression in dental pulp stem cells exposed to inflammatory cytokines.

    Directory of Open Access Journals (Sweden)

    Neda S T Hozhabri

    Full Text Available Inflammatory response in the dental pulp can alter the collagen matrix formation by dental pulp stem cells and lead to a delay or poor healing of the pulp. This inflammatory response is mediated by cytokines, including interleukin-1β and tumor necrosis factor-α. In this study, it is hypothesized that suppressing the actions of these inflammatory cytokines by knocking down the activity of transcription factor Nuclear Factor-κB will lead to dental pulp stem cell differentiation into odontoblasts and the production of collagen. Here, the role of Nuclear Factor-κB signaling and its reduction was examined during odontogenic behavior in the presence of these cytokines. The results showed a significant increase in Nuclear Factor-κB gene expression and p65 protein expression by interleukin-1β and tumor necrosis factor-α. Nuclear Factor-κB activation in the presence of these cytokines decreased significantly in a dose-dependent manner by a Nuclear Factor-κB inhibitor (MG132 and p65 siRNA. Down-regulation of Nuclear Factor-κB activity also enhanced the gene expression of the odontoblastic markers (dentin sialophosphoprotein, Nestin, and alkaline phosphatase and displayed an odontoblastic cell morphology indicating the promotion of odontogenic differentiation of dental pulp stem cells. Finally, dental pulp stem cells exposed to reduced Nuclear Factor-κB activity resulted in a significant increase in collagen (I-α1 expression in the presence of these cytokines. In conclusion, a decrease in Nuclear Factor-κB in dental pulp stem cells in the presence of inflammatory cytokines enhanced odontoblastic differentiation and collagen matrix formation.

  20. Reduced LAK cytotoxicity of peripheral blood mononuclear cells in patients with bladder cancer: Decreased LAK cytotoxicity caused by a low incidence of CD56+ and CD57+ mononuclear blood cells

    International Nuclear Information System (INIS)

    Hermann, G.G.; Petersen, K.R.; Steven, K.; Zeuthen, J.

    1990-01-01

    The cytotoxicity of unstimulated peripheral blood mononuclear cells (US-PBMC), phytohemagglutinin (PHA)-stimulated PBMC (PS-PBMC) and interleukin-2 (IL-2)-activated PBMC (LAK cells) was assessed in patients with noninvasive and invasive transitional-cell bladder cancer and compared with those determined in healthy controls. The differences in the cytotoxicities were correlated with specific changes in the subsets of peripheral blood mononuclear cells (PBMC). PBMC from 37 patients and 13 healthy controls were tested against the bladder cancer cell line T24 in 51 Cr-release assays. The PBMC subsets were analyzed using monoclonal antibodies against T cells, natural killer (NK) -cells, monocytes, and activation markers. The cytotoxicities of US-PBMC, PS-PBMC, and LAK cells were all significantly lower in the cancer patients than in the controls (P less than 0.05). The percentages of PBMC positive for the NK-cell markers CD56 and CD57 were lowest in the patients and were correlated to the decrease in cytotoxicity. Depletion of CD56+ or CD57+ cells from PBMC prior to or after 2 days stimulation with IL-2 demonstrated that these cells are the major source of LAK-cell cytotoxicity and showed that the reduced ability of bladder cancer patient PBMC to develop LAK-cell cytotoxicity is a result of a low incidence of CD56+ and CD57+ cells in the blood. These findings indicate that IL-2 therapy alone might not be a sufficient therapy of bladder cancer patients

  1. Dithiolethione modified valproate and diclofenac increase E-cadherin expression and decrease proliferation of non-small cell lung cancer cells.

    Science.gov (United States)

    Moody, Terry W; Switzer, Christopher; Santana-Flores, Wilmarie; Ridnour, Lisa A; Berna, Marc; Thill, Michelle; Jensen, Robert T; Sparatore, Anna; Del Soldato, Piero; Yeh, Grace C; Roberts, David D; Giaccone, Giuseppe; Wink, David A

    2010-05-01

    The effects of dithiolethione modified valproate, diclofenac and sulindac on non-small cell lung cancer (NSCLC) cells were investigated. Sulfur(S)-valproate and S-diclofenac at 1 microg/ml concentrations significantly reduced prostaglandin (PG)E(2) levels in NSCLC cell lines A549 and NCI-H1299 as did the COX-2 inhibitor DuP-697. In vitro, S-valproate, S-diclofenac and S-sulindac half-maximally inhibited the clonal growth of NCI-H1299 cells at 6, 6 and 15 microg/ml, respectively. Using the MTT assay, 10 microg/ml S-valproate, NO-aspirin and Cay10404, a selective COX-2 inhibitor, but not SC-560, a selective COX-1 inhibitor, inhibited the growth of A549 cells. In vivo, 18mg/kg i.p. of S-valproate and S-diclofenac, but not S-sulindac, significantly inhibited A549 or NCI-H1299 xenograft proliferation in nude mice, but had no effect on the nude mouse body weight. The mechanism by which S-valproate and S-diclofenac inhibited the growth of NSCLC cells was investigated. Nitric oxide-aspirin but not S-valproate caused apoptosis of NSCLC cells. By Western blot, S-valproate and S-diclofenac increased E-cadherin but reduced vimentin and ZEB1 (a transcriptional suppressor of E-cadherin) protein expression in NSCLC cells. Because S-valproate and S-diclofenac inhibit the growth of NSCLC cells and reduce PGE(2) levels, they may prove beneficial in the chemoprevention and/or therapy of NSCLC. Published by Elsevier Ireland Ltd.

  2. Attempts to predict the long-term decrease in lung function due to radiotherapy of non-small cell lung cancer

    International Nuclear Information System (INIS)

    Nielsen, Morten; Hansen, Olfred; Vach, Werner

    2006-01-01

    Purpose: To obtain a model which can predict long-term decrease in lung function due to radiation damage from dose-volume data for patients with non-small cell lung cancer. Patients and methods: 27 patients were included, all long-term survivors after radical radiation therapy. For each patient a regression analysis was performed on a post-RT succession of measurements of FEV1 in order to estimate the decrease after 2 years and a standard error (SE) on this regression estimate. The modelling was based on dose-volume histograms (DVH) exported from the treatment planning system, and involved fits of threshold models, a mean lung dose model as well as more complex models based on the relative damaged volume (rdV). Results: Decreases after 2 years of up to 28% in FEV1 was measured (median 10%), with significant day-to-day variation in FEV1 for the individual patient. The threshold models predicted the long-term decrease in FEV1 well when the SE was interpreted as the uncertainty of the measured decrease. The best threshold value, marginally, was 30 Gy with an R 2 of 0.46. The mean lung dose model did not perform so well. A complex model based on rdV performed better than any of the other models (R 2 =0.52). Conclusion: The long-term decrease in FEV1 could be predicted from a simple dose-volume model when the SE was interpreted as the uncertainty of the measured decrease

  3. Decrease in the red cell cofactor 2,3-diphosphoglycerate increases hemoglobin oxygen affinity in the hibernating brown bear Ursus arctos.

    Science.gov (United States)

    Revsbech, Inge G; Malte, Hans; Fröbert, Ole; Evans, Alina; Blanc, Stéphane; Josefsson, Johan; Fago, Angela

    2013-01-01

    During winter hibernation, brown bears (Ursus arctos) reduce basal O(2) consumption rate to ∼25% compared with the active state, while body temperature decreases moderately (to ∼30°C), suggesting a temperature-independent component in their metabolic depression. To establish whether changes in O(2) consumption during hibernation correlate with changes in blood O(2) affinity, we took blood samples from the same six individuals of hibernating and nonhibernating free-ranging brown bears during winter and summer, respectively. A single hemoglobin (Hb) component was detected in all samples, indicating no switch in Hb synthesis. O(2) binding curves measured on red blood cell lysates at 30°C and 37°C showed a less temperature-sensitive O(2) affinity than in other vertebrates. Furthermore, hemolysates from hibernating bears consistently showed lower cooperativity and higher O(2) affinity than their summer counterparts, regardless of the temperature. We found that this increase in O(2) affinity was associated with a significant decrease in the red cell Hb-cofactor 2,3-diphosphoglycerate (DPG) during hibernation to approximately half of the summer value. Experiments performed on purified Hb, to which DPG had been added to match summer and winter levels, confirmed that the low DPG content was the cause of the left shift in the Hb-O(2) equilibrium curve during hibernation. Levels of plasma lactate indicated that glycolysis is not upregulated during hibernation and that metabolism is essentially aerobic. Calculations show that the increase in Hb-O(2) affinity and decrease in cooperativity resulting from decreased red cell DPG may be crucial in maintaining a fairly constant tissue oxygen tension during hibernation in vivo.

  4. Decreased neural precursor cell pool in NADPH oxidase 2-deficiency: From mouse brain to neural differentiation of patient derived iPSC

    Directory of Open Access Journals (Sweden)

    Zeynab Nayernia

    2017-10-01

    Full Text Available There is emerging evidence for the involvement of reactive oxygen species (ROS in the regulation of stem cells and cellular differentiation. Absence of the ROS-generating NADPH oxidase NOX2 in chronic granulomatous disease (CGD patients, predominantly manifests as immune deficiency, but has also been associated with decreased cognition. Here, we investigate the role of NOX enzymes in neuronal homeostasis in adult mouse brain and in neural cells derived from human induced pluripotent stem cells (iPSC. High levels of NOX2 were found in mouse adult neurogenic regions. In NOX2-deficient mice, neurogenic regions showed diminished redox modifications, as well as decrease in neuroprecursor numbers and in expression of genes involved in neural differentiation including NES, BDNF and OTX2. iPSC from healthy subjects and patients with CGD were used to study the role of NOX2 in human in vitro neuronal development. Expression of NOX2 was low in undifferentiated iPSC, upregulated upon neural induction, and disappeared during neuronal differentiation. In human neurospheres, NOX2 protein and ROS generation were polarized within the inner cell layer of rosette structures. NOX2 deficiency in CGD-iPSCs resulted in an abnormal neural induction in vitro, as revealed by a reduced expression of neuroprogenitor markers (NES, BDNF, OTX2, NRSF/REST, and a decreased generation of mature neurons. Vector-mediated NOX2 expression in NOX2-deficient iPSCs rescued neurogenesis. Taken together, our study provides novel evidence for a regulatory role of NOX2 during early stages of neurogenesis in mouse and human.