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  1. 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.

  2. Ethanol decreases radiosensitivity of human breast cancer MCF-7 cells

    International Nuclear Information System (INIS)

    Objective: To investigate the effect of ethanol on radiosensitivity of human breast cancer MCF-7 cells. Methods: Human breast cancer MCF-7 cells were divided into four groups including control group,ethanol treatment group, X-ray exposed group,and ethanol combined with X-ray group. Clonogenic assay was used to determine cell survival. Flow cytometry was employed to analyze cell cycle progression. Annexin V-FITC kit was used to determine cell apoptosis induction.Results Ethanol (50 and 100 mmol/L, 50 h) had no influence on MCF-7 cell growth (t=0.82, 1.15, P>0.05). The radiosensitivity of MCF-7 cells was reduced when the cells were pretreated with 50 mmol/L ethanol (t=4.15, P<0.05) and 100 mmol/L ethanol (t=10.28, P<0.05) for 2 h. Compared with irradiation with X-ray alone, ethanol treatment decreased G2/M phase arrest (t=7.18, P<0.05) and sub-G1 population (an indicator of apoptosis induction) (t=5.39, P<0.05). A decrease of advanced and early apoptosis in the cells pretreated with ethanol was also confirmed by Annexin V-FITC apoptosis assay (t=4.86, 7.59, P<0.05). Conclusions: Ethanol causes radioresistance in human breast cancer MCF-7 cells, where the decreases of radiation-induced G2/M phase arrest and apoptosis may be involved. (authors)

  3. 4-Aminopyridine Decreases Progesterone Production by Porcine Granulosa Cells

    Directory of Open Access Journals (Sweden)

    Mitchell Brianna M

    2003-04-01

    Full Text Available Abstract Background Ion channels occur as large families of related genes with cell-specific expression patterns. Granulosa cells have been shown to express voltage-gated potassium channels from more than one family. The purpose of this study was to determine the effects of 4-aminopyridine (4-AP, an antagonist of KCNA but not KCNQ channels. Methods Granulosa cells were isolated from pig follicles and cultured with 4-AP, alone or in combination with FSH, 8-CPT-cAMP, estradiol 17β, and DIDS. Complimentary experiments determined the effects of 4-AP on the spontaneously established pig granulosa cell line PGC-2. Granulosa cell or PGC-2 function was assessed by radio-immunoassay of media progesterone accumulation. Cell viability was assessed by trypan blue exclusion. Drug-induced changes in cell membrane potential and intracellular potassium concentration were documented by spectrophotometric determination of DiBAC4(3 and PBFI fluorescence, respectively. Expression of proliferating cell nuclear antigen (PCNA and steroidogenic acute regulatory protein (StAR was assessed by immunoblotting. Flow cytometry was also used to examine granulosa cell viability and size. Results 4-AP (2 mM decreased progesterone accumulation in the media of serum-supplemented and serum-free granulosa cultures, but inhibited cell proliferation only under serum-free conditions. 4-AP decreased the expression of StAR, the production of cAMP and the synthesis of estradiol by PGC-2. Addition of either 8-CPT-cAMP or estradiol 17β to serum-supplemented primary cultures reduced the inhibitory effects of 4-AP. 4-AP treatment was also associated with increased cell size, increased intracellular potassium concentration, and hyperpolarization of resting membrane potential. The drug-induced hyperpolarization of resting membrane potential was prevented either by decreasing extracellular chloride or by adding DIDS to the media. DIDS also prevented 4-AP inhibition of progesterone production

  4. Fibronectin Fiber Extension Decreases Cell Spreading and Migration.

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    Hubbard, Brant; Buczek-Thomas, Jo Ann; Nugent, Matthew A; Smith, Michael L

    2016-08-01

    The extracellular matrix (ECM) is present in a range of molecular conformations and intermolecular arrangements. Fibronectin (Fn) molecules that constitute fibers within the ECM can exist in a variety of conformations that result from both mechanical stress and chemical factors such as allosteric binding partners. The long-standing hypothesis that conformational changes regulate the binding of cells to Fn fibers has only been tested for mutated molecules of Fn and has yet to be fully evaluated with Fn fibers. Using time-lapse microscopy we examined how mechanical extension of single fibers of Fn affects the adhesion and migration of endothelial cells. Using this single fiber adhesion technique, we show that high levels of mechanical strain applied to Fn fibers decreases the rates of both cell spreading and cell migration. These data indicate a fundamental cellular response to mechanical strain in the ECM that might have important implications for understanding how cells are recruited during tissue development and repair. J. Cell. Physiol. 231: 1728-1736, 2016. © 2015 Wiley Periodicals, Inc. PMID:26621030

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

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    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. PMID:25499819

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

    International Nuclear Information System (INIS)

    Arecoline, the most abundant areca alkaloid, has been reported to decrease interleukin-6 (IL-6) levels in epithelial cancer cells. Since IL-6 overexpression contributes to the tumorigenic potency of basal cell carcinoma (BCC), this study was designed to investigate whether arecoline altered IL-6 expression and its downstream regulation of apoptosis and the cell cycle in cultured BCC-1/KMC cells. BCC-1/KMC cells and a human keratinocyte cell line, HaCaT, were treated with arecoline at concentrations ranging from 10 to 100 μg/ml, then IL-6 production and expression of apoptosis- and cell cycle progress-related factors were examined. After 24 h exposure, arecoline inhibited BCC-1/KMC cell growth and decreased IL-6 production in terms of mRNA expression and protein secretion, but had no effect on HaCaT cells. Analysis of DNA fragmentation and chromatin condensation showed that arecoline induced apoptosis of BCC-1/KMC cells in a dose-dependent manner, activated caspase-3, and decreased expression of the anti-apoptotic protein Bcl-2. In addition, arecoline induced progressive and sustained accumulation of BCC-1/KMC cells in G2/M phase as a result of reducing checkpoint Cdc2 activity by decreasing Cdc25C phosphatase levels and increasing p53 levels. Furthermore, subcutaneous injection of arecoline led to decreased BCC-1/KMC tumor growth in BALB/c mice by inducing apoptosis. This study demonstrates that arecoline has potential for preventing BCC tumorigenesis by reducing levels of the tumor cell survival factor IL-6, increasing levels of the tumor suppressor factor p53, and eliciting cell cycle arrest, followed by apoptosis. Highlights: ► Arecoline has potential to prevent against basal cell carcinoma tumorigenesis. ► It has more effectiveness on BCC as compared with a human keratinocyte cell line. ► Mechanisms involved including reducing tumor cells’ survival factor IL-6, ► Decreasing Cdc25C phosphatase, enhancing tumor suppressor factor p53, ► Eliciting G2/M

  7. 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

  8. Acetylcholinesterase is associated with a decrease in cell proliferation of hepatocellular carcinoma cells.

    Science.gov (United States)

    Pérez-Aguilar, Benjamín; Vidal, Cecilio J; Palomec, Guillermina; García-Dolores, Fernando; Gutiérrez-Ruiz, María Concepción; Bucio, Leticia; Gómez-Olivares, José Luis; Gómez-Quiroz, Luis Enrique

    2015-07-01

    Acetylcholinesterase (AChE), the enzyme that rapidly splits acetylcholine into acetate and choline, presents non-cholinergic functions through which may participate in the control of cell proliferation and apoptosis. These two features are relevant in cancer, particularly in hepatocellular carcinoma (HCC), a very aggressive liver tumor with high incidence and poor prognosis in advanced stages. Here we explored the relation between acetylcholinesterase and HCC growth by testing the influence of AChE on proliferation of Huh-7 and HepG2 cell lines, addressed in monolayer cultures, spheroid formation and human liver tumor samples. Results showed a clear relation in AChE expression and cell cycle progression, an effect which depended on cell confluence. Inhibition of AChE activity led to an increase in cell proliferation, which was associated with downregulation of p27 and cyclins. The fact that Huh-7 and HepG2 cell lines provided similar results lent weight to the relationship of AChE expression with cell cycle progression in hepatoma cell lines at least. Human liver tumor samples exhibited a decrease in AChE activity as compared with normal tissue. The evidence presented herein provides additional support for the proposed tumor suppressor role of AChE, which makes it a potential therapeutic target in therapies against hepatocellular carcinoma. PMID:25869328

  9. Contrast Adaptation Decreases Complexity in Retinal Ganglion Cell Spike Train

    Institute of Scientific and Technical Information of China (English)

    WANG Guang-Li; HUANG Shi-Yong; ZHANG Ying-Ying; LIANG Pei-Ji

    2007-01-01

    @@ The difference in temporal structures of retinal ganglion cell spike trains between spontaneous activity and firing activity after contrast adaptation is investigated. The Lempel-Ziv complexity analysis reveals that the complexity of the neural spike train decreases after contrast adaptation. This implies that the behaviour of the neuron becomes ordered, which may carry relevant information about the external stimulus. Thus, during the neuron activity after contrast adaptation, external information could be encoded in forms of some certain patterns in the temporal structure of spike train that is significantly different, compared to that of the spike train during spontaneous activity, although the firing rates in spontaneous activity and firing activity after contrast adaptation are sometime similar.

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

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    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 < 0.01), while 3-MA considerably reduced it in melamine-treated PC12 cells (P < 0.01). Furthermore, flow cytometry assay showed that rapamycin considerably reduced the reactive oxygen species (ROS) level of the cells (P < 0.01), but 3-MA increased the generation of ROS (P < 0.01). Additionally, the superoxide dismutase (SOD) activity was notably increased by rapamycin in melamine-treated PC12 cells (P < 0.01), while the activity of which was prominently decreased by 3-MA (P < 0.01). Malondialdehyde (MDA) assay showed that rapamycin remarkably decreased the MDA level of the cells (P < 0.05), while 3-MA increased it (P < 0.01). Consequently, this study demonstrated that autophagy protected 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. PMID:25724280

  11. Cigarette Smoke Decreases the Maturation of Lung Myeloid Dendritic Cells

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    Calero-Acuña, Carmen; Moreno-Mata, Nicolás; Gómez-Izquierdo, Lourdes; Sánchez-López, Verónica; López-Ramírez, Cecilia; Tobar, Daniela; López-Villalobos, José Luis; Gutiérrez, Cesar; Blanco-Orozco, Ana; López-Campos, José Luis

    2016-01-01

    Background Conflicting data exist on the role of pulmonary dendritic cells (DCs) and their maturation in patients with chronic obstructive pulmonary disease (COPD). Herein, we investigated whether disease severity and smoking status could affect the distribution and maturation of DCs in lung tissues of patients undergoing elective pneumectomy or lobectomy for suspected primary lung cancer. Materials and Methods A total of 75 consecutive patients were included. Spirometry testing was used to identify COPD. Lung parenchyma sections anatomically distant from the primary lesion were examined. We used flow cytometry to identify different DCs subtypes—including BDCA1-positive myeloid DCs (mDCs), BDCA3-positive mDCs, and plasmacytoid DCs (pDCs)—and determine their maturation markers (CD40, CD80, CD83, and CD86) in all participants. We also identified follicular DCs (fDCs), Langerhans DCs (LDCs), and pDCs in 42 patients by immunohistochemistry. Results COPD was diagnosed in 43 patients (16 current smokers and 27 former smokers), whereas the remaining 32 subjects were classified as non-COPD (11 current smokers, 13 former smokers, and 8 never smokers). The number and maturation of DCs did not differ significantly between COPD and non-COPD patients. However, the results of flow cytometry indicated that maturation markers CD40 and CD83 of BDCA1-positive mDCs were significantly decreased in smokers than in non-smokers (P = 0.023 and 0.013, respectively). Immunohistochemistry also revealed a lower number of LDCs in COPD patients than in non-COPD subjects. Conclusions Cigarette smoke, rather than airflow limitation, is the main determinant of impaired DCs maturation in the lung. PMID:27058955

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

    Directory of Open Access Journals (Sweden)

    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.

  13. Resolvins Decrease Oxidative Stress Mediated Macrophage and Epithelial Cell Interaction through Decreased Cytokine Secretion

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    Cox, Ruan; Phillips, Oluwakemi; Fukumoto, Jutaro; Fukumoto, Itsuko; Tamarapu Parthasarathy, Prasanna; Mandry, Maria; Cho, Young; Lockey, Richard; Kolliputi, Narasaiah

    2015-01-01

    Background Inflammation is a key hallmark of ALI and is mediated through ungoverned cytokine signaling. One such cytokine, interleukin-1beta (IL-1β) has been demonstrated to be the most bioactive cytokine in ALI patients. Macrophages are the key players responsible for IL-1β secretion into the alveolar space. Following the binding of IL-1β to its receptor, “activated” alveolar epithelial cells show enhanced barrier dysfunction, adhesion molecule expression, cytokine secretion, and leukocyte a...

  14. Decreased serum cell-free DNA levels in rheumatoid arthritis

    OpenAIRE

    Dunaeva, Marina; Buddingh’, Bastiaan C.; René E M Toes; Luime, Jolanda J.; Lubberts, Erik; Pruijn, Ger J. M.

    2015-01-01

    Purpose Recent studies have demonstrated that serum/plasma DNA and RNA molecules in addition to proteins can serve as biomarkers. Elevated levels of these nucleic acids have been found not only in acute, but also in chronic conditions, including autoimmune diseases. The aim of this study was to assess cell-free DNA (cfDNA) levels in sera of rheumatoid arthritis (RA) patients compared to controls. Methods cfDNA was extracted from sera of patients with early and established RA, relapsing-remitt...

  15. CHLORAL HYDRATE DECREASES GAP JUNCTION COMMUNICATION IN RAT LIVER EPITHELIAL CELLS

    Science.gov (United States)

    Chloral hydrate decreases gap junction communication in rat liver epithelial cells Gap junction communication (GJC) is involved in controlling cell proliferation and differentiation. Connexins (Cx) that make up these junctions are composed of a closely related group of m...

  16. Pravastatin induces cell cycle arrest and decreased production of VEGF and bFGF in multiple myeloma cell line.

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    Trojan, P J J; Bohatch-Junior, M S; Otuki, M F; Souza-Fonseca-Guimarães, F; Svidnicki, P V; Nogaroto, V; Fernandes, D; Krum, E A; Favero, G M

    2016-02-01

    Multiple myeloma (MM) is a B cell bone marrow neoplasia characterized by inflammation with an intense secretion of growth factors that promote tumor growth, cell survival, migration and invasion. The aim of this study was to evaluate the effects of pravastatin, a drug used to reduce cholesterol, in a MM cell line.Cell cycle and viability were determinate by Trypan Blue and Propidium Iodide. IL6, VEGF, bFGF and TGFβ were quantified by ELISA and qRT-PCR including here de HMG CoA reductase. It was observed reduction of cell viability, increase of cells in G0/G1 phase of the cell cycle and reducing the factors VEGF and bFGF without influence on 3-Methyl-Glutaryl Coenzyme A reductase expression.The results demonstrated that pravastatin induces cell cycle arrest in G0/G1 and decreased production of growth factors in Multiple Myeloma cell line. PMID:26909624

  17. Glucose and Inflammatory Cells Decrease Adiponectin in Epicardial Adipose Tissue Cells: Paracrine Consequences on Vascular Endothelium.

    Science.gov (United States)

    Fernández-Trasancos, Ángel; Guerola-Segura, Raquel; Paradela-Dobarro, Beatriz; Álvarez, Ezequiel; García-Acuña, José María; Fernández, Ángel Luis; González-Juanatey, José Ramón; Eiras, Sonia

    2016-05-01

    Epicardial adipose tissue (EAT) is a source of energy for heart that expresses the insulin-sensitizer, anti-inflammatory and anti-atherogenic protein, adiponectin. But, in coronary artery disease, adiponectin production declines. Our objective was to determine its regulation by glucose and inflammation in stromal cells from EAT and subcutaneous adipose tissue (SAT) and its paracrine effect on endothelial cells. Stromal cells of EAT and SAT were obtained from patients who underwent cardiac surgery. Adipogenesis was induced at 117, 200, or 295 mg/dl glucose, with or without macrophage-conditioned medium (MCM). Expression of adiponectin, GLUT-4 and the insulin receptor was analyzed by real-time PCR. The paracrine effect of stromal cells was determined in co-cultures with endothelial cells, by exposing them to high glucose and/or MCM, and, additionally, to leukocyte-conditioned medium from patients with myocardial infarction. The endothelial response was determined by analyzing vascular adhesion molecule expression. Our results showed a U-shaped dose-response curve of glucose on adiponectin in EAT, but not in SAT stromal cells. Conversely, MCM reduced the adipogenesis-induced adiponectin expression of EAT stromal cells. The presence of EAT stromal increased the inflammatory molecules of endothelial cells. This deleterious effect was emphasized in the presence of inflammatory cell-conditioned medium from patients with myocardial infarction. Thus, high glucose and inflammatory cells reduced adipogenesis-induced adiponectin expression of EAT stromal cells, which induced an inflammatory paracrine process in endothelial cells. This inflammatory effect was lower in presence of mature adipocytes, producers of adiponectin. These results contribute to understanding the role of EAT dysfunction on coronary atherosclerosis progression. PMID:26406271

  18. EGCG decreases binding of calcium oxalate monohydrate crystals onto renal tubular cells via decreased surface expression of alpha-enolase.

    Science.gov (United States)

    Kanlaya, Rattiyaporn; Singhto, Nilubon; Thongboonkerd, Visith

    2016-06-01

    Crystal retention on tubular cell surface inside renal tubules is considered as the earliest and crucial step for kidney stone formation. Therapeutics targeting this step would cease the development of kidney stone. This study thus aimed to investigate the potential role of epigallocatechin-3-gallate (EGCG), a major antioxidant found in green tea leaves, in the reduction of calcium oxalate monohydrate (COM) crystal binding onto renal tubular cells. Pretreatment of the cells with EGCG for up to 6 h significantly diminished crystal-binding capability in a dose-dependent manner. Indirect immunofluorescence assay without and with cell permeabilization followed by laser-scanning confocal microscopy revealed that EGCG significantly reduced surface expression of alpha-enolase, whereas its intracellular level was increased. Western blot analysis confirmed such contradictory changes in membrane and cytosolic fractions of EGCG-treated cells, whereas the total level in whole cell lysate remained unchanged. Moreover, overexpression of surface alpha-enolase and enhancement of cell-crystal adhesion induced by 10 mM sodium oxalate were completely abolished by EGCG. Taken together, these data indicate that EGCG decreases binding of COM crystals onto renal tubular cells by decreasing the surface expression of alpha-enolase via re-localization or inhibition of alpha-enolase shuttling from the cytoplasm to the plasma membrane. These findings may also explain the effects of EGCG in reducing COM crystal deposition in previous animal models of kidney stone disease. Thus, EGCG may be useful for the prevention of new or recurrent stone formation. PMID:26898643

  19. Feline immunodeficiency virus decreases cell-cell communication and mitochondrial membrane potential.

    OpenAIRE

    Danave, I R; Tiffany-Castiglioni, E; Zenger, E; Barhoumi, R.; Burghardt, R C; Collisson, E W

    1994-01-01

    The in vitro effects of viral replication on mitochondrial membrane potential (MMP) and gap junctional intercellular communication (GJIC) were evaluated as two parameters of potential cellular injury. Two distinct cell types were infected with the Petaluma strain of feline immunodeficiency virus (FIV). Primary astroglia supported acute FIV infection, resulting in syncytia within 3 days of infection, whereas immortalized Crandell feline kidney (CRFK) cells of epithelial origin supported persis...

  20. T315 Decreases Acute Myeloid Leukemia Cell Viability through a Combination of Apoptosis Induction and Autophagic Cell Death

    Science.gov (United States)

    Chiu, Chang-Fang; Weng, Jing-Ru; Jadhav, Appaso; Wu, Chia-Yung; Sargeant, Aaron M.; Bai, Li-Yuan

    2016-01-01

    T315, an integrin-linked kinase (ILK) inhibitor, has been shown to suppress the proliferation of breast cancer, stomach cancer and chronic lymphocytic leukemia cells. Here we demonstrate that T315 decreases cell viability of acute myeloid leukemia (AML) cell lines (HL-60 and THP-1) and primary leukemia cells from AML patients in a dose-responsive manner. Normal human bone marrow cells are less sensitive than leukemia cells to T315. T315 down regulates protein kinase B (Akt) and p-Akt and induces caspase activation, poly-ADP-ribose polymerase (PARP) cleavage, apoptosis and autophagy through an ILK-independent manner. Interestingly, pretreatment with autophagy inhibitors rescues cells from apoptosis and concomitant PARP cleavage, which implicates a key role of autophagic cell death in T315-mediated cytotoxicity. T315 also demonstrates efficacy in vivo, suppressing the growth of THP-1 xenograft tumors in athymic nude mice when administered intraperitoneally. This study shows that autophagic cell death and apoptosis cooperatively contribute to the anticancer activity of T315 in AML cells. In conclusion, the complementary roles of apoptotic and autophagic cell death should be considered in the future assessment of the translational value of T315 in AML therapy. PMID:27537872

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

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    Sofia Baptista; Charlène Lasgi; Caroline Benstaali; Nuno Milhazes; Fernanda Borges; Carlos Fontes-Ribeiro; Fabienne Agasse; Ana Paula Silva

    2014-01-01

    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...

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

    International Nuclear Information System (INIS)

    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

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

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    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.

  4. Connexin50D47A Decreases Levels of Fiber Cell Connexins and Impairs Lens Fiber Cell Differentiation

    OpenAIRE

    Berthoud, Viviana M.; Minogue, Peter J.; Yu, Helena; Schroeder, Richard; Snabb, Joseph I.; Beyer, Eric C.

    2013-01-01

    Both heterozygous and homozygous Cx50D47A-expressing mice develop cataracts due to a severe decrease in the abundance of lens fiber connexins. The mutant mouse lenses also show impaired degradation of organelles and decreased levels of some crystallins implicating Cx50 in fiber cell differentiation.

  5. β-elemene decreases cell invasion by upregulating E-cadherin expression in MCF-7 human breast cancer cells.

    Science.gov (United States)

    Zhang, Xian; Zhang, Yang; Li, Yinghua

    2013-08-01

    Inactivation of E-cadherin results in cell migration and invasion, hence leading to cancer aggressiveness and metastasis. Downregulation of E-cadherin is closely correlated with a poor prognosis in invasive breast cancer. Thus, re-introducing E-cadherin is a novel strategy for cancer therapy. The aim of the present study was to determine the effects of the traditional Chinese medicine, β-elemene (ELE), on E-cadherin expression, cell migration and invasion in the breast cancer cell line MCF-7. MCF-7 cells were treated with 50 and 100 µg/ml ELE. E-cadherin mRNA was analyzed by reverse transcription‑polymerase chain reaction. E-cadherin protein levels were determined by immunofluorescence and western blot assays. Cell motility was measured by a Transwell assay. ELE increased both the protein and mRNA levels of E-cadherin, accompanied by decreased cell migration and invasion. Further analysis demonstrated that ELE upregulated estrogen receptor‑α (ERα) and metastasis-associated protein 3 (MTA3), and decreased the nuclear transcription factor Snail. In conclusion, our results demonstrate that ELE decreases cell migration and invasion by upregulating E-cadherin expression via controlling the ERα/MTA3/Snail signaling pathway. PMID:23732279

  6. Involvement of regulatory volume decrease in the migration of nasopharyngeal carcinoma cells

    Institute of Scientific and Technical Information of China (English)

    Jian Wen MAO; Li Xin CHEN; Li Wei WANG; Tim JACOB; Xue Rong SUN; Hui LI; Lin Yan ZHU; Pan LI; Ping ZHONG; Si Huai NIE

    2005-01-01

    The transwell chamber migration assay and CCD digital camera imaging techniques were used to investigate the relationship between regulatory volume decrease (RVD) and cell migration in nasopharyngeal carcinoma cells (CNE-2Z cells). Both migrated and non-migrated CNE-2Z cells, when swollen by 47% hypotonic solution, exhibited RVD which was inhibited by extracellular application of chloride channel blockers adenosine 5'-triphosphate (ATP), 5-nitro-2-(3-phenylpropylamino) benzoic acid (NPPB) and tamoxifen. However, RVD rate in migrated CNE-2Z cells was bigger than that of non-migrated cells and the sensitivity of migrated cells to NPPB and tamoxifen was higher than that of nonmigrated cells. ATP, NPPB and tamoxifen also inhibited migration of CNE-2Z cells. The inhibition of migration was positively correlated to the blockage of RVD, with a correlation coefficient (r) = 0.99, suggesting a functional relationship between RVD and cell migration. We conclude that RVD is involved in cell migration and RVD may play an important role in migratory process in CNE-2Z cells.

  7. LeftyA decreases Actin Polymerization and Stiffness in Human Endometrial Cancer Cells

    Science.gov (United States)

    Salker, Madhuri S.; Schierbaum, Nicolas; Alowayed, Nour; Singh, Yogesh; Mack, Andreas F.; Stournaras, Christos; Schäffer, Tilman E.; Lang, Florian

    2016-01-01

    LeftyA, a cytokine regulating stemness and embryonic differentiation, down-regulates cell proliferation and migration. Cell proliferation and motility require actin reorganization, which is under control of ras-related C3 botulinum toxin substrate 1 (Rac1) and p21 protein-activated kinase 1 (PAK1). The present study explored whether LeftyA modifies actin cytoskeleton, shape and stiffness of Ishikawa cells, a well differentiated endometrial carcinoma cell line. The effect of LeftyA on globular over filamentous actin ratio was determined utilizing Western blotting and flow cytometry. Rac1 and PAK1 transcript levels were measured by qRT-PCR as well as active Rac1 and PAK1 by immunoblotting. Cell stiffness (quantified by the elastic modulus), cell surface area and cell volume were studied by atomic force microscopy (AFM). As a result, 2 hours treatment with LeftyA (25 ng/ml) significantly decreased Rac1 and PAK1 transcript levels and activity, depolymerized actin, and decreased cell stiffness, surface area and volume. The effect of LeftyA on actin polymerization was mimicked by pharmacological inhibition of Rac1 and PAK1. In the presence of the Rac1 or PAK1 inhibitor LeftyA did not lead to significant further actin depolymerization. In conclusion, LeftyA leads to disruption of Rac1 and Pak1 activity with subsequent actin depolymerization, cell softening and cell shrinkage. PMID:27404958

  8. FAK and HAS Inhibition Synergistically Decrease Colon Cancer Cell Viability and Affect Expression of Critical Genes

    OpenAIRE

    Heffler, Melissa; Golubovskaya, Vita; Conroy, Jeffrey; Liu, Song; Wang, Dan; Cance, William; Dunn, Kelli B.

    2013-01-01

    Focal adhesion kinase (FAK), hyaluronan (HA), and hyaluronan synthase-3 (HAS3) have been implicated in cancer growth and progression. FAK inhibition with the small molecule inhibitor Y15 decreases colon cancer cell growth in vitro and in vivo. HAS3 inhibition in colon cancer cells decreases FAK expression and activation, and exogenous HA increases FAK activation. We sought to determine the genes affected by HAS and FAK inhibition and hypothesized that dual inhibition would synergistically inh...

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

    Directory of Open Access Journals (Sweden)

    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.

  10. Localized decrease of β-catenin contributes to the differentiation of human embryonic stem cells

    International Nuclear Information System (INIS)

    Human embryonic stem cells (hESC) are pluripotent, and can be directed to differentiate into different cell types for therapeutic applications. To expand hESCs, it is desirable to maintain hESC growth without differentiation. As hESC colonies grow, differentiated cells are often found at the periphery of the colonies, but the underlying mechanism is not well understood. Here, we utilized micropatterning techniques to pattern circular islands or strips of matrix proteins, and examined the spatial pattern of hESC renewal and differentiation. We found that micropatterned matrix restricted hESC differentiation at colony periphery but allowed hESC growth into multiple layers in the central region, which decreased hESC proliferation and induced hESC differentiation. In undifferentiated hESCs, β-catenin primarily localized at cell-cell junctions but not in the nucleus. The amount of β-catenin in differentiating hESCs at the periphery of colonies or in multiple layers decreased significantly at cell-cell junctions. Consistently, knocking down β-catenin decreased Oct-4 expression in hESCs. These results indicate that localized decrease of β-catenin contributes to the spatial pattern of differentiation in hESC colonies

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

    Directory of Open Access Journals (Sweden)

    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.

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

    Directory of Open Access Journals (Sweden)

    Ade Kallas

    Full Text Available Nocodazole is a known destabiliser of microtubule dynamics and arrests cell-cycle at the G2/M phase. In the context of the human embryonic stem cell (hESC it is important to understand how this arrest influences the pluripotency of cells. Here we report for the first time the changes 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 hESC 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, the presence of ROCK-2 inhibitor Y-27632 in the medium had no effect on increasing the expression of pluripotency markers Nanog and Oct4 or decreasing apoptosis or the level of p53. The expression of SSEA-3 and SSEA-4 increased in Nanog-positive cells after wash-out of nocodazole in the presence and in the absence of Y-27632. Our data show that in hESC nocodazole reversible blocks cell cycle, which is accompanied by irreversible loss of expression of pluripotency markers Nanog and Oct4.

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

    International Nuclear Information System (INIS)

    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). (paper)

  14. Decreased origin usage and initiation of DNA replication in haploinsufficient HCT116 Ku80+/- cells.

    Science.gov (United States)

    Sibani, Sahar; Price, Gerald B; Zannis-Hadjopoulos, Maria

    2005-08-01

    One of the functions of the abundant heterodimeric nuclear protein, Ku (Ku70/Ku80), is its involvement in the initiation of DNA replication through its ability to bind to chromosomal replication origins in a sequence-specific and cell cycle dependent manner. Here, using HCT116 Ku80+/- cells, the effect of Ku80 deficiency on cell cycle progression and origin activation was examined. Western blot analyses revealed a 75% and 36% decrease in the nuclear expression of Ku80 and Ku70, respectively. This was concomitant with a 33% and 40% decrease in chromatin binding of both proteins, respectively. Cell cycle analysis of asynchronous and late G1 synchronized Ku80+/- cells revealed a prolonged G1 phase. Furthermore, these Ku-deficient cells had a 4.5-, 3.4- and 4.3-fold decrease in nascent strand DNA abundance at the lamin B2, beta-globin and c-myc replication origins, respectively. Chromatin immunoprecipitation (ChIP) assays showed that the association of Ku80 with the lamin B2, beta-globin and c-myc origins was decreased by 1.5-, 2.3- and 2.5-fold, respectively, whereas that of Ku70 was similarly decreased (by 2.1-, 1.5- and 1.7-fold, respectively) in Ku80+/- cells. The results indicate that a deficiency of Ku80 resulted in a prolonged G1 phase, as well as decreased Ku binding to and activation of origins of DNA replication. PMID:16014376

  15. Human Adipose-Derived Mesenchymal Stem Cells Cryopreservation and Thawing Decrease α4-Integrin Expression

    Science.gov (United States)

    Irioda, Ana Carolina; Cassilha, Rafael; Zocche, Larissa; Francisco, Julio Cesar; Cunha, Ricardo Correa; Ferreira, Priscila Elias; Guarita-Souza, Luiz Cesar; Ferreira, Reginaldo Justino; Mogharbel, Bassam Felipe; Garikipati, Venkata Naga Srikanth; Souza, Daiany; Beltrame, Mirian Perlingeiro; de Carvalho, Katherine Athayde Teixeira

    2016-01-01

    Aim. The effects of cryopreservation on adipose tissue-derived mesenchymal stem cells are not clearly documented, as there is a growing body of evidence about the importance of adipose-derived mesenchymal stem cells for regenerative therapies. The aim of this study was to analyze human adipose tissue-derived mesenchymal stem cells phenotypic expression (CD34, CD45, CD73, CD90, CD105, and CD49d), colony forming unit ability, viability, and differentiation potential before and after cryopreservation. Materials and Methods. 12 samples of the adipose tissue were collected from a healthy donor using the liposuction technique. The cell isolation was performed by enzymatic digestion and then the cells were cultured up to passage 2. Before and after cryopreservation the immunophenotype, cellular viability analysis by flow cytometer, colony forming units ability, differentiation potential into adipocytes and osteoblasts as demonstrated by Oil Red O and Alizarin Red staining, respectively. Results. The immunophenotypic markers expression was largely preserved, and their multipotency was maintained. However, after cryopreservation, the cells decreased α4-integrin expression (CD49d), cell viability, and number of colony forming units. Conclusions. These findings suggest that ADMSC transplanted after cryopreservation might compromise the retention of transplanted cells in the host tissue. Therefore, further studies are warranted to standardize protocols related to cryopreservation to attain full benefits of stem cell therapy. PMID:26981129

  16. Human Adipose-Derived Mesenchymal Stem Cells Cryopreservation and Thawing Decrease α4-Integrin Expression

    Directory of Open Access Journals (Sweden)

    Ana Carolina Irioda

    2016-01-01

    Full Text Available Aim. The effects of cryopreservation on adipose tissue-derived mesenchymal stem cells are not clearly documented, as there is a growing body of evidence about the importance of adipose-derived mesenchymal stem cells for regenerative therapies. The aim of this study was to analyze human adipose tissue-derived mesenchymal stem cells phenotypic expression (CD34, CD45, CD73, CD90, CD105, and CD49d, colony forming unit ability, viability, and differentiation potential before and after cryopreservation. Materials and Methods. 12 samples of the adipose tissue were collected from a healthy donor using the liposuction technique. The cell isolation was performed by enzymatic digestion and then the cells were cultured up to passage 2. Before and after cryopreservation the immunophenotype, cellular viability analysis by flow cytometer, colony forming units ability, differentiation potential into adipocytes and osteoblasts as demonstrated by Oil Red O and Alizarin Red staining, respectively. Results. The immunophenotypic markers expression was largely preserved, and their multipotency was maintained. However, after cryopreservation, the cells decreased α4-integrin expression (CD49d, cell viability, and number of colony forming units. Conclusions. These findings suggest that ADMSC transplanted after cryopreservation might compromise the retention of transplanted cells in the host tissue. Therefore, further studies are warranted to standardize protocols related to cryopreservation to attain full benefits of stem cell therapy.

  17. Wuweizisu C from Schisandra chinensis decreases membrane potential in C6 glioma cells

    Institute of Scientific and Technical Information of China (English)

    Young-whan CHOI; Kyeok KIM; Ji-yeong JO; Hyo-lim KIM; You-jin LEE; Woo-jung SHIN; Santosh J SACKET; Mijin HAN; Dong-soon IM

    2008-01-01

    Aim:To study the effects of dibenzocyclooctadiene lignans isolated from Schi-sandra chinensis, such as wuweizisu C, gomisin N, gomisin A, and schisandrin, on the membrane potential in C6 glioma cells. Methods: The membrane po-tential was estimated by measuring the fluorescence change in DiBAC-loaded glioma cells. Results: Wuweizisu C decreased the membrane potential in a concentration-dependent manner. Gomisin N and gomisin A, however, showed differential modulation and no change was induced by schisandrin or dimethyl-4,4'-dimethoxy-5,6,5',6'-dimethylene dioxybiphenyl-2,2'-dicarboxylate, a syn-thetic drug derived from dibenzocyclooctadiene lignans. We found no involve-ment of Gi/o proteins, phospholipase C, and extracellular Na+ on the wuweizisu C-indueed decrease of the membrane potential. Wuweizisu C by itself did not change the intracellular Ca2+ [Ca2+]I concentration, but decreased the ATP-indu-ted Ca2+ increase in C6 glioma cells. The 4 lignans at all concentrations used in this study did not induce any effect on cell viability. Furthermore, we found a similar decrease of the membrane potential by wuweizisu C in PC12 neuronal cells. Conclusion: Our results suggest that the decrease in the membrane poten-tial and the modulation of [Ca2+]I concentration by wuweizisu C could be impor-tant action mechanisms ofwuweizisu C.

  18. Carbon nanotubes enhance the internalization of drugs by cancer cells and decrease their chemoresistance to cytostatics

    International Nuclear Information System (INIS)

    Etoposide is a semisynthetic, chemotherapeutic drug widely recommended to treat an extensive range of human cancers. Our studies indicate that, while etoposide is capable of killing human cancer cells, exposure to single-walled carbon nanotubes (SWCNTs) and etoposide results in enhanced cell death that appears to be synergistic and not merely additive. In this study, we used high pressure liquid chromatography and mass spectrometry to quantify the internal effective dose of etoposide when the human pancreatic cancer cell (PANC-1) was exposed to the combination of these agents. Our results unequivocally indicate that SWCNTs improve etoposide uptake and increase its capacity to kill cancer cells. We suggest that a combination of SWCNTs and etoposide may prove to be a more efficient chemotherapeutic protocol, especially because of the potential to lower toxic drug doses to levels that may be useful in decreasing adverse side effects, as well as in lowering the probability of inducing chemoresistance in exposed cancer cells. (paper)

  19. Endothelin receptor B antagonists decrease glioma cell viability independently of their cognate receptor

    International Nuclear Information System (INIS)

    Endothelin receptor antagonists inhibit the progression of many cancers, but research into their influence on glioma has been limited. We treated glioma cell lines, LN-229 and SW1088, and melanoma cell lines, A375 and WM35, with two endothelin receptor type B (ETRB)-specific antagonists, A-192621 and BQ788, and quantified viable cells by the capacity of their intracellular esterases to convert non-fluorescent calcein AM into green-fluorescent calcein. We assessed cell proliferation by labeling cells with carboxyfluorescein diacetate succinimidyl ester and quantifying the fluorescence by FACS analysis. We also examined the cell cycle status using BrdU/propidium iodide double staining and FACS analysis. We evaluated changes in gene expression by microarray analysis following treatment with A-192621 in glioma cells. We examined the role of ETRB by reducing its expression level using small interfering RNA (siRNA). We report that two ETRB-specific antagonists, A-192621 and BQ788, reduce the number of viable cells in two glioma cell lines in a dose- and time-dependent manner. We describe similar results for two melanoma cell lines. The more potent of the two antagonists, A-192621, decreases the mean number of cell divisions at least in part by inducing a G2/M arrest and apoptosis. Microarray analysis of the effects of A-192621 treatment reveals up-regulation of several DNA damage-inducible genes. These results were confirmed by real-time RT-PCR. Importantly, reducing expression of ETRB with siRNAs does not abrogate the effects of either A-192621 or BQ788 in glioma or melanoma cells. Furthermore, BQ123, an endothelin receptor type A (ETRA)-specific antagonist, has no effect on cell viability in any of these cell lines, indicating that the ETRB-independent effects on cell viability exhibited by A-192621 and BQ788 are not a result of ETRA inhibition. While ETRB antagonists reduce the viability of glioma cells in vitro, it appears unlikely that this effect is mediated by

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

    OpenAIRE

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

    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...

  1. Plakophilin3 downregulation leads to a decrease in cell adhesion and promotes metastasis.

    Science.gov (United States)

    Kundu, Samrat T; Gosavi, Prajakta; Khapare, Nileema; Patel, Rachana; Hosing, Amol S; Maru, Girish B; Ingle, Arvind; Decaprio, James A; Dalal, Sorab N

    2008-11-15

    Plakophilin3 is a desmosomal plaque protein whose levels are reduced in poorly differentiated tumors of the oropharyngeal cavity and in invasive colon carcinomas. To test the hypothesis that plakophilin3 loss stimulates neoplastic progression, plakophilin3 expression was inhibited by DNA vector driven RNA interference in 3 epithelial cell lines, HCT116, HaCaT and fetal buccal mucosa. The plakophilin3-knockdown clones showed a decrease in cell-cell adhesion as assessed in a hanging drop assay, which was accompanied by an increase in cell migration. The HCT116 plakophilin3-knockdown clones showed a decrease in desmosome size as revealed by electron microscopy. These altered desmosomal properties were accompanied by colony formation in soft agar and growth to high density in culture. The HCT116-derived clones showed accelerated tumor formation in nude mice and increased metastasis to the lung, a phenotype consistent with the increased migration observed in vitro and is consistent with data from human tumors that suggests that plakophililn3 is lost in invasive and metastatic tumors. These data indicate that plakophilin3 loss leads to a decrease in cell-cell adhesion leading to the stimulation of neoplastic progression and metastasis. PMID:18729189

  2. 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.

  3. Sudden collapse of vacuoles in Saintpaulia sp. palisade cells induced by a rapid temperature decrease.

    Science.gov (United States)

    Kadohama, Noriaki; Goh, Tatsuaki; Ohnishi, Miwa; Fukaki, Hidehiro; Mimura, Tetsuro; Suzuki, Yoshihiro

    2013-01-01

    It is well known that saintpaulia leaf is damaged by the rapid temperature decrease when cold water is irrigated onto the leaf surface. We investigated this temperature sensitivity and the mechanisms of leaf damage in saintpaulia (Saintpaulia sp. cv. 'Iceberg') and other Gesneriaceae plants. Saintpaulia leaves were damaged and discolored when subjected to a rapid decrease in temperature, but not when the temperature was decreased gradually. Sensitivity to rapid temperature decrease increased within 10 to 20 min during pre-incubation at higher temperature. Injury was restricted to the palisade mesophyll cells, where there was an obvious change in the color of the chloroplasts. During a rapid temperature decrease, chlorophyll fluorescence monitored by a pulse amplitude modulated fluorometer diminished and did not recover even after rewarming to the initial temperature. Isolated chloroplasts were not directly affected by the rapid temperature decrease. Intracellular pH was monitored with a pH-dependent fluorescent dye. In palisade mesophyll cells damaged by rapid temperature decrease, the cytosolic pH decreased and the vacuolar membrane collapsed soon after a temperature decrease. In isolated chloroplasts, chlorophyll fluorescence declined when the pH of the medium was lowered. These results suggest that a rapid temperature decrease directly or indirectly affects the vacuolar membrane, resulting in a pH change in the cytosol that subsequently affects the chloroplasts in palisade mesophyll cells. We further confirmed that the same physiological damage occurs in other Gesneriaceae plants. These results strongly suggested that the vacuoles of palisade mesophyll cells collapsed during the initial phase of leaf injury. PMID:23451194

  4. Sudden collapse of vacuoles in Saintpaulia sp. palisade cells induced by a rapid temperature decrease.

    Directory of Open Access Journals (Sweden)

    Noriaki Kadohama

    Full Text Available It is well known that saintpaulia leaf is damaged by the rapid temperature decrease when cold water is irrigated onto the leaf surface. We investigated this temperature sensitivity and the mechanisms of leaf damage in saintpaulia (Saintpaulia sp. cv. 'Iceberg' and other Gesneriaceae plants. Saintpaulia leaves were damaged and discolored when subjected to a rapid decrease in temperature, but not when the temperature was decreased gradually. Sensitivity to rapid temperature decrease increased within 10 to 20 min during pre-incubation at higher temperature. Injury was restricted to the palisade mesophyll cells, where there was an obvious change in the color of the chloroplasts. During a rapid temperature decrease, chlorophyll fluorescence monitored by a pulse amplitude modulated fluorometer diminished and did not recover even after rewarming to the initial temperature. Isolated chloroplasts were not directly affected by the rapid temperature decrease. Intracellular pH was monitored with a pH-dependent fluorescent dye. In palisade mesophyll cells damaged by rapid temperature decrease, the cytosolic pH decreased and the vacuolar membrane collapsed soon after a temperature decrease. In isolated chloroplasts, chlorophyll fluorescence declined when the pH of the medium was lowered. These results suggest that a rapid temperature decrease directly or indirectly affects the vacuolar membrane, resulting in a pH change in the cytosol that subsequently affects the chloroplasts in palisade mesophyll cells. We further confirmed that the same physiological damage occurs in other Gesneriaceae plants. These results strongly suggested that the vacuoles of palisade mesophyll cells collapsed during the initial phase of leaf injury.

  5. Essential oil of clove (Eugenia caryophyllata) augments the humoral immune response but decreases cell mediated immunity.

    Science.gov (United States)

    Halder, Sumita; Mehta, Ashish K; Mediratta, Pramod K; Sharma, Krishna K

    2011-08-01

    The present study was undertaken to explore the effect of the essential oil isolated from the buds of Eugenia caryophyllata on some immunological parameters. Humoral immunity was assessed by measuring the hemagglutination titre to sheep red blood cells and delayed type hypersensitivity was assessed by measuring foot pad thickness. Clove oil administration produced a significant increase in the primary as well as secondary humoral immune response. In addition, it also produced a significant decrease in foot pad thickness compared with the control group. Thus, these results suggest that clove oil can modulate the immune response by augmenting humoral immunity and decreasing cell mediated immunity. PMID:21796701

  6. 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.

  7. Analysis of Hereditary Elliptocytosis with Decreased Binding of Eosin-5-maleimide to Red Blood Cells

    Directory of Open Access Journals (Sweden)

    Shin-ichiro Suemori

    2015-01-01

    Full Text Available Flow cytometric test for analyzing the eosin-5-maleimide (EMA binding to red blood cells has been believed to be a specific method for diagnosing hereditary spherocytosis (HS. However, it has been reported that diseases other than HS, such as hereditary pyropoikilocytosis (HPP and Southeast Asian ovalocytosis (SAO, which are forms in the category of hereditary elliptocytosis (HE, show decreased EMA binding to red blood cells. We analyzed EMA binding to red blood cells in 101 healthy control subjects and 42 HS patients and obtained a mean channel fluorescence (MCF cut-off value of 36.4 (sensitivity 0.97, specificity 0.95. Using this method, we also analyzed 12 HE patients. Among them, four HE patients showed the MCF at or below the cut-off value. It indicates that some HE patients have decreased EMA binding to red blood cells. Two of these four HE patients were classified as common HE, and two were spherocytic HE with reduced spectrin. This study demonstrates that, in addition to patients with HPP or SAO, some HE patients have decreased EMA binding to red blood cells.

  8. Decreased expression of mucin 18 is associated with unfavorable postoperative prognosis in patients with clear cell renal cell carcinoma

    OpenAIRE

    Bai, Qi; Liu, Li; Long, Qilai; Xia, Yu; Wang, Jiajun; Xu, Jiejie; Guo, Jianming

    2015-01-01

    Background: MUC18 is correlated with tumor progression and metastasis in types of malignancy. But the role of MUC18 in clear cell renal cell carcinoma remains unclear. In this study, we aimed to investigate the expression of MUC18 and its correlation with clinical outcomes in clear cell renal cell carcinoma. Patients and Methods: Immunohistochemical staining was performed in samples from 288 patients with clear cell renal cell carcinoma. We used Kaplan-Meier method and Cox proportional hazard...

  9. Ethanol Extracts of Selected Cyathea Species Decreased Cell Viability and Inhibited Growth in MCF 7 Cell Line Cultures.

    Science.gov (United States)

    Janakiraman, Narayanan; Johnson, Marimuthu

    2016-06-01

    Cancer is the cause of more than 6 million deaths worldwide every year. For centuries, medicinal plants have been used in the treatment of cancer. Chemotherapy, radiotherapy, surgery and acupuncture point stimulation are also used to treat cancer. The present study was intended to reveal the cytotoxic and anticancer potential of selected Cyathea species and to highlight their importance in the pharmaceutical industry for the development of cost-effective drugs. Cytotoxic studies using brine shrimp lethality bioassays and MCF 7 cell line cultures were carried out. Compared to petroleum ether, chloroform and acetone extracts, the ethanol extracts of selected Cyathea species were found to be more effective against brine shrimps. The ethanol extracts were further subjected to 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) cell proliferation assays. A decrease in cell viability and an increase in growth inhibition were observed for the MCF 7 cell line. The maximum percentage of cell inhibition was observed in Cyathea crinit, followed by Cyathea nilgirensis and Cyathea gigantea. The results of the present study suggest that Cyathea species are an effective source of cytotoxic compounds. PMID:27342889

  10. Low-dose testosterone treatment decreases oxidative damage in TM3 Leydig cells

    Institute of Scientific and Technical Information of China (English)

    Thomas IS Hwang; Tien-Ling Liao; Ji-Fan Lin; Yi-Chia Lin; Shu-Yu Lee; Yen-Chun Lai; Shu-Huei Kao

    2011-01-01

    Testosterone replacement therapy has benefits for aging men and those with hypogonadism. However, the effects of exogenous testosterone on Leydig cells are still unclear and need to be clarified. In this report, we demonstrate that testosterone supplementation can reduce oxidative damage in Leydig cells. The TM3 Leydig cell line was used as an in vitro cell model in this study. Cytoprotective effects were identified with 100-nmol l-1 testosterone treatment, but cytotoxic effects were found with ≥ 500-nmol l-1 testosterone supplementation. Significantly reduced reactive oxygen species (ROS) generation, lipid peroxide contents and hypoxia induction factor (HIF)-1α stabilization and activation were found with 100-nmol l-1 testosterone treatment. There was a 1.72-fold increase in ROS generation in the 500-nmol l-1 compared to the 100-nmol l-1 testosterone treatment. A 1.58-fold increase in steroidogenic acute regulatory protein (StAR) expression was found in 50-nmol l-1 testosterone-treated cells (P<0.01). Chemically induced hypoxia was attenuated by testosterone supplementation. Leydig cells treated with low-dose testosterone supplementation showed cytoprotection by decreasing ROS and lipid peroxides, increasing StAR expression and relieving hypoxia stress as demonstrated by HIF-1α stabilization. Increased oxidative damage was found with ≥ 500-nmol l-1 testosterone manipulation. The mechanism governing the differential dose effects of testosterone on Leydig cells needs further investigation in order to shed light on testosterone replacement therapy.

  11. 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 maxi...

  12. Impact of selection for decreased somatic cell score on productive life and culling for mastitis

    Science.gov (United States)

    Impact of continued selection for decreased somatic cell score (SCS) was examined to determine if such selection resulted in greater mastitis susceptibility and shorter productive life (PL). Holstein artificial-insemination bulls with a predicted transmitting ability (PTA) for SCS based on >=35 daug...

  13. Decreased chicken ovalbumin upstream promoter transcription factor II expression in tamoxifen-resistant breast cancer cells.

    Science.gov (United States)

    Riggs, Krista A; Wickramasinghe, Nalinie S; Cochrum, Renate K; Watts, Mary Beth; Klinge, Carolyn M

    2006-10-15

    Tamoxifen (TAM) is successfully used for the treatment and prevention of breast cancer. However, many patients that are initially TAM responsive develop tumors that are antiestrogen/TAM resistant (TAM-R). The mechanism behind TAM resistance in estrogen receptor alpha (ERalpha)-positive tumors is not understood. The orphan nuclear receptor chicken ovalbumin upstream promoter transcription factor (COUP-TF)-I interacts directly with 4-hydroxytamoxifen (4-OHT)- and estradiol (E(2))-occupied ERalpha, corepressors NCoR and SMRT, and inhibit E(2)-induced gene transcription in breast cancer cells. Here we tested the hypothesis that reduced COUP-TFI and COUP-TFII correlate with TAM resistance. We report for the first time that COUP-TFII, but not COUP-TFI, is reduced in three antiestrogen/TAM-R cell lines derived from TAM-sensitive (TAM-S) MCF-7 human breast cancer cells and in MDA-MB-231 cells compared with MCF-7. ERalpha and ERbeta protein expression was not different between TAM-S and TAM-R cells, but progesterone receptor (PR) was decreased in TAM-R cells. Further, E(2) increased COUP-TFII transcription in MCF-7, but not TAM-R, cells. Importantly, reexpression of COUP-TFII in TAM-S cells to levels comparable to those in MCF-7 was shown to increase 4-OHT-mediated growth inhibition and increased apoptosis. Conversely, knockdown of COUP-TFII in TAM-S MCF-7 cells blocked growth inhibitory activity and increased 4-OHT agonist activity. 4-OHT increased COUP-TFII-ERalpha interaction approximately 2-fold in MCF-7 cells. COUP-TFII expression in TAM-R cells also inhibited 4-OHT-induced endogenous PR and pS2 mRNA expression. These data indicate that reduced COUP-TFII expression correlates with acquired TAM resistance in human breast cancer cell lines and that COUP-TFII plays a role in regulating the growth inhibitory activity of TAM in breast cancer cells. PMID:17047084

  14. Radiation causes increased production and decreased utilization of IL-2 in human mononuclear cells

    International Nuclear Information System (INIS)

    The effects of radiation on the kinetics of Interleukin-2 (IL-2) production and utilization by mononuclear cells (MNCs) were studied. Mononuclear cells from normal, healthy individuals were subjected to various doses of radiation ranging from 0 to 2,000 rad and cultured in the presence of PHA. Supernatants from these cultures were harvested at various periods and their IL-2 contents determined by both the standard bioassay and ELISA. A radiation dose of 800 rad and higher had a marked effect on both IL-2 production and consumption. Although the supernatants from both the irradiated and non-irradiated MNCs contained maximal concentrations of IL-2 between 8 and 24 h of culture, the former had three times as much IL-2 as the latter. An increase in IL-2-mRNA levels was also noticed in irradiated, PHA-stimulated cells. Moreover, the supernatants from irradiated MNCs collected as late as 72 h after the initiation of culture contained more than 30% of the total IL-2 produced compared to less than 8% in supernatants from non-irradiated cells. Supernatants from non-irradiated cells incubated further with irradiated cells contained relatively higher quantities of IL-2 than those incubated continuously with non-irradiated cells. Supernatants from co-cultures of irradiated and non-irradiated MNCs contained less than expected amounts of IL-2 in two of the three subjects. Despite a difference in both the production and consumption of IL-2 between the irradiated and non-irradiated cells, there was no difference in their ability to generate IL-2 receptors. The results indicate that inactivation of radiosensitive suppressor T cells is associated with superinduction of IL-2 mRNA, increased production and decreased consumption of IL-2 by MNCs, thereby resulting in increased accumulation of IL-2

  15. 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.

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

    Directory of Open Access Journals (Sweden)

    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.

  17. Age-related decrease in rod bipolar cell density of the human retina: an immunohistochemical study

    Indian Academy of Sciences (India)

    P Aggarwal; T C Nag; S Wadhwa

    2007-03-01

    During normal ageing, the rods (and other neurones) undergo a significant decrease in density in the human retina from the fourth decade of life onward. Since the rods synapse with the rod bipolar cells in the outer plexiform layer, a decline in rod density (mainly due to death) may ultimately cause an associated decline of the neurones which, like the rod bipolar cells, are connected to them. The rod bipolar cells are selectively stained with antibodies to protein kinase C-. This study examined if rod bipolar cell density changes with ageing of the retina, utilizing donor human eyes (age: 6–91 years). The retinas were fixed and their temporal parts from the macula to the mid-periphery sectioned and processed for protein kinase C- immunohistochemistry. The density of the immunopositive rod bipolar cells was estimated in the mid-peripheral retina (eccentricity: 3–5 mm) along the horizontal temporal axis. The results show that while there is little change in the density of the rod bipolar cells from 6 to 35 years (2.2%), the decline during the period from 35 to 62 years is about 21% and between seventh and tenth decades, it is approximately 27%.

  18. Mesenchymal Stem Cells Promote Mammosphere Formation and Decrease E-Cadherin in Normal and Malignant Breast Cells

    OpenAIRE

    Klopp, Ann H; Lacerda, Lara; Gupta, Anshul; Debeb, Bisrat G; Solley, Travis; Li, Li; Spaeth, Erika; Xu, Wei; Zhang, Xiaomei; Lewis, Michael T.; Reuben, James M.; Krishnamurthy, Savitri; Ferrari, Mauro; Gaspar, Rogério; Buchholz, Thomas A.

    2010-01-01

    Introduction Normal and malignant breast tissue contains a rare population of multi-potent cells with the capacity to self-renew, referred to as stem cells, or tumor initiating cells (TIC). These cells can be enriched by growth as “mammospheres” in three-dimensional cultures. Objective We tested the hypothesis that human bone-marrow derived mesenchymal stem cells (MSC), which are known to support tumor growth and metastasis, increase mammosphere formation. Results We found that MSC increased ...

  19. 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.

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

    International Nuclear Information System (INIS)

    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

  1. 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.

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

    International Nuclear Information System (INIS)

    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

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

    Directory of Open Access Journals (Sweden)

    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.

  4. 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....

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

    Directory of Open Access Journals (Sweden)

    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.

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

    International Nuclear Information System (INIS)

    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, ∼ 25 and ∼ 40 μg/dL, respectively, on PN10 and by PN30 all were ≤ 1 μg/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: ► Peak [BPb] in control, low-, moderate- and high-dose newborn mice with gestational lead exposure: ≤ 1, ≤ 10, 25 and 40 μg/dL ► Gestational lead exposure dose-dependently decreased the number of TH

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

    Energy Technology Data Exchange (ETDEWEB)

    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

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

    Directory of Open Access Journals (Sweden)

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

    Science.gov (United States)

    Ercan, Ertugrul; Bozdemir, Huseyin; Tengiz, Istemihan; Sekuri, Cevad; Aliyev, Emil; Akilli, Azem; Akin, Mustafa

    2004-01-01

    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. PMID:15059285

  10. Decrease in Circulating Dendritic Cell Precursors in Patients with Peripheral Artery Disease

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    Daniel Kretzschmar

    2015-01-01

    Full Text Available Peripheral artery disease (PAD is a common manifestation of atherosclerosis. Inflammation is important for initiation and progression of the disease. Dendritic cells (DCs as antigen-presenting cells play an important role in the immune system. Therefore, we hypothesize that, in patients with PAD, DCPs might be reduced in blood due to their recruitment into the vascular wall and induce a proinflammatory response. The numbers of myeloid DCPs, plasmacytoid DCPs, and total DCPs were analyzed by flow cytometry in blood of patients with PAD (n=52 compared to controls (n=60. Femoralis plaques (n=12 of patients who underwent surgery were immunostained for CD209 and CD83 (mDCs as well as CD304, CD123 (pDCs, and HLA-DR. In patients with PAD, a significant decrease in mDCPs, pDCPs, and tDCPs was observed. In immunostaining, markers indicative for mDCs (CD209: 16 versus 8 cells/0.1 mm2, P=0.02; CD83: 19 versus 5 cells/0.1 mm2, P=0.03 were significantly elevated in femoralis plaques compared to control vessels. We show for the first time that mDCPs, pDCPs, and tDCPs are significantly reduced in patients with PAD. Immunohistochemical analysis unraveled that the decrease in DCPs might be due to their recruitment into atherosclerotic plaques.

  11. Electroporation transiently decreases GJB2 (connexin 26) expression in B16/BL6 melanoma cell line.

    Science.gov (United States)

    Rangel, Marcelo Monte Mór; Chaible, Lucas Martins; Nagamine, Marcia Kazumi; Mennecier, Gregory; Cogliati, Bruno; de Oliveira, Krishna Duro; Fukumasu, Heidge; Sinhorini, Idércio Luiz; Mir, Lluis Maria; Dagli, Maria Lúcia Zaidan

    2015-02-01

    Connexins are proteins that form gap junctions. Perturbations in the cell membrane reportedly promote changes in the expression profile of connexins. Electroporation promotes destabilization by applying electrical pulses, and this procedure is used in electrochemotherapy and gene therapy, among others. This in vitro work aimed to study the interference of electroporation on the expression profile of GJB2 (Cx26 gene) and Connexin 26 in melanoma cell line B16/BL6. The techniques of immunocytochemistry, Western blot, and real-time PCR were used. After electroporation, cells showed a transient decrease in GJB2 mRNA. The immunostaining of Cx26 showed no noticeable change after electroporation at different time points. However, Western blot showed a significant reduction in Cx26 30 min after electroporation. Our results showed that electroporation interferes transiently in the expression of Connexin 26 in melanoma and are consistent with the idea that electroporation is a process of intense stress that promotes cell homeostatic imbalance and results in disruption of cell physiological processes such as transcription and translation. PMID:25298064

  12. Enhanced survival and decreased mutation frequency after photoreactivation of UV damage in rat kangaroo cells

    International Nuclear Information System (INIS)

    The effect of pyrimidine dimers on cytotoxicity, DNA repair and mutagenesis was studied in cells, derived from the rat kangaroo, which possess photoreactivating capabilities. A significant enhancement in colony-forming ability was achieved after UV irradiation in exponentially growing cells if photoreactivating light treatment followed the UV irradiation. If photoreactivation treatment was delayed 24h after UV irradiation, no significant increase in survival was observed. Assays of pyrimidine dimers, unscheduled DNA synthesis, and survival in contact-inhibited cells all confirmed a minor role of dark excision repair and a major role of photoreactivation. Photoreactivation decreased the frequency of mutations to 6-thioguanine resistance to a greater extent than the alteration seen in survival. Approximately 1.6 times the dose must be given to get equal killing in photoreactivated cells, whereas 4 times the dose must be given to obtain equal mutation frequencies in light-treated cells. This suggests that the removal of dimers is more effective in mutant reduction than enhancement of survival. (orig.)

  13. Enhanced survival and decreased mutation frequency after photoreactivation of UV damage in rat kangaroo cells

    Energy Technology Data Exchange (ETDEWEB)

    Wade, M.H.; Trosko, J.E. (Michigan State Univ., East Lansing (USA). Dept. of Pediatrics and Human Development)

    1983-08-01

    The effect of pyrimidine dimers on cytotoxicity, DNA repair and mutagenesis was studied in cells, derived from the rat kangaroo, which possess photoreactivating capabilities. A significant enhancement in colony-forming ability was achieved after UV irradiation in exponentially growing cells if photoreactivating light treatment followed the UV irradiation. If photoreactivation treatment was delayed 24h after UV irradiation, no significant increase in survival was observed. Assays of pyrimidine dimers, unscheduled DNA synthesis, and survival in contact-inhibited cells all confirmed a minor role of dark excision repair and a major role of photoreactivation. Photoreactivation decreased the frequency of mutations to 6-thioguanine resistance to a greater extent than the alteration seen in survival. Approximately 1.6 times the dose must be given to get equal killing in photoreactivated cells, whereas 4 times the dose must be given to obtain equal mutation frequencies in light-treated cells. This suggests that the removal of dimers is more effective in mutant reduction than enhancement of survival.

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

    International Nuclear Information System (INIS)

    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.

  15. Decreased Polysaccharide Feruloylation Compromises Plant Cell Wall Integrity and Increases Susceptibility to Necrotrophic Fungal Pathogens

    Science.gov (United States)

    Reem, Nathan T.; Pogorelko, Gennady; Lionetti, Vincenzo; Chambers, Lauran; Held, Michael A.; Bellincampi, Daniela; Zabotina, Olga A.

    2016-01-01

    The complexity of cell wall composition and structure determines the strength, flexibility, and function of the primary cell wall in plants. However, the contribution of the various components to cell wall integrity (CWI) and function remains unclear. Modifications of cell wall composition can induce plant responses known as CWI control. In this study, we used transgenic expression of the fungal feruloyl esterase AnFAE to examine the effect of post-synthetic modification of Arabidopsis and Brachypodium cell walls. Transgenic Arabidopsis plants expressing AnFAE showed a significant reduction of monomeric ferulic acid, decreased amounts of wall-associated extensins, and increased susceptibility to Botrytis cinerea, compared with wild type. Transgenic Brachypodium showed reductions in monomeric and dimeric ferulic acids and increased susceptibility to Bipolaris sorokiniana. Upon infection, transgenic Arabidopsis and Brachypodium plants also showed increased expression of several defense-related genes compared with wild type. These results demonstrate a role, in both monocot and dicot plants, of polysaccharide feruloylation in plant CWI, which contributes to plant resistance to necrotrophic pathogens. PMID:27242834

  16. Decreased cell adhesion promotes angiogenesis in a Pyk2-dependent manner

    International Nuclear Information System (INIS)

    Angiogenesis is regulated by both soluble growth factors and cellular interactions with the extracellular matrix (ECM). While cell adhesion via integrins has been shown to be required for angiogenesis, the effects of quantitative changes in cell adhesion and spreading against the ECM remain less clear. Here, we show that angiogenic sprouting in natural and engineered three-dimensional matrices exhibited a biphasic response, with peak sprouting when adhesion to the matrix was limited to intermediate levels. Examining changes in global gene expression to determine a genetic basis for this response, we demonstrate a vascular endothelial growth factor (VEGF)-induced upregulation of genes associated with vascular invasion and remodeling when cell adhesion was limited, whereas cells on highly adhesive surfaces upregulated genes associated with proliferation. To explore a mechanistic basis for this effect, we turned to focal adhesion kinase (FAK), a central player in adhesion signaling previously implicated in angiogenesis, and its homologue, proline-rich tyrosine kinase 2 (Pyk2). While FAK signaling had some impact, our results suggested that Pyk2 can regulate both gene expression and endothelial sprouting through its enhanced activation by VEGF in limited adhesion contexts. We also demonstrate decreased sprouting of tissue explants from Pyk2-null mice as compared to wild type mice as further confirmation of the role of Pyk2 in angiogenic sprouting. These results suggest a surprising finding that limited cell adhesion can enhance endothelial responsiveness to VEGF and demonstrate a novel role for Pyk2 in the adhesive regulation of angiogenesis.

  17. Decreased Polysaccharide Feruloylation Compromises Plant Cell Wall Integrity and Increases Susceptibility to Necrotrophic Fungal Pathogens.

    Science.gov (United States)

    Reem, Nathan T; Pogorelko, Gennady; Lionetti, Vincenzo; Chambers, Lauran; Held, Michael A; Bellincampi, Daniela; Zabotina, Olga A

    2016-01-01

    The complexity of cell wall composition and structure determines the strength, flexibility, and function of the primary cell wall in plants. However, the contribution of the various components to cell wall integrity (CWI) and function remains unclear. Modifications of cell wall composition can induce plant responses known as CWI control. In this study, we used transgenic expression of the fungal feruloyl esterase AnFAE to examine the effect of post-synthetic modification of Arabidopsis and Brachypodium cell walls. Transgenic Arabidopsis plants expressing AnFAE showed a significant reduction of monomeric ferulic acid, decreased amounts of wall-associated extensins, and increased susceptibility to Botrytis cinerea, compared with wild type. Transgenic Brachypodium showed reductions in monomeric and dimeric ferulic acids and increased susceptibility to Bipolaris sorokiniana. Upon infection, transgenic Arabidopsis and Brachypodium plants also showed increased expression of several defense-related genes compared with wild type. These results demonstrate a role, in both monocot and dicot plants, of polysaccharide feruloylation in plant CWI, which contributes to plant resistance to necrotrophic pathogens. PMID:27242834

  18. Force via integrins but not E-cadherin decreases Oct3/4 expression in embryonic stem cells

    International Nuclear Information System (INIS)

    Highlights: ► Force via integrins or cadherins induces similar cell stiffening responses. ► Force via integrins but not cadherins induces cell spreading. ► Force via integrins but not cadherins induces differentiation of embryonic stem cells. -- Abstract: Increasing evidence suggests that mechanical factors play a critical role in fate decisions of stem cells. Recently we have demonstrated that a local force applied via Arg-Gly-Asp (RGD) peptides coated magnetic beads to mouse embryonic stem (ES) cells increases cell spreading and cell stiffness and decreases Oct3/4 (Pou5f1) gene expression. However, it is not clear whether the effects of the applied stress on these functions of ES cells can be extended to natural extracellular matrix proteins or cell–cell adhesion molecules. Here we show that a local cyclic shear force applied via fibronectin or laminin to integrin receptors increased cell spreading and stiffness, downregulated Oct3/4 gene expression, and decreased cell proliferation rate. In contrast, the same cyclic force applied via cell–cell adhesion molecule E-cadherin (Cdh1) had no effects on cell spreading, Oct3/4 gene expression, and the self-renewal of mouse ES cells, but induced significant cell stiffening. Our findings demonstrate that biological responses of ES cells to force applied via integrins are different from those to force via E-cadherin, suggesting that mechanical forces might play different roles in different force transduction pathways to shape early embryogenesis.

  19. Role of mitochondrial DNA decrease in apoptosis of human bronchial epithelial cells induced by radon and its progeny

    International Nuclear Information System (INIS)

    Human bronchia epithelia with mtDNA decrease (ρ-HBE) cells generated by treatment of ethidium bromide (EB) were exposed to radon gas in a special inhalation chamber. Cell proliferation was determined by cell survival assay Cell apoptosis and membrane potential of mitochondria were analyzed by flow cytometry. The results showed that the survival fraction of ρ-HBE cells significantly increased compared with that of ρ+ HBE cells after irradiation with radon and its progeny. Although the apoptosis rate of p HBE cells was lower than that of the ρ+ HBE cells at early period, the total apoptosis rate was increased, along with the membrane potential decrease of mitochondria in ρ-HBE cells. The results indicate that the increased potential of ρ-HBE proliferation correlates with the total apoptotic rate and mitochondrial membrane potential. (authors)

  20. Serum metabolites of proanthocyanidin-administered rats decrease lipid synthesis in HepG2 cells.

    Science.gov (United States)

    Guerrero, Ligia; Margalef, Maria; Pons, Zara; Quiñones, Mar; Arola, Lluis; Arola-Arnal, Anna; Muguerza, Begoña

    2013-12-01

    The regular consumption of flavonoids has been associated with reduced mortality and a decreased risk of cardiovascular diseases. The proanthocyanidins found in plasma are very different from the original flavonoids in food sources. The use of physiologically appropriate conjugates of proanthocyanidins is essential for the in vitro analysis of flavonoid bioactivity. In this study, the effect of different proanthocyanidin-rich extracts, which were obtained from cocoa (CCX), French maritime pine bark (Pycnogenol extract, PYC) and grape seed (GSPE), on lipid homeostasis was evaluated. Hepatic human cells (HepG2 cells) were treated with 25 mg/L of CCX, PYC or GSPE. We also performed in vitro experiments to assess the effect on lipid synthesis that is induced by the bioactive GSPE proanthocyanidins using the physiological metabolites that are present in the serum of GSPE-administered rats. For this, Wistar rats were administered 1 g/kg of GSPE, and serum was collected after 2 h. The semipurified serum of GSPE-administered rats was fully characterized by liquid chromatography tandem triple quadrupole mass spectrometry (LC-QqQ/MS(2)). The lipids studied in the analyses were free cholesterol (FC), cholesterol ester (CE) and triglycerides (TG). All three proanthocyanidin-rich extracts induced a remarkable decrease in the de novo lipid synthesis in HepG2 cells. Moreover, GSPE rat serum metabolites reduced the total percentage of CE, FC and particularly TG; this reduction was significantly higher than that observed in the cells directly treated with GSPE. In conclusion, the bioactivity of the physiological metabolites that are present in the serum of rats after their ingestion of a proanthocyanidin-rich extract was demonstrated in Hep G2 cells. PMID:24231101

  1. Red blood cell aquaporin-1 expression is decreased in hereditary spherocytosis.

    Science.gov (United States)

    Crisp, Renée L; Maltaneri, Romina E; Vittori, Daniela C; Solari, Liliana; Gammella, Daniel; Schvartzman, Gabriel; García, Eliana; Rapetti, María C; Donato, Hugo; Nesse, Alcira

    2016-10-01

    Aquaporin-1 (AQP1) is the membrane water channel responsible for changes in erythrocyte volume in response to the tonicity of the medium. As the aberrant distribution of proteins in hereditary spherocytosis (HS) generates deficiencies of proteins other than those codified by the mutated gene, we postulated that AQP1 expression might be impaired in spherocytes. AQP1 expression was evaluated through flow cytometry in 5 normal controls, 1 autoimmune hemolytic anemia, 10 HS (2 mild, 3 moderate, 2 severe, and 3 splenectomized), and 3 silent carriers. The effect of AQP1 inhibitors was evaluated through water flow-based tests: osmotic fragility and hypertonic cryohemolysis. Serum osmolality was measured in 20 normal controls and 13 HS. The effect of erythropoietin (Epo) on AQP1 expression was determined in cultures of erythroleukemia UT-7 cells, dependent on Epo to survive. Independent of erythrocyte size, HS patients showed a lower content of AQP1 in erythrocyte membranes which correlated with the severity of the disease. Accordingly, red blood cells from HS subjects were less sensitive to cryohemolysis than normal erythrocytes after inhibition of the AQP1 water channel. A lower serum osmolality in HS with respect to normal controls suggests alterations during reticulocyte remodeling. The decreased AQP1 expression could contribute to explain variable degrees of anemia in hereditary spherocytosis. The finding of AQP1 expression induced by Epo in a model of erythroid cells may be interpreted as a mechanism to restore the balance of red cell water fluxes. PMID:27465156

  2. Decreased Regulatory T Cells in Vulnerable Atherosclerotic Lesions: Imbalance between Pro- and Anti-Inflammatory Cells in Atherosclerosis

    Directory of Open Access Journals (Sweden)

    Ilonka Rohm

    2015-01-01

    Full Text Available Atherosclerosis is a chronic inflammatory disease of the arterial wall in which presentation of autoantigens by dendritic cells (DCs leads to the activation of T cells. Anti-inflammatory cells like Tregs counterbalance inflammation in atherogenesis. In our study, human carotid plaque specimens were classified as stable (14 and unstable (15 according to established morphological criteria. Vessel specimens (n=12 without any signs of atherosclerosis were used as controls. Immunohistochemical staining was performed to detect different types of DCs (S100, fascin, CD83, CD209, CD304, and CD123, proinflammatory T cells (CD3, CD4, CD8, and CD161, and anti-inflammatory Tregs (FoxP3. The following results were observed: in unstable lesions, significantly higher numbers of proinflammatory cells like DCs, T helper cells, cytotoxic T cells, and natural killer cells were detected compared to stable plaques. Additionally, there was a significantly higher expression of HLA-DR and more T cell activation (CD25, CD69 in unstable lesions. On the contrary, unstable lesions contained significantly lower numbers of Tregs. Furthermore, a significant inverse correlation between myeloid DCs and Tregs was shown. These data suggest an increased inflammatory state in vulnerable plaques resulting from an imbalance of the frequency of local pro- and anti-inflammatory immune cells.

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

    Directory of Open Access Journals (Sweden)

    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.'

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

    Directory of Open Access Journals (Sweden)

    Justin T Seil

    2008-11-01

    Full Text Available Justin T Seil, Thomas J WebsterLaboratory for Nanomedicine Research, Division of Engineering, Brown University, Providence, RI, USAAbstract: 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.Keywords: zinc oxide, nanoparticles, astrocytes, neural tissue, nervous system, biomaterials

  5. Decreased GSSG reductase activity enhances cellular zinc toxicity in three human lung cell lines

    Energy Technology Data Exchange (ETDEWEB)

    Walther, U.I.; Czermak, A.; Mueckter, H.; Walther, S.C.; Fichtl, B. [Walther-Straub-Institut fuer Pharmakologie und Toxikologie, Ludwig-Maximilians-Universitaet Muenchen, Nussbaumstr. 26, 80336, Muenchen (Germany)

    2003-03-01

    Cellular reduced glutathione (GSH) levels have been identified as an essential determinant in zinc-induced cytotoxicity. However, cytotoxic effects of zinc have also been observed without depletion of GSH stores. In a previous study, the intracellular activity of GSSG reductase (GR) has come into focus (Walther et al. 2000, Biol Trace Elem Res 78:163-177). In the present paper we have tried to address this issue more deeply by inhibiting the activity of cellular GR without any appreciable decreases of cellular glutathione. In three pulmonary cell lines, GR activity was inhibited in a dose-dependent manner by the alkylating agent carmustine (BCNU), a known inhibitor of GR. Cells were pretreated with BCNU for 14 h, followed by exposure to various concentrations of zinc chloride. Then we determined the incorporation of radiolabelled methionine (to assess protein synthesis), and measured the GSH and oxidized glutathione (GSSG) levels. Additionally, GR activity of controls was measured. IC{sub 50} values for zinc-induced inhibition of methionine incorporation, as well as GSH contents, was strongly correlated to the decreased GR activity. These results firmly suggest that GR is an important factor in the event chain of zinc cytotoxicity. Together with the results from our previously cited study where impaired regeneration of GSH levels were accompanied by a decrease in total cellular glutathione (GSH + GSSG) we conclude that GSSG itself is an important effector in zinc cytotoxicity. (orig.)

  6. Decreased expression of dual-specificity phosphatase 9 is associated with poor prognosis in clear cell renal cell carcinoma

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    Liu Zhuowei

    2011-09-01

    Full Text Available Abstract Background The molecular mechanisms involved in the development and progression of clear cell renal cell carcinomas (ccRCCs are poorly understood. The objective of this study was to analyze the expression of dual-specificity phosphatase 9 (DUSP-9 and determine its clinical significance in human ccRCCs. Methods The expression of DUSP-9 mRNA was determined in 46 paired samples of ccRCCs and adjacent normal tissues by using real-time qPCR. The expression of the DUSP-9 was determined in 211 samples of ccRCCs and 107 paired samples of adjacent normal tissues by immunohistochemical analysis. Statistical analysis was performed to define the relationship between the expression of DUSP-9 and the clinical features of ccRCC. Results The mRNA level of DUSP-9, which was determined by real-time RT-PCR, was found to be significantly lower in tumorous tissues than in the adjacent non-tumorous tissues (p Conclusion To our knowledge, this is the first study that determines the relationship between DUSP-9 expression and prognosis in ccRCC. We found that decreased expression of DUSP-9 is associated with poor prognosis in ccRCC. DUSP-9 may represent a novel and useful prognostic marker for ccRCC.

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

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    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

  8. Decreased circulating endothelial progenitor cell levels and function in patients with nonalcoholic fatty liver disease.

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    Chia-Hung Chiang

    Full Text Available OBJECTIVES: Nonalcoholic fatty liver disease (NAFLD is associated with advanced atherosclerosis and a higher risk of cardiovascular disease. Increasing evidence suggests that injured endothelial monolayer is regenerated by circulating bone marrow derived-endothelial progenitor cells (EPCs, and levels of circulating EPCs reflect vascular repair capacity. However, the relation between NAFLD and EPC remains unclear. Here, we tested the hypothesis that patients with nonalcoholic fatty liver disease (NAFLD might have decreased endothelial progenitor cell (EPC levels and attenuated EPC function. METHODS AND RESULTS: A total of 312 consecutive patients undergoing elective coronary angiography because of suspected coronary artery disease were screened and received examinations of abdominal ultrasonography between July 2009 and November 2010. Finally, 34 patients with an ultrasonographic diagnosis of NAFLD, and 68 age- and sex-matched controls without NAFLD were enrolled. Flow cytometry with quantification of EPC markers (defined as CD34(+, CD34(+KDR(+, and CD34(+KDR(+CD133(+ in peripheral blood samples was used to assess circulating EPC numbers. The adhesive function, and migration, and tube formation capacities of EPCs were also determined in NAFLD patients and controls. Patients with NAFLD had a significantly higher incidence of metabolic syndrome, previous myocardial infarction, hyperuricemia, and higher waist circumference, body mass index, fasting glucose and triglyceride levels. In addition, patients with NAFLD had significantly decreased circulating EPC levels (all P<0.05, attenuated EPC functions, and enhanced systemic inflammation compared to controls. Multivariate logistic regression analysis showed that circulating EPC level (CD34(+KDR(+ [cells/10(5 events] was an independent reverse predictor of NAFLD (Odds ratio: 0.78; 95% confidence interval: 0.69-0.89, P<0.001. CONCLUSIONS: NAFLD patients have decreased circulating EPC numbers and

  9. Light-induced Voc increase and decrease in high-efficiency amorphous silicon solar cells

    OpenAIRE

    Stuckelberger, Michael; Riesen, Yannick Samuel; Despeisse, Matthieu; Schüttauf, Jan-Willem Alexander; Haug, Franz-Josef; Ballif, Christophe

    2014-01-01

    High-efficiency amorphous silicon (a-Si:H) solar cells were deposited with different thicknesses of the p-type amorphous silicon carbide layer on substrates of varying roughness. We observed a light-induced open-circuit voltage (Voc) increase upon light soaking for thin p-layers, but a decrease for thick p-layers. Further, the Voc increase is enhanced with increasing substrate roughness. After correction of the p-layer thickness for the increased surface area of rough substrates, we can exclu...

  10. 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

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    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.

  11. 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...

  12. Adherence and Blocking of Candida Albicans to Cultured Vaginal Epithelial Cells: Treatments to Decrease Adherence

    Directory of Open Access Journals (Sweden)

    Bryan Larsen

    2006-06-01

    Full Text Available Background. Pathogenesis of mucosal microorganisms depends on adherence to the tissues they colonize and infect. For Candida albicans, cell surface hydrophobicity may play a significant role in tissue binding ability. Methods. A continuous cell line of vaginal epithelial cells (VEC was grown in keratinocyte serum-free medium (KSFM with supplements and harvested by trypsinization. VEC were combined with yeast cells to evaluate adherence and inhibition of adherence. In this experimental setup, yeast stained with fluorescein isothiocyanate were allowed to attach to VEC and the resulting fluorescent VEC were detected by flow cytometry. Results. VEC were cultured and examined daily after plating and showed morphology similar to basal epithelial cells. Culture media supplemented with estradiol showed increased VEC proliferation initially (first 24 h but cell morphology was not altered. Fluorescinated Candida cells bound effectively to the cultured VEC. Using fresh cells exposed to various preparations of K-Y, we showed that all formulations of the product reduced Candida binding to VEC by 25% to 50%. While VEC were generally harvested for use in experiments when they were near confluent growth, we allowed some cultures to grow beyond that point and discovered that cells allowed to become overgrown or stressed appeared to bind yeast cells more effectively. Conclusion. Flow cytometry is a useful method for evaluating binding of stained yeast cells to cultured VEC and has demonstrated that commercially available products have the ability to interfere with the process of yeast adherence to epithelial cells.

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

    International Nuclear Information System (INIS)

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

  14. Nutrient Starvation Decreases Cx43 Levels and Limits Intercellular Communication in Primary Bovine Corneal Endothelial Cells.

    Science.gov (United States)

    D'hondt, Catheleyne; Iyyathurai, Jegan; Welkenhuyzen, Kirsten; Himpens, Bernard; Leybaert, Luc; Bultynck, Geert

    2016-06-01

    Connexin (Cx) proteins form large conductance channels which function as regulators of communication between neighboring cells via gap junctions and/or hemichannels. Intercellular communication is essential to coordinate cellular responses in tissues and organs, thereby fulfilling an essential role in the spreading of signaling, survival and death processes. Connexin 43 (Cx43), a major connexin isoform in brain and heart, is rapidly turned over. Recent studies implicated that autophagy, a lysosomal degradation pathway induced upon nutrient starvation, mediates connexins, including Cx43, degradation. Here, we examined the impact of nutrient starvation on endogenous Cx43-protein levels and endogenous Cx43-driven intercellular communication in primary bovine corneal endothelial cells (BCECs). Hank's Balanced Salt Solution (HBSS) was used as a starvation condition that induces autophagic flux without impacting the survival of the BCECs. Nutrient starvation of BCECs caused a rapid decline in Cx43-protein levels, both as gap junctions and as hemichannels. The time course of the decline in Cx43-protein levels coincided with the time course of the decline in intercellular communication, assessed as intercellular Ca(2+)-wave propagation in BCECs exposed to a single-cell mechanical stimulus. The decline in Cx43-protein levels, both as gap junctions and as hemichannels, could be prevented by the addition of bafilomycin A1, a lysosomal inhibitor, during the complete nutrient starvation period. Consistent with this, bafilomycin A1 significantly alleviated the decrease in intercellular Ca(2+)-wave propagation. This study further underpins the importance of autophagy as an important degradation pathway for Cx43 proteins during periods of nutrient deprivation, thereby impacting the ability of cells to perform intercellular communication. PMID:26873723

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

    DEFF Research Database (Denmark)

    Schlütter, Jacob Mørup; Hatt, Lotte; Bach, Cathrine; Kirkegaard, Ida; Kølvraa, Steen; Uldbjerg, Niels

    2014-01-01

    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 < 0.01), a decrease varying......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...

  16. Elevated temperature treatment as a novel method for decreasing p57 on the cell surface of Renibacterium salmoninarum.

    Science.gov (United States)

    Piganelli, J D; Wiens, G D; Kaattari, S L

    1999-04-15

    Renibacterium salmoninarum is a Gram-positive diplo-bacillus and the causative agent of bacterial kidney disease, a prevalent disease of salmonid fish. Virulent isolates of R. salmoninarum have a hydrophobic cell surface and express the 57-58 kDa protein (p57). Here we have investigated parameters which effect cell hydrophobicity and p57 degradation. Incubation of R. salmoninarum cells at 37 degrees C for > 4 h decreased cell surface hydrophobicity as measured by the salt aggregation assay, and decreased the amount of cell associated p57. Incubation of cells at lower temperatures (22, 17, 4 or -20 degrees C) for up to 16 h did not reduce hydrophobicity or the amount of cell associated p57. Both the loss of cell surface hydrophobicity and the degradation of p57 were inhibited by pre-incubation with the serine protease inhibitor phenylmethylsulfonyl fluoride (PMSF). Cell surface hydrophobicity was specifically reconstituted by incubation with extracellular protein (ECP) concentrated from culture supernatant and was correlated with the reassociation of p57 onto the bacterial cell surface as determined by western blot and total protein stain analyses. The ability of p57 to reassociate suggests that the bacterial cell surface is not irreversibly modified by the 37 degrees C treatment and that p57 contributes to the hydrophobic nature of R. salmoninarum. In summary, we describe parameters effecting the removal of the p57 virulence factor and suggest the utility of this modification for generating a whole cell vaccine against bacterial kidney disease. PMID:10349550

  17. 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.

  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. Light-induced Voc increase and decrease in high-efficiency amorphous silicon solar cells

    Science.gov (United States)

    Stuckelberger, M.; Riesen, Y.; Despeisse, M.; Schüttauf, J.-W.; Haug, F.-J.; Ballif, C.

    2014-09-01

    High-efficiency amorphous silicon (a-Si:H) solar cells were deposited with different thicknesses of the p-type amorphous silicon carbide layer on substrates of varying roughness. We observed a light-induced open-circuit voltage (Voc) increase upon light soaking for thin p-layers, but a decrease for thick p-layers. Further, the Voc increase is enhanced with increasing substrate roughness. After correction of the p-layer thickness for the increased surface area of rough substrates, we can exclude varying the effective p-layer thickness as the cause of the substrate roughness dependence. Instead, we explain the observations by an increase of the dangling-bond density in both the p-layer—causing a Voc increase—and in the intrinsic absorber layer, causing a Voc decrease. We present a mechanism for the light-induced increase and decrease, justified by the investigation of light-induced changes of the p-layer and supported by Advanced Semiconductor Analysis simulation. We conclude that a shift of the electron quasi-Fermi level towards the conduction band is the reason for the observed Voc enhancements, and poor amorphous silicon quality on rough substrates enhances this effect.

  20. Niemann Pick type C cells show cholesterol dependent decrease of APP expression at the cell surface its increased processing through the ?-secretase pathway

    OpenAIRE

    Malnar, Martina; KOŠIČEK, MARKO; Mitterreiter, Stefan; Omerbašić, Damir; Lichtenthaler, Stefan F.; Goate, Alison; Hećimović, Silva

    2010-01-01

    Abstract The link between cholesterol and Alzheimer's disease has recently been revealed in Niemann Pick type C disease. We found that NPC1-/- cells show decreased expression of APP at the cell surface and increased processing of APP through the ?-secretase pathway resulting in increased C99, sAPP? and intracellular A?40 levels. This effect is dependent on increased cholesterol levels, since cholesterol depletion reversed cell surface APP expression and lowered A?/C99 levels in NPC...

  1. Cyclic AMP--dependent aggregation of Swiss 3T3 cells on a cellulose substratum (Cuprophan) and decreased cell membrane Rho A.

    Science.gov (United States)

    Faucheux, N; Nagel, M D

    2002-06-01

    Cell surface integrin receptors and Rho family GTPases function together to mediate adhesion-dependent events in cells. We have shown that the attachment of Swiss 3T3 cells to a cellulose substratum (Cuprophan, CU) activates adenylyl cyclase, which catalyses cyclic AMP (cAMP) production. CU adsorbs vitronectin poorly, prevents cell spreading and causes cells to aggregate. By contrast, spread cells on polystyrene (PS) contain low cAMP concentrations. We have now investigated the shift between integrin signalling-Rho A and the cAMP pathway. CU did not support the formation of focal contacts and stress fibres. The plasma membranes of cells on CU had less Rho A than those of cells on PS. Also, blocking vitronectin (VN) or fibronectin (FN)-integrin receptors with echistatin, which activates cAMP production, decreased Rho A in the plasma membrane of cells attached to PS. But adsorption of VN or FN onto CU, which limits the production of the cAMP, increased the cell membrane Rho A. Adding an inhibitor of cAMP-dependent protein kinase PKA to the medium also increased the plasma membrane Rho A in aggregated cells attached to CU. These results highlight the importance of cAMP, generated by cell attachment to substratum, as a gating element in integrin-Rho A signalling. PMID:12013176

  2. 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.

  3. Cardiolipin deficiency leads to decreased cardiolipin peroxidation and increased resistance of cells to apoptosis

    OpenAIRE

    Huang, Zhentai; Jiang, Jianfei; Tyurin, Vladimir A.; Zhao, Qing; Mnuskin, Alexandra; Ren, Jin; Belikova, Natalia A.; Feng, Weihong; Kurnikov, Igor V.; Kagan, Valerian E.

    2008-01-01

    Cardiolipin (CL), a unique mitochondrial phospholipid synthesized by CL synthase (CLS), plays important, yet not fully understood, roles in mitochondria-dependent apoptosis. We manipulated CL levels in HeLa cells by knocking down CLS using RNA interference and selected a clone of CL-deficient cells with ~45% of its normal content. ESI-MS analysis showed that CL molecular species were the same in deficient and sufficient cells. CL deficiency did not change mitochondrial functions (membrane pot...

  4. Erythropoietin reduces storage lesions and decreases apoptosis indices in blood bank red blood cells

    OpenAIRE

    Oscar Andrés Penuela; Fernando Palomino; Lina Andrea Gómez

    2015-01-01

    ABSTRACT Background: Recent evidence shows a selective destruction of the youngest circulating red blood cells (neocytolysis) trigged by a drop in erythropoietin levels. Objective: The aim of this study was to evaluate the effect of recombinant human erythropoietin beta on the red blood cell storage lesion and apoptosis indices under blood bank conditions. Methods: Each one of ten red blood cell units preserved in additive solution 5 was divided in two volumes of 100 mL and assigned to one...

  5. Exosomes: Decreased Sensitivity of Lung Cancer A549 Cells to Cisplatin

    OpenAIRE

    Xia Xiao; Shaorong Yu; Shuchun Li; Jianzhong Wu; Rong Ma; Haixia Cao; Yanliang Zhu; Jifeng Feng

    2014-01-01

    Exosomes are small extracellular membrane vesicles of endocytic origin released by many cells that could be found in most body fluids. The main functions of exosomes are cellular communication and cellular waste clean-up. This study was conducted to determine the involvement of exosomes in the regulation of sensitivity of the lung cancer cell line A549 to cisplatin (DDP). When DDP was added to A549 cells, exosomes secretion was strengthened. Addition of the secreted exosomes to other A549 cel...

  6. High expression of osteoglycin decreases the metastatic capability of mouse hepatocarcinoma Hca-F cells to lymph nodes

    Institute of Scientific and Technical Information of China (English)

    Xiaonan Cui; Bo Song; Li Hou; Zhiyi Wei; Jianwu Tang

    2008-01-01

    Osteoglycin, one of the matrix molecules, belongs to the small leucine-rich proteoglycan gene family and might play important roles in cell growth and differentiation and in pathological processes such as fibrosis and cancer growth.In this study, a eukaryotic expression plasmid pIRESpuro3 osteoglycin(+) was constructed and transfected into mouse hepatocarcinoma Hca-F cells to evaluate the contribution of osteoglycin to the malignant behavior of Hca-F. It was foundthat Hca-F cells transfected with pIRESpuro3 osteoglycin(+) showed significantly decreased potential for both migration and invasion. Furthermore, Hca-F cells transfected with osteoglycin showed decreased metastatic potential to peripheral lymph nodes. However, proliferation potential and adhesive capacity of Hca-F cells to different protein substrates were not influenced by osteoglycin transfection. In summary,high expression of osteoglycin decreases the metastatic capability of Hca-F to lymph nodes.

  7. Decreased Infections in Recipients of Unrelated Donor Hematopoietic Cell Transplantation from Donors with an Activating KIR Genotype

    OpenAIRE

    Tomblyn, Marcie; Young, Jo-Anne H.; Haagenson, Michael D.; Klein, John P.; Trachtenberg, Elizabeth A.; Storek, Jan; Spellman, Stephen R.; Cooley, Sarah; Miller, Jeffrey S.; Weisdorf, Daniel J.

    2010-01-01

    Infectious complications following allogeneic hematopoietic cell transplantation (HCT) from unrelated donors (URD) result in significant morbidity. We hypothesized that recipients of an URD with an activating natural killer cell immunoglobulin-like receptor (KIR) (B/x) genotype would have decreased infectious complications due to enhanced NK cell function. We compared the infectious complications in 116 recipients of a graft from a donor with an A/A KIR (n = 44) genotype and a B/x KIR (n = 72...

  8. Decreased expression of dual-specificity phosphatase 9 is associated with poor prognosis in clear cell renal cell carcinoma

    International Nuclear Information System (INIS)

    The molecular mechanisms involved in the development and progression of clear cell renal cell carcinomas (ccRCCs) are poorly understood. The objective of this study was to analyze the expression of dual-specificity phosphatase 9 (DUSP-9) and determine its clinical significance in human ccRCCs. The expression of DUSP-9 mRNA was determined in 46 paired samples of ccRCCs and adjacent normal tissues by using real-time qPCR. The expression of the DUSP-9 was determined in 211 samples of ccRCCs and 107 paired samples of adjacent normal tissues by immunohistochemical analysis. Statistical analysis was performed to define the relationship between the expression of DUSP-9 and the clinical features of ccRCC. The mRNA level of DUSP-9, which was determined by real-time RT-PCR, was found to be significantly lower in tumorous tissues than in the adjacent non-tumorous tissues (p < 0.001). An immunohistochemical analysis of 107 paired tissue specimens showed that the DUSP-9 expression was lower in tumorous tissues than in the adjacent non-tumorous tissues (p < 0.001). Moreover, there was a significant correlation between the DUSP-9 expression in ccRCCs and gender (p = 0.031), tumor size (p = 0.001), pathologic stage (p = 0.001), Fuhrman grade (p = 0.002), T stage (p = 0.001), N classification (p = 0.012), metastasis (p = 0.005), and recurrence (p < 0.001). Patients with lower DUSP-9 expression had shorter overall survival time than those with higher DUSP-9 expression (p < 0.001). Multivariate analysis indicated that low expression of the DUSP-9 was an independent predictor for poor survival of ccRCC patients. To our knowledge, this is the first study that determines the relationship between DUSP-9 expression and prognosis in ccRCC. We found that decreased expression of DUSP-9 is associated with poor prognosis in ccRCC. DUSP-9 may represent a novel and useful prognostic marker for ccRCC

  9. Decreased prolactin-inducible protein expression exhibits inhibitory effects on the metastatic potency of breast cancer cells

    Institute of Scientific and Technical Information of China (English)

    Zhendong Zheng; Xiaodong Xie

    2013-01-01

    Objective: The aim of the research was to study the effects of prolactin-inducible protein (PIP) downregulation on metastatic abilities of human breast cancer MDA-MB-453 cells. Methods: PIP-siRNA was transfected into human breast cancer MDA-MB-453 cells through liposome. Reverse transcription PCR and immunocytochemistry were employed to detect the downregulated expression of PIP. Cell migration, adhesion and invasion assays were performed to assess the impacts of PIP downregulation on cell migration, adhesion and invasion respectively. Results: Knockdown of PIP obviously inhibited cell migration, the migrated cells were decreased by 83.1% compared with the negative control group. Cell adhesion was also reduced, the adhesion rates at 30 min and 60 min were decreased by 42.6% and 48.5% respectively compared with the negative control group. Moreover, PIP downregulation resulted in decreased invasion rate by 73.9%. Conclusion: Reduced PIP expression in MDA-MB-453 cells can inhibit the abilities of migration, adhesion and invasion, which suggests that PIP plays an important role in the metastatic potency of breast cancer cells.

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

    Directory of Open Access Journals (Sweden)

    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.

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

    International Nuclear Information System (INIS)

    In radioiodine therapy the ''stunning phenomenon'' is defined as a reduction of radioiodine uptake after diagnostic application of 131I. 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 131I. Time-dependent 131I 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 131I concentration. Sodium iodide symporter (NIS) and the housekeeping gene (β-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 131I 24-48 h after application. Significantly decreased 131I 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 131I long-term incubation. It could be demonstrated that during 131I 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.)

  12. 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…

  13. Inhibition of macrophage migration inhibitory factor decreases proliferation and cytokine expression in bladder cancer cells

    International Nuclear Information System (INIS)

    The importance of various inflammatory cytokines in maintaining tumor cell growth and viability is well established. Increased expression of the proinflammatory cytokine macrophage migration inhibitory factor (MIF) has previously been associated with various types of adenocarcinoma. MIF IHC was used to localize MIF in human bladder tissue. ELISA and Western blot analysis determined the synthesis and secretion of MIF by human bladder transitional cell carcinoma cells. The effects of MIF inhibitors (high molecular weight hyaluronate (HA), anti-MIF antibody or MIF anti-sense) on cell growth and cytokine expression were analyzed. Human bladder cancer cells (HT-1376) secrete detectable amounts of MIF protein. Treatment with HA, anti-MIF antibody and MIF anti-sense reduced HT-1376 cell proliferation, MIF protein secretion, MIF gene expression and secreted inflammatory cytokines. Our evidence suggests MIF interacts with the invariant chain, CD74 and the major cell surface receptor for HA, CD44. This study is the first to report MIF expression in the human bladder and these findings support a role for MIF in tumor cell proliferation. Since MIF participates in the inflammatory response and bladder cancer is associated with chronic inflammatory conditions, these new findings suggest that neutralizing bladder tumor MIF may serve as a novel therapeutic treatment for bladder carcinoma

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

    International Nuclear Information System (INIS)

    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

  15. 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. PMID:25672417

  16. Myrtucommulone-A treatment decreases pluripotency- and multipotency-associated marker expression in bladder cancer cell line HTB-9.

    Science.gov (United States)

    Iskender, Banu; Izgi, Kenan; Karaca, Halit; Canatan, Halit

    2015-10-01

    Cancer and stem cells exhibit similar features, including self-renewal, differentiation and immortality. The expression of stem-cell-related genes in cancer cells is demonstrated to be potentially correlated with cancer cell behaviour, affecting both drug response and tumor recurrence. There is an emerging body of evidence that subpopulations of tumors carry a distinct molecular sign and are selectively resistant to chemotherapy. Therefore, it is important to find novel therapeutic agents that could suppress the stem-like features of cancer cells while inhibiting their proliferation. Myrtucommulone-A (MC-A) is an active compound of a nonprenylated acylphloroglucinol isolated from the leaves of myrtle. Here we have investigated the potential of MC-A in inhibiting the expression of self-renewal regulatory factors and cancer stem cell markers in a bladder cancer cell line HTB-9. We used RT-PCR, immunocytochemistry, flow cytometry and western blotting to examine the expression of pluripotency- and multipotency-associated markers with or without treatment with MC-A. Treatment with MC-A not only decreased cancer cell viability and proliferation but also resulted in a decrease in the expression of pluripotency- and multipotency-associated markers such as NANOG, OCT-4, SOX-2, SSEA-4, TRA-1-60, CD90, CD73 and CD44. MC-A treatment was also observed to decrease the sphere-forming ability of HTB-9 cells. In summary, this study provides valuable information on the presence of stem-cell marker expression in HTB-9 cells and our results imply that MC-A could be utilized to target cancer cells with stem-like characteristics. PMID:26054707

  17. Silencing CTL1 mRNA decreases choline transport and cell growth in NG108-15 cells

    Czech Academy of Sciences Publication Activity Database

    Machová, Eva; Michal, Pavel; Lisá, Věra; Doležal, Vladimír

    Praha, 2005. s. 138-138. [Conference of the Czech Neuroscience Society /5./, The Annual Meeting of the Network of European Neuroscience Institutes. 19.11.2005-21.11.2005, Prague] R&D Projects: GA AV ČR(CZ) IAA5011206; GA MŠk(CZ) LC554 Grant ostatní: GA-(XE) QLK1-CT-2002-00172 Institutional research plan: CEZ:AV0Z50110509 Keywords : CTL1 protein knockdown * RNA interference * NG108-15 cells * cell growth * choline transport Subject RIV: ED - Physiology

  18. 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...

  19. Leccinum vulpinum Watling induces DNA damage, decreases cell proliferation and induces apoptosis on the human MCF-7 breast cancer cell line.

    Science.gov (United States)

    Reis, Filipa S; Sousa, Diana; Barros, Lillian; Martins, Anabela; Morales, Patricia; Ferreira, Isabel C F R; Vasconcelos, M Helena

    2016-04-01

    The current work aimed to study the antitumour activity of a phenolic extract of the edible mushroom Leccinum vulpinum Watling, rich essentially in hydroxybenzoic acids. In a first approach, the mushroom extract was tested against cancer cell growth by using four human tumour cell lines. Given the positive results obtained in these initial screening experiments and the evidence of some studies for an inverse relationship between mushroom consumption and breast cancer risk, a detailed study of the bioactivity of the extract was carried out on MCF-7 cells. Once the selected cell line to precede the work was the breast adenocarcinoma cell line, the human breast non-malignant cell line MCF-10A was used as control. Overall, the extract decreased cellular proliferation and induced apoptosis. Furthermore, the results also suggest that the extract causes cellular DNA damage. Data obtained highlight the potential of mushrooms as a source of biologically active compounds, particularly with antitumour activity. PMID:26854920

  20. Targeting CD138-/CD20+ Clonogenic Myeloma Precursor Cells Decreases These Cells and Induces Transferable Antimyeloma Immunity.

    Science.gov (United States)

    Lum, Lawrence G; Thakur, Archana; Kondadasula, Sri Vidya; Al-Kadhimi, Zaid; Deol, Abhinav; Tomaszewski, Elyse N; Yano, Hiroshi; Schalk, Dana L; Ayash, Lois; Zonder, Jeffrey A; Uberti, Joseph P; Abidi, Muneer H; Ratanatharathorn, Voravit

    2016-05-01

    This phase Ib clinical trial evaluated whether pretargeting of CD20(+) clonogenic myeloma precursor cells (CMPCs) with anti-CD3 × anti-CD20 bispecific antibody-armed T cells (BATs) before autologous stem cell transplantation (SCT) in patients with standard-risk and high-risk multiple myeloma would induce antimyeloma immunity that could be detected and boosted after SCT. All 12 patients enrolled in this study received 2 BATs infusions before SCT, and 4 patients received a booster infusion of BATs after SCT. Pretargeting CD138(-)/CD20(+) CMPCs with BATs before SCT was safe and reduced levels of CMPCs by up to 58% in the postinfusion bone marrow in patients who remained in remission. Four of 5 patients who remained in remission had a >5-fold increase in IFN-γ enzyme-linked immunospot responses. SOX2 antibody increased after BATs infusions and persisted after SCT. The median anti-SOX2 level at 3 months after SCT was 28.1 ng/mL (range, 4.6 to 256 ng/mL) in patients who relapsed and 46 ng/mL (range, 28.3 to 73.3 ng/mL) in patients who remained in remission. The immune correlates suggest that infusions of targeted T cells given before SCT were able to reduce CMPC levels and induced cellular and humoral antimyeloma immunity that could be transferred and boosted after SCT. PMID:26827660

  1. 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. PMID:27294113

  2. Decreased Polysaccharide Feruloylation Compromises Plant Cell Wall Integrity and Increases Susceptibility to Necrotrophic Fungal Pathogens

    Directory of Open Access Journals (Sweden)

    Nathan T Reem

    2016-05-01

    Full Text Available The complexity of cell wall composition and structure determines the strength, flexibility, and function of the primary cell wall in plants. However, the contribution of the various components to cell wall integrity and function remains unclear. Modifications of cell wall composition can induce plant responses known as Cell Wall Integrity control. In this study, we used transgenic expression of the fungal feruloyl esterase AnFAE to examine the effect of post-synthetic modification of Arabidopsis and Brachypodium cell walls. Transgenic Arabidopsis plants expressing AnFAE showed a significant reduction of monomeric ferulic acid, increased amounts of wall-associated extensins, and increased susceptibility to Botrytis cinerea, compared with wild type. Transgenic Brachypodium showed reductions in monomeric and dimeric ferulic acids and increased susceptibility to Bipolaris sorokiniana. Upon infection, transgenic Arabidopsis and Brachypodium plants also showed increased expression of several defense-related genes compared with wild type. These results demonstrate a role, in both monocot and dicot plants, of polysaccharide feruloylation in plant cell wall integrity, which contributes to plant resistance to necrotrophic pathogens.

  3. 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

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

    Directory of Open Access Journals (Sweden)

    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.

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

    Directory of Open Access Journals (Sweden)

    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.

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

    Directory of Open Access Journals (Sweden)

    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.

  7. Decreased HPV-specific T cell responses and accumulation of immunosuppressive influences in oropharyngeal cancer patients following radical therapy.

    Science.gov (United States)

    Al-Taei, Saly; Banner, Russell; Powell, Ned; Evans, Mererid; Palaniappan, Nachi; Tabi, Zsuzsanna; Man, Stephen

    2013-12-01

    Oropharyngeal cancer (OPC) is a type of squamous cell head and neck cancer that is often associated with human papillomavirus (HPV) infection, suggesting the potential for immunotherapeutic targeting of HPV antigens. This study aimed to determine the effect of radical therapy on HPV-specific T cells and other immune parameters in 20 OPC patients, as a prelude to future immunotherapy studies. HPV DNA could be detected in 9/12 available tissue samples (8/9 HPV(+) samples were also p16(+)). HPV-specific T cell responses against HPV16 E6 and E7 peptides were detected by enzyme-linked immunoSPOT in 10/13 and 8/13 evaluable patients, respectively, but did not appear to correlate with HPV status. Post-treatment, both HPV E6 and E7 T cell responses were decreased (4/13 and 2/13 patients, respectively). These reductions in T cell response could not be explained by a concurrent decrease in memory T cells whose absolute numbers were relatively unaffected by radical therapy (27,975 vs. 25,661/10(5) PBMC) despite a significant decrease in overall lymphocyte counts (1.74 vs. 0.69 × 10(9)/L). Instead, there were significant increases in regulatory T cells (3.7 vs. 6.8 %) and a population of myeloid-derived suppressor cells (CD14(-)HLA-DR(-)CD15(hi), 12.38 vs. 21.92 %). This suggests that immunosuppression may contribute to the reduction in HPV-specific T cell responses post-treatment, although study of larger patient cohorts will be required to test whether this affects clinical outcome. Overall these findings suggest that HPV-targeted immunotherapy in post-therapy OPC patients will require multiple strategies to boost T cell immunity and to overcome the influence of immunosuppressive cells. PMID:24146146

  8. Fractalkine restores the decreased expression of StAR and progesterone in granulosa cells from patients with polycystic ovary syndrome.

    Science.gov (United States)

    Huang, Shuo; Pang, Yanli; Yan, Jie; Lin, Shengli; Zhao, Yue; Lei, Li; Yan, Liying; Li, Rong; Ma, Caihong; Qiao, Jie

    2016-01-01

    Low progesterone levels are associated with luteal phase deficiency in women with polycystic ovary syndrome (PCOS). The mechanisms regulating progesterone biosynthesis in the granulosa cells from women with PCOS is largely unknown. Fractalkine is expressed in human ovaries, and is reported to regulate progesterone production in granulosa cells of healthy women. In the current study, we aimed to examine the role of fractalkine in women with PCOS. Reduced fractalkine levels were found in follicular fluid and granulosa cells, accompanied by decreased progesterone production and reduced steroidogenic acute regulatory protein (StAR) expression in the granulosa cells of patients with PCOS. Administration of fractalkine reversed the inhibition of progesterone and StAR expression. The mechanism mediating these effects may be associated with the inhibition of ERK activity in the granulosa cells from women with PCOS. Our findings revealed that fractalkine regulated steroidogenesis in follicular granulosa cells of women with PCOS. PMID:27386819

  9. Decreased autocrine EGFR signaling in metastatic breast cancer cells inhibits tumor growth in bone and mammary fat pad.

    Science.gov (United States)

    Nickerson, Nicole K; Mohammad, Khalid S; Gilmore, Jennifer L; Crismore, Erin; Bruzzaniti, Angela; Guise, Theresa A; Foley, John

    2012-01-01

    Breast cancer metastasis to bone triggers a vicious cycle of tumor growth linked to osteolysis. Breast cancer cells and osteoblasts express the epidermal growth factor receptor (EGFR) and produce ErbB family ligands, suggesting participation of these growth factors in autocrine and paracrine signaling within the bone microenvironment. EGFR ligand expression was profiled in the bone metastatic MDA-MB-231 cells (MDA-231), and agonist-induced signaling was examined in both breast cancer and osteoblast-like cells. Both paracrine and autocrine EGFR signaling were inhibited with a neutralizing amphiregulin antibody, PAR34, whereas shRNA to the EGFR was used to specifically block autocrine signaling in MDA-231 cells. The impact of these was evaluated with proliferation, migration and gene expression assays. Breast cancer metastasis to bone was modeled in female athymic nude mice with intratibial inoculation of MDA-231 cells, and cancer cell-bone marrow co-cultures. EGFR knockdown, but not PAR34 treatment, decreased osteoclasts formed in vitro (p<0.01), reduced osteolytic lesion tumor volume (p<0.01), increased survivorship in vivo (p<0.001), and resulted in decreased MDA-231 growth in the fat pad (p<0.01). Fat pad shEGFR-MDA-231 tumors produced in nude mice had increased necrotic areas and decreased CD31-positive vasculature. shEGFR-MDA-231 cells also produced decreased levels of the proangiogenic molecules macrophage colony stimulating factor-1 (MCSF-1) and matrix metalloproteinase 9 (MMP9), both of which were decreased by EGFR inhibitors in a panel of EGFR-positive breast cancer cells. Thus, inhibiting autocrine EGFR signaling in breast cancer cells may provide a means for reducing paracrine factor production that facilitates microenvironment support in the bone and mammary gland. PMID:22276166

  10. Decreased autocrine EGFR signaling in metastatic breast cancer cells inhibits tumor growth in bone and mammary fat pad.

    Directory of Open Access Journals (Sweden)

    Nicole K Nickerson

    Full Text Available Breast cancer metastasis to bone triggers a vicious cycle of tumor growth linked to osteolysis. Breast cancer cells and osteoblasts express the epidermal growth factor receptor (EGFR and produce ErbB family ligands, suggesting participation of these growth factors in autocrine and paracrine signaling within the bone microenvironment. EGFR ligand expression was profiled in the bone metastatic MDA-MB-231 cells (MDA-231, and agonist-induced signaling was examined in both breast cancer and osteoblast-like cells. Both paracrine and autocrine EGFR signaling were inhibited with a neutralizing amphiregulin antibody, PAR34, whereas shRNA to the EGFR was used to specifically block autocrine signaling in MDA-231 cells. The impact of these was evaluated with proliferation, migration and gene expression assays. Breast cancer metastasis to bone was modeled in female athymic nude mice with intratibial inoculation of MDA-231 cells, and cancer cell-bone marrow co-cultures. EGFR knockdown, but not PAR34 treatment, decreased osteoclasts formed in vitro (p<0.01, reduced osteolytic lesion tumor volume (p<0.01, increased survivorship in vivo (p<0.001, and resulted in decreased MDA-231 growth in the fat pad (p<0.01. Fat pad shEGFR-MDA-231 tumors produced in nude mice had increased necrotic areas and decreased CD31-positive vasculature. shEGFR-MDA-231 cells also produced decreased levels of the proangiogenic molecules macrophage colony stimulating factor-1 (MCSF-1 and matrix metalloproteinase 9 (MMP9, both of which were decreased by EGFR inhibitors in a panel of EGFR-positive breast cancer cells. Thus, inhibiting autocrine EGFR signaling in breast cancer cells may provide a means for reducing paracrine factor production that facilitates microenvironment support in the bone and mammary gland.

  11. Leukemia Inhibitory Factor Decreases the Arylamine N-Acetyltransferase Activity in Human Cumulus Granulosa Cells

    OpenAIRE

    Chang, Chi-Chen; Hsieh, Yao-Yuan; CHUNG, JING-GUNG; Tsai, Horng-Der; Tsai, Chang-Hai

    2001-01-01

    Purpose: To evaluate the activities of acetyl coenzyme A (AcCoA): arylamine N-acetyltransferase (NAT) of intact cumulus granulosa cells and the role of leukemia inhibitory factor (LIF) upon their NAT activities.

  12. 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

  13. Chloral hydrate decreases gap junction communications in rat liver epithelial cells

    Science.gov (United States)

    Gap junction communication (GJC) is involved in controlling cell proliferation and differentiation. Alterations in GJC are associated with carcinogenesis, but the mechanisms involvedareunknown.Chloralhydrate(CH), a by-productofchlorinedisinfection ofwater,is carcinogenic in mice,...

  14. A new method for decreasing cell-load variation in dynamic cellular manufacturing systems

    OpenAIRE

    Aidin Delgoshaei; Mohd Khairol Mohd Ariffin,; Btht Hang Tuah Bin Baharudin; Zulkiflle Leman

    2016-01-01

    Cell load variation is considered a significant shortcoming in scheduling of cellular manufacturing systems. In this article, a new method is proposed for scheduling dynamic cellular manufacturing systems in the presence of bottleneck and parallel machines. The aim of this method is to control cell load variation during the process of determining the best trading off values between in-house manufacturing and outsourcing. A genetic algorithm (GA) is developed because of the high potential of t...

  15. Gradient sensing by a bistable regulatory motif enhances signal amplification but decreases accuracy in individual cells

    Science.gov (United States)

    Sharma, Rati; Roberts, Elijah

    2016-06-01

    Many vital eukaryotic cellular functions require the cell to respond to a directional gradient of a signaling molecule. The first two steps in any eukaryotic chemotactic/chemotropic pathway are gradient detection and cell polarization. Like many processes, such chemotactic and chemotropic decisions are made using a relatively small number of molecules and are thus susceptible to internal and external fluctuations during signal transduction. Large cell-to-cell variations in the magnitude and direction of a response are therefore possible and do, in fact, occur in natural systems. In this work we use three-dimensional probabilistic modeling of a simple gradient sensing pathway to study the capacity for individual cells to accurately determine the direction of a gradient, despite fluctuations. We include a stochastic external gradient in our simulations using a novel gradient boundary condition modeling a point emitter a short distance away. We compare and contrast three different variants of the pathway, one monostable and two bistable. The simulation data show that an architecture combining bistability with spatial positive feedback permits the cell to both accurately detect and internally amplify an external gradient. We observe strong polarization in all individual cells, but in a distribution of directions centered on the gradient. Polarization accuracy in our study was strongly dependent upon a spatial positive feedback term that allows the pathway to trade accuracy for polarization strength. Finally, we show that additional feedback links providing information about the gradient to multiple levels in the pathway can help the cell to refine initial inaccuracy in the polarization direction.

  16. 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...... and neurons in embryonic cortical primary cell cultures, which was accompanied by changes in mRNA expression of specific receptors and transporters for glutamatergic and monoaminergic neurotransmitters....

  17. Decrease in Circulating Dendritic Cell Precursors in Patients with Peripheral Artery Disease

    OpenAIRE

    Daniel Kretzschmar; Ilonka Rohm; Sebastian Schäller; Stefan Betge; Rudin Pistulli; Yevgeniya Atiskova; Hans-R. Figulla; Atilla Yilmaz

    2015-01-01

    Peripheral artery disease (PAD) is a common manifestation of atherosclerosis. Inflammation is important for initiation and progression of the disease. Dendritic cells (DCs) as antigen-presenting cells play an important role in the immune system. Therefore, we hypothesize that, in patients with PAD, DCPs might be reduced in blood due to their recruitment into the vascular wall and induce a proinflammatory response. The numbers of myeloid DCPs, plasmacytoid DCPs, and total DCPs were analyzed by...

  18. 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.

  19. 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

  20. T helper 17 and T helper 1 cells are increased but regulatory T cells are decreased in subchondral bone marrow microenvironment of patients with rheumatoid arthritis

    Science.gov (United States)

    Wang, Ting; Li, Shufeng; Yang, Yun; Zhang, Kaining; Dong, Shixiao; Wang, Xiuhua; Liu, Xinguang; Ren, Yanjun; Zhang, Ming; Yan, Xinfeng; Li, Jianmin; Zhang, Lei

    2016-01-01

    Objectives: The present study is to investigate the profiles of Th17, Th1 and Treg cells in bone marrow of patients with rheumatoid arthritis (RA). Methods: Flow cytometry was used to analyze the frequencies of Th17, Th1 and Treg cells in paired peripheral blood and bone marrow of 26 RA patients and 11 osteoarthritis (OA) patients, as well as 10 healthy controls. In addition, the disease activity was analyzed by the 28-joint disease activity score (DAS28). Results: The frequencies of Th17 and Th1 cells were significantly elevated in bone marrow of RA patients. Importantly, Th17 and Th1 cells were significantly elevated in bone marrow compared with the matched peripheral blood from RA patients. However, Treg cells were significantly decreased in bone marrow of RA patients compared with the matched peripheral blood of RA patients and bone marrow of osteoarthritis patients and healthy controls. Moreover, the frequencies of tumor necrosis factor-α-producing T cells were significantly elevated in bone marrow from RA patients. Additionally, Th17 and Th1 cells in bone marrow were positively correlated with DAS28, while Treg cells were negatively correlated with DAS28. Conclusions: The present study demonstrates that Th17 and Th1 cells are markedly increased in bone marrow from RA patients. By contrast, Treg cells are significantly decreased in bone marrow from RA patients. These results suggest that local abnormality of Th17, Th1 and Treg cells in bone marrow of RA patients may contribute to bone destruction in skeletal system.

  1. Decreased frequencies of circulating follicular helper T cell counterparts and plasmablasts in ankylosing spondylitis patients Naive for TNF blockers.

    Directory of Open Access Journals (Sweden)

    María-Belén Bautista-Caro

    Full Text Available Follicular helper T cells (Tfh, localized in lymphoid organs, promote B cell differentiation and function. Circulating CD4 T cells expressing CXCR5, ICOS and/or PD-1 are counterparts of Tfh. Three subpopulations of circulating CD4+CXCR5+ cells have been described: CXCR3+CCR6- (Tfh-Th1, CXCR3-CCR6+ (Tfh-Th17, and CXCR3-CCR6- (Tfh-Th2. Only Tfh-Th17 and Tfh-Th2 function as B cell helpers. Our objective was to study the frequencies of circulating Tfh (cTfh, cTfh subsets and plasmablasts (CD19+CD20-CD27+CD38high cells, and the function of cTfh cells, in patients with Ankylosing Spondylitis (AS. To this end, peripheral blood was drawn from healthy controls (HC (n = 50, AS patients naïve for TNF blockers (AS/nb (n = 25 and AS patients treated with TNF blockers (AS/b (n = 25. The frequencies of cTfh and plasmablasts were determined by flow cytometry. Cocultures of magnetically sorted CD4+CXCR5+ T cells with autologous CD19+CD27- naïve B cells were established from 3 AS/nb patients and 3 HC, and concentrations of IgG, A and M were measured in supernatants. We obseved that AS/nb but not AS/b patients, demonstrated decreased frequencies of circulating CD4+CXCR5+ICOS+PD-1+ cells and plasmablasts, together with a decreased (Tfh-Th17+Tfh-Th2/Tfh-Th1 ratio. The amounts of IgG and IgA produced in cocultures of CD4+CXCR5+ T cells with CD19+CD27- B cells of AS/nb patients were significantly lower than observed in cocultures established from HC. In summary, AS/nb but not AS/b patients, demonstrate a decreased frequency of cTfh and plasmablasts, and an underrepresentation of cTfh subsets bearing a B helper phenotype. In addition, peripheral blood CD4+CXCR5+ T cells of AS/nb patients showed a decreased capacity to help B cells ex vivo.

  2. Curcumin (Diferuloylmethane) Inhibits Cell Proliferation, Induces Apoptosis, and Decreases Hormone Levels and Secretion in Pituitary Tumor Cells

    OpenAIRE

    Miller, Matthew; Chen, Shenglin; Woodliff, Jeffrey; Kansra, Sanjay

    2008-01-01

    Prolactinomas are the most prevalent functional pituitary adenomas. Dopamine D2 receptor (D2R) agonists, such as bromocriptine are the first line of therapy; however, drug intolerance/resistance to D2R agonists exists. Apart from D2R agonists, there is no established medical therapy for prolactinomas; therefore, identifying novel therapeutics is warranted. Curcumin, a commonly used food additive in South Asian cooking, inhibits proliferation of several tumor cell lines; however, its effect on...

  3. 17β-Estradiol treatment inhibits breast cell proliferation, migration and invasion by decreasing MALAT-1 RNA level

    International Nuclear Information System (INIS)

    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

  4. 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.

  5. Cyclosporine A inhibits transcription of cytokine genes and decreases the frequencies of IL-2 producing cells in feline mononuclear cells.

    Science.gov (United States)

    Kuga, Kazufumi; Nishifuji, Koji; Iwasaki, Toshiroh

    2008-10-01

    Cyclosporine A (CsA) has been widely used for suppression of transplant rejection and controlling pruritus in allergic dermatitis in humans, dogs and cats. CsA is known to suppress the expression of inflammatory cytokines, including IL-2, IL-4, IFN-gamma and TNF-alpha in humans, dogs and experimental mice. However, little is known about the immunomodulating effect of CsA in cats. The aim of this study was to evaluate the effects of CsA on the expression of inflammatory cytokines in feline peripheral blood mononuclear cells (PBMC). Real-time PCR analyses with Concanavalin A (ConA)-stimulated PBMC obtained from 5 cats revealed that the expression of mRNAs for IL-2, IL-4, IFN- gamma and TNF-alpha was inhibited by CsA in a dose-dependent manner. Moreover, an enzyme-linked immunospot (ELISPOT) assay, which is capable of detecting IL-2 secreting cells as single spots, revealed that the frequency of IL-2 secreting cells in ConA-stimulated feline PBMC was significantly reduced in the presence of CsA. These results might provide an explanation for the mechanisms of action of CsA in the suppression of transplant rejection and the control of pruritus in cats. PMID:18981654

  6. MDSC-decreasing chemotherapy increases the efficacy of cytokine-induced killer cell immunotherapy in metastatic renal cell carcinoma and pancreatic cancer.

    Science.gov (United States)

    Wang, Zibing; Liu, Yuqing; Zhang, Yong; Shang, Yiman; Gao, Quanli

    2016-01-26

    Adoptive immunotherapy using cytokine-induced killer (CIK) cells is a promising cancer treatment, but its efficacy is restricted by various factors, including the accumulation of myeloid-derived suppressor cells (MDSCs). In this study, we determine whether chemotherapeutic drugs that reduce MDSC levels enhance the efficacy of CIK cell therapy in the treatment of solid tumors. Fifty-three patients were included in this study; 17 were diagnosed with metastatic renal cell carcinoma (MRCC), 10 with advanced pancreatic cancer (PC), and 26 with metastatic melanoma (MM). These patients were divided into two groups: CIK cell therapy alone and CIK cell therapy combined with chemotherapy. Combining CIK cell therapy and chemotherapy increased 1-year survival rates and median survival times in MRCC and PC patients, but not in MM patients. The disease control rate did not differ between treatment groups for MRCC or MM patients, but was higher in PC patients receiving combined treatment than CIK cell treatment alone. These data suggest that addition of MDSC-decreasing chemotherapy to CIK cell therapy improves survival in MRCC and PC patients. PMID:26716894

  7. Targeting choline phospholipid metabolism: GDPD5 and GDPD6 silencing decrease breast cancer cell proliferation, migration, and invasion.

    Science.gov (United States)

    Cao, Maria Dung; Cheng, Menglin; Rizwan, Asif; Jiang, Lu; Krishnamachary, Balaji; Bhujwalla, Zaver M; Bathen, Tone F; Glunde, Kristine

    2016-08-01

    Abnormal choline phospholipid metabolism is associated with oncogenesis and tumor progression. We have investigated the effects of targeting choline phospholipid metabolism by silencing two glycerophosphodiesterase genes, GDPD5 and GDPD6, using small interfering RNA (siRNA) in two breast cancer cell lines, MCF-7 and MDA-MB-231. Treatment with GDPD5 and GDPD6 siRNA resulted in significant increases in glycerophosphocholine (GPC) levels, and no change in the levels of phosphocholine or free choline, which further supports their role as GPC-specific regulators in breast cancer. The GPC levels were increased more than twofold during GDPD6 silencing, and marginally increased during GDPD5 silencing. DNA laddering was negative in both cell lines treated with GDPD5 and GDPD6 siRNA, indicating absence of apoptosis. Treatment with GDPD5 siRNA caused a decrease in cell viability in MCF-7 cells, while GDPD6 siRNA treatment had no effect on cell viability in either cell line. Decreased cell migration and invasion were observed in MDA-MB-231 cells treated with GDPD5 or GDPD6 siRNA, where a more pronounced reduction in cell migration and invasion was observed under GDPD5 siRNA treatment as compared with GDPD6 siRNA treatment. In conclusion, GDPD6 silencing increased the GPC levels in breast cancer cells more profoundly than GDPD5 silencing, while the effects of GDPD5 silencing on cell viability/proliferation, migration, and invasion were more severe than those of GDPD6 silencing. Our results suggest that silencing GDPD5 and GDPD6 alone or in combination may have potential as a new molecular targeting strategy for breast cancer treatment. Copyright © 2016 John Wiley & Sons, Ltd. PMID:27356959

  8. Targeting sphingosine kinase 1 in carcinoma cells decreases proliferation and survival by compromising PKC activity and cytokinesis.

    Directory of Open Access Journals (Sweden)

    Nataliya Kotelevets

    Full Text Available Sphingosine kinases (SK catalyze the phosphorylation of proapoptotic sphingosine to the prosurvival factor sphingosine 1-phosphate (S1P, thereby promoting oncogenic processes. Breast (MDA-MB-231, lung (NCI-H358, and colon (HCT 116 carcinoma cells were transduced with shRNA to downregulate SK-1 expression or treated with a pharmacologic SK-1 inhibitor. The effects of SK-1 targeting were investigated by measuring the level of intracellular sphingosine, the activity of protein kinase C (PKC and cell cycle regulators, and the mitotic index. Functional assays included measurement of cell proliferation, colony formation, apoptosis, and cell cycle analysis. Downregulation of SK-1 or its pharmacologic inhibition increased intracellular sphingosine and decreased PKC activity as shown by reduced phosphorylation of PKC substrates. In MDA-MB-231 cells this effect was most pronounced and reduced cell proliferation and colony formation, which could be mimicked using exogenous sphingosine or the PKC inhibitor RO 31-8220. SK-1 downregulation in MDA-MB-231 cells increased the number of cells with 4N and 8N DNA content, and similar effects were observed upon treatment with sphingosine or inhibitors of SK-1 or PKC. Examination of cell cycle regulators unveiled decreased cdc2 activity and expression of Chk1, which may compromise spindle checkpoint function and cytokinesis. Indeed, SK-1 kd cells entered mitosis but failed to divide, and in the presence of taxol also failed to sustain mitotic arrest, resulting in further increased endoreduplication and apoptosis. Our findings delineate an intriguing link between SK-1, PKC and components of the cell cycle machinery, which underlines the significance of SK-1 as a target for cancer therapy.

  9. n-3 PUFAs reduce T-helper 17 cell differentiation by decreasing responsiveness to interleukin-6 in isolated mouse splenic CD4⁺ T cells.

    Science.gov (United States)

    Allen, M Jeannie; Fan, Yang-Yi; Monk, Jennifer M; Hou, Tim Y; Barhoumi, Rola; McMurray, David N; Chapkin, Robert S

    2014-08-01

    Cluster of differentiation 4(+) (CD4(+)) effector T-cell subsets [e.g., T-helper (Th) 1 and Th17] are implicated in autoimmune and inflammatory disorders such as multiple sclerosis, psoriasis, and rheumatoid arthritis. Interleukin (IL)-6 is a pleiotropic cytokine that induces Th17 polarization via signaling through the membrane-bound transducer glycoprotein 130 (GP130). Previously, we demonstrated that n-3 (ω-3) polyunsaturated fatty acids (PUFAs) reduce CD4(+) T-cell activation and differentiation into pathogenic Th17 cells by 25-30%. Here we report that n-3 PUFAs alter the response of CD4(+) T cells to IL-6 in a lipid raft membrane-dependent manner. Naive splenic CD4(+) T cells from fat-1 transgenic mice exhibited 30% lower surface expression of the IL-6 receptor. This membrane-bound receptor is known to be shed during cellular activation, but the release of soluble IL-6 receptor after treatment with anti-CD3 and anti-CD28 was not changed in the CD4(+) T cells from fat-1 mice, suggesting that the decrease in surface expression was not due to ectodomain release. We observed a significant 20% decrease in the association of GP130 with lipid rafts in activated fat-1 CD4(+) T cells and a 35% reduction in GP130 homodimerization, an obligate requirement for downstream signaling. The phosphorylation of signal transducer and activator of transcription 3 (STAT3), a downstream target of IL-6-dependent signaling, was also decreased by 30% in response to exogenous IL-6 in fat-1 CD4(+) T cells. Our results suggest that n-3 PUFAs suppress Th17 cell differentiation in part by reducing membrane raft-dependent responsiveness to IL-6, an essential polarizing cytokine. PMID:24944284

  10. Japanese encephalitis virus infects neural progenitor cells and decreases their proliferation.

    Science.gov (United States)

    Das, Sulagna; Basu, Anirban

    2008-08-01

    Japanese encephalitis virus (JEV), a common cause of encephalitis in humans, especially in children, leads to substantial neuronal injury. The survivors of JEV infection have severe cognitive impairment, motor and behavioral disorders. We hypothesize that depletion of neural progenitor cells (NPCs) by the virus culminates in neurological sequelae in survivors of Japanese encephalitis (JE). We utilized both in vivo model of JEV infection and in vitro neurosphere cultures to study progressive JEV infection. Cellular infection and cell death was determined by flow cytometry. BrdU administration in animals and in neurospheres was used to determine the proliferative ability of NPCs. JEV leads to massive loss of actively proliferating NPC population from the subventricular zone (SVZ). The ability of JEV infected subventricular zone cells to form neurospheres is severely compromised. This can be attributed to JEV infection in NPCs, which however do not result in robust death of the resilient NPC cells. Instead, JEV suppresses the cycling ability of these cells, preventing their proliferation. JEV primarily targets at a critical postnatal age and severely diminishes the NPC pool in SVZ, thus impairing the process of recovery after the insult. This arrested growth and proliferation of NPCs might have an effect on the neurological consequences in JE survivors. PMID:18540995

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

    Science.gov (United States)

    Ushijima, Hironori; Horyozaki, Akiko; Maeda, Masatomo

    2016-09-01

    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. PMID:27404124

  12. Fluid shear stress enhances the cell volume decrease of osteoblast cells by increasing the expression of the ClC-3 chloride channel

    OpenAIRE

    Liu, Li; Cai, Siyi; Qiu, Guixing; Lin, Jin

    2016-01-01

    ClC-3 is a volume-sensitive chloride channel that is responsible for cell volume adjustment and regulatory cell volume decrease (RVD). In order to evaluate the effects of fluid shear stress (FSS) stimulation on the osteoblast ClC-3 chloride channel, MC3T3-E1 cells were stimulated by FSS in the experimental group. Fluorescence quantitative polymerase chain reaction was used to detect changes in ClC-3 mRNA expression, the chloride ion fluorescent probe N-(ethoxycarbonylmethyl)-6-methoxyquinolin...

  13. Reduced cardiolipin content decreases respiratory chain capacities and increases ATP synthesis yield in the human HepaRG cells.

    Science.gov (United States)

    Peyta, Laure; Jarnouen, Kathleen; Pinault, Michelle; Guimaraes, Cyrille; Pais de Barros, Jean-Paul; Chevalier, Stephan; Dumas, Jean-François; Maillot, François; Hatch, Grant M; Loyer, Pascal; Servais, Stephane

    2016-04-01

    Cardiolipin (CL) is a unique mitochondrial phospholipid potentially affecting many aspects of mitochondrial function/processes, i.e. energy production through oxidative phosphorylation. Most data focusing on implication of CL content and mitochondrial bioenergetics were performed in yeast or in cellular models of Barth syndrome. Previous work reported that increase in CL content leads to decrease in liver mitochondrial ATP synthesis yield. Therefore the aim of this study was to determine the effects of moderate decrease in CL content on mitochondrial bioenergetics in human hepatocytes. For this purpose, we generated a cardiolipin synthase knockdown (shCLS) in HepaRG hepatoma cells showing bioenergetics features similar to primary human hepatocytes. shCLS cells exhibited a 55% reduction in CLS gene and a 40% decrease in protein expression resulting in a 45% lower content in CL compared to control (shCTL) cells. Oxygen consumption was significantly reduced in shCLS cells compared to shCTL regardless of substrate used and energy state analyzed. Mitochondrial low molecular weight supercomplex content was higher in shCLS cells (+60%) compared to shCTL. Significant fragmentation of the mitochondrial network was observed in shCLS cells compared to shCTL cells. Surprisingly, mitochondrial ATP synthesis was unchanged in shCLS compared to shCTL cells but exhibited a higher ATP:O ratio (+46%) in shCLS cells. Our results suggest that lowered respiratory chain activity induced by moderate reduction in CL content may be due to both destabilization of supercomplexes and mitochondrial network fragmentation. In addition, CL content may regulate mitochondrial ATP synthesis yield. PMID:26768115

  14. Erythropoietin reduces storage lesions and decreases apoptosis indices in blood bank red blood cells

    Directory of Open Access Journals (Sweden)

    Oscar Andrés Penuela

    2016-02-01

    Full Text Available ABSTRACT Background: Recent evidence shows a selective destruction of the youngest circulating red blood cells (neocytolysis trigged by a drop in erythropoietin levels. Objective: The aim of this study was to evaluate the effect of recombinant human erythropoietin beta on the red blood cell storage lesion and apoptosis indices under blood bank conditions. Methods: Each one of ten red blood cell units preserved in additive solution 5 was divided in two volumes of 100 mL and assigned to one of two groups: erythropoietin (addition of 665 IU of recombinant human erythropoietin and control (isotonic buffer solution was added. The pharmacokinetic parameters of erythropoietin were estimated and the following parameters were measured weekly, for six weeks: Immunoreactive erythropoietin, hemolysis, percentage of non-discocytes, adenosine triphosphate, glucose, lactate, lactate dehydrogenase, and annexin-V/esterase activity. The t-test or Wilcoxon's test was used for statistical analysis with significance being set for a p-value 6 weeks under blood bank conditions, with persistent supernatant concentrations of erythropoietin during the entire storage period. Adenosine triphosphate was higher in the Erythropoietin Group in Week 6 (4.19 ± 0.05 µmol/L vs. 3.53 ± 0.02 µmol/L; p-value = 0.009. The number of viable cells in the Erythropoietin Group was higher than in the Control Group (77% ± 3.8% vs. 71% ± 2.3%; p-value <0.05, while the number of apoptotic cells was lower (9.4% ± 0.3% vs. 22% ± 0.8%; p-value <0.05. Conclusions: Under standard blood bank conditions, an important proportion of red blood cells satisfy the criteria of apoptosis. Recombinant human erythropoietin beta seems to improve storage lesion parameters and mitigate apoptosis.

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

    Directory of Open Access Journals (Sweden)

    Edward Śpiewla

    2014-02-01

    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.

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

    Institute of Scientific and Technical Information of China (English)

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

    2010-01-01

    AIM: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.METHODS:Two HCC cell lines,HuH7 and HepG2,were used.Expression of RAGE was investigated by poly-merase chain reaction,Western blotting,and flow cytemetry(FACS).The effect of MK615 on RAGE expression was also evaluated by FACS.The proliferat...

  17. Breast tumors in PyMT transgenic mice expressing mitochondrial catalase have decreased labeling for macrophages and endothelial cells

    Directory of Open Access Journals (Sweden)

    Sy Fatemie

    2012-05-01

    Full Text Available We show by immunohistochemical labeling that prominent cell types in the tumor microenvironment of PyMT transgenic mice are tumor-associated macrophages (TAMs and endothelial cells, and that both populations are decreased in the presence of mitochondrial targeted catalase (mCAT. This observation suggests that mitochondrial ROS can drive tumor invasiveness in conjunction with the presence of TAMs and increased angiogenesis. Since primary PyMT tumor cells expressing mCAT undergo increased apoptosis, mitochondrial antioxidants might be attractive anti-tumor agents.

  18. Suppressed histamine release from rat peritoneal mast cells by ultraviolet B irradiation: decreased diacylglycerol formation as a possible mechanism

    International Nuclear Information System (INIS)

    This study was designed to investigate the effect of ultraviolet B (UVB) irradiation on mast cell functions. Purified mast cells obtained from rat peritoneal cavity were irradiated with UVB and subsequently exposed to a degranulator, compound 48/80, or the calcium ionophore A-23187. The amount of histamine released from mast cells measured by the enzyme isotopic assay was significantly decreased by UVB irradiation (100-400 mJ/cm2). Within this dose range, UVB alone was not cytotoxic to the cells because it did not induce histamine release. The suppression was observed when mast cells were subjected to degranulation without intervals after UVB irradiation, and even after 5 h postirradiation. The wavelength of 300 nm from a monochromatic light source showed the maximum effect. When mast cells prelabeled with [3H]arachidonate were irradiated and challenged by compound 48/80, label accumulation in diacylglycerol produced by the phosphatidylinositol cycle was considerably decreased by UVB irradiation. From these results, we hypothesize that, within an adequate irradiation dose, UVB irradiation suppresses histamine release from mast cells, probably by causing noncytotoxic damage to the membrane phospholipid metabolism, which is tied to the degranulation mechanisms

  19. 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.

  20. The high dosage of earthworm (Eisenia andrei) extract decreases cell proliferation and neuroblast differentiation in the mouse hippocampal dentate gyrus

    Science.gov (United States)

    Yan, Bing Chun; Yoo, Ki-Yeon; Park, Joon Ha; Lee, Choong Hyun; Choi, Jung Hoon

    2011-01-01

    Earthworm extract has shown anticancer characteristics. In the present study, we examined the effect of chronic treatment with a high dose of earthworm (Eisenia andrei) extract (EE) on cell proliferation and neuroblast differentiation in the hippocampal dentate gyrus (DG) of 3-week-old mice using 5-bromo-2'-deoxyuridine (BrdU) and Ki-67 immunohistochemistry for cell proliferation and doublecortin (DCX) immunohistochemistry for neuroblast differentiation, respectively. BrdU-, Ki-67-, and DCX-immunoreactive cells were easily detected in the subgranular zone of the DG in vehicle (saline)-treated mice. However, BrdU-, Ki-67-, and DCX-immunoreactive cells in the 500 mg/kg EE-treated mice decreased distinctively compared to those in the vehicle-treated mice. In addition, brain-derived neurotrophic factor (BDNF) immunoreactivity and its protein level decreased markedly in the DG of the EE-treated group compared to those in the vehicle-treated group. These results indicate that chronic treatment with high dose EE decreased cell proliferation and neuroblast differentiation, and that BDNF immunoreactivity decreased in the DG of EE-treated mice. PMID:22025974

  1. 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

    that received local pre-treatment with αGalCer prior to intra-vaginal HSV-2 infection had a lower mean disease score, mortality and viral load in the vagina following infection, compared to mice that did not receive αGalCer pre-treatment. Further, we found increased numbers of CD45 and NK1.1 positive cells...

  2. 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) for...

  3. 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

    % of maximal oxygen consumption. The intracellular expression of cytokines was detected following stimulation with ionomycin and phorbol 12-myristate 13-acetate in blood obtained before, during, and after exercise. The percentage of type 1 T cells in the circulation was suppressed at the end of...

  4. 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)

    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 3H-thymidine incorporation; lower IFNγ and TNFα production; higher CD4+CD27-CD28- and CD8+CD27-CD28- frequencies; lower CD4+CD25hi; and higher FoxP3 expression in CD8+CD25hi 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+CD25hi frequencies and their FoxP3 expression. These data indicate that FoxP3- and TGFβ1-expressing regulatory T cells contribute to decreased immunity in HAART recipients

  5. 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

  6. Differentiation of glioblastoma multiforme stem-like cells leads to downregulation of EGFR and EGFRvIII and decreased tumorigenic and stem-like cell potential

    DEFF Research Database (Denmark)

    Stockhausen, Marie-Thérése; Kristoffersen, Karina; Stobbe, Louise;

    2014-01-01

    Glioblastoma multiforme (GBM) is the most common and devastating primary brain tumor among adults. Despite recent treatment progress, most patients succumb to their disease within 2 years of diagnosis. Current research has highlighted the importance of a subpopulation of cells, assigned brain...... cancer stem-like cells (bCSC), to play a pivotal role in GBM malignancy. bCSC are identified by their resemblance to normal neural stem cells (NSC), and it is speculated that the bCSC have to be targeted in order to improve treatment outcome for GBM patients. One hallmark of GBM is aberrant expression......, differentiation is induced. Furthermore, we show that differentiation leads to decreased tumorigenic and stem cell-like potential of the neurosphere cultures and that by specifically inhibiting EGFR signaling it is possible to target the bCSC population. Our results suggest that differentiation therapy, possibly...

  7. Axonal outgrowth is associated with increased ERK 1/2 activation but decreased caspase 3 linked cell death in Schwann cells after immediate nerve repair in rats

    Directory of Open Access Journals (Sweden)

    Kanje Martin

    2011-01-01

    Full Text Available Abstract Background Extracellular-signal regulated kinase (ERK1/2 is activated by nerve damage and its activation precedes survival and proliferation of Schwann cells. In contrast, activation of caspase 3, a cysteine protease, is considered as a marker for apoptosis in Schwann cells. In the present study, axonal outgrowth, activation of ERK1/2 by phosphorylation (p-ERK 1/2 and immunoreactivity of cleaved caspase 3 were examined after immediate, delayed, or no repair of transected rat sciatic nerves. Results Axonal outgrowth, detected by neurofilament staining, was longer after immediate repair than after either the delayed or no repair conditions. Immediate repair also showed a higher expression of p-ERK 1/2 and a lower number of cleaved caspase 3 stained Schwann cells than after delayed nerve repair. If the transected nerve was not repaired a lower level of p-ERK 1/2 was found than in either the immediate or delayed repair conditions. Axonal outgrowth correlated to p-ERK 1/2, but not clearly with cleaved caspase 3. Contact with regenerating axons affected Schwann cells with respect to p-ERK 1/2 and cleaved caspase 3 after immediate nerve repair only. Conclusion The decreased regenerative capacity that has historically been observed after delayed nerve repair may be related to impaired activation of Schwann cells and increased Schwann cell death. Outgrowing axons influence ERK 1/2 activation and apoptosis of Schwann cells.

  8. Altered connexin 43 expression underlies age-dependent decrease of regulatory T cell suppressor function in nonobese diabetic mice.

    Science.gov (United States)

    Kuczma, Michal; Wang, Cong-Yi; Ignatowicz, Leszek; Gourdie, Robert; Kraj, Piotr

    2015-06-01

    Type 1 diabetes is one of the most extensively studied autoimmune diseases, but the cellular and molecular mechanisms leading to T cell-mediated destruction of insulin-producing β cells are still not well understood. In this study, we show that regulatory T cells (T(regs)) in NOD mice undergo age-dependent loss of suppressor functions exacerbated by the decreased ability of activated effector T cells to upregulate Foxp3 and generate T(regs) in the peripheral organs. This age-dependent loss is associated with reduced intercellular communication mediated by gap junctions, which is caused by impaired upregulation and decreased expression of connexin 43. Regulatory functions can be corrected, even in T cells isolated from aged, diabetic mice, by a synergistic activity of retinoic acid, TGF-β, and IL-2, which enhance connexin 43 and Foxp3 expression in T(regs) and restore the ability of conventional CD4(+) T cells to upregulate Foxp3 and generate peripherally derived T(regs). Moreover, we demonstrate that suppression mediated by T(regs) from diabetic mice is enhanced by a novel reagent, which facilitates gap junction aggregation. In summary, our report identifies gap junction-mediated intercellular communication as an important component of the T(reg) suppression mechanism compromised in NOD mice and suggests how T(reg) mediated immune regulation can be improved. PMID:25911751

  9. Modulation of oxidative stress and subsequent induction of apoptosis and endoplasmic reticulum stress allows citral to decrease cancer cell proliferation.

    Science.gov (United States)

    Kapur, Arvinder; Felder, Mildred; Fass, Lucas; Kaur, Justanjot; Czarnecki, Austin; Rathi, Kavya; Zeng, San; Osowski, Kathryn Kalady; Howell, Colin; Xiong, May P; Whelan, Rebecca J; Patankar, Manish S

    2016-01-01

    The monoterpenoid, citral, when delivered through PEG-b-PCL nanoparticles inhibits in vivo growth of 4T1 breast tumors. Here, we show that citral inhibits proliferation of multiple human cancer cell lines. In p53 expressing ECC-1 and OVCAR-3 but not in p53-deficient SKOV-3 cells, citral induces G1/S cell cycle arrest and apoptosis as determined by Annexin V staining and increased cleaved caspase3 and Bax and decreased Bcl-2. In SKOV-3 cells, citral induces the ER stress markers CHOP, GADD45, EDEM, ATF4, Hsp90, ATG5, and phospho-eIF2α. The molecular chaperone 4-phenylbutyric acid attenuates citral activity in SKOV-3 but not in ECC-1 and OVCAR-3 cells. In p53-expressing cells, citral increases phosphorylation of serine-15 of p53. Activation of p53 increases Bax, PUMA, and NOXA expression. Inhibition of p53 by pifithrin-α, attenuates citral-mediated apoptosis. Citral increases intracellular oxygen radicals and this leads to activation of p53. Inhibition of glutathione synthesis by L-buthionine sulfoxamine increases potency of citral. Pretreatment with N-acetylcysteine decreases phosphorylation of p53 in citral-treated ECC-1 and OVCAR-3. These results define a p53-dependent, and in the absence of p53, ER stress-dependent mode of action of citral. This study indicates that citral in PEG-b-PCL nanoparticle formulation should be considered for treatment of breast and other tumors. PMID:27270209

  10. Aspiration, but not injection, decreases cultured equine mesenchymal stromal cell viability

    OpenAIRE

    Williams, Lynn B.; Russell, Keith A.; Koenig, Judith B.; Thomas G. Koch

    2016-01-01

    Background Recently, equine multipotent mesenchymal stromal cells (MSC) have received significant attention as therapy for various conditions due to their proposed regenerative and immune-modulating capacity. MSC are commonly administered to the patient through a hypodermic needle. Currently, little information is available on the effect of such injection has on equine MSC immediate and delayed viability. We hypothesize that viability of equine MSC is not correlated with needle diameter durin...

  11. gamma-Aminobutyric acid antagonists decrease junctional communication between L-horizontal cells of the retina.

    OpenAIRE

    Piccolino, M; Neyton, J; Witkovsky, P; Gerschenfeld, H M

    1982-01-01

    The antagonists of gamma-aminobutyric acid, bicuculline and picrotoxin, were found to narrow the receptive field profile of the large field horizontal cell (L1HC) in the turtle retina when added to the perfusion medium in micromolar concentrations. The coupling resistance between neighboring L1HCs was increased by bicuculline or picrotoxin. Under control conditions, the dye Lucifer yellow injected into one L1HC diffused into a large number of neighboring L1HCs; bicuculline or picrotoxin great...

  12. A Decrease in Ambient Temperature Induces Post-Mitotic Enlargement of Palisade Cells in North American Lake Cress.

    Directory of Open Access Journals (Sweden)

    Rumi Amano

    Full Text Available In order to maintain organs and structures at their appropriate sizes, multicellular organisms orchestrate cell proliferation and post-mitotic cell expansion during morphogenesis. Recent studies using Arabidopsis leaves have shown that compensation, which is defined as post-mitotic cell expansion induced by a decrease in the number of cells during lateral organ development, is one example of such orchestration. Some of the basic molecular mechanisms underlying compensation have been revealed by genetic and chimeric analyses. However, to date, compensation had been observed only in mutants, transgenics, and γ-ray-treated plants, and it was unclear whether it occurs in plants under natural conditions. Here, we illustrate that a shift in ambient temperature could induce compensation in Rorippa aquatica (Brassicaceae, a semi-aquatic plant found in North America. The results suggest that compensation is a universal phenomenon among angiosperms and that the mechanism underlying compensation is shared, in part, between Arabidopsis and R. aquatica.

  13. 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.

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

    International Nuclear Information System (INIS)

    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

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

    International Nuclear Information System (INIS)

    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

  16. Eckol inhibits ultraviolet B-induced cell damage in human keratinocytes via a decrease in oxidative stress

    International Nuclear Information System (INIS)

    In previous reports, the antioxidant effects of eckol were shown to protect cells against hydrogen peroxide- and gamma ray-induced oxidative stress. In this study, the role of eckol in protecting human skin keratinocytes (HaCaT) against UVB-induced oxidative cell damage was investigated. Also, triphlorethol-A, one of the chemical components in Ecklonia cava, and quercetin a well known antioxidant, were compared with eckol in terms of antioxidant activity based on chemical structure. Eckol decreased UVB-induced intracellular reactive oxygen species (ROS), decreased injury to cellular components resulting from UVB-induced oxidative stress, and restored cell viability. In addition, eckol reduced UVB-induced apoptosis by inhibiting the disruption of mitochondrial membranes. These results suggest that eckol protects human keratinocytes against UVB-induced oxidative stress by scavenging ROS, thereby lessening injury to cellular components. (author)

  17. Telomerase inhibition decreases alpha-fetoprotein expression and secretion by hepatocellular carcinoma cell lines: in vitro and in vivo study.

    Science.gov (United States)

    Tahtouh, Roula; Azzi, Anne-Sophie; Alaaeddine, Nada; Chamat, Soulaima; Bouharoun-Tayoun, Hasnaa; Wardi, Layal; Raad, Issam; Sarkis, Riad; Antoun, Najibe Abou; Hilal, George

    2015-01-01

    Alpha-fetoprotein (AFP) is a diagnostic marker for hepatocellular carcinoma (HCC). A direct relationship between poor prognosis and the concentration of serum AFP has been observed. Telomerase, an enzyme that stabilizes the telomere length, is expressed by 90% of HCC. The aim of this study was to investigate the effect of telomerase inhibition on AFP secretion and the involvement of the PI3K/Akt/mTOR signaling pathway. Proliferation and viability tests were performed using tetrazolium salt. Apoptosis was determined through the Annexin V assay using flow cytometry. The concentrations of AFP were measured using ELISA kits. The AFP mRNA expression was evaluated using RT-PCR, and cell migration was evaluated using a Boyden chamber assay. The in vivo effect of costunolide on AFP production was tested in NSG mice. Telomerase inhibition by costunolide and BIBR 1532 at 5 and 10 μM decreased AFP mRNA expression and protein secretion by HepG2/C3A cells. The same pattern was obtained with cells treated with hTERT siRNA. This treatment exhibited no apoptotic effect. The AFP mRNA expression and protein secretion by PLC/PRF/5 was decreased after treatment with BIBR1532 at 10 μM. In contrast, no effect was obtained for PLC/PRF/5 cells treated with costunolide at 5 or 10 μM. Inhibition of the PI3K/Akt/mTOR signaling pathway decreased the AFP concentration. In contrast, the MAPK/ERK pathway appeared to not be involved in HepG2/C3A cells, whereas ERK inhibition decreased the AFP concentration in PLC/PRF/5 cells. Modulation of the AFP concentration was also obtained after the inhibition or activation of PKC. Costunolide (30 mg/kg) significantly decreased the AFP serum concentration of NSG mice bearing HepG2/C3A cells. Both the inhibition of telomerase and the inhibition of the PI3K/Akt/mTOR signaling pathway decreased the AFP production of HepG2/C3A and PLC/PRF/5 cells, suggesting a relationship between telomerase and AFP expression through the PI3K/Akt/mTOR pathway. PMID:25822740

  18. Telomerase inhibition decreases alpha-fetoprotein expression and secretion by hepatocellular carcinoma cell lines: in vitro and in vivo study.

    Directory of Open Access Journals (Sweden)

    Roula Tahtouh

    Full Text Available Alpha-fetoprotein (AFP is a diagnostic marker for hepatocellular carcinoma (HCC. A direct relationship between poor prognosis and the concentration of serum AFP has been observed. Telomerase, an enzyme that stabilizes the telomere length, is expressed by 90% of HCC. The aim of this study was to investigate the effect of telomerase inhibition on AFP secretion and the involvement of the PI3K/Akt/mTOR signaling pathway. Proliferation and viability tests were performed using tetrazolium salt. Apoptosis was determined through the Annexin V assay using flow cytometry. The concentrations of AFP were measured using ELISA kits. The AFP mRNA expression was evaluated using RT-PCR, and cell migration was evaluated using a Boyden chamber assay. The in vivo effect of costunolide on AFP production was tested in NSG mice. Telomerase inhibition by costunolide and BIBR 1532 at 5 and 10 μM decreased AFP mRNA expression and protein secretion by HepG2/C3A cells. The same pattern was obtained with cells treated with hTERT siRNA. This treatment exhibited no apoptotic effect. The AFP mRNA expression and protein secretion by PLC/PRF/5 was decreased after treatment with BIBR1532 at 10 μM. In contrast, no effect was obtained for PLC/PRF/5 cells treated with costunolide at 5 or 10 μM. Inhibition of the PI3K/Akt/mTOR signaling pathway decreased the AFP concentration. In contrast, the MAPK/ERK pathway appeared to not be involved in HepG2/C3A cells, whereas ERK inhibition decreased the AFP concentration in PLC/PRF/5 cells. Modulation of the AFP concentration was also obtained after the inhibition or activation of PKC. Costunolide (30 mg/kg significantly decreased the AFP serum concentration of NSG mice bearing HepG2/C3A cells. Both the inhibition of telomerase and the inhibition of the PI3K/Akt/mTOR signaling pathway decreased the AFP production of HepG2/C3A and PLC/PRF/5 cells, suggesting a relationship between telomerase and AFP expression through the PI3K

  19. Decreased T Follicular Regulatory Cell/T Follicular Helper Cell (TFH) in Simian Immunodeficiency Virus-Infected Rhesus Macaques May Contribute to Accumulation of TFH in Chronic Infection.

    Science.gov (United States)

    Chowdhury, Ankita; Del Rio Estrada, Perla Mariana; Del Rio, Perla Maria Estrada; Tharp, Greg K; Trible, Ronald P; Amara, Rama R; Chahroudi, Ann; Reyes-Teran, Gustavo; Bosinger, Steven E; Silvestri, Guido

    2015-10-01

    T follicular helper cells (TFH) are critical for the development and maintenance of germinal center (GC) and humoral immune responses. During chronic HIV/SIV infection, TFH accumulate, possibly as a result of Ag persistence. The HIV/SIV-associated TFH expansion may also reflect lack of regulation by suppressive follicular regulatory CD4(+) T cells (TFR). TFR are natural regulatory T cells (TREG) that migrate into the follicle and, similar to TFH, upregulate CXCR5, Bcl-6, and PD1. In this study, we identified TFR as CD4(+)CD25(+)FOXP3(+)CXCR5(+)PD1(hi)Bcl-6(+) within lymph nodes of rhesus macaques (RM) and confirmed their localization within the GC by immunohistochemistry. RNA sequencing showed that TFR exhibit a distinct transcriptional profile with shared features of both TFH and TREG, including intermediate expression of FOXP3, Bcl-6, PRDM1, IL-10, and IL-21. In healthy, SIV-uninfected RM, we observed a negative correlation between frequencies of TFR and both TFH and GC B cells, as well as levels of CD4(+) T cell proliferation. Post SIV infection, the TFR/TFH ratio was reduced with no change in the frequency of TREG or TFR within the total CD4(+) T cell pool. Finally, we examined whether higher levels of direct virus infection of TFR were responsible for their relative depletion post SIV infection. We found that TFH, TFR, and TREG sorted from SIV-infected RM harbor comparable levels of cell-associated viral DNA. Our data suggest that TFR may contribute to the regulation and proliferation of TFH and GC B cells in vivo and that a decreased TFR/TFH ratio in chronic SIV infection may lead to unchecked expansion of both TFH and GC B cells. PMID:26297764

  20. Ferulic acid decreases cell viability and colony formation while inhibiting migration of MIA PaCa-2 human pancreatic cancer cells in vitro.

    Science.gov (United States)

    Fahrioğlu, Umut; Dodurga, Yavuz; Elmas, Levent; Seçme, Mücahit

    2016-01-15

    Novel and combinatorial treatment methods are becoming sought after entities in cancer treatment and these treatments are even more valuable for pancreatic cancer. The scientists are always on the lookout for new chemicals to help them in their fight against cancer. In this study, we examine the effects of ferulic acid (FA), a phenolic compound, on gene expression, viability, colony formation and migration/invasion in the cultured MIA PaCa-2 human pancreatic cancer cell. Cytotoxic effects of FA were determined by using trypan blue dye exclusion test and Cell TiterGlo (CTG) assay. IC50 dose in MIA PaCa-2 cells was detected as 500μM/ml at the 72nd hour. Expression profiles of certain cell cycle and apoptosis genes such as CCND1 (cyclin D1),CDK4, CDK6, RB, p21, p16, p53, caspase-3, caspase-9, caspase-8, caspase-10, Bcl-2, BCL-XL,BID, DR4,DR5,FADD,TRADD,PARP, APAF, Bax, Akt, PTEN, PUMA, NOXA, MMP2, MMP9, TIMP1 and TIMP2 were determined by real-time PCR. The effect of FA on cell viability was determined by CellTiter-Glo® Luminescent Cell Viability Assay. Additionally, effects of FA on colony formation and invasion were also investigated. It was observed that FA caused a significant decrease in the expression of CCND1, CDK 4/6, Bcl2 and caspase 8 and 10 in the MIA PaCa-2 cells while causing an increase in the expression of p53, Bax, PTEN caspase 3 and 9. FA was observed to decrease colony formation while inhibiting cell invasion and migration as observed by the BioCoat Matrigel Invasion Chamber guide and colony formation assays. In conclusion, FA is thought to behave as an anti-cancer agent by affecting cell cycle, apoptotic, invasion and colony formation behavior of MIA PaCa-2 cells. Therefore, FA is placed as a strong candidate for further studies aimed at finding a better, more effective treatment approach for pancreatic cancer. PMID:26516023

  1. 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.

  2. 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. PMID:27369448

  3. Combinatorial PX-866 and Raloxifene Decrease Rb Phosphorylation, Cyclin E2 Transcription, and Proliferation of MCF-7 Breast Cancer Cells.

    Science.gov (United States)

    Peek, Gregory W; Tollefsbol, Trygve O

    2016-07-01

    As a potential means to reduce proliferation of breast cancer cells, a multiple-pathway approach with no effect on control cells was explored. The human interactome being constructed by the Center for Cancer Systems Biology will prove indispensable to understanding composite effects of multiple pathways, but its discovered protein-protein interactions require characterization. Accordingly, we explored the effects of regulators of one protein on downstream targets of the other protein. MCF-7 estrogen receptor-positive (ER+) breast cancer cells were treated with raloxifene to upregulate the TGF-β pathway and PX-866 to down-regulate the PI3K/Akt pathway. This resulted in highly significant downstream reduction of cell cycle proliferation in breast cancer cells with no significant proliferation reduction following similar treatment of noncancerous MCF10A breast epithelial cells. Reduced phosphorylation of p107 and substantial reduction of Rb phosphorylation were observed in response. The effects of reduced Rb and p107 phosphorylation were reflected in significant decline in E2F-1 transcriptional activity, which is dependent on pocket protein phosphorylation status. The reduced proliferation was related to decreased expression of cyclins, including E2F-1-regulated Cyclin E2, which was also in response to raloxifene and PX-866. All combinations of raloxifene and PX-866 produced significant or highly significant results for reduced MCF-7 cell proliferation, reduced Cyclin E2 transcription, and reduced Rb phosphorylation. These studies demonstrated that uncontrolled proliferation of ER+ breast cancer cells can be significantly reduced by combinational targeting of two relevant pathways. J. Cell. Biochem. 117: 1688-1696, 2016. © 2015 Wiley Periodicals, Inc. PMID:26660119

  4. Moderate Hypothermia Significantly Decreases Hippocampal Cell Death Involving Autophagy Pathway after Moderate Traumatic Brain Injury.

    Science.gov (United States)

    Jin, Yichao; Lin, Yingying; Feng, Jun-feng; Jia, Feng; Gao, Guo-yi; Jiang, Ji-yao

    2015-07-15

    Here, we evaluated changes in autophagy after post-traumatic brain injury (TBI) followed by moderate hypothermia in rats. Adult male Sprague-Dawley rats were randomly divided into four groups: sham injury with normothermia group (37 °C); sham injury with hypothermia group (32 °C); TBI with normothermia group (TNG; 37 °C); and TBI with hypothermia group (THG; 32 °C). Injury was induced by a fluid percussion TBI device. Moderate hypothermia (32 °C) was achieved by partial immersion in a water bath (0 °C) under general anesthesia for 4 h. All rats were killed at 24 h after fluid percussion TBI. The ipsilateral hippocampus in all rats was analyzed with hematoxylin and eosin staining; terminal deoxynucleoitidyl transferase-mediated nick end labeling staining was used to determine cell death in ipsilateral hippocampus. Immunohistochemistry and western blotting of microtubule-associated protein light chain 3 (LC3), Beclin-1, as well as transmission electron microscopy performed to assess changes in autophagy. At 24 h after TBI, the cell death index was 27.90 ± 2.36% in TNG and 14.90 ± 1.52% in THG. Expression level of LC3 and Beclin-1 were significantly increased after TBI and were further up-regulated after post-TBI hypothermia. Further, ultrastructural observations showed that there was a marked increase of autophagosomes and autolysosomes in ipsilateral hippocampus after post-TBI hypothermia. Our data demonstrated that moderate hypothermia significantly attenuated cell death and increased autophagy in ipsilateral hippocampus after fluid percussion TBI. In conclusion, autophagy pathway may participate in the neuroprotective effect of post-TBI hypothermia. PMID:25942484

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

    Science.gov (United States)

    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. PMID:23353780

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

    International Nuclear Information System (INIS)

    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

  7. 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.

  8. Exercise-induced norepinephrine decreases circulating hematopoietic stem and progenitor cell colony-forming capacity.

    Directory of Open Access Journals (Sweden)

    Julia M Kröpfl

    Full Text Available A recent study showed that ergometry increased circulating hematopoietic stem and progenitor cell (CPC numbers, but reduced hematopoietic colony forming capacity/functionality under normoxia and normobaric hypoxia. Herein we investigated whether an exercise-induced elevated plasma free/bound norepinephrine (NE concentration could be responsible for directly influencing CPC functionality. Venous blood was taken from ten healthy male subjects (25.3+/-4.4 yrs before and 4 times after ergometry under normoxia and normobaric hypoxia (FiO2<0.15. The circulating hematopoietic stem and progenitor cell numbers were correlated with free/bound NE, free/bound epinephrine (EPI, cortisol (Co and interleukin-6 (IL-6. Additionally, the influence of exercise-induced NE and blood lactate (La on CPC functionality was analyzed in a randomly selected group of subjects (n = 6 in vitro under normoxia by secondary colony-forming unit granulocyte macrophage assays. Concentrations of free NE, EPI, Co and IL-6 were significantly increased post-exercise under normoxia/hypoxia. Ergometry-induced free NE concentrations found in vivo showed a significant impairment of CPC functionality in vitro under normoxia. Thus, ergometry-induced free NE was thought to trigger CPC mobilization 10 minutes post-exercise, but as previously shown impairs CPC proliferative capacity/functionality at the same time. The obtained results suggest that an ergometry-induced free NE concentration has a direct negative effect on CPC functionality. Cortisol may further influence CPC dynamics and functionality.

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

    International Nuclear Information System (INIS)

    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

  10. Decreased heat shock protein 27 expression and altered autophagy in human cells harboring A8344G mitochondrial DNA mutation.

    Science.gov (United States)

    Chen, Chin-Yi; Chen, Hsueh-Fu; Gi, Siao-Jhen; Chi, Tang-Hao; Cheng, Che-Kun; Hsu, Chi-Fu; Ma, Yi-Shing; Wei, Yau-Huei; Liu, Chin-Shan; Hsieh, Mingli

    2011-09-01

    Mitochondrial DNA (mtDNA) mutations are responsible for human neuromuscular diseases caused by mitochondrial dysfunction. Myoclonus epilepsy associated with ragged-red fibers (MERRF) is a maternally inherited mitochondrial encephalomyopathy with various syndromes involving both muscular and nervous systems. The most common mutation in MERRF syndrome, A8344G mutation in mtDNA, has been associated with severe defects in protein synthesis. This defect impairs assembly of complexes in electron transport chain and results in decreased respiratory function of mitochondria. In this study, we showed a significant decrease of the heat shock protein 27 (Hsp27) in lymphoblastoid cells derived from a MERRF patient and in cybrid cells harboring MERRF A8344G mutation. However, normal cytoplasmic distributions of Hsp27 and normal heat shock responses were observed in both wild type and mutant cybrids. Furthermore, overexpression of wild type Hsp27 in mutant MERRF cybrids significantly decreased cell death under staurosporine (STS) treatment, suggesting a protective function of Hsp27 in cells harboring the A8344G mutation of mtDNA. Meanwhile, reverse transcriptase PCR showed no difference in the mRNA level between normal and mutant cybrids, indicating that alterations may occur at the protein level. Evidenced by the decreased levels of Hsp27 upon treatment with proteasome inhibitor, starvation and rapamycin and the accumulation of Hsp27 upon lysosomal inhibitor treatment; Hsp27 may be degraded by the autophagic pathway. In addition, the increased formation of LC3-II and autophagosomes was found in MERRF cybrids under the basal condition, indicating a constitutively-activated autophagic pathway. It may explain, at least partially, the faster turnover of Hsp27 in MERRF cybrids. This study provides information for us to understand that Hsp27 is degraded through the autophagic pathway and that Hsp27 may have a protective role in MERRF cells. Regulating Hsp27 and the autophagic pathway

  11. Human placenta-derived stromal cells decrease inflammation, placental injury and blood pressure in hypertensive pregnant mice.

    Science.gov (United States)

    Chatterjee, Piyali; Chiasson, Valorie L; Pinzur, Lena; Raveh, Shani; Abraham, Eytan; Jones, Kathleen A; Bounds, Kelsey R; Ofir, Racheli; Flaishon, Liat; Chajut, Ayelet; Mitchell, Brett M

    2016-04-01

    Pre-eclampsia, the development of hypertension and proteinuria or end-organ damage during pregnancy, is a leading cause of both maternal and fetal morbidity and mortality, and there are no effective clinical treatments for pre-eclampsia aside from delivery. The development of pre-eclampsia is characterized by maladaptation of the maternal immune system, excessive inflammation and endothelial dysfunction. We have reported that detection of extracellular RNA by the Toll-like receptors (TLRs) 3 and 7 is a key initiating signal that contributes to the development of pre-eclampsia. PLacental eXpanded (PLX-PAD) cells are human placenta-derived, mesenchymal-like, adherent stromal cells that have anti-inflammatory, proangiogenic, cytoprotective and regenerative properties, secondary to paracrine secretion of various molecules in response to environmental stimulation. We hypothesized that PLX-PAD cells would reduce the associated inflammation and tissue damage and lower blood pressure in mice with pre-eclampsia induced by TLR3 or TLR7 activation. Injection of PLX-PAD cells on gestational day 14 significantly decreased systolic blood pressure by day 17 in TLR3-induced and TLR7-induced hypertensive mice (TLR3 144-111 mmHg; TLR7 145-106 mmHg; both Presponses improved significantly in TLR3-induced and TLR7-induced hypertensive mice that received PLX-PAD cells on gestational day 14 (TLR3 35-65%; TLR7 37-63%; both Psystemic inflammation and placental injury, increased markedly in both groups of TLR-induced hypertensive mice, were reduced by PLX-PAD cells. Importantly, PLX-PAD cell therapy had no effects on these measures in pregnant control mice or on the fetuses. These data demonstrate that PLX-PAD cell therapy can safely reverse pre-eclampsia-like features during pregnancy and have a potential therapeutic role in pre-eclampsia treatment. PMID:26685104

  12. Expression of FK506 binding protein 65 (FKBP65) is decreased in epithelial ovarian cancer cells compared to benign tumor cells and to ovarian epithelium

    DEFF Research Database (Denmark)

    Henriksen, Rudi; Sørensen, Flemming Brandt; Ørntoft, Torben Falck; Birkenkamp-Demtroder, Karin

    2011-01-01

    followed by a strongly increased risk of ovarian cysts. We performed the present study to reveal how FKBP65 is expressed in the ovary and in ovarian tumors and to see if this expression might be related to ovarian tumor development, a relationship we have found in colorectal cancer. Biopsies from...... prospectively collected samples from ovaries and benign, borderline, and invasive ovarian tumors were analyzed for expression of FKBP65 by immunohistochemistry. The expression was compared to survival and several clinicopathological parameters. FKBP65 is strongly expressed in ovarian epithelium and in benign...... ovarian tumor cells. In the ovary, a positive staining was also found in endothelial cells of blood vessels. In non-invasive and in invasive malignant tumor cells, a decreased staining was observed, which was not correlated to stage, histology, or survival. A significant inversed correlation to expression...

  13. Decreased [18F]fluoro-2-deoxy-D-glucose incorporation and increased glucose transport are associated with resistance to 5FU in MCF7 cells in vitro

    International Nuclear Information System (INIS)

    Introduction: Tumor refractoriness to chemotherapy is frequently due to the acquisition of resistance. Resistant cells selected by exposure to chemotherapy agents may exhibit differences in [18F]fluoro-2-deoxy-D-glucose (FDG) incorporation, as compared with sensitive cells. Methods: FDG incorporation, hexokinase (HK) activity, glucose transport and ATP content were determined in clones of 5-fluorouracil (5FU)-resistant MCF7 cells, established by long-term exposure to increasing 5FU concentrations, and in parental MCF7 cells. Results: FDG incorporation was decreased in MCF7 cells resistant to 5FU; HK activity was similar in the resistant and sensitive cells, while glucose transport was increased, as compared with sensitive cells. Treatment of cells with the glucose efflux inhibitor phloretin increased FDG incorporation to similar levels in the resistant and sensitive cells. Analysis of microarray data demonstrated the expression of GLUT1, 8 and 10 transporters in MCF7 cells. GLUT8 and 10 expression was decreased in the resistant cells, while GLUT1 was only increased in cells resistant to the lowest 5FU concentration. Conclusion: FDG incorporation in 5FU-resistant MCF7 cells is decreased, as compared with sensitive cells. Our findings also suggest that this may be due to high rates of membrane glucose transport in the resistant cells resulting in enhanced efflux of FDG. We believe that this is the first demonstration that facilitative glucose transporters can actually decrease the incorporation of FDG

  14. Decreasing methylation of pectin caused by nitric oxide leads to higher aluminium binding in cell walls and greater aluminium sensitivity of wheat roots

    OpenAIRE

    Sun, Chengliang; Lu, Lingli; Yu, Yan; Liu, Lijuan; Hu, Yan; Ye, Yiquan; Jin, Chongwei; Lin, Xianyong

    2015-01-01

    Highlight Aluminium-induced nitric oxide production enhances the aluminium sensitivity of wheat by decreasing pectin methylation of root cell-wall pectin, resulting in greater aluminium binding in root cell walls.

  15. Aspirin and salicylic acid decrease c-Myc expression in cancer cells: a potential role in chemoprevention.

    Science.gov (United States)

    Ai, Guoqiang; Dachineni, Rakesh; Muley, Pratik; Tummala, Hemachand; Bhat, G Jayarama

    2016-02-01

    Epidemiological studies have demonstrated a significant correlation between regular aspirin use and reduced colon cancer incidence and mortality; however, the pathways by which it exerts its anti-cancer effects are still not fully explored. We hypothesized that aspirin's anti-cancer effect may occur through downregulation of c-Myc gene expression. Here, we demonstrate that aspirin and its primary metabolite, salicylic acid, decrease the c-Myc protein levels in human HCT-116 colon and in few other cancer cell lines. In total cell lysates, both drugs decreased the levels of c-Myc in a concentration-dependent fashion. Greater inhibition was observed in the nucleus than the cytoplasm, and immunofluorescence studies confirmed these observations. Pretreatment of cells with lactacystin, a proteasome inhibitor, partially prevented the downregulatory effect of both aspirin and salicylic acid, suggesting that 26S proteasomal pathway is involved. Both drugs failed to decrease exogenously expressed DDK-tagged c-Myc protein levels; however, under the same conditions, the endogenous c-Myc protein levels were downregulated. Northern blot analysis showed that both drugs caused a decrease in c-Myc mRNA levels in a concentration-dependent fashion. High-performance liquid chromatography (HPLC) analysis showed that aspirin taken up by cells was rapidly metabolized to salicylic acid, suggesting that aspirin's inhibitory effect on c-Myc may occur through formation of salicylic acid. Our result suggests that salicylic acid regulates c-Myc level at both transcriptional and post-transcription levels. Inhibition of c-Myc may represent an important pathway by which aspirin exerts its anti-cancer effect and decrease the occurrence of cancer in epithelial tissues. PMID:26314861

  16. Cadmium Induced Cell Apoptosis, DNA Damage, Decreased DNA Repair Capacity, and Genomic Instability during Malignant Transformation of Human Bronchial Epithelial Cells

    OpenAIRE

    Zhou, Zhiheng; Wang, Caixia; Liu, Haibai; Huang, Qinhai; Wang, Min; Lei, Yixiong

    2013-01-01

    Cadmium and its compounds are well-known human carcinogens, but the mechanisms underlying the carcinogenesis are not entirely understood. Our study was designed to elucidate the mechanisms of DNA damage in cadmium-induced malignant transformation of human bronchial epithelial cells. We analyzed cell cycle, apoptosis, DNA damage, gene expression, genomic instability, and the sequence of exons in DNA repair genes in several kinds of cells. These cells consisted of untreated control cells, cells...

  17. C-reactive protein decreases expression of VEGF receptors and neuropilins and inhibits VEGF165-induced cell proliferation in human endothelial cells

    International Nuclear Information System (INIS)

    C-reactive protein (CRP) is associated with cardiovascular disease. However, its biological functions for the vascular system are largely unknown. The objective of this study was to determine whether CRP could affect endothelial cell proliferation and expression of VEGF receptors (VEGFRs) and/or neuropilins. Human coronary artery endothelial cells (HCAECs) treated with CRP showed a significant reduction of mRNA levels of VEGFR-2, VEGFR-3, NRP-1, and NRP-2 by 34%, 63%, 41%, and 43%, respectively, as compared to untreated control cells (p 165-induced cell proliferation was determined by [3H]thymidine incorporation and MTS assay as well as capillary-like tube formation on Matrigel. HCAECs pretreated with CRP significantly decreased VEGF165-induced [3H]thymidine incorporation by 73%, MTS absorbance by 44%, and capillary-like tube formation by 54% as compared to CRP-untreated cells (p 165-induced HCAEC proliferation and capillary-like tube formation through downregulation of expression of VEGFRs and NRPs. This study suggests a new molecular mechanism underlying the adverse effect of CRP on the vascular system

  18. 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.

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

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    Madalina Rujoi

    Full Text Available BACKGROUND: Niemann-Pick type C (NPC disease is a genetically inherited multi-lipid storage disorder with impaired efflux of cholesterol from lysosomal storage organelles. METHODOLOGY/PRINCIPAL FINDINGS: 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. CONCLUSIONS/SIGNIFICANCE: 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.

  20. Human intestinal dendritic cells decrease cytokine release against Salmonella infection in the presence of Lactobacillus paracasei upon TLR activation.

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    Miriam Bermudez-Brito

    Full Text Available Probiotic bacteria have been shown to modulate immune responses and could have therapeutic effects in allergic and inflammatory disorders. However, little is known about the signalling pathways that are engaged by probiotics. Dendritic cells (DCs are antigen-presenting cells that are involved in immunity and tolerance. Monocyte-derived dendritic cells (MoDCs and murine DCs are different from human gut DCs; therefore, in this study, we used human DCs generated from CD34+ progenitor cells (hematopoietic stem cells harvested from umbilical cord blood; those DCs exhibited surface antigens of dendritic Langerhans cells, similar to the lamina propria DCs in the gut. We report that both a novel probiotic strain isolated from faeces of exclusively breast-fed newborn infants, Lactobacillus paracasei CNCM I-4034, and its cell-free culture supernatant (CFS decreased pro-inflammatory cytokines and chemokines in human intestinal DCs challenged with Salmonella. Interestingly, the supernatant was as effective as the bacteria in reducing pro-inflammatory cytokine expression. In contrast, the bacterium was a potent inducer of TGF-β2 secretion, whereas the supernatant increased the secretion of TGF-β1 in response to Salmonella. We also showed that both the bacteria and its supernatant enhanced innate immunity through the activation of Toll-like receptor (TLR signalling. These treatments strongly induced the transcription of the TLR9 gene. In addition, upregulation of the CASP8 and TOLLIP genes was observed. This work demonstrates that L. paracasei CNCM I-4034 enhanced innate immune responses, as evidenced by the activation of TLR signalling and the downregulation of a broad array of pro-inflammatory cytokines. The use of supernatants like the one described in this paper could be an effective and safe alternative to using live bacteria in functional foods.

  1. Decreased Circulating Interleukin-35 Levels Are Related to Interleukin-4-Producing CD8+ T Cells in Patients with Allergic Asthma.

    Science.gov (United States)

    Wang, Wei; Li, Ping; Yang, Jiong

    2015-08-01

    Interleukin (IL)-35 is a newly discovered suppressive cytokine and has been shown to alleviate inflammatory and autoimmune diseases. The purpose of this study was to investigate immunomodulatory capacity of IL-35 in patients with allergic asthma. IL-35 mRNA expression levels in peripheral blood mononuclear cells (PBMCs) were detected by quantitative real-time PCR (qPCR). The frequencies of cytotoxic T cells (Tc)1, Tc2 and Tc17 cells were measured by flow cytometry. Plasma levels of IL-35, interferon (IFN)-γ, IL-4, and IL-17 were examined by enzyme-linked immunosorbent assay (ELISA). The correlations between plasma IL-35 levels and Tc1, Tc2, and Tc17 cytokine production in allergic asthmatics (n = 25) and healthy controls (n = 12) were analyzed by Pearson's test. IL-35 protein and mRNA expression levels were down-regulated in allergic asthmatics compared with healthy controls. The frequencies of Tc2 and Tc17 cells were significantly increased in patients with asthma, and the frequency of Tc1 cells did not differ between asthmatic patients and healthy controls. Similarly, plasma levels of IL-4 and IL-17 were significantly increased in asthmatic patients, while there was no difference in IFN-γ levels between allergic asthma patients and healthy controls. More importantly, plasma IL-35 protein levels were negatively correlated with the frequency of IL-4-producing CD8+ T (Tc2) cells and with the IL-4 level in patients with allergic asthma. Our results suggest that decreased circulating IL-35 levels could contribute to the pathogenesis of allergic asthma by regulating CD8+ T cells. PMID:26547705

  2. Decreased reactive oxygen species production in cells with mitochondrial haplogroups associated with longevity.

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    Ai Chen

    Full Text Available Mitochondrial DNA (mtDNA is highly polymorphic, and its variations in humans may contribute to individual differences in function. Zhang and colleagues found a strikingly higher frequency of a C150T transition in the D-loop of mtDNA from centenarians and twins of an Italian population, and also demonstrated that this base substitution causes a remodeling of the mtDNA 151 replication origin in human leukocytes and fibroblasts [1]. The C150T transition is a polymorphism associated with several haplogroups. To determine whether haplogroups that carry the C150T transition display any phenotype that may be advantageous for longevity, we analyzed cybrids carrying or not the C150T transition. These cybrids were obtained by fusing cytoplasts derived from human fibroblasts with human mtDNA-less cells (ρ(0 cells. We chose for cybrid construction and analysis haplogroup-matched pairs of fibroblast strains containing or not the C150T transition. In particular, we used, as one pair of mtDNA donors, a fibroblast strain of the U3a haplogroup, carrying the C150T transition and a strain of the U-K2 haplogroup, without the C150T transition, and as another pair, fibroblasts of the J2b haplogroup, carrying the C150T transition and of the J1c haplogroup, without the C150T transition. We have found no association of respiratory capacity, mtDNA level, mitochondrial gene expression level, or growth rate with the presence of the C150T transition. However, we have found that the cybrids with haplogroups that include the C150T transition have in common a lower reactive oxygen species (ROS production rate than the haplogroup-matched cybrids without that transition. Thus, the lower ROS production rate may be a factor in the increased longevity associated with the U and the J2 haplogroups. Of further interest, we found that cybrids with the U3a haplogroup exhibited a higher respiration rate than the other cybrids examined.

  3. Exercise-induced norepinephrine decreases circulating hematopoietic stem and progenitor cell colony-forming capacity.

    Science.gov (United States)

    Kröpfl, Julia M; Stelzer, Ingeborg; Mangge, Harald; Pekovits, Karin; Fuchs, Robert; Allard, Nathalie; Schinagl, Lukas; Hofmann, Peter; Dohr, Gottfried; Wallner-Liebmann, Sandra; Domej, Wolfgang; Müller, Wolfram

    2014-01-01

    A recent study showed that ergometry increased circulating hematopoietic stem and progenitor cell (CPC) numbers, but reduced hematopoietic colony forming capacity/functionality under normoxia and normobaric hypoxia. Herein we investigated whether an exercise-induced elevated plasma free/bound norepinephrine (NE) concentration could be responsible for directly influencing CPC functionality. Venous blood was taken from ten healthy male subjects (25.3+/-4.4 yrs) before and 4 times after ergometry under normoxia and normobaric hypoxia (FiO2exercise-induced NE and blood lactate (La) on CPC functionality was analyzed in a randomly selected group of subjects (n = 6) in vitro under normoxia by secondary colony-forming unit granulocyte macrophage assays. Concentrations of free NE, EPI, Co and IL-6 were significantly increased post-exercise under normoxia/hypoxia. Ergometry-induced free NE concentrations found in vivo showed a significant impairment of CPC functionality in vitro under normoxia. Thus, ergometry-induced free NE was thought to trigger CPC mobilization 10 minutes post-exercise, but as previously shown impairs CPC proliferative capacity/functionality at the same time. The obtained results suggest that an ergometry-induced free NE concentration has a direct negative effect on CPC functionality. Cortisol may further influence CPC dynamics and functionality. PMID:25180783

  4. Men with Sickle Cell Anemia and Priapism Exhibit Increased Hemolytic Rate, Decreased Red Blood Cell Deformability and Increased Red Blood Cell Aggregate Strength

    Science.gov (United States)

    Cita, Kizzy-Clara; Brureau, Laurent; Lemonne, Nathalie; Billaud, Marie; Connes, Philippe; Ferdinand, Séverine; Tressières, Benoit; Tarer, Vanessa; Etienne-Julan, Maryse; Blanchet, Pascal; Elion, Jacques; Romana, Marc

    2016-01-01

    Objectives To investigate the association between priapism in men with sickle cell anemia (SCA) and hemorheological and hemolytical parameters. Materials and Methods 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. Results 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. Conclusion 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. PMID:27145183

  5. Men with Sickle Cell Anemia and Priapism Exhibit Increased Hemolytic Rate, Decreased Red Blood Cell Deformability and Increased Red Blood Cell Aggregate Strength.

    Directory of Open Access Journals (Sweden)

    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.

  6. Cyclosporine restores hematopoietic function by compensating for decreased Tregs in patients with pure red cell aplasia and acquired aplastic anemia.

    Science.gov (United States)

    Dao, An T T; Yamazaki, Hirohito; Takamatsu, Hiroyuki; Sugimori, Chiharu; Katagiri, Takamasa; Maruyama, Hiroyuki; Zaimoku, Yoshitaka; Maruyama, Kana; Ly, Trung Q; Espinoza, Luis; Nakao, Shinji

    2016-04-01

    Most patients with acquired pure red cell aplasia (PRCA) and some with acquired aplastic anemia (AA) respond well to cyclosporine (CsA), but thereafter often show CsA dependency. The mechanism underlying this dependency remains unknown. We established a reliable method for measuring the regulatory T cell (Treg) count using FoxP3 and Helios expression as markers and determined the balance between Tregs and other helper T cell subsets in 16 PRCA and 29 AA patients. The ratios of interferon-γ-producing CD4(+) (Th1) T cells to Tregs in untreated patients and CsA-dependent patients were significantly higher (PRCA 5.77 ± 1.47 and 7.38 ± 2.58; AA 6.18 ± 2.35 and 8.94 ± 4.06) than in healthy volunteers (HVs; 3.33 ± 0.90) due to the profound decrease in the percentage of Tregs. In contrast, the ratios were comparable to HVs in convalescent CsA-treated AA patients (4.74 ± 2.10) and AA patients in remission after the cessation of CsA treatment (4.24 ± 1.67). Low-dose CsA (100 ng/ml) inhibited the proliferation of conventional T cells (Tconv) to a similar degree to the inhibition by Tregs in a co-culture with a 1:1 Treg/Tconv ratio. The data suggest that CsA may reverse the hematopoietic suppression in PRCA and AA patients by compensating for the inadequate immune regulatory function that occurs due to a profound decrease in the Treg count. PMID:26968551

  7. Decreased N-TAF1 expression in X-linked dystonia-parkinsonism patient-specific neural stem cells

    Science.gov (United States)

    Ito, Naoto; Hendriks, William T.; Dhakal, Jyotsna; Vaine, Christine A.; Liu, Christina; Shin, David; Shin, Kyle; Wakabayashi-Ito, Noriko; Dy, Marisela; Multhaupt-Buell, Trisha; Sharma, Nutan; Breakefield, Xandra O.; Bragg, D. Cristopher

    2016-01-01

    ABSTRACT X-linked dystonia-parkinsonism (XDP) is a hereditary neurodegenerative disorder involving a progressive loss of striatal medium spiny neurons. The mechanisms underlying neurodegeneration are not known, in part because there have been few cellular models available for studying the disease. The XDP haplotype consists of multiple sequence variations in a region of the X chromosome containing TAF1, a large gene with at least 38 exons, and a multiple transcript system (MTS) composed of five unconventional exons. A previous study identified an XDP-specific insertion of a SINE-VNTR-Alu (SVA)-type retrotransposon in intron 32 of TAF1, as well as a neural-specific TAF1 isoform, N-TAF1, which showed decreased expression in post-mortem XDP brain compared with control tissue. Here, we generated XDP patient and control fibroblasts and induced pluripotent stem cells (iPSCs) in order to further probe cellular defects associated with this disease. As initial validation of the model, we compared expression of TAF1 and MTS transcripts in XDP versus control fibroblasts and iPSC-derived neural stem cells (NSCs). Compared with control cells, XDP fibroblasts exhibited decreased expression of TAF1 transcript fragments derived from exons 32-36, a region spanning the SVA insertion site. N-TAF1, which incorporates an alternative exon (exon 34′), was not expressed in fibroblasts, but was detectable in iPSC-differentiated NSCs at levels that were ∼threefold lower in XDP cells than in controls. These results support the previous findings that N-TAF1 expression is impaired in XDP, but additionally indicate that this aberrant transcription might occur in neural cells at relatively early stages of development that precede neurodegeneration. PMID:26769797

  8. Expansion of CD8+ T cells lacking Sema4D/CD100 during HIV-1 infection identifies a subset of T cells with decreased functional capacity

    OpenAIRE

    Eriksson, Emily M.; Milush, Jeffrey M.; Ho, Emily L.; Batista, Mariana D.; Holditch, Sara J.; Keh, Chris E.; Norris, Philip J.; Keating, Sheila M.; Deeks, Steven G; Hunt, Peter W.; Martin, Jeffrey N.; Rosenberg, Michael G.; Hecht, Frederick M.; Nixon, Douglas F.

    2012-01-01

    Sema4D, also known as CD100, is a constitutively expressed immune semaphorin on T cells and NK cells. CD100 has important immune regulatory functions that improve antigen-specific priming by antigen-presenting cells, and can also act as a costimulatory molecule on T cells. We investigated the consequence of HIV-1 infection on CD100 expression by T cells, and whether CD100 expression signifies functionally competent effector cells. CD100 expression on T cells from healthy individuals was compa...

  9. Nucleolin antagonist triggers autophagic cell death in human glioblastoma primary cells and decreased in vivo tumor growth in orthotopic brain tumor model.

    Science.gov (United States)

    Benedetti, Elisabetta; Antonosante, Andrea; d'Angelo, Michele; Cristiano, Loredana; Galzio, Renato; Destouches, Damien; Florio, Tiziana Marilena; Dhez, Anne Chloé; Astarita, Carlo; Cinque, Benedetta; Fidoamore, Alessia; Rosati, Floriana; Cifone, Maria Grazia; Ippoliti, Rodolfo; Giordano, Antonio; Courty, José; Cimini, Annamaria

    2015-12-01

    Nucleolin (NCL) is highly expressed in several types of cancer and represents an interesting therapeutic target. It is expressed at the plasma membrane of tumor cells, a property which is being used as a marker for several human cancer including glioblastoma. In this study we investigated targeting NCL as a new therapeutic strategy for the treatment of this pathology. To explore this possibility, we studied the effect of an antagonist of NCL, the multivalent pseudopeptide N6L using primary culture of human glioblastoma cells. In this system, N6L inhibits cell growth with different sensitivity depending to NCL localization. Cell cycle analysis indicated that N6L-induced growth reduction was due to a block of the G1/S transition with down-regulation of the expression of cyclin D1 and B2. By monitoring autophagy markers such as p62 and LC3II, we demonstrate that autophagy is enhanced after N6L treatment. In addition, N6L-treatment of mice bearing tumor decreased in vivo tumor growth in orthotopic brain tumor model and increase mice survival. The results obtained indicated an anti-proliferative and pro-autophagic effect of N6L and point towards its possible use as adjuvant agent to the standard therapeutic protocols presently utilized for glioblastoma. PMID:26540346

  10. The decreased expression of electron transfer flavoprotein β is associated with tubular cell apoptosis in diabetic nephropathy.

    Science.gov (United States)

    Wang, Hua; Zhang, Haojun; Chen, Xiaohong; Zhao, Tingting; Kong, Qin; Yan, Meihua; Zhang, Bingxuan; Sun, Sifan; Lan, Hui-Yao; Li, Ning; Li, Ping

    2016-05-01

    Tubular injury is closely correlated with the development of progressive diabetic nephropathy (DN), particularly in cases of type 2 diabetes. The apoptosis of tubular cells has been recognized as a major cause of tubular atrophy, followed by tubulointerstitial fibrosis. Electron transfer flavoprotein β (ETFβ) is known as an important electron acceptor in energy metabolism, but its role in DN was not fully understood. In the present study, we examined the expression pattern of ETFβ using diabetic kidney samples and further investigated ETFβ involvement in tubular epithelial cell (TEC) apoptosis. Human renal biopsy specimens from patients with DN as well as a spontaneous rat model of diabetes using Otsuka Long-Evans Tokushima fatty (OLETF) rats, were employed in order to examine the expression of ETFβ and cell apoptosis in kidneys during the development of DN (for the rats, at 36 and 56 weeks of age respectively). Moreover, ETFβ siRNA was used to investigate the role of ETFβ in the apoptosis of renal tubular cells. Our present results showed that the expression of ETFβ in the kidneys was progressively decreased both in patients with DN and OLETF rats, which coincided with progressive renal injury and TEC apoptosis. In addition, the in vitro study demonstrated that knockdown of ETFβ caused apoptosis in tubular cells, as proven by the increased expression of pro‑apoptotic proteins and TUNEL assay. Therefore, the findings of our present study suggest that ETFβ plays an important role in renal tubular cell apoptosis during the progression of DN. PMID:27035869

  11. Decreased Laminin Expression by Human Lung Epithelial Cells and Fibroblasts Cultured in Acellular Lung Scaffolds from Aged Mice.

    Directory of Open Access Journals (Sweden)

    Lindsay M Godin

    Full Text Available The lung changes functionally and structurally with aging. However, age-related effects on the extracellular matrix (ECM and corresponding effects on lung cell behavior are not well understood. We hypothesized that ECM from aged animals would induce aging-related phenotypic changes in healthy inoculated cells. Decellularized whole organ scaffolds provide a powerful model for examining how ECM cues affect cell phenotype. The effects of age on ECM composition in both native and decellularized mouse lungs were assessed as was the effect of young vs old acellular ECM on human bronchial epithelial cells (hBECs and lung fibroblasts (hLFs. Native aged (1 year lungs demonstrated decreased expression of laminins α3 and α4, elastin and fibronectin, and elevated collagen, compared to young (3 week lungs. Proteomic analyses of decellularized ECM demonstrated similar findings, and decellularized aged lung ECM contained less diversity in structural proteins compared to young ECM. When seeded in old ECM, hBECs and hLFs demonstrated lower gene expression of laminins α3 and α4, respectively, as compared to young ECM, paralleling the laminin deficiency of aged ECM. ECM changes appear to be important factors in potentiating aging-related phenotypes and may provide clues to mechanisms that allow for aging-related lung diseases.

  12. Placental insufficiency decreases pancreatic vascularity and disrupts hepatocyte growth factor signaling in the pancreatic islet endothelial cell in fetal sheep.

    Science.gov (United States)

    Rozance, Paul J; Anderson, Miranda; Martinez, Marina; Fahy, Anna; Macko, Antoni R; Kailey, Jenai; Seedorf, Gregory J; Abman, Steven H; Hay, William W; Limesand, Sean W

    2015-02-01

    Hepatocyte growth factor (HGF) and vascular endothelial growth factor A (VEGFA) are paracrine hormones that mediate communication between pancreatic islet endothelial cells (ECs) and β-cells. Our objective was to determine the impact of intrauterine growth restriction (IUGR) on pancreatic vascularity and paracrine signaling between the EC and β-cell. Vessel density was less in IUGR pancreata than in controls. HGF concentrations were also lower in islet EC-conditioned media (ECCM) from IUGR, and islets incubated with control islet ECCM responded by increasing insulin content, which was absent with IUGR ECCM. The effect of ECCM on islet insulin content was blocked with an inhibitory anti-HGF antibody. The HGF receptor was not different between control and IUGR islets, but VEGFA was lower and the high-affinity VEGF receptor was higher in IUGR islets and ECs, respectively. These findings show that paracrine actions from ECs increase islet insulin content, and in IUGR ECs, secretion of HGF was diminished. Given the potential feed-forward regulation of β-cell VEGFA and islet EC HGF, these two growth factors are highly integrated in normal pancreatic islet development, and this regulation is decreased in IUGR fetuses, resulting in lower pancreatic islet insulin concentrations and insulin secretion. PMID:25249573

  13. Decreased SIRT3 in aged human mesenchymal stromal/stem cells increases cellular susceptibility to oxidative stress.

    Science.gov (United States)

    Wang, Xue-Qing; Shao, Yong; Ma, Chong-Yi; Chen, Wei; Sun, Lu; Liu, Wei; Zhang, Dong-Yang; Fu, Bi-Cheng; Liu, Kai-Yu; Jia, Zhi-Bo; Xie, Bao-Dong; Jiang, Shu-Lin; Li, Ren-Ke; Tian, Hai

    2014-11-01

    Sirtuin3 (SIRT3) is an important member of the sirtuin family of protein deacetylases that is localized to mitochondria and linked to lifespan extension in organisms ranging from yeast to humans. As aged cells have less regenerative capacity and are more susceptible to oxidative stress, we investigated the effect of ageing on SIRT3 levels and its correlation with antioxidant enzyme activities. Here, we show that severe oxidative stress reduces SIRT3 levels in young human mesenchymal stromal/stem cells (hMSCs). Overexpression of SIRT3 improved hMSCs resistance to the detrimental effects of oxidative stress. By activating manganese superoxide dismutase (MnSOD) and catalase (CAT), SIRT3 protects hMSCs from apoptosis under stress. SIRT3 expression, levels of MnSOD and CAT, as well as cell survival showed little difference in old versus young hMSCs under normal growth conditions, whereas older cells had a significantly reduced capacity to withstand oxidative stress compared to their younger counterparts. Expression of the short 28 kD SIRT3 isoform was higher, while the long 44 kD isoform expression was lower in young myocardial tissues compared with older ones. These results suggest that the active short isoform of SIRT3 protects hMSCs from oxidative injury by increasing the expression and activity of antioxidant enzymes. The expression of this short isoform decreases in cardiac tissue during ageing, leading to a reduced capacity for the heart to withstand oxidative stress. PMID:25210848

  14. Suppression of dynamin GTPase decreases α-synuclein uptake by neuronal and oligodendroglial cells: a potent therapeutic target for synucleinopathy

    Directory of Open Access Journals (Sweden)

    Konno Masatoshi

    2012-08-01

    Full Text Available Abstract Background The intracellular deposition of misfolded proteins is a common neuropathological hallmark of most neurodegenerative disorders. Increasing evidence suggests that these pathogenic proteins may spread to neighboring cells and induce the propagation of neurodegeneration. Results In this study, we have demonstrated that α-synuclein (αSYN, a major constituent of intracellular inclusions in synucleinopathies, was taken up by neuronal and oligodendroglial cells in both a time- and concentration-dependent manner. Once incorporated, the extracellular αSYN was immediately assembled into high-molecular-weight oligomers and subsequently formed cytoplasmic inclusion bodies. Furthermore, αSYN uptake by neurons and cells of the oligodendroglial lineage was markedly decreased by the genetic suppression and pharmacological inhibition of the dynamin GTPases, suggesting the involvement of the endocytic pathway in this process. Conclusions Our findings shed light on the mode of αSYN uptake by neuronal and oligodendroglial cells and identify therapeutic strategies aimed at reducing the propagation of protein misfolding.

  15. Decreased ERCC1 Expression After Platinum-Based Neoadjuvant Chemotherapy in non-Small Cell Lung Cancer.

    Science.gov (United States)

    Podmaniczky, Eszter; Fábián, Katalin; Pápay, Judit; Puskás, Rita; Gyulai, Márton; Furák, József; Tiszlavicz, László; Losonczy, György; Tímár, József; Moldvay, Judit

    2015-04-01

    We have already demonstrated in a small cohort of 17 non-small cell lung cancer patients that ERCC1 (excision repair cross-complementation group 1) protein expression decreased after platinum-based treatment, however, certain clinicopathological parameters, such as histologic subtypes, ERCC1 expression scores, chemotherapeutic combinations, response rate, gender and smoking history were not analyzed. The aim of our present study was to extend the studied cohort and analyze those parameters. ERCC1 protein expression was examined in 46 patients treated with neoadjuvant chemotherapy. 46 bronchoscopic biopsy samples (27 squamous cell carcinomas /SCC/ and 19 adenocarcinomas /ADC/) together with their corresponding surgical biopsies were studied. ERCC1 immunohistochemistry was performed on formalin-fixed, paraffin-embedded tissues. Staining scores were calculated by multiplying the percentage of positive tumor cells (0-100) by the staining intensity (0-3). 24/27 bronchoscopic SCC tissues expressed ERCC1. Thirteen of these cases became negative after neoadjuvant therapy and the expression differences between pre- and postchemotherapy samples were highly significant (p affects the ERCC1 expression probably referring to an induction of tumor cell selection. PMID:25194563

  16. Age-Dependent Decrease in Chaperone Activity Impairs MANF Expression, Leading to Purkinje Cell Degeneration in Inducible SCA17 Mice

    Science.gov (United States)

    Yang, Su; Huang, Shanshan; Gaertig, Marta A.; Li, Xiao-Jiang; Li, Shihua

    2016-01-01

    SUMMARY Although protein-misfolding-mediated neurodegenerative diseases have been linked to aging, how aging contributes to selective neurodegeneration remains unclear. We established spinocerebellar ataxia 17 (SCA17) knockin mice that inducibly express one copy of mutant TATA box binding protein (TBP) at different ages by tamoxifen-mediated Cre recombination. We find that more mutant TBP accumulates in older mouse and that this accumulation correlates with age-related decreases in Hsc70 and chaperone activity. Consistently, older SCA17 mice experienced earlier neurological symptom onset and more severe Purkinje cell degeneration. Mutant TBP shows decreased association with XBP1s, resulting in the reduced transcription of mesencephalic astrocyte-derived neurotrophic factor (MANF), which is enriched in Purkinje cells. Expression of Hsc70 improves the TBP-XBP1s interaction and MANF transcription, and overexpression of MANF ameliorates mutant TBP-mediated Purkinje cell degeneration via protein kinase C (PKC)-dependent signaling. These findings suggest that the age-related decline in chaperone activity affects polyglutamine protein function that is important for the viability of specific types of neurons. PMID:24462098

  17. Thalidomide's effectiveness in erythema nodosum leprosum is associated with a decrease in CD4+ cells in the peripheral blood.

    Science.gov (United States)

    Shannon, E J; Ejigu, M; Haile-Mariam, H S; Berhan, T Y; Tasesse, G

    1992-03-01

    Thalidomide is well documented as being an effective drug in the treatment of erythema nodosum leprosum (ENL). The mechanism of action of thalidomide in ENL as well as the pathogenesis of ENL are yet to be fully determined. Lepromatous leprosy patients experiencing ENL have been reported to have an increase in the ratio of CD4+ to CD8+ cells in their blood and ENL skin lesions. Thalidomide has been shown to cause a decrease in the ratio of CD4+ to CD8+ lymphocytes in the blood of healthy males. This decrease was due to a significant reduction in the numbers of Cd4+ lymphocytes and an apparent increase in the numbers of CD8+ lymphocytes. In this study, thalidomide's effectiveness in halting chronic ENL and arresting a relapse into ENL was consistently associated with a decrease in the numbers of CD4+ lymphocytes in the blood of 2 male lepromatous leprosy patients. PMID:1569817

  18. 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.

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

    International Nuclear Information System (INIS)

    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

  20. 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...... 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...... AVD, primarily via modulation of NaCl movements, contribute to protection against apoptosis in MDR EATC....

  1. The 2-oxoglutarate analog 3-oxoglutarate decreases normoxic hypoxia-inducible factor-1α in cancer cells, induces cell death, and reduces tumor xenograft growth

    Directory of Open Access Journals (Sweden)

    Koivunen P

    2016-03-01

    Full Text Available Peppi Koivunen,1 Stuart M Fell,2,3 Wenyun Lu,4 Joshua D Rabinowitz,4 Andrew L Kung,5,6 Susanne Schlisio,2,7 1Biocenter Oulu, Faculty of Biochemistry and Molecular Medicine, Oulu Center for Cell-Matrix Research, University of Oulu, Oulu, Finland; 2Ludwig Institute for Cancer Research Ltd, Stockholm, Sweden; 3Department of Cell and Molecular Biology, Karolinska Institutet, Stockholm, Sweden; 4Department of Chemistry and Integrative Genomics, Princeton University, Princeton, NJ, 5Department of Medical Oncology, Dana Farber Cancer Institute, Harvard Medical School, Boston, MA, 6Department of Pediatrics, Columbia University Medical Center, New York, NY, USA; 7Department of Microbiology and Tumor and Cell Biology, Karolinska Institutet, Stockholm, Sweden Abstract: The cellular response to hypoxia is primarily regulated by the hypoxia-inducible factors (HIFs. HIF-1α is also a major mediator of tumor physiology, and its abundance is correlated with therapeutic resistance in a broad range of cancers. Accumulation of HIF-1α under hypoxia is mainly controlled by the oxygen-sensing HIF prolyl 4-hydroxylases (EGLNs, also known as PHDs. Here, we identified a high level of normoxic HIF-1α protein in various cancer cell lines. EGLNs require oxygen and 2-oxoglutarate for enzymatic activity. We tested the ability of several cell-permeable 2-oxoglutarate analogs to regulate the abundance of HIF-1α protein. We identified 3-oxoglutarate as a potent regulator of HIF-1α in normoxic conditions. In contrast to 2-oxoglutarate, 3-oxoglutarate decreased the abundance of HIF-1α protein in several cancer cell lines in normoxia and diminished HIF-1α levels independent of EGLN enzymatic activity. Furthermore, we observed that 3-oxoglutarate was detrimental to cancer cell survival. We show that esterified 3-oxoglutarate, in combination with the cancer chemotherapeutic drug vincristine, induces apoptosis and inhibits tumor growth in vitro and in vivo. Our data

  2. 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.

  3. Glucose restriction decreases telomerase activity and enhances its inhibitor response on breast cancer cells: possible extra-telomerase role of BIBR 1532

    OpenAIRE

    Wardi, Layal; Alaaeddine, Nada; Raad, Issam; Sarkis, Riad; Serhal, Rim; Khalil, Charbel; Hilal, George

    2014-01-01

    Background Considerable progress has been made to understand the association between lifestyle and diet in cancer initiation and promotion. Because excessive glucose consumption is a key metabolic hallmark of cancer cells, glucose restriction (GR) decreases the proliferation, and promotes the differentiation and transformation of cancer cells to quiescent cells. The immortality of cancerous cells is largely assured by telomerase, which is an interesting target for inhibition by BIBR 1532. In ...

  4. Decreased Circulating Interleukin-35 Levels Are Related to Interleukin-4-Producing CD8+ T Cells in Patients with Allergic Asthma

    Directory of Open Access Journals (Sweden)

    Wei Wang

    2015-10-01

    Full Text Available  Interleukin (IL-35 is a newly discovered suppressive cytokine and has been shown to alleviate  inflammatory  and  autoimmune  diseases.  The  purpose  of  this  study  was  to investigate immunomodulatory capacity of IL-35 in patients with allergic asthma.IL-35 mRNA expression levels in peripheral blood mononuclear cells (PBMCs were detected  by  quantitative  real-time  PCR  (qPCR.  The  frequencies  of  cytotoxic  T  cells (Tc1,Tc2  and  Tc17  cells  were  measured  by  flow  cytometry.  Plasma  levels  of  IL-35, interferon (IFN-γ, IL-4, and IL-17 were examined by enzyme-linked immunosorbent assay (ELISA. The correlations between plasma IL-35 levels and Tc1, Tc2, and Tc17 cytokine production in allergic asthmatics (n = 25 and healthy controls (n = 12 were analyzed by Pearson’s test.IL-35 protein and mRNA expression levels were down-regulated in allergic asthmaticscompared with healthy controls. The frequencies of Tc2 and Tc17 cells were significantly increased in patients with asthma, and the frequency of Tc1 cells did not differ between asthmatic patients and healthy controls. Similarly, plasma levels of IL-4 and IL-17 were significantly increased in asthmatic patients, while there was no difference in IFN-γ levels between allergic asthma patients  and  healthy  controls.  More importantly,  plasma  IL-35 protein levels were negatively correlated with the frequency of IL-4-producing CD8+ T (Tc2 cells and with the IL-4 level in patients with allergic asthma.Our results suggest that decreased circulating IL-35 levels could contribute to the pathogenesis of allergic asthma by regulating CD8+ T cells

  5. 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.

  6. Cell-free culture supernatant of Bifidobacterium breve CNCM I-4035 decreases pro-inflammatory cytokines in human dendritic cells challenged with Salmonella typhi through TLR activation.

    Directory of Open Access Journals (Sweden)

    Miriam Bermudez-Brito

    Full Text Available Dendritic cells (DCs constitute the first point of contact between gut commensals and our immune system. Despite growing evidence of the immunomodulatory effects of probiotics, the interactions between the cells of the intestinal immune system and bacteria remain largely unknown. Indeed,, the aim of this work was to determine whether the probiotic Bifidobacterium breve CNCM I-4035 and its cell-free culture supernatant (CFS have immunomodulatory effects in human intestinal-like dendritic cells (DCs and how they respond to the pathogenic bacterium Salmonella enterica serovar Typhi, and also to elucidate the molecular mechanisms involved in these interactions. Human DCs were directly challenged with B. breve/CFS, S. typhi or a combination of these stimuli for 4 h. The expression pattern of genes involved in Toll-like receptor (TLR signaling pathway and cytokine secretion was analyzed. CFS decreased pro-inflammatory cytokines and chemokines in human intestinal DCs challenged with S. typhi. In contrast, the B. breve CNCM I-4035 probiotic strain was a potent inducer of the pro-inflammatory cytokines and chemokines tested, i.e., TNF-α, IL-8 and RANTES, as well as anti-inflammatory cytokines including IL-10. CFS restored TGF-β levels in the presence of Salmonella. Live B.breve and its supernatant enhanced innate immune responses by the activation of TLR signaling pathway. These treatments upregulated TLR9 gene transcription. In addition, CFS was a more potent inducer of TLR9 expression than the probiotic bacteria in the presence of S. typhi. Expression levels of CASP8 and IRAK4 were also increased by CFS, and both treatments induced TOLLIP gene expression. Our results indicate that the probiotic strain B. breve CNCM I-4035 affects the intestinal immune response, whereas its supernatant exerts anti-inflammatory effects mediated by DCs. This supernatant may protect immune system from highly infectious agents such as Salmonella typhi and can down

  7. Naproxen sodium decreases prostaglandins secretion from cultured human endometrial stromal cells modulating metabolizing enzymes mRNA expression.

    Science.gov (United States)

    Carrarelli, Patrizia; Funghi, Lucia; Bruni, Simone; Luisi, Stefano; Arcuri, Felice; Petraglia, Felice

    2016-04-01

    Dysmenorrhea, defined as painful cramps occurring immediately before or during the menstrual period, is a common symptom of different gynecological diseases. An acute uterine inflammatory response driven by prostaglandins (PGs) is responsible for painful symptoms. Progesterone withdrawal is responsible for activation of cyclooxygenase (COX-2) enzyme and decrease of hydroxyprostaglandin dehydrogenase (HPDG) with consequent increased secretion of PGs secretion, inducing uterine contractility and pain. The most widely used drugs for the treatment of pelvic pain associated with menstrual cycle are non steroidal anti-inflammatory drugs (NSAIDs). The uterine site of action of these drugs is still not defined and the present study evaluated the effect of naproxen sodium in cultured human endometrial stromal cells (HESC) collected from healthy women. PGE2 release was measured by ELISA; COX-2 and HPDG mRNA expression were assessed by qRT-PCR. Naproxen sodium did not affect HESC vitality. Naproxen sodium significantly decreased PGE2 secretion (p dysmenorrhea. PMID:26634864

  8. Regular Exercise Training Increases the Number of Endothelial Progenitor Cells and Decreases Homocysteine Levels in Healthy Peripheral Blood

    OpenAIRE

    Choi, Jeong Kyu; Moon, Ki Myung; Jung, Seok Yun; Kim, Ji Yong; Choi, Sung Hyun; Kim, Da Yeon; Kang, Songhwa; Chu, Chong Woo; Kwon, Sang Mo

    2014-01-01

    Endothelial progenitor cells (EPCs) are known to play an important role in the repair of damaged blood vessels. We used an endothelial progenitor cell colony-forming assay (EPC-CFA) to determine whether EPC numbers could be increased in healthy individuals through regular exercise training. The number of functional EPCs obtained from human peripheral blood-derived AC133 stem cells was measured after a 28-day regular exercise training program. The number of total endothelial progenitor cell co...

  9. Carvacrol modulates oxidative stress and decreases cell injury in pancreas of rats with acute pancreatitis.

    Science.gov (United States)

    Kılıç, Yeliz; Geyikoglu, Fatime; Çolak, Suat; Turkez, Hasan; Bakır, Murat; Hsseinigouzdagani, Mirkhalil

    2016-08-01

    Acute pancreatitis (AP) is considered as major problem around the world and the incidence of AP is increasing. Carvacrol (CAR), a monoterpenic phenol, has good antioxidant activity. This in vivo study was designed to evaluate whether CAR provide protection against AP that developed by pancreas injury. The rats were randomised into groups to receive (I) no therapy; (II) 50 µg/kg cerulein at 1 h intervals by four intraperitonally (i.p.) injections; (III) 50, 100 and 200 mg/kg CAR by one i.p. injection; and (IV) cerulein plus CAR after 2 h of cerulein administration. 12 h later, serum samples were obtained to assess pancreatic function, the lipase and amylase values. The oxidative stress markers were evaluated by changes in the amount of lipid peroxides measured as malondialdehyde (MDA) and changes in main tissue antioxidant enzyme levels including SOD, CAT and GSH-PX. Histopathological examination was performed using scoring systems. Additionally, oxidative DNA damage was determined by measuring the increases of 8-hydroxy-deoxyguanosine (8-OH-dG) formations. We found that the increasing doses of CAR decreased AP-induced MDA and 8-OH-dG levels. Moreover, the pancreas antioxidant enzyme activities were higher than that of the rats in the AP group when compared to the AP plus CAR group. In the treatment groups, the lipase and amylase were reduced. Besides, histopathological findings in the pancreatic tissue were alleviated (p < 0.05). We suggest that CAR could be a safe and potent new drug candidate for treating AP through its antioxidative mechanism of action for the treatment of a wide range of disorders related to pancreas. PMID:26093481

  10. C-reactive protein decreases interleukin-8 production in human endothelial progenitor cells by inhibition of p38 MAPK pathway

    Institute of Scientific and Technical Information of China (English)

    NAN Jing-long; LI Jian-jun; HE Jian-guo

    2009-01-01

    Background C-reactive protein (CRP) has been reported to damage the vascular wall by inducing endothelial dysfunction and inflammation,and it is also speculated to have a role in attenuating angiogenic functions of human endothelial progenitor cells (EPCs).Interleukin-8 (IL-8) is an important mediator of the paracrine mitogenic effect of EPCs,which has direct angiogenic effects on mature endothelial cells.We,herein,investigated the direct effect of CRP on IL-8 production and gene expression in cultured human EPCs.Methods EPCs were isolated from the peripheral venous blood of healthy male volunteers.Cells were cultured in EndoCultTM liquid medium in the absence and presence of CRP at clinically relevant concentrations (5 to 25 μg/ml) for different durations (3 to 48 hours).IL-8 protein and mRNA of cultured EPCs were evaluated using ELISA and real-time PCR.Results The results showed that CRP at a concentration of 10 pg/ml significantly reduced IL-8 secretion of cultured EPCs with a peak at 25 μg/ml,and also decreased mRNA expression in EPCs with a peak at 12 hours.In addition,preincubation of EPCs with SB203580,an inhibitor of p38 mitogen-activated protein kinase (MAPK) decreased CRP inhibition of IL-8 mRNA expression at 12 hours in EPCs.Conclusions Our study,for the first time,demonstrates that CRP directly inhibits EPCs IL-8 secretion,a key cytokine player of angiogenesis induced by EPCs.Inhibition occurred in part via an effect of CRP to active the p38 MAPK signal transduction pathway in EPC.The ability of CRP to inhibit EPCs IL-8 secretion may represent an important mechanism that further links inflammation to cardiovascular disease.

  11. RNAi-mediated knockdown of inhibin α subunit increased apoptosis in granulosa cells and decreased fertility in mice.

    Science.gov (United States)

    Kadariya, Ishwari; Wang, Jiaxing; ur Rehman, Zia; Ali, Hamid; Riaz, Hasan; He, JiuYa; Bhattarai, Dinesh; Liu, Jia Jia; Zhang, Shu Jun

    2015-08-01

    Inhibin α (INHα), a member of TGFβ superfamily, is an important modulator of reproductive function that plays a vital role in follicular changes, cell differentiation, oocyte development, and ultimately in mammalian reproduction. However, the role of inhibin α in female fertility and ovarian function remains largely unknown. To define its role in reproduction, transgenic mice of RNAi-INHα that knock down the INHα expression by shRNAi were used. Inhibin α subunit gene was knocked down successfully at both transcriptional and translational levels by RNAi PiggyBac transposon (Pbi) mediated recombinant pshRNA vectors and purified DNA fragments were microinjected into mouse zygotes. Results showed that transgenic female mice were sub-fertile and exhibited 35.28% reduction in litter size in F1 generation relative to wild type. The decreased litter size associated with the reduction in the number of oocytes ovulated after puberty. Serum INHα level was significantly decreased in both 3 and 6 weeks; whereas, FSH was significantly increased in 3 weeks but not in 6 weeks. Furthermore, suppression of INHα expression significantly promoted apoptosis by up-regulating Caspase-3, bcl2, INHβB and GDF9 and down regulated Kitl and TGFβRIII genes both at transcriptional and translational levels. Moreover, it also dramatically reduced the progression of G1 phase of cell cycle and the number of cells in S phase as determined by flow cytometer. These results indicate that suppression of INHα expression in RNAi-transgenic mice leads to disruption of normal ovarian regulatory mechanism and causes reproductive deficiencies by promoting cellular apoptosis, arresting cellular progression and altering hormonal signaling. PMID:25998417

  12. TLR9-induced miR-155 and Ets-1 decrease expression of CD1d on B cells in SLE.

    Science.gov (United States)

    Liu, Fei; Fan, Hongye; Ren, Deshan; Dong, Guanjun; Hu, Erling; Ji, Jianjian; Hou, Yayi

    2015-07-01

    B cells present lipid antigens to CD1d-restricted invariant natural killer T (iNKT) cells to maintain autoimmune tolerance, and this process is disrupted in systemic lupus erythematosus (SLE). Inflammation may inhibit CD1d expression to exacerbate the pathology of lupus. However, how inflammation regulates CD1d expression on B cells is unclear in SLE. In the present study, we showed that the surface expression of CD1d on B cells from SLE mice was decreased and that stimulation of inflammatory responses through TLR9 decreased the membrane and total CD1d levels of CD1d on B cells. Moreover, inflammation-related microRNA-155 (miR-155) negatively correlated with the expression of CD1d in B cells. miR-155 directly targeted the 3'-untranslated region (3'-UTR) of CD1d upon TLR9 activation in both humans and mice. The inhibitory effects of miR-155 on CD1d expression in B cells impaired their antigen-presenting capacity to iNKT cells. In addition, Ets-1, a susceptibility gene of SLE, also directly regulated the expression of the CD1d gene at the transcriptional level. These findings provide new insight into the mechanism underlying decreased CD1d expression on B cells in SLE, suggesting that inhibition of inflammation may increase CD1d expression in B cells to ameliorate SLE via modulating iNKT cells. PMID:25929465

  13. 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.

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

    International Nuclear Information System (INIS)

    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 PGE2. •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 E2 (PGE2) 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. PGE2 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, PGE2, cellular proliferation and MEK/MAPK activation, implicating this pathway in regulating miR-143 expression

  15. Peripheral blood non-MAIT CD8+CD161hi cells are decreased in relapsing-remitting multiple sclerosis patients treated with interferon beta.

    Science.gov (United States)

    Negrotto, Laura; Cantó, Ester; Río, Jordi; Tintoré, Mar; Montalban, Xavier; Comabella, Manuel

    2015-11-15

    CD8+CD161hi cells, comprising MAIT and non-MAIT cells, have been involved in multiple sclerosis (MS) pathogenesis. Here, we investigated the frequency of CD8+CD161hi, MAIT and non-MAIT cells by flow cytometry in peripheral blood samples from 41 untreated MS patients, 48 patients receiving disease modifying therapies, and 17 healthy controls (HC). IFNβ treatment was associated with a decrease in the frequency of Tc17 cells compared to untreated patients (p=0.019). No significant differences were observed between untreated MS patients and HC for any of the study cell populations. These results suggest previously unknown mechanisms of action of IFNβ. PMID:26531701

  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. 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)

    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)

  18. A novel oxygen carrier “YQ23” suppresses the liver tumor metastasis by decreasing circulating endothelial progenitor cells and regulatory T cells

    International Nuclear Information System (INIS)

    Surgical therapies are the first-line treatments for hepatocellular carcinoma (HCC) patients. However, the high incidence of tumor metastasis after liver surgery remains a severe problem. We aim to investigate the roles and the underlying mechanism of YQ23, stabilized non-polymeric diaspirin cross-linked tetrameric hemoglobin, in liver tumor metastasis after major hepatectomy and partial hepatic ischemia reperfusion (I/R) injury. An orthotopic liver tumor model in Buffalo rat was established using the hepatocellular carcinoma cell line McA-RH7777. Major hepatectomy for tumor-bearing lobe and partial hepatic I/R injury were performed at two weeks after orthotopic liver tumor implantation. YQ23 (0.2 g/kg) was administered at 1 hour before ischemia and immediately after reperfusion. Blood samples were collected at day 0, 1, 7, 14, 21 and 28 for detection of circulating endothelial progenitor cells (EPCs) and regulatory T cells (Tregs). Our results showed that YQ23 treatment effectively inhibited intrahepatic and lung metastases together with less tumor angiogenesis at 4 weeks after major hepatectomy and partial hepatic I/R injury. The levels of circulating EPCs and Tregs were significantly decreased in YQ23 treatment group. Furthermore, YQ23 treatment also increased liver tissue oxygenation during hepatic I/R injury. Up-regulation of HO1 and down-regulation of CXCR3, TNF-α and IL6 were detected after YQ23 treatment. YQ23 treatment suppressed liver tumor metastasis after major hepatectomy and partial hepatic I/R injury in a rat liver tumor model through increasing liver oxygen and reducing the populations of circulating EPCs and Tregs

  19. Decrease in transient receptor potential melastatin 6 mRNA stability caused by rapamycin in renal tubular epithelial cells.

    Science.gov (United States)

    Ikari, Akira; Sanada, Ayumi; Sawada, Hayato; Okude, Chiaki; Tonegawa, Chie; Sugatani, Junko

    2011-06-01

    Rapamycin, an inhibitor of mammalian target of rapamycin (mTOR), is used in treatments for transplantation and cancer. Rapamycin causes hypomagnesemia, although precisely how has not been examined. Here, we investigated the effect of rapamycin on the expression of transient receptor potential melastatin 6 (TRPM6), a Mg2+ channel. Rapamycin and LY-294002, an inhibitor of phosphatidilinositol-3 kinase (PI3K) located upstream of mTOR, inhibited epidermal growth factor (EGF)-induced expression of the TRPM6 protein without affecting TRPM7 expression in rat renal NRK-52E epithelial cells. Both rapamycin and LY-294002 decreased EGF-induced Mg2+ influx. U0126, a MEK inhibitor, inhibited EGF-induced increases in c-Fos, p-ERK, and TRPM6 levels. In contrast, neither rapamycin nor LY-294002 inhibited EGF-induced increases in p-ERK and c-Fos levels. EGF increased p-Akt level, an effect inhibited by LY-294002 and 1L-6-hydroxymethyl-chiro-inositol2-[(R)-2-O-methyl-3-O-octadecylcarbonate] (Akt inhibitor). Akt inhibitor decreased TRPM6 level similar to rapamycin and LY-294002. These results suggest that a PI3K/Akt/mTOR pathway is involved in the regulation of TRPM6 expression. Rapamycin inhibited the EGF-induced increase in TRPM6 mRNA but did not inhibit human TRPM6 promoter activity. In the presence of actinomycin D, a transcriptional inhibitor, rapamycin accelerated the decrease in TRPM6 mRNA. Rapamycin decreased the expression and activity of a luciferase linked with the 3'-untranslated region of human TRPM6 mRNA. These results suggest that TRPM6 expression is up-regulated by a PI3K/Akt/mTOR pathway and rapamycin reduces TRPM6 mRNA stability, resulting in a decrease in the reabsorption of Mg2+. PMID:21073857

  20. Valproic acid exposure decreases Cbp/p300 protein expression and histone acetyltransferase activity in P19 cells.

    Science.gov (United States)

    Lamparter, Christina L; Winn, Louise M

    2016-09-01

    The teratogenicity of the antiepileptic drug valproic acid (VPA) is well established and its inhibition of histone deacetylases (HDAC) is proposed as an initiating factor. Recently, VPA-mediated HDAC inhibition was demonstrated to involve transcriptional downregulation of histone acetyltransferases (HATs), which was proposed to compensate for the increased acetylation resulting from HDAC inhibition. Cbp and p300 are HATs required for embryonic development and deficiencies in either are associated with congenital malformations and embryolethality. The objective of the present study was to characterize Cbp/p300 following VPA exposure in P19 cells. Consistent with previous studies, exposure to 5mM VPA over 24h induced a moderate decrease in Cbp/p300 mRNA, which preceded a strong decrease in total cellular protein mediated by ubiquitin-proteasome degradation. Nuclear Cbp/p300 protein was also decreased following VPA exposure, although to a lesser extent. Total cellular and nuclear p300 HAT activity was reduced proportionately to p300 protein levels, however while total cellular HAT activity also decreased, nuclear HAT activity was unaffected. Using the Cbp/p300 HAT inhibitor C646, we demonstrated that HAT inhibition similarly affected many of the same endpoints as VPA, including increased reactive oxygen species and caspase-3 cleavage, the latter of which could be attenuated by pre-treatment with the antioxidant catalase. C646 exposure also decreased NF-κB/p65 protein, which was not due to reduced mRNA and was not attenuated with catalase pre-treatment. This study provides support for an adaptive HAT response following VPA exposure and suggests that reduced Cbp/p300 HAT activity could contribute to VPA-mediated alterations. PMID:27381264

  1. Decreased GATA5 mRNA expression associates with CpG island methylation and shortened recurrence-free survival in clear cell renal cell carcinoma

    International Nuclear Information System (INIS)

    GATA-5, a zinc-finger transcription factor and member of the GATA family proteins 1–6, is known to be involved in cellular differentiation. We recently found that tumor-specific hypermethylation of the GATA5 CpG island (CGI) occurs in renal cell carcinoma (RCC) and is associated with an adverse clinical outcome. In this study, we investigated whether epigenetic GATA5 alterations may result in changes in GATA5 mRNA expression levels and correlate with the observed prognostic impact of epigenetic changes in GATA5 in RCC. Quantitative real-time reverse-transcribed polymerase chain reaction was applied to measure relative GATA5 mRNA expression levels in 135 kidney tissue samples, including 77 clear cell RCC (ccRCC) tissues and 58 paired adjacent normal renal tissue samples. Relative GATA5 expression levels were determined using the ΔΔCt method and detection of three endogenous control genes then compared to previously measured values of relative methylation. The mean relative GATA5 mRNA expression level exhibited an approximately 31-fold reduction in tumor specimens compared with corresponding normal tissues (p < 0.001, paired t-test). Decreased GATA5 mRNA expression was inversely correlated with increased GATA5 CGI methylation (p < 0.001) and was associated with shortened recurrence-free survival in ccRCC patients (p = 0.023, hazard ratio = 0.25). GATA5 mRNA expression is decreased in ccRCC, likely due to gene silencing by methylation of the GATA5 CGI. Moreover, reduced GATA5 mRNA levels were associated with a poor clinical outcome, indicating a possible role of GATA5 for the development of aggressive ccRCC phenotypes

  2. LEDGF1-326 Decreases P23H and Wild Type Rhodopsin Aggregates and P23H Rhodopsin Mediated Cell Damage in Human Retinal Pigment Epithelial Cells

    OpenAIRE

    Rinku Baid; Scheinman, Robert I; Toshimichi Shinohara; Singh, Dhirendra P; Kompella, Uday B.

    2011-01-01

    BACKGROUND: P23H rhodopsin, a mutant rhodopsin, is known to aggregate and cause retinal degeneration. However, its effects on retinal pigment epithelial (RPE) cells are unknown. The purpose of this study was to determine the effect of P23H rhodopsin in RPE cells and further assess whether LEDGF(1-326), a protein devoid of heat shock elements of LEDGF, a cell survival factor, reduces P23H rhodopsin aggregates and any associated cellular damage. METHODS: ARPE-19 cells were transiently transfect...

  3. Decreased Expression of Inhibitor of Caspase-Activated DNase (ICAD in Renal Cell Carcinoma – Tissue Microarray of Human Samples

    Directory of Open Access Journals (Sweden)

    Retnagowri Rajandram

    2016-03-01

    Full Text Available Although primary localised tumours of renal cell carcinoma (RCC can be treated relatively successfully with surgery, metastatic RCC has poor prognosis because of late diagnosis and resistance to therapies. In the present study, we were interested in profiling the protein expression of “inhibitor of caspase-activated DNase” (ICAD, an apoptosis inhibitor, in kidney cancer and its paired normal kidney. Immunohistochemistry with automated batch staining and morphometry using digital pathology were used to compare ICAD in 121 RCC specimens with their paired normal kidney tissue. Tissue microarray of formalin-fixed, paraffin-embedded archival tissue was used. Intensity and localisation of ICAD were compared between normal and cancer samples, and against grading within the cancers. The results demonstrated that, in this cohort, ICAD was highly expressed in the proximal tubular epithelium of normal kidney, and significantly decreased in clear cell RCC tissue (p < 0.05 as well as other subtypes of RCC (p < 0.01 compared with normal kidney. There was a tendency towards nuclear localisation of ICAD in clear cell RCC, but not in other subtypes of RCC. No significant association was found between ICAD intensity and grade of RCC. In summary, down-regulation of ICAD occurs in RCC. ICAD normally inhibits DNA fragmentation and apoptosis; thus, its down-regulation was unexpected in a cancer known for its resistance to apoptosis. However, these RCC samples were from primary, not metastatic, RCC sites, and down-regulated ICAD may be part of a progressive pathway that promotes RCC metastasis.

  4. Selective decrease in colonic CD56+ T and CD161+ T cells in the inflamed mucosa of patients with ulcerative colitis

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    AIM: To investigate the role of local colonic mucosal NK receptor-positive T (NKR+ T) cells in the regulation of intestinal inflammation, we analyzed the population and function of these cells in ulcerative colitis (UC).METHODS: Colonic mucosal tissues were obtained from colonoscopic biopsies of the descending colon from 96 patients with UC (51 endoscopically uninflamed, 45 inflamed) and 18 normal controls. Endoscopic appearance and histologic score at the biopsied site were determined by Matts' classification. A single cell suspension was prepared from each biopsy by collagenase digestion.Two NKR+ T cell subsets, CD56+ (CD56+CD3+) T cells and CD161+(CD161+CD3+) T cells, were detected by flow cytometric analysis.Intracellular cytokine analysis for anti-inflammatory cytokine interleukin-10 (IL-10) was performed by in vitro stimulation with phorbol-myristate acetate (PMA) and ionomycin.RESULTS: CD56+ T cells and CD161+ T cells are present in the normal human colon and account for 6.7% and 21.3% of all mononuclear cells, respectively. The populations of both CD56+ T cells and CD161+ T cells were decreased significantly in the inflamed mucosa of UC. In contrast, the frequency of conventional T cells CD56-CD3+cells and CD161-CD3+ cells) was similar among the patient and control groups. The populations of NKR+ T cells were correlated inversely with the severity of inflammation, which was classified according to the endoscopic and histologic Matts' criteria.Interestingly, approximately 4% of mucosal NKR+ T cells expressing IL-10 were detected by in vitro stimulation with PMA and ionomycin.CONCLUSION: Selective reduction in the population of colonic mucosal NKR+T cells may contribute to the development of intestinal inflammation in UC.

  5. Decreased production of TNF-alpha by lymph node cells indicates experimental autoimmune encephalomyelitis remission in Lewis rats

    Directory of Open Access Journals (Sweden)

    Juliana Seger

    2010-05-01

    Full Text Available Experimental autoimmune encephalomyelitis (EAE is mediated by CD4+ Th1 cells that mainly secrete IFN-γ and TNF-α, important cytokines in the pathophysiology of the disease. Spontaneous remission is, in part, attributed to the down regulation of IFN-γ and TNF-α by TGF-β. In the current paper, we compared weight, histopathology and immunological parameters during the acute and recovery phases of EAE to establish the best biomarker for clinical remission. Female Lewis rats were immunised with myelin basic protein (MBP emulsified with complete Freund's adjuvant. Animals were evaluated daily for clinical score and weight prior to euthanisation. All immunised animals developed the expected characteristics of EAE during the acute phase, including significant weight loss and high clinical scores. Disease remission was associated with a significant reduction in clinical scores, although immunised rats did not regain their initial weight values. Brain inflammatory infiltrates were higher during the acute phase. During the remission phase, anti-myelin antibody levels increased, whereas TNF-α and IFN-γ production by lymph node cells cultured with MBP or concanavalin A, respectively, decreased. The most significant difference observed between the acute and recovery phases was in the induction of TNF-α levels in MBP-stimulated cultures. Therefore, the in vitro production of this cytokine could be used as a biomarker for EAE remission.

  6. Decrease of back recombination rate in CdS quantum dots sensitized solar cells using reduced graphene oxide

    Institute of Scientific and Technical Information of China (English)

    Ali Badawi

    2015-01-01

    The photovoltaic performance of CdS quantum dots sensitized solar cells (QDSSCs) using the 0.2 wt%of reduced graphene oxide and TiO2 nanoparticles (RGO+TiO2 nanocomposite) photoanode is investigated. CdS QDs are adsorbed onto RGO+TiO2 nanocomposite films by the successive ionic layer adsorption and reaction (SILAR) technique for several cycles. The current density–voltage (J–V ) characteristic curves of the assembled QDSSCs are measured at AM1.5 sim-ulated sunlight. The optimal photovoltaic performance for CdS QDSSC was achieved for six SILAR cycles. Solar cells based on the RGO+TiO2 nanocomposite photoanode achieve a 33%increase in conversion efficiency (η) compared with those based on plain TiO2 nanoparticle (NP) photoanodes. The electron back recombination rates decrease significantly for CdS QDSSCs based on RGO+TiO2 nanocomposite photoanodes. The lifetime constant (τ) for CdS QDSSC based on the RGO+TiO2 nanocomposite photoanode is at least one order of magnitude larger than that based on the bare TiO2NPs photoanode.

  7. Indirubins Decrease Glioma Invasion by Blocking Migratory Phenotypes in Both the Tumor and Stromal Endothelial Cell Compartments

    OpenAIRE

    Williams, Shanté P.; Nowicki, Michal O.; Liu, Fang; Press, Rachael; Godlewski, Jakub; Abdel-Rasoul, Mahmoud; Kaur, Balveen; Fernandez, Soledad A.; Chiocca, E. Antonio; Lawler, Sean E.

    2011-01-01

    Invasion and proliferation in neoplasia require the cooperation of tumor cell and endothelial compartments. Glycogen synthase kinase-3 (GSK-3) is increasingly recognized as a major contributor to signaling pathways that modulate invasion and proliferation. Here we show that GSK-3 inhibitors of the indirubin family reduce invasion of glioma cells and glioma-initiating cell-enriched neurospheres both in vitro and in vivo, and we show that β-catenin signaling plays an important role in mediating...

  8. Impaired progenitor cell function in HIV-negative infants of HIV-positive mothers results in decreased thymic output and low CD4 counts

    DEFF Research Database (Denmark)

    Nielsen, S D; Jeppesen, D L; Kolte, L;

    2001-01-01

    and fetal thymic organ cultures (FTOCs). Lower naive CD4 counts (459.3 +/- 68.9 vs 1128.9 +/- 146.8 cells/microL, P <.001) and reduced thymic output in infants of HIV-positive mothers were found (frequency of CD4(+) cells with TRECs was 3.6% +/- 0.7% compared with 14.3% +/- 2.2% in controls, P <.001......). In combination with lower red blood cell counts in infants of HIV-positive mothers, this finding suggested impairment of progenitor cell function. Indeed, progenitors from infants of HIV-positive mothers had decreased cloning efficiency (15.7% +/- 2.6% vs 55.8% +/- 15.9%, P =.009) and seemed to...... generate fewer T cells in FTOCs. In conclusion, lower numbers of naive CD4(+) cells and reduced thymic output in HIV-negative infants of HIV-positive mothers may be due to impaired progenitor cell function....

  9. Association between decreasing trend in the mortality of adult T-cell leukemia/lymphoma and allogeneic hematopoietic stem cell transplants in Japan: analysis of Japanese vital statistics and Japan Society for Hematopoietic Cell Transplantation (JSHCT)

    International Nuclear Information System (INIS)

    Adult T-cell leukemia/lymphoma (ATLL) is a peripheral T-cell neoplasm with a very poor outcome. However, several studies have shown a progress in the treatment. To evaluate the effect of the progress in the treatment of ATLL in a whole patient population, we used vital statistics data and estimated age-adjusted mortality and trends in the mortality from 1995 to 2009. Since allogeneic hematopoietic stem-cell transplantation (allo-HSCT) has been introduced as a modality with curative potential during study period, we also evaluated the association of the annual number of allo-HSCT and the trend of the mortality of ATLL. Endemic (Kyushu) and non-endemic areas (others) were evaluated separately. Significance in the trend of mortality was evaluated by joinpoint regression analysis. During the study period, a total of 14 932 patients died of ATLL in Japan, and mortality decreased significantly in both areas (annual percent change (95% confidence interval (CI)): Kyushu, −3.1% (−4.3, −1.9); others, −3.4% (−5.3, −1.5)). This decreasing trend in mortality seems to be associated with an increase in the number of allo-HSCTs (Kyushu, R-squared=0.70, P=0.003; and others, R-squared=0.55, P=0.058). This study reveals that the mortality of ATLL is now significantly decreasing in Japan and this decreasing trend might be associated with allo-HSCT

  10. 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 investigate...

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

    DEFF Research Database (Denmark)

    Bonfils, Linéa; Ellervik, Christina; Friedrich, Nele;

    2015-01-01

    Aims/hypothesis: Elevated serum ferritin levels are associated with an increased risk of type 2 diabetes, but the nature of this association remains elusive. The aim of this study was to test the hypothesis that an elevated fasting serum ferritin level is associated with an increased risk of type 2...... 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 <20th percentile. Elevated serum ferritin levels were associated with elevated plasma...... glucose levels at 0, 30 and 120 min (p < 0.001), elevated serum insulin levels at 0 and 120 min (p = 0.02 and p < 0.001), decreased beta cell function estimated as the insulinogenic index and corrected insulin response (p < 0.001), and decreased insulin sensitivity estimated by the Matsuda index of...

  12. 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)

    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

  13. Intratumoral application of standardized mistletoe extracts down regulates tumor weight via decreased cell proliferation, increased apoptosis and necrosis in a murine model.

    Science.gov (United States)

    Beuth, J; Ko, H L; Schneider, H; Tawadros, S; Kasper, H U; Zimst, H; Schierholz, J M

    2006-01-01

    The cytotoxic in vitro activity of standardized mistletoe extracts (ME) was examined by established assays towards the human ductal breast carcinoma cell line BT474. A dose-dependent (optimum 25 mg/mL medium) and significantly (p control mice with intratumoral phosphate-buffered saline (PBS) injections, tumors of the ME-A and ME-M treated groups showed a decreased cell proliferation rate, as well as an increased cell necrosis and apoptosis rate. Standardized mistletoe extracts, interfering with defined tumor cell functions, e.g., proliferation, necrosis and apoptosis, may have an impact on local cancer treatment. PMID:17201168

  14. Astaxanthin Normalizes Epigenetic Modifications of Bovine Somatic Cell Cloned Embryos and Decreases the Generation of Lipid Peroxidation.

    Science.gov (United States)

    Li, R; Wu, H; Zhuo, W W; Mao, Q F; Lan, H; Zhang, Y; Hua, S

    2015-10-01

    Astaxanthin is an extremely common antioxidant scavenging reactive oxygen species (ROS) and blocking lipid peroxidation. This study was conducted to investigate the effects of astaxanthin supplementation on oocyte maturation, and development of bovine somatic cell nuclear transfer (SCNT) embryos. Cumulus-oocyte complexes were cultured in maturation medium with astaxanthin (0, 0.5, 1.0, or 1.5 mg/l), respectively. We found that 0.5 mg/l astaxanthin supplementation significantly increased the proportion of oocyte maturation. Oocytes cultured in 0.5 mg/l astaxanthin supplementation were used to construct SCNT embryos and further cultured with 0, 0.5, 1.0 or 1.5 mg/l astaxanthin. The results showed that the supplementation of 0.5 mg/l astaxanthin significantly improved the proportions of cleavage and blastulation, as well as the total cell number in blastocysts compared with the control group, yet this influence was not concentration dependent. Chromosomal analyses revealed that more blastomeres showed a normal chromosomal complement in 0.5 mg/l astaxanthin treatment group, which was similar to that in IVF embryos. The methylation levels located on the exon 1 of the imprinted gene H19 and IGF2, pluripotent gene OCT4 were normalized, and global DNA methylation, H3K9 and H4K12 acetylation were also improved significantly, which was comparable to that in vitro fertilization (IVF) embryos. Moreover, we also found that astaxanthin supplementation significantly decreased the level of lipid peroxidation. Our findings showed that the supplementation of 0.5 mg/l astaxanthin to oocyte maturation medium and embryo culture medium improved oocyte maturation, SCNT embryo development, increased chromosomal stability and normalized the epigenetic modifications, as well as inhibited overproduction of lipid peroxidation. PMID:26280670

  15. Notch signaling mediates the age-associated decrease in adhesion of germline stem cells to the niche.

    Directory of Open Access Journals (Sweden)

    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.

  16. Electrical Stimulation Decreases Coupling Efficiency Between Beta-Adrenergic Receptors and Cyclic AMP Production in Cultured Muscle Cells

    Science.gov (United States)

    Young, R. B.; Bridge, K. Y.

    1999-01-01

    Electrical stimulation of skeletal muscle cells in culture is an effective way to simulate the effects of muscle contraction and its effects on gene expression in muscle cells. Expression of the beta-adrenergic receptor and its coupling to cyclic AMP synthesis are important components of the signaling system that controls muscle atrophy and hypertrophy, and the goal of this project was to determine if electrical stimulation altered the beta-adrenergic response in muscle cells. Chicken skeletal muscle cells that had been grown for seven days in culture were subjected to electrical stimulation for an additional two days at a pulse frequency of 0.5 pulses/sec and a pulse duration of 200 msec. At the end of this two-day stimulation period, beta-adrenergic receptor population was measured by the binding of tritium-labeled CGP-12177 to muscle cells, and coupling to cAMP synthesis was measured by Radioimmunoassay (RIA) after treating the cells for 10 min with the potent (beta)AR agonist, isoproterenol. The number of beta adrenergic receptors and the basal levels of intracellular cyclic AMP were not affected by electrical stimulation. However, the ability of these cells to synthesize cyclic AMP was reduced by approximately 50%. Thus, an enhanced level of contraction reduces the coupling efficiency of beta-adrenergic receptors for cyclic AMP production.

  17. Notch signaling mediates the age-associated decrease in adhesion of germline stem cells to the niche.

    Science.gov (United States)

    Tseng, Chen-Yuan; Kao, Shih-Han; Wan, Chih-Ling; Cho, Yueh; Tung, Shu-Yun; Hsu, Hwei-Jan

    2014-12-01

    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. PMID:25521289

  18. 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

    2016-01-01

    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. PMID:27245508

  19. TLR ligands of ryegrass pollen microbial contaminants enhance Th1 and Th2 responses and decrease induction of Foxp3(hi) regulatory T cells.

    Science.gov (United States)

    Mittag, Diana; Varese, Nirupama; Scholzen, Anja; Mansell, Ashley; Barker, Gillian; Rice, Gregory; Rolland, Jennifer M; O'Hehir, Robyn E

    2013-03-01

    Microbial contamination of grass pollens could affect sensitization, subsequent allergic response, and efficacy of allergen-specific immunotherapy. We investigated whether bacterial immunomodulatory substances can direct PBMC responses of allergic and nonatopic subjects against ryegrass pollen (RGP) toward Th1, Th2, or regulatory T (Treg) cells. Aqueous extracts of RGP with high or low LPS were fractionated into large and small molecular weight (MW) components by diafiltration. CFSE-labeled PBMCs from allergic and nonatopic subjects were stimulated with RGP extracts (RGPEs) and analyzed for cytokine secretion and T-cell responses. High LPS RGPE increased IFN-γ(+) Th1 and IL-4(+) Th2 effector cell induction and consistently decreased CD4(+) Foxp3(hi) Treg-cell induction. IL-10-producing T-cell frequency was unaltered, but IL-10 secretion was increased by high LPS RGPE. RGPE-stimulation of TLR-transfected cell lines revealed that high LPS pollen also contained a TLR2-ligand, and both batches a TLR9-ligand. Beta-1,3-glucans were detected in large and small MW fractions and were also T-cell stimulatory. In conclusion, coexposure to allergen and proinflammatory microbial stimuli does not convert an established Th2- into a Th1-response. Instead, proinflammatory responses are exacerbated and Foxp3(hi) Treg-cell induction is decreased. These findings show that adjuvants for specific immunotherapy should enhance Treg cells rather than target immune deviation from Th2 to Th1. PMID:23238878

  20. 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

  1. 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

  2. Accumulation of sigmaS due to enhanced synthesis and decreased degradation in acidic phospholipid-deficient Escherichia coli cells.

    Science.gov (United States)

    Uchiyama, Junji; Sasaki, Yu; Nagahama, Hideki; Itou, Aya; Matsuoka, Satoshi; Matsumoto, Kouji; Hara, Hiroshi

    2010-06-01

    The Escherichia coli pgsA3 mutation, which causes deficiency in acidic phospholipids, leads to a significant accumulation of sigma(S). This accumulation is partly accounted for by the higher transcription level of rpoS; however, it has also been suggested that the cells accumulate sigma(S) post-transcriptionally. We found that the level of the small regulatory RNA RprA, which is involved in the promotion of rpoS translation, is higher in pgsA3 cells than in pgsA(+) cells. Induction of altered rpoS mRNA that does not depend on RprA in pgsA(+) cells did not increase the level of sigma(S) to the high level observed in pgsA3 cells, suggesting post-translational sigma(S) accumulation in the latter. The mRNA levels of clpX and clpP, whose products form a ClpXP protease that degrades sigma(S), were much reduced in pgsA3 cells. Consistent with the reduced mRNA levels, the half-life of sigma(S) in pgsA3 cells was much longer than in pgsA(+) cells, indicating that downregulation of the degradation is a major cause for the high sigma(S) content. We show that the downregulation can be partially attributed to activated CpxAR in the mutant cells, which causes repression of rpoE on whose gene product the expression of clpPX depends. PMID:20455949

  3. n–3 PUFAs Reduce T-Helper 17 Cell Differentiation by Decreasing Responsiveness to Interleukin-6 in Isolated Mouse Splenic CD4+ T Cells123

    OpenAIRE

    Allen, M. Jeannie; Fan, Yang-Yi; Monk, Jennifer M.; Hou, Tim Y.; Barhoumi, Rola; McMurray, David N.; Chapkin, Robert S.

    2014-01-01

    Cluster of differentiation 4+ (CD4+) effector T-cell subsets [e.g., T-helper (Th) 1 and Th17] are implicated in autoimmune and inflammatory disorders such as multiple sclerosis, psoriasis, and rheumatoid arthritis. Interleukin (IL)-6 is a pleiotropic cytokine that induces Th17 polarization via signaling through the membrane-bound transducer glycoprotein 130 (GP130). Previously, we demonstrated that n–3 (ω-3) polyunsaturated fatty acids (PUFAs) reduce CD4+ T-cell activation and differentiation...

  4. Scorpion venom heat-resistant protein decreases immunoreactivity of OX-42-positive microglia cells in MPTP-treated mice

    Institute of Scientific and Technical Information of China (English)

    Shengming Yin; Deqin Yu; Xi Gao; Yan Peng; Yanhui Feng; Jie Zhao; Yiyuan Tang; Wanqin Zhang

    2008-01-01

    BACKGROUND: Microglia function as the immune surveyors of the brain under normal physiological conditions. However, microglia become activated in response to brain injuries and immunological stimulation. OBJECTIVE: To explore the influence of scorpion venom (SV) heat-resistant protein on frontal cortex and hippocampal microglia cells in a mice model of Parkinson's disease. DESIGN, TIME AND SETTING: Randomized, controlled, cellular immunity study. The experiment was performed at the Physiology Department Laboratory in Dalian Medical University between June 2005 and July 2008. MATERIALS: Ninety-six healthy, C57BI/6 mice; 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) from Sigma, USA; SV heat-resistant protein (Experimental Base Institute in Dalian Medical University). The mice were randomly divided into tour groups (n = 24): normal control, negative control, model, and SV heat-resistant protein. METHODS: Mice in the model and SV heat-resistant protein groups were subcutaneously injected with MPTP (20 mg/kg) to model Parkinson's disease, while the normal control and negative control groups were injected with physiological saline in the neck for 8 successive days. In addition, mice in the model and normal control groups were intraperitoneally injected with physiological saline 2 hours following administration, while SV heat-resistant protein and negative control groups were injected SV heat-resistant protein (0.01 mg/kg). MAIN OUTCOME MEASURES: Immunoreactivity of microglia cells in MPTP-treated mice. RESULTS: Compared with normal control mice, MPTP-treated mice displayed increased OX-42 expression in the brain. However, in the SV heat-resistant protein-treated mice, OX-42 expression was decreased, compared to the model group. In the model mouse group, the number of OX-42-positive microglia was increased in the frontal cortex, caudatum, and hippocampal hilus, compared to the normal control mice (P < 0.01). However, in the SV heat-resistant protein-treated mice

  5. 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. PMID:27183584

  6. 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

    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...... [myosin-light chain-2 (MLC2) phosphorylation], loss of cell polarity, loss of cell-cell junctions, luminal infiltration and basal invasion induced by AGE-modified basal lamina matrix in PEC acini. Our in vitro results were concordant with luminal occlusion of acini in the prostate glands of adult Endo180...

  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. Increased RANKL expression in peripheral T cells is associated with decreased bone mineral density in patients with COPD.

    Science.gov (United States)

    Chen, Ying; Bai, Peng; Liu, Lili; Han, Junyan; Zeng, Hui; Sun, Yongchang

    2016-08-01

    Receptor activator of nuclear factor-κB ligand (RANKL)-expressing adaptive T cells contribute to bone damage in autoimmune arthritis, although their role in chronic obstructive pulmonary disease (COPD)-associated osteoporosis is unknown. In the present study, the functional expression of RANKL in CD4+/CD8+ T cells and Th17 cells, and the potential role of these cells in COPD-associated bone loss was investigated. A total of 36 non-smokers, 38 smokers with normal lung function and 57 patients with COPD were enrolled. Femoral and vertebral bone mineral density (BMD) was assessed by dual energy X-ray absorptiometry. RANKL expression in peripheral CD4+ and CD8+ T cells and Th17 cells was evaluated by flow cytometry. For in vitro experiments, CD4+ and CD8+ T cells from 17 non-smokers were evaluated for RANKL expression following dose-dependent culture with cigarette smoke extract (CSE) for 5 days. The frequencies of RANKL-positive CD4+ and CD8+ T cells were higher in the patients with COPD than in the non-smokers (P=0.001 and P=0.002, respectively). The proportion of CD4+ T cells positive for both RANKL and interleukin-17 (IL-17) was higher in the patients with COPD than in the non-smokers (P=0.010). However, the frequency of RANKL-expressing Th17 cells was similar among all groups (P=0.508). The frequency of RANKL+CD4+ T cells inversely correlated with BMD of the lumbar vertebrae (P=0.01, r=-0.229), and that of the femoral neck (Pdiseases of the lung and bone in patients with COPD. PMID:27279356

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

    OpenAIRE

    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...

  10. Decreased SCF/c-kit signaling pathway contributes to loss of interstitial cells of Cajal in gallstone disease

    OpenAIRE

    Tan, Yu-Yan; Ji, Zhen-Ling; ZHAO, GANG; Jiang, Jia-Rui; Wang, Dong; Wang, Jing-Min

    2014-01-01

    Cholecystolithiasis is a common disease, and gallbladder dysmotility is considered as a pivotal pathogenesis. Interstitial cells of Cajal (ICCs) serve as pacemakers and mediators of neuromuscular transmission for gastrointestinal motility. Reduction of ICCs has been reported in gallstone diseases. However, there are no reasonable mechanisms for the cholecystolithiasis-associated loss of ICCs in humans. Stem cell factor (SCF) and its ligand c-kit are essential for normal development and surviv...

  11. Decreased Autocrine EGFR Signaling in Metastatic Breast Cancer Cells Inhibits Tumor Growth in Bone and Mammary Fat Pad

    OpenAIRE

    Nickerson, Nicole K.; Mohammad, Khalid S.; Gilmore, Jennifer L.; Crismore, Erin; Bruzzaniti, Angela; Guise, Theresa A.; Foley, John

    2012-01-01

    Breast cancer metastasis to bone triggers a vicious cycle of tumor growth linked to osteolysis. Breast cancer cells and osteoblasts express the epidermal growth factor receptor (EGFR) and produce ErbB family ligands, suggesting participation of these growth factors in autocrine and paracrine signaling within the bone microenvironment. EGFR ligand expression was profiled in the bone metastatic MDA-MB-231 cells (MDA-231), and agonist-induced signaling was examined in both breast cancer and oste...

  12. The high dosage of earthworm (Eisenia andrei) extract decreases cell proliferation and neuroblast differentiation in the mouse hippocampal dentate gyrus

    OpenAIRE

    Yan, Bing Chun; Yoo, Ki-Yeon; Park, Joon Ha; Lee, Choong Hyun; Choi, Jung Hoon; Won, Moo-Ho

    2011-01-01

    Earthworm extract has shown anticancer characteristics. In the present study, we examined the effect of chronic treatment with a high dose of earthworm (Eisenia andrei) extract (EE) on cell proliferation and neuroblast differentiation in the hippocampal dentate gyrus (DG) of 3-week-old mice using 5-bromo-2'-deoxyuridine (BrdU) and Ki-67 immunohistochemistry for cell proliferation and doublecortin (DCX) immunohistochemistry for neuroblast differentiation, respectively. BrdU-, Ki-67-, and DCX-i...

  13. Double targeting of Survivin and XIAP radiosensitizes 3D grown human colorectal tumor cells and decreases migration

    International Nuclear Information System (INIS)

    Background and purpose: In the present study, we aimed to investigate the effect of single and double knockdown of the inhibitor of apoptosis proteins (IAP) Survivin and X-linked IAP (XIAP) on three-dimensional (3D) clonogenic survival, migration capacity and underlying signaling pathways. Materials and methods: Colorectal cancer cell lines (HCT-15, SW48, SW480, SW620) were subjected to siRNA-mediated single or Survivin/XIAP double knockdown followed by 3D colony forming assays, cell cycle analysis, Caspase activity assays, migration assays, matrigel transmigration assays and Western blotting (Survivin, XIAP, Focal adhesion kinase (FAK), p-FAK Y397, Akt1, p-Akt1 S473, Extracellular signal-regulated kinase (ERK1/2), p-ERK1/2 T202/Y204, Glycogen synthase kinase (GSK)3β, p-GSK3β S9, nuclear factor (NF)-κB p65). Results: While basal cell survival was altered cell line-dependently, Survivin or XIAP single and Survivin/XIAP double knockdown enhanced cellular radiosensitivity of all tested cancer cell lines grown in 3D. Particularly double knockdown conditions revealed accumulation of cells in G2/M, increased subG1 fraction, elevated Caspase 3/7 activity, and reduced migration. Intracellular signaling showed dephosphorylation of FAK and Akt1 upon Survivin and/or Survivin/XIAP silencing. Conclusions: Our results strengthen the notion of Survivin and XIAP to act as radiation resistance factors and further indicate that these apoptosis-regulating proteins are also functioning in cell cycling and cell migration

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

    International Nuclear Information System (INIS)

    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

  15. 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.

  16. Knockdown of autophagy-related protein 5, ATG5, decreases oxidative stress and has an opposing effect on camptothecin-induced cytotoxicity in osteosarcoma cells

    International Nuclear Information System (INIS)

    Autophagy induction can increase or decrease anticancer drug efficacy. Anticancer drug-induced autophagy induction is poorly characterized in osteosarcoma (OS). In this study, we investigated the impact of autophagy inhibition on camptothecin (CPT)-induced cytotoxicity in OS. Autophagy-inhibited DLM8 and K7M3 metastatic murine OS cell lines were generated by infection with lentiviral shRNA directed against the essential autophagy protein ATG5. Knockdown of ATG5 protein expression and inhibition of autophagy was confirmed by immunoblot of ATG5 and LC3II proteins, respectively. Metabolic activity was determined by MTT assay and cell viability was determined by trypan blue exclusion. Acridine orange staining and immunoblotting for LC3II protein expression were used to determine autophagy induction. Oxidative stress was assessed by staining cells with HE and DCFH-DA followed by flow cytometry analysis. Mitochondrial membrane potential was determined by staining cells with TMRE followed by flow cytometry analysis. Immunoblotting was used to detect caspase activation, Parp cleavage and p53 phosphorylation. Autophagy inhibition caused a greater deficit in metabolic activity and cell growth in K7M3 cells compared to DLM8 cells. K7M3 cells exhibited higher basal autophagy levels than DLM8 cells and non-transformed murine MCT3 osteoblasts. Autophagy inhibition did not affect CPT-induced DNA damage. Autophagy inhibition decreased CPT-induced cell death in DLM8 cells while increasing CPT-induced cell death in K7M3 cells. Autophagy inhibition reduced CPT-induced mitochondrial damage and CPT-induced caspase activation in DLM8 cells. Buthionine sulfoximine (BSO)-induced cell death was greater in autophagy-competent DLM8 cells and was reversed by antioxidant pretreatment. Camptothecin-induced and BSO-induced autophagy induction was also reversed by antioxidant pretreatment. Significantly, autophagy inhibition not only reduced CPT-induced oxidative stress but also reduced basal

  17. 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. PMID:24916076

  18. Decreased proliferative, migrative and neuro-differentiative potential of postnatal rat enteric neural crest-derived cells during culture in vitro.

    Science.gov (United States)

    Yu, Hui; Pan, Wei-Kang; Zheng, Bai-Jun; Wang, Huai-Jie; Chen, Xin-Lin; Liu, Yong; Gao, Ya

    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(+) 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. PMID:27068376

  19. Decreased expression of the mannose 6- phosphate/insulin-like growth factor-II receptor promotes growth of human breast cancer cells

    International Nuclear Information System (INIS)

    Loss or mutation of the mannose 6-phosphate/insulin-like growth factor-II receptor (M6P/IGF2R) has been found in breast cancer. However, whether or not decreased levels of functional M6P/IGF2R directly contribute to the process of carcinogenesis needs to be further verified by functional studies. In this study, using viral and ribozyme strategies we reduced the expression of M6P/IGF2R in human breast cancer cells and then examined the effect on growth and apoptosis of these cells. Our results showed that infection of MCF-7 cells with the adenovirus carrying a ribozyme targeted against the M6P/IGF2R mRNA dramatically reduced the level of transcripts and the functional activity of M6P/IGF2R in these cells. Accordingly, cells treated with the ribozyme exhibited a higher growth rate and a lower apoptotic index than control cells (infected with a control vector). Furthermore, decreased expression of M6P/IGF2R enhanced IGF-II-induced proliferation and reduced cell susceptibility to TNF-induced apoptosis. These results suggest that M6P/IGF2R functions as a growth suppressor and its loss or mutation may contribute to development and progression of cancer. This study also demonstrates that adenoviral delivery of the ribozyme provides a useful tool for investigating the role of M6P/IGF2R in regulation of cell growth

  20. Effect of ulinastatin on growth inhibition, apoptosis of breast carcinoma cells is related to a decrease in signal conduction of JNk-2 and NF-κB

    Directory of Open Access Journals (Sweden)

    Wang Hong

    2012-01-01

    Full Text Available Abstract Objective This study aims to investigate the in vitro effects of Ulinastatin (UTI and Taxotere (TXT on cell proliferation; cell apoptosis; xenografted tumor growth; and expression of insulin-like growth factor receptor 1 (IGF-1R, platelet-derived growth factor A (PDGFA, nerve growth factor (NGF, c-Jun N-terminal kinase 2 (JNk-2, and NF-κB in a human primary breast cancer cells and breast cancer cell line MDA-MB-231. Methods The cell lines cultured were divided into four groups: 1 control group, 2 UTI group, 3 TXT group, and 4 UTI+TXT group. The method of MTT essay, flow cytometry, and RT-PCR were used to detect cell proliferation, cell apoptosis, and expression of IGF-1R, PDGFA, NGF, NF-κB, JNk-2, respectively. The growth of xenografted tumor in nude mice was used to calculate the anti-tumor rate. Immunohistochemistry staining (SP was used to detect the expression of IGF-1R, PDGFA, NGF, ki-67, caspase-3, JNk-2, and NF-κB. Results Proliferation of human breast cancer cells and MDA-MB-231 cell lines, and growth rate of xenografted tumor decreased in order of UTI+TXT > TXT > UTI > control, apoptosis increased in the order control Conclusions UTI 1 inhibits the proliferation of human breast cancer cells and the growth of xenografted tumors, 2 induces cancer cell apoptosis, and 3 enhances the anti-tumor effect of TXT. This mechanism might be related to decreasing signal transduction of JNk-2 and NF-κB, and then expression of IGF-1R, PDGFA, NGF.

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

    Energy Technology Data Exchange (ETDEWEB)

    Bagge, Annika [Department of Science, Systems and Models, Roskilde University, Roskilde (Denmark); Clausen, Trine R. [Diabetes Biology, Novo Nordisk, Maaloev (Denmark); Larsen, Sylvester [Department of Science, Systems and Models, Roskilde University, Roskilde (Denmark); Ladefoged, Mette [Diabetes Biology, Novo Nordisk, Maaloev (Denmark); Rosenstierne, Maiken W. [Department of Science, Systems and Models, Roskilde University, Roskilde (Denmark); Department of Virology, Statens Serum Institut (Denmark); Larsen, Louise [Department of Biomedical Sciences, University of Copenhagen, Copenhagen (Denmark); Vang, Ole [Department of Science, Systems and Models, Roskilde University, Roskilde (Denmark); Nielsen, Jens H. [Department of Biomedical Sciences, University of Copenhagen, Copenhagen (Denmark); Dalgaard, Louise T., E-mail: ltd@ruc.dk [Department of Science, Systems and Models, Roskilde University, Roskilde (Denmark)

    2012-09-21

    Highlights: Black-Right-Pointing-Pointer MicroRNA-29a (miR-29a) levels are increased by glucose in human and rat islets and INS-1E cells. Black-Right-Pointing-Pointer miR-29a increases proliferation of INS-1E beta-cells. Black-Right-Pointing-Pointer Forced expression of miR-29a decreases glucose-stimulated insulin secretion (GSIS). Black-Right-Pointing-Pointer Depletion of beta-cell miR-29a improves GSIS. Black-Right-Pointing-Pointer 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.

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

    International Nuclear Information System (INIS)

    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.

  3. 4-IBP, a σ1 Receptor Agonist, Decreases the Migration of Human Cancer Cells, Including Glioblastoma Cells, In Vitro and Sensitizes Them In Vitro and In Vivo to Cytotoxic Insults of Proapoptotic and Proautophagic Drugs

    Directory of Open Access Journals (Sweden)

    Veronique Mégalizzi

    2007-05-01

    Full Text Available Although the molecular function of cr receptors has not been fully defined and the natural ligand(s is still not known, there is increasing evidence that these receptors and their ligands might play a significant role in cancer biology. 4-(N-tibenzylpiperidin-4-yl-4iodobenzamide (4-IBP, a selective σ1, agonist, has been used to investigate whether this compound is able to modify: 1 in vitro the migration and proliferation of human cancer cells; 2 in vitro the sensitivity of human glioblastoma cells to cytotoxic drugs; and 3 in vivo in orthotopic glioblastoma and non-small cell lung carcinoma (NSCLC models the survival of mice coadministered cytotoxic agents. 4-IBP has revealed weak anti proliferative effects on human U373-MG glioblastoma and C32 melanoma cells but induced marked concentration-dependent decreases in the growth of human A549 NSCLC and PC3 prostate cancer cells. The compound was also significantly antimigratory in all four cancer cell lines. This may result, at least in U373-MG cells, from modifications to the actin cytoskeleton. 4-IBP modified the sensitivity of U373-MG cells in vitro to proapoptotic lomustin and proautophagic temozolomide, and markedly decreased the expression of two proteins involved in drug resistance: glucosylceramide synthase and Rho guanine nucleotide dissociation inhibitor. In vivo, 4-IBP increased the antitumor effects of temozolomide and irinotecan in immunodeficient mice that were orthotopically grafted with invasive cancer cells.

  4. Decrease in radio-sensitivity of the tumor by radiation-induced damage to immuno-related cells

    Energy Technology Data Exchange (ETDEWEB)

    Makidono, Reiko

    1987-08-01

    Immunological competence plays an important role in response of patients to radiation therapy and dose of radiation required for tumor control depends also on the immunocompetence of the individual patient. Radiation therapy (even localized irradiation) can, however, cause lymphopenia and induce an immunodeficient state. This may facilitate growth of residual tumor cells or metastatic foci, this negating benefits of the therapy. A brief overview of damage to T and B lymphocytes as well as macrophages and natural killer (NK) cells by radiation therapy was presented. The restoration and potentiation of the immunological competence of the patients by biological response modifiers (BRM) such as OK432 (a bacterial preparation), recombinant interferon (rIFN-..gamma..) and recombinant interleukin-2 (rIL-2) with or without lymphokine activated killer (LAK) cells, were discussed. (author) 61 refs.

  5. Decreased Circulating Interleukin-35 Levels Are Related to Interleukin-4-Producing CD8+ T Cells in Patients with Allergic Asthma

    OpenAIRE

    Wei Wang; Ping Li; Jiong Yang

    2015-01-01

     Interleukin (IL)-35 is a newly discovered suppressive cytokine and has been shown to alleviate  inflammatory  and  autoimmune  diseases.  The  purpose  of  this  study  was  to investigate immunomodulatory capacity of IL-35 in patients with allergic asthma.IL-35 mRNA expression levels in peripheral blood mononuclear cells (PBMCs) were detected  by  quantitative  real-time  PCR  (qPCR).  The  frequencies  of  cytotoxic  T  cells (Tc)1,Tc2  and  Tc17  cells  were  measured  by  flow  cytometry...

  6. Dietary supplementation with zinc oxide decreases expression of the stem cell factor in the small intestine of weanling pigs.

    Science.gov (United States)

    Ou, Deyuan; Li, Defa; Cao, Yunhe; Li, Xilong; Yin, Jingdong; Qiao, Shiyan; Wu, Guoyao

    2007-12-01

    Dietary supplementation with a high level of zinc oxide (ZnO) has been shown to reduce the incidence of diarrhea in weanling pigs, but the underlying mechanisms remain largely unknown. Intestinal-mucosal mast cells, whose maturation and proliferation is under the control of the stem cell factor (SCF), play an important role in the etiology of diarrhea by releasing histamine. The present study was conducted to test the novel hypothesis that supplementing ZnO to the diet for weanling piglets may inhibit SCF expression in the small intestine, thereby reducing the number of mast cells, histamine release, and diarrhea. In Experiment 1, 32 piglets (28 days of age) were weaned and fed diets containing 100 or 3000 mg zinc/kg (as ZnO) for 10 days (16 piglets per group). In Experiment 2, two groups of 28-day-old piglets (8 piglets per group) were fed the 100- or 3000-mg zinc/kg diet as in Experiment 1, except that they were pair-fed the same amounts of feed. Supplementation with a high level of ZnO reduced the incidence of diarrhea in weanling piglets. Dietary Zn supplementation reduced expression of the SCF gene at both mRNA and protein levels, the number of mast cells in the mucosa and submucosa of the small intestine and histamine release from mucosal mast cells. Collectively, our results indicate that dietary supplementation with ZnO inhibits SCF expression in the small intestine, leading to reductions in the number of mast cells and histamine release. These findings may have important implications for the prevention of weaning-associated diarrhea in piglets. PMID:17475461

  7. Insulin-like growth factors decrease oxygen-regulated erythropoietin production by human hepatoma cells (Hep G2)

    OpenAIRE

    Scholz, H; Baier, W.; Ratcliffe, P.; Eckardt, K.; Zapf, J.; Kurtz, Armin; Bauer, C

    1992-01-01

    We examined the effects of insulin-like growth factors (IGFs) and insulin on erythropoietin (EPO) production by human hepatoma cells (Hep G2). Compared with normoxia (20% O2), EPO production by Hep G2 cells during a 72-h incubation was stimulated fivefold by exposure to low oxygen tension (1% O2) and nearly threefold by exposure to cobalt chloride (100 microM). IGF-I caused a concentration-dependent attenuation of EPO formation under normoxic conditions and inhibited (maximally 50%) EPO produ...

  8. Downregulation of β-catenin decreases the tumorigenicity, but promotes epithelial-mesenchymal transition in breast cancer cells

    OpenAIRE

    Kai Cai; Longwei Jiang; Jing Wang; Hongyi Zhang; Xiaoying Wang; Dengyu Cheng; Jun Dou

    2014-01-01

    Background: Wnt/β-catenin signaling pathway plays a key role in human breast cancer progression. In this study, we down regulated β-catenin expression in human breast cancer MDA-MB-231 cells and investigated the effect of β-catenin knockdown on the cell biological characteristics. Materials and Methods: The recombinant plasmids of pSUPER-enhancement green fluorescent protein 1 (EGFP1)-scrabble-β-catenin-short hairpin ribonucleic acid (shRNA) and pSUPER-EGFP1-β-catenin-shRNA-1 were transfe...

  9. Etanercept decreases HMGB1 expression in dorsal root ganglion neuron cells in a rat chronic constriction injury model

    OpenAIRE

    WANG, RUI-KE; Zhang, Qin-Qin; PAN, YUN-DAN; Guo, Qu-Lian

    2012-01-01

    In the present study, we examined the effect of etanercept on high mobility group box 1 (HMGB1) expression in dorsal root ganglion (DRG) neuron cells in a rat model of chronic constriction injury (CCI) of the sciatic nerve, with the aim of exploring the molecular mechanism underlying the therapeutic effect of etanercept on sciatica-related nociception and the potential interaction between tumor necrosis factor-α (TNF-α) and HMGB1 in DRG neuron cells. A rat CCI model was employed and the anima...

  10. Experimental evaluation of decrease in the activities of polyphosphate/glycogen-accumulating organisms due to cell death and activity decay in activated sludge.

    Science.gov (United States)

    Hao, Xiaodi; Wang, Qilin; Cao, Yali; van Loosdrecht, Mark C M

    2010-06-15

    Decrease in bacterial activity (biomass decay) in activated sludge can result from cell death (reduction in the amount of active bacteria) and activity decay (reduction in the specific activity of active bacteria). The goal of this study was to experimentally differentiate between cell death and activity decay as the cause of decrease in bacterial activity. By means of measuring maximal anaerobic phosphate release rates, verifying membrane integrity by live/dead staining and verifying presence of 16S rRNA with fluorescence in situ hybridization (FISH), the decay rates and death rates of polyphosphate-accumulating organisms (PAOs) in a biological nutrient removal (BNR) system and a laboratory phosphate removing sequencing batch reactor (SBR) system were determined, respectively, under famine conditions. In addition, the decay rate and death rate of glycogen-accumulating organisms (GAOs) in a SBR system with an enrichment culture of GAOs were also measured under famine conditions. Hereto the maximal anaerobic volatile fatty acid uptake rates, live/dead staining, and FISH were used. The experiments revealed that in the BNR and enriched PAO-SBR systems, activity decay contributed 58% and 80% to the decreased activities of PAOs, and that cell death was responsible for 42% and 20% of decreases in their respective activities. In the enriched GAOs system, activity decay constituted a proportion of 74% of the decreased activity of GAOs, and cell death only accounted for 26% of the decrease of their activity. PMID:20178124

  11. 2,2',4,4'-Tetrabromodiphenyl ether injures cell viability and mitochondrial function of mouse spermatocytes by decreasing mitochondrial proteins Atp5b and Uqcrc1.

    Science.gov (United States)

    Huang, Shaoping; Wang, Jing; Cui, Yiqiang

    2016-09-01

    Our object was to explore direct effects and mechanism of BDE47 on GC2 (immortalized mouse spermatocyte). GC2 were exposed to DMSO, 0.1, 1, 10, 100μM BDE47 for 48h. Cell viability was detected by trypan-blue exclusion; ultrastructure by electron-microscopy; cell cycle, mitochondrial membrane motential (MMP), reactive oxygen species (ROS) by flow-cytometry; ATP production by luminometer; Atp5b, Uqcrc1, Bcl-2 level by WB. To explore whether the decreased mitochondrial proteins play an important role in apoptosis, MMP and apoptosis were detected after Atp5b or Uqcrc1 knockdown in GC2. Results showed BDE47 reduced cell viability, caused condensation of nuclear and vacuolated mitochondria, decreased MMP and ATP, induced ROS, cell cycle arrest at S and G2/M phase, reduced Atp5b, Uqcrc1, Bcl-2 in GC2. Knockdown of Atp5b or Uqcrc1 decreased MMP, induced apoptosis in GC2. Results suggested that BDE47 reduced cell viability, injured mitochondria in spermatocytes probably by decreasing mitochondrial protein Atp5b and Uqcrc1. PMID:27525561

  12. Decreased gene expression of human beta-defensin-1 in the development of squamous cell carcinoma of the oral cavity.

    NARCIS (Netherlands)

    Wenghoefer, M.H.; Pantelis, A.; Dommisch, H.; Reich, R.; Martini, M.; Allam, J.P.; Novak, N.; Berge, S.; Jepsen, S.; Winter, J.

    2008-01-01

    The aim of this study was to investigate the gene expression of human beta-defensin-1, -2, -3 (hBD-1, -2, -3), interleukin-1beta, tumour necrosis factor-alpha and cyclooxygenase-2 in oral squamous cell carcinoma (OSCC) compared to benign and premalignant lesions as well as healthy controls. Biopsies

  13. Dietary phenethyl isothiocyanate inhibition of androgen-responsive LNCaP prostate cancer cell tumor growth correlates with decreased angiogenesis

    Science.gov (United States)

    Phenethyl isothiocyanate (PEITC), found in certain cruciferous vegetables, has antitumor activity in several cancer models, including prostate cancer. In our xenograft model, dietary administration of PEITC (100-150 mg/kg/d) inhibited androgen-responsive LNCaP human prostate cancer cell tumor growth...

  14. Decrease of FOXP3 mRNA in CD4~+ T cells in latent autoimmune diabetes in adult

    Institute of Scientific and Technical Information of China (English)

    杨治芳

    2006-01-01

    Objective To study the percentage of peripheral blood CD4+ CD25+ T cells and the expression of F0XP3 mRNA in patients with latent autoimmune diabetes in adult (LADA). Methods Fresh peripheral blood samples were obtained from 60 patients with LADA,30 patients with type 2 diabetes and 30 age- and sex-matched

  15. Optimization of temperature, sugar concentration, and inoculum size to maximize ethanol production without significant decrease in yeast cell viability

    Energy Technology Data Exchange (ETDEWEB)

    Laluce, Cecilia; Morais, Meline Rezende [Inst. de Quimica de Araraquara-UNESP, SP (Brazil). Dept. of Biochemistry and Biotechnological Chemistry; Tognolli, Joao Olimpio [Inst. de Quimica de Araraquara-UNESP, SP (Brazil). Dept. of Analytical Chemistry; Oliveira, Karen Fernanda de; Souza, Crisla Serra [Cidade Universitaria, Sao Paulo, SP (Brazil). Programa de Pos-Graduacao Interunidades em Biotecnologia

    2009-06-15

    Aiming to obtain rapid fermentations with high ethanol yields and a retention of high final viabilities (responses), a 2{sup 3} full-factorial central composite design combined with response surface methodology was employed using inoculum size, sucrose concentration, and temperature as independent variables. From this statistical treatment, two well-fitted regression equations having coefficients significant at the 5% level were obtained to predict the viability and ethanol production responses. Three-dimensional response surfaces showed that increasing temperatures had greater negative effects on viability than on ethanol production. Increasing sucrose concentrations improved both ethanol production and viability. The interactions between the inoculum size and the sucrose concentrations had no significant effect on viability. Thus, the lowering of the process temperature is recommended in order to minimize cell mortality and maintain high levels of ethanol production when the temperature is on the increase in the industrial reactor. Optimized conditions (200 g/l initial sucrose, 40 g/l of dry cell mass, 30 C) were experimentally confirmed and the optimal responses are 80.8{+-}2.0 g/l of maximal ethanol plus a viability retention of 99.0{+-}3.0% for a 4-h fermentation period. During consecutive fermentations with cell reuse, the yeast cell viability has to be kept at a high level in order to prevent the collapse of the process. (orig.)

  16. In vivo selection for spine-derived highly metastatic lung cancer cells is associated with increased migration, inflammation and decreased adhesion.

    Science.gov (United States)

    Cai, Xiaopan; Luo, Jian; Yang, Xinghai; Deng, Huayun; Zhang, Jishen; Li, Shichang; Wei, Haifeng; Yang, Cheng; Xu, Leqin; Jin, Rongrong; Li, Zhenxi; Zhou, Wang; Ding, JianDong; Chu, Jianjun; Jia, Lianshun; Jia, Qi; Tan, Chengjun; Liu, Mingyao; Xiao, Jianru

    2015-09-01

    We developed a murine spine metastasis model by screening five metastatic non-small cell lung cancer cell lines (PC-9, A549, NCI-H1299, NCI-H460, H2030). A549 cells displayed the highest tendency towards spine metastases. After three rounds of selection in vivo, we isolated a clone named A549L6, which induced spine metastasis in 80% of injected mice. The parameters of the A549L6 cell spinal metastatic mouse models were consistent with clinical spine metastasis features. All the spinal metastatic mice developed symptoms of nerve compression after 40 days. A549L6 cells had increased migration, invasiveness and decreased adhesion compared to the original A549L0 cells. In contrast, there was no significant differences in cell proliferation, apoptosis and sensitivity to chemotherapeutic agents such as cisplatin. Comparative transcriptomic analysis and real-time PCR analysis showed that expression of signaling molecules regulating several tumor properties including migration (MYL9), metastasis (CEACAM6, VEGFC, CX3CL1, CST1, CCL5, S100A9, IGF1, NOTCH3), adhesion (FN1, CEACAM1) and inflammation (TRAF2, NFκB2 and RelB) were altered in A549L6 cells. We suggest that migration, adhesion and inflammation related genes contribute to spine metastatic capacity. PMID:26090868

  17. Time dependence of micronucleus frequency in germinating onion root tip cells and growth decrease of plant's roots irradiated by accelerated heavy ions

    International Nuclear Information System (INIS)

    Micronucleus induction of germinating onion seed root tip cells delayed with increase of radiation dose. The frequency at the time the cells showed the maximum frequency indicated a dose relationship of upward bell shape. Growth decrease of germinating green sorgo seed roots by radiations were severer than onion and it showed also linear energy transfer (LET) dependence like onion. Alphalfa seeds are very strong for radiation, however, finally showed recognizable growth decrease at high dose of 60 Gy, and no LET dependence was found. (author)

  18. Acidosis Decreases c-Myc Oncogene Expression in Human Lymphoma Cells: A Role for the Proton-Sensing G Protein-Coupled Receptor TDAG8

    OpenAIRE

    Zhigang Li; Lixue Dong; Eric Dean; Yang, Li V.

    2013-01-01

    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 partia...

  19. 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.

  20. Prolactin receptor attenuation induces zinc pool redistribution through ZnT2 and decreases invasion in MDA-MB-453 breast cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    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.

  1. Prolactin receptor attenuation induces zinc pool redistribution through ZnT2 and decreases invasion in MDA-MB-453 breast cancer cells

    International Nuclear Information System (INIS)

    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

  2. Lipoic acid decreases Mcl-1, Bcl-xL and up regulates Bim on ovarian carcinoma cells leading to cell death.

    OpenAIRE

    Kafara, Perrine; Icard, Philippe; Guillamin, Marilyne; Schwartz, Laurent; Lincet, Hubert

    2014-01-01

    Background Ovarian carcinoma is the leading cause of death from gynecological cancer because there is risk of chemoresistance. As previously demonstrated in our laboratory, Alpha-lipoic acid (LA), a co-factor for metabolic enzymes, suppresses the tumor growth. In this study, we have researched the mechanisms that are responsible for the activity of LA. Methods We have studied the mechanisms of LA in two ovarian cancer cell lines, a cisplatin sensitive one, IGROV1 and its resistant counterpart...

  3. Decreased numbers of peripheral blood dendritic cells in patients with coronary artery disease are associated with diminished plasma Flt3 ligand levels and impaired plasmacytoid dendritic cell function

    OpenAIRE

    Van Brussel, Ilse; Van Vré, Emily A; De Meyer, Guido RY; Vrints, Christiaan J.; Bosmans, Johan M; Bult, Hidde

    2011-01-01

    Abstract Background: We investigate whether activation of circulating DCs or levels of FMS-like tyrosine kinase 3 ligand (Flt3L) and granulocyte-macrophage colony stimulating factor (GM-CSF), haematopoietic growth factors important for dendritic cell (DC) differentiation, could account for reduced blood DC numbers in coronary artery disease (CAD) patients. Methods: Concentrations of Flt3L and GM-CSF were measured in plasma from CAD patients (n=15) and controls (n=12). Frequenc...

  4. Global Decrease of Histone H3K27 Acetylation in ZEB1-Induced Epithelial to Mesenchymal Transition in Lung Cancer Cells

    International Nuclear Information System (INIS)

    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

  5. 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. PMID:19444642

  6. Increased level of both CD4+FOXP3+ regulatory T cells and CD14+HLA-DR⁻/low myeloid-derived suppressor cells and decreased level of dendritic cells in patients with multiple myeloma

    DEFF Research Database (Denmark)

    Brimnes, M K; Vangsted, Annette Juul; Knudsen, L M;

    2010-01-01

    immune status in patients with MM seems crucial prior to active immune therapy. We evaluated the proportion of both, DC, Treg cells and myeloid-derived suppressor cells (MDSC) in peripheral blood from patients with MM at diagnosis and in remission as well as patients with monoclonal gammopathy of......+FOXP3+ Treg cells was increased in patients at diagnosis and not in patients in remission or with MGUS. Also, Treg cells from patients with MM were functionally intact as they were able to inhibit proliferation of both CD4 and CD8 T cells. Finally, we observed an increase in the proportion of CD14+HLA......-DR¿/low MDSC in patients with MM at diagnosis, illustrating that this cell fraction is also distorted in patients with MM. Taken together, our results illustrate that, both mDC, pDC, Treg cells and MDSC are affected in patients with MM underlining the fact that the immune system is dysregulated as a...

  7. Dietary omega-3 fatty acids decrease mortality and Kupffer cell prostaglandin E2 production in a rat model of chronic sepsis.

    Science.gov (United States)

    Barton, R G; Wells, C L; Carlson, A; Singh, R; Sullivan, J J; Cerra, F B

    1991-06-01

    We tested the hypothesis that substitution of omega-3 fat for dietary omega-6 fat would reduce mortality and decrease Kupffer cell prostaglandin E2 (PGE2) production in a rat model of chronic sepsis. Rats were fed via gastrostomy for 12 days with isonitrogenous, isocaloric diets containing 15% of calories as either safflower oil (omega-6) or a 10:1 mixture of menhaden oil (omega-3) and safflower oil. After five days of feeding, animals received an intra-abdominal abscess of defined bacterial content. Survivors were killed on post-laparotomy day 6 in conjunction with liver perfusion and protease liver digestion for Kupffer cell isolation. Kupffer cell PGE2 production was measured by radioimmunoassay after 18 hours of cell culture and again after stimulation with 0 LPS, 10 ng/ml LPS, and 10 micrograms/LPS. Mortality was decreased in menhaden oil-fed animals compared with safflower oil-fed animals (16% vs. 35%). Kupffer cell PGE2 production was decreased in menhaden oil-fed animals at 18 hours (354 +/- 54 vs. 570 +/- 95 pg/0.1 ml; p = 0.09) and after stimulation with 10 micrograms/ml LPS (140 +/- 41 vs. 288 +/- 45 pg/0.1 ml; p = 0.03) compared with safflower oil-fed animals. PMID:2056540

  8. 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

    identification and immunoprecipitation studies were used for DNA binding studies. Results: Microarray transcript profiling of 89 bladder biopsies showed a significant up-regulation of TOX3 (p<10-4) in non-muscle invasive (Ta-T1) bladder tumors compared to muscle-invasive (T2-T4) bladder tumors and normal...... 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...... expressing cell extracts with an artificial “GAS”- DNA element resulted in an enrichment of the GAS containing DNA-sequence, providing evidence for a potential interaction of TOX3 with the GAS-sequence of STAT1. Conclusions: These results provide evidence for an alternative activation of the downstream...

  9. Decreased sensitivity of multidrug-resistant tumor cells to cisplatin is correlated with sorcin gene co-amplification

    International Nuclear Information System (INIS)

    A set of multidrug resistant (MDR) murine leukemia P388 sublines possessing 30-50-fold mdr1 gene amplification was obtained as a result of experimental chemotherapy with rubomycin, ruboxyl, vinblastine, vincristine, or combination of rubomycin and vincristine. Significant differences of developed MDR sublines in response to treatment with cisplatin, tiophosphamide, sarcolysin, and dopad were found. Strong correlation between drug sensitivity and a copy number of gene coding for 19-22 kDa calcium-binding sorcin gene co-amplification were hypersensitive to cisplatin and alkylating agents, the cell sublines showing amplification of sorcin DNA sequences did not possess such collateral sensitivity and even acquired cross-resistance. The dependence of sensitivity to cisplatin on sorcin gene co-amplification was confirmed by analysis of Djungarian hamster DM15 cell sublines that selected for MDR in vitro by colchicine. (author)

  10. Impaired Telomere Maintenance and Decreased Canonical WNT Signaling but Normal Ribosome Biogenesis in Induced Pluripotent Stem Cells from X-Linked Dyskeratosis Congenita Patients.

    Science.gov (United States)

    Gu, Bai-Wei; Apicella, Marisa; Mills, Jason; Fan, Jian-Meng; Reeves, Dara A; French, Deborah; Podsakoff, Gregory M; Bessler, Monica; Mason, Philip J

    2015-01-01

    Dyskeratosis congenita (DC) is an inherited bone marrow failure syndrome characterized by the presence of short telomeres at presentation. Mutations in ten different genes, whose products are involved in the telomere maintenance pathway, have been shown to cause DC. The X-linked form is the most common form of the disease and is caused by mutations in the gene DKC1, encoding the protein dyskerin. Dyskerin is required for the assembly and stability of telomerase and is also involved in ribosomal RNA (rRNA) processing where it converts specific uridines to pseudouridine. DC is thought to result from failure to maintain tissues, like blood, that are renewed by stem cell activity, but research into pathogenic mechanisms has been hampered by the difficulty of obtaining stem cells from patients. We reasoned that induced pluripotent stem (iPS) cells from X-linked DC patients may provide information about the mechanisms involved. Here we describe the production of iPS cells from DC patients with DKC1 mutations Q31E, A353V and ΔL37. In addition we constructed "corrected" lines with a copy of the wild type dyskerin cDNA expressed from the AAVS1 safe harbor locus. We show that in iPS cells with DKC1 mutations telomere maintenance is compromised with short telomere lengths and decreased telomerase activity. The degree to which telomere lengths are affected by expression of telomerase during reprograming, or with ectopic expression of wild type dyskerin, is variable. The recurrent mutation A353V shows the most severe effect on telomere maintenance. A353V cells but not Q31E or ΔL37 cells, are refractory to correction by expression of wild type DKC1 cDNA. Because dyskerin is involved in both telomere maintenance and ribosome biogenesis it has been postulated that defective ribosome biogenesis and translation may contribute to the disease phenotype. Evidence from mouse and zebra fish models has supported the involvement of ribosome biogenesis but primary cells from human

  11. Epidermal growth factor decreases PEPT2 transport capacity and expression in the rat kidney proximal tubule cell line SKPT0193 cl.2

    DEFF Research Database (Denmark)

    Bravo, Silvina A; Nielsen, Carsten Uhd; Amstrup, Jan; Frokjaer, Sven; Brodin, Birger

    2004-01-01

    suggests that this might be disadvantageous when studying PEPT2-mediated transport phenomena. These findings demonstrate for the first time EGF-mediated regulation of PEPT2 expression in a kidney cell line. The relevance for kidney regulation of peptide transport activity in physiological and......The renal peptide transporter PEPT2 plays an important role in absorption of di- and tripetides in the proximal tubule; however, knowledge of regulation of PEPT2 by growth factors and hormones is limited. In the present study, we examined the effects of epidermal growth factor (EGF) on PEPT2...... transport capacity and expression in the rat proximal tubule cell line SKPT0193 cl.2 (SKPT), which expresses rat PEPT2 (rPEPT2) in the apical membrane. Treatment of SKPT cells with EGF during cell culture growth caused a dose-dependent decrease in rPEPT2 transport capacity and expression, as determined by...

  12. Ion-neutral gas reactions in a collision/reaction cell in inductively coupled plasma mass spectrometry: Correlation of ion signal decrease to kinetic rate constants

    International Nuclear Information System (INIS)

    Reaction gas flow rate dependent Ar2+ and Ar+ signals are correlated to fundamental kinetic rate coefficients. A simple calculation, assuming that gas exits the reaction cell due only to effusion, is described to estimate the gas pressure in the reaction cell. The value of the product of the kinetic rate constant and the ion residence time in the reaction cell can be determined from experimental measurement of the decrease in an ion signal as a function of reaction gas flow rate. New kinetic rate constants are determined for the reaction of CH3F with Ar+ and Ar2+. - Highlights: • How to determine pressure and the product of the kinetic rate constant times the ion residence time in reaction cell • Relate measured ICP-DRC-MS signals versus gas flow rate to kinetic rate constants measured previously using SIFT-MS • Describe how to determine previously unmeasured kinetic rate constants using ICP-DRC-MS

  13. 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.

  14. Nitric oxide decreases the sensitivity of pulmonary endothelial cells to LPS-induced apoptosis in a zinc-dependent fashion.

    Science.gov (United States)

    Tang, Zi-Lue; Wasserloos, Karla J; Liu, Xianghong; Stitt, Molly S; Reynolds, Ian J; Pitt, Bruce R; St Croix, Claudette M

    2002-01-01

    We hypothesized that: (a) S-nitrosylation of metallothionein (MT) is a component of pulmonary endothelial cell nitric oxide (NO) signaling that is associated with an increase in labile zinc; and (b) NO mediated increases in labile zinc in turn reduce the sensitivity of pulmonary endothelium to LPS-induced apoptosis. We used microspectrofluorometric techniques to show that exposing mouse lung endothelial cells (MLEC) to the NO-donor, S-nitrosocysteine, resulted in a 45% increase in fluorescence of the Zn2+-specific fluorophore, Zinquin, that was rapidly reversed by exposure to the Zn2+ chelator, NNN'N'-tetrakis-(2-pyridylmethyl)ethylenediamine; TPEN). The absence of a NO-mediated increase in labile Zn2+ in MLEC from MT-I and -II knockout mice inferred a critical role for MT in the regulation of Zn2+ homeostasis by NO. Furthermore, we found that prior exposure of cultured endothelial cells from sheep pulmonary artery (SPAEC), to the NO-donor, S-nitroso-N-acetylpenicillamine (SNAP) reduced their sensitivity to lipopolysaccharide (LPS) induced apoptosis. The anti-apoptotic effects of NO were significantly inhibited by Zn2+ chelation with low doses of TPEN (10 microM). Collectively, these data suggest that S-nitrosylation of MT is associated with an increase in labile (TPEN chelatable) zinc and NO-mediated MT dependent zinc release is associated with reduced sensitivity to LPS-induced apoptosis in pulmonary endothelium. PMID:12162436

  15. Fluoxetine reduces CES1, CES2, and CYP3A4 expression through decreasing PXR and increasing DEC1 in HepG2 cells.

    Science.gov (United States)

    Shang, Wei; Liu, Jie; Chen, Ruini; Ning, Rui; Xiong, Jing; Liu, Wei; Mao, Zhao; Hu, Gang; Yang, Jian

    2016-05-01

    1. This study investigated the mechanisms of the decreases of carboxylesterases (CES) and cytochrome P4503A4 (CYP3A4) and the enzymatic activities induced by fluoxetine (FLX) in HepG2 cells. We found that FLX decreased the carboxylesterase 1 (CES1) and carboxylesterase 2 (CES2) expression and the hydrolytic activity. 2. FLX decreased the pregnane X receptor (PXR) expression which regulated the target genes such as CYP3A4, whereas increased the differentiated embryonic chondrocyte-expressed gene 1 (DEC1) expression. 3. FLX repressed the PXR at transcriptional level. 4. Overexpression of PXR alone increased the expression of CES1, CES2, and CYP3A4 and attenuated the decreases of CES1, CES2, and CYP3A4 induced by FLX. On the contrary, knockdown of PXR alone decreased the expression of CES1, CES2, and CYP3A4 and almost abolished the decreases of CES1, CES2, and CYP3A4 induced by FLX. 5. Knockdown of DEC1 alone increased the expression of PXR and CYP3A4 and almost abolished the decreases of CES1, CES2, and CYP3A4 induced by FLX. 6. Taken together, the decreases of CES and CYP3A4 expression and enzymatic activities induced by FLX are through decreasing PXR and increasing DEC1 in HepG2 cells. PMID:26340669

  16. Adenovirus-mediated hAQP1 expression in irradiated mouse salivary glands causes recovery of saliva secretion by enhancing acinar cell volume decrease.

    Science.gov (United States)

    Teos, L Y; Zheng, C-Y; Liu, X; Swaim, W D; Goldsmith, C M; Cotrim, A P; Baum, B J; Ambudkar, I S

    2016-07-01

    Head and neck irradiation (IR) during cancer treatment causes by-stander effects on the salivary glands leading to irreversible loss of saliva secretion. The mechanism underlying loss of fluid secretion is not understood and no adequate therapy is currently available. Delivery of an adenoviral vector encoding human aquaporin-1 (hAQP1) into the salivary glands of human subjects and animal models with radiation-induced salivary hypofunction leads to significant recovery of saliva secretion and symptomatic relief in subjects. To elucidate the mechanism underlying loss of salivary secretion and the basis for AdhAQP1-dependent recovery of salivary gland function we assessed submandibular gland function in control mice and mice 2 and 8 months after treatment with a single 15-Gy dose of IR (delivered to the salivary gland region). Salivary secretion and neurotransmitter-stimulated changes in acinar cell volume, an in vitro read-out for fluid secretion, were monitored. Consistent with the sustained 60% loss of fluid secretion following IR, a carbachol (CCh)-induced decrease in acinar cell volume from the glands of mice post IR was transient and attenuated as compared with that in cells from non-IR age-matched mice. The hAQP1 expression in non-IR mice induced no significant effect on salivary fluid secretion or CCh-stimulated cell volume changes, except in acinar cells from 8-month group where the initial rate of cell shrinkage was increased. Importantly, the expression of hAQP1 in the glands of mice post IR induced recovery of salivary fluid secretion and a volume decrease in acinar cells to levels similar to those in cells from non-IR mice. The initial rates of CCh-stimulated cell volume reduction in acinar cells from hAQP1-expressing glands post IR were similar to those from control cells. Altogether, the data suggest that expression of hAQP1 increases the water permeability of acinar cells, which underlies the recovery of fluid secretion in the salivary glands

  17. Decreased expression of the mannose 6- phosphate/insulin-like growth factor-II receptor promotes growth of human breast cancer cells

    Directory of Open Access Journals (Sweden)

    Landman Natalie

    2002-07-01

    Full Text Available Abstract Background Loss or mutation of the mannose 6-phosphate/insulin-like growth factor-II receptor (M6P/IGF2R has been found in breast cancer. However, whether or not decreased levels of functional M6P/IGF2R directly contribute to the process of carcinogenesis needs to be further verified by functional studies. Methods In this study, using viral and ribozyme strategies we reduced the expression of M6P/IGF2R in human breast cancer cells and then examined the effect on growth and apoptosis of these cells. Results Our results showed that infection of MCF-7 cells with the adenovirus carrying a ribozyme targeted against the M6P/IGF2R mRNA dramatically reduced the level of transcripts and the functional activity of M6P/IGF2R in these cells. Accordingly, cells treated with the ribozyme exhibited a higher growth rate and a lower apoptotic index than control cells (infected with a control vector. Furthermore, decreased expression of M6P/IGF2R enhanced IGF-II-induced proliferation and reduced cell susceptibility to TNF-induced apoptosis. Conclusions These results suggest that M6P/IGF2R functions as a growth suppressor and its loss or mutation may contribute to development and progression of cancer. This study also demonstrates that adenoviral delivery of the ribozyme provides a useful tool for investigating the role of M6P/IGF2R in regulation of cell growth.

  18. Acidosis decreases c-Myc oncogene expression in human lymphoma cells: a role for the proton-sensing G protein-coupled receptor TDAG8.

    Science.gov (United States)

    Li, Zhigang; Dong, Lixue; Dean, Eric; Yang, Li V

    2013-01-01

    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. PMID:24152439

  19. 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.

  20. 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

  1. 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.

  2. 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.

  3. Baicalein induces G1 arrest in oral cancer cells by enhancing the degradation of cyclin D1 and activating AhR to decrease Rb phosphorylation

    International Nuclear Information System (INIS)

    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.

  4. Ectopic expression of clusterin/apolipoprotein J or Bcl-2 decreases the sensitivity of HaCaT cells to toxic effects of ropivacaine

    Institute of Scientific and Technical Information of China (English)

    2004-01-01

    Local anesthetics inhibit cell proliferation and induce apoptosis in various cell types. Ropivacaine, a unique, novel tertiary amine-type anesthetic, was shown to inhibit the proliferation of several cell types including keratinocytes. We found that Ropivacaine could inhibit the proliferation and induce apoptosis in an immortalized human keratinocyte line,HaCaT, in a dose- and time-dependent manner and with the deprivation of serum. The dose-dependent induction of apoptosis by ropivacaine was demonstrated by DNA fragmentation analysis and the proteolytic cleavage of a caspase-3substrate - poly (ADP-ribose) polymerase (PARP). In addition, ropivacaine downregulated the expression of clusterin/apoliporotein J, a protein with anti-apoptotic properties, in a dose-dependent manner, which well correlated with the induction of apoptosis of HaCaT cells. To investigate the role of clusterin/apoliporotein J in ropivacaine-induced apoptosis,HaCaT cells overexpressing clusterin/apoliporotein J were generated and compared to cells expressing the well established anti-apoptotic Bcl-2 protein. Ectopic overexpression of the secreted form of clusterin/apoliporotein J or Bcl-2decreased the sensitivity of HaCaT cells to toxic effects of ropivacaine as demonstrated by DNA fragmentation, the proteolytic cleavage of PARP and by a reduction in procaspase-3 expression. Furthermore, the downregulation of endogenous clusterin/apolipoprotein J levels by ropivacaine suggested that this might be one mechanism by which ropivacaine induced cell death in HaCaT cells. In conclusion, the ability of ropivacaine to induce antiproliferative responses and to suppress the expression of the anti-apoptotic protein clusterin/apolipoprotein J, combined with previously reported anti-inflammatory activity and analgesic property of the drug, suggests that ropivacaine may have potential utility in the local treatment of tumors.

  5. Stem Cells

    Science.gov (United States)

    Stem cells are cells with the potential to develop into many different types of cells in the body. ... the body. There are two main types of stem cells: embryonic stem cells and adult stem cells. Stem ...

  6. 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.

  7. Long none coding RNA HOTTIP/HOXA13 act as synergistic role by decreasing cell migration and proliferation in Hirschsprung disease

    International Nuclear Information System (INIS)

    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

  8. Age-related decrease in the responsiveness of rat articular chondrocytes to EGF is associated with diminished number and affinity for the ligand of cell surface binding sites.

    Science.gov (United States)

    Ribault, D; Habib, M; Abdel-Majid, K; Barbara, A; Mitrovic, D

    1998-01-12

    The effect of age on the responsiveness of articular chondrocytes (AC) to epidermal growth factor (EGF) was examined. Cells were isolated by digesting cartilage fragments from the humeral and femoral heads of 21-day old, 8- and 14-month old rats with collagenase. The cells were cultured under standard conditions, as monolayers. DNA synthesis was measured by [3H]thymidine incorporation and cell proliferation by the DNA content of subconfluent cultures. [125I]EGF binding and the amounts of EGF and EGF-receptor mRNAs were determined using confluent cells. DNA synthesis was decreased with age of animals. EGF stimulated DNA synthesis in cultures in 1- and 8-month old rats at low serum concentrations (< 5%), and in cultures in 14-month old animals at high serum concentrations. It also increased 5-day DNA content of cultures compared to serum-treated controls but this effect was weak in cultures in 14-month old rats. The number of high affinity binding sites for [125I]EGF decreased from 37,800 in the 1-month old to 1950 in the 14-month old rat AC. The apparent dissociation constant (Kd) also decreased with age: 0.18 nmol/l in the 1-month old; 0.12 nmol/l in the 8-month old; and 0.07 nmol/l in the 14-month old cells. AC in older rats contained more EGF mRNA and less EGF-receptor mRNA. Incubation of the cells with EGF resulted in down regulation of the EGF- and upregulation of EGF-receptor mRNA expressions. These findings show the age-related quantitative and qualitative alterations in EGF and EGF-receptor which may account, at least in part, for the diminished responsiveness of senescent AC to EGF. PMID:9509392

  9. 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.

  10. Ornithine and Homocitrulline Impair Mitochondrial Function, Decrease Antioxidant Defenses and Induce Cell Death in Menadione-Stressed Rat Cortical Astrocytes: Potential Mechanisms of Neurological Dysfunction in HHH Syndrome.

    Science.gov (United States)

    Zanatta, Ângela; Rodrigues, Marília Danyelle Nunes; Amaral, Alexandre Umpierrez; Souza, Débora Guerini; Quincozes-Santos, André; Wajner, Moacir

    2016-09-01

    Hyperornithinemia-hyperammonemia-homocitrullinuria (HHH) syndrome is caused by deficiency of ornithine translocase leading to predominant tissue accumulation and high urinary excretion of ornithine (Orn), homocitrulline (Hcit) and ammonia. Although affected patients commonly present neurological dysfunction manifested by cognitive deficit, spastic paraplegia, pyramidal and extrapyramidal signs, stroke-like episodes, hypotonia and ataxia, its pathogenesis is still poorly known. Although astrocytes are necessary for neuronal protection. Therefore, in the present study we investigated the effects of Orn and Hcit on cell viability (propidium iodide incorporation), mitochondrial function (thiazolyl blue tetrazolium bromide-MTT-reduction and mitochondrial membrane potential-ΔΨm), antioxidant defenses (GSH) and pro-inflammatory response (NFkB, IL-1β, IL-6 and TNF-α) in unstimulated and menadione-stressed cortical astrocytes that were previously shown to be susceptible to damage by neurotoxins. We first observed that Orn decreased MTT reduction, whereas both amino acids decreased GSH levels, without altering cell viability and the pro-inflammatory factors in unstimulated astrocytes. Furthermore, Orn and Hcit decreased cell viability and ΔΨm in menadione-treated astrocytes. The present data indicate that the major compounds accumulating in HHH syndrome impair mitochondrial function and reduce cell viability and the antioxidant defenses in cultured astrocytes especially when stressed by menadione. It is presumed that these mechanisms may be involved in the neuropathology of this disease. PMID:27161368

  11. The Over-expression of the β2 Catalytic Subunit of the Proteasome Decreases Homologous Recombination and Impairs DNA Double-Strand Break Repair in Human Cells

    Directory of Open Access Journals (Sweden)

    Anita Collavoli

    2011-01-01

    Full Text Available By a human cDNA library screening, we have previously identified two sequences coding two different catalytic subunits of the proteasome which increase homologous recombination (HR when overexpressed in the yeast Saccharomyces cerevisiae. Here, we investigated the effect of proteasome on spontaneous HR and DNA repair in human cells. To determine if the proteasome has a role in the occurrence of spontaneous HR in human cells, we overexpressed the β2 subunit of the proteasome in HeLa cells and determined the effect on intrachromosomal HR. Results showed that the overexpression of β2 subunit decreased HR in human cells without altering the cell proteasome activity and the Rad51p level. Moreover, exposure to MG132 that inhibits the proteasome activity reduced HR in human cells. We also found that the expression of the β2 subunit increases the sensitivity to the camptothecin that induces DNA double-strand break (DSB. This suggests that the β2 subunit has an active role in HR and DSB repair but does not alter the intracellular level of the Rad51p.

  12. 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)

    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

  13. 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

  14. 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...... thinner in ADOA than in healthy controls (CRAE (mean ± 2 standard deviations (SD)) 153.9 ± 41.0 μm and CRVE 236.1 ± 42.0 μm in ADOA, CRAE 172.5 ± 25.0 μm (p = 0.0004) and CRVE 254.2 ± 37.6 μm (p = 0.0019) in healthy controls). MABP was comparable in the two groups (p = 0.18), and in both groups, CRAE...... 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...

  15. The acquired radioresistance in HeLa cells under conditions mimicking hypoxia was attenuated by a decreased expression of HIF subunit genes induced by RNA interference

    International Nuclear Information System (INIS)

    The cancer cells residing in the hypoxic layer are resistant to radiation and these are ones responsible for cancer recurrence after radiation therapy. One of the reasons why hypoxic cancer cells acquire radioresistance may be attributable to changes in the gene expression profile by the activation of hypoxia inducible factors (HIFs). However, the details underlying this process remain unknown. In this study, we investigated the effects of knockdown of HIF subunit genes to elucidate how HIF subunit genes may be involved in the radioresistance acquired by HeLa cells following exposure to a hypoxia mimic. Interestingly, HIF-1α and HIF-2α seemed mutually complementary for each other when either of them was suppressed. We thus suppressed the expression of both genes simultaneously. To do this, we developed a short hairpin RNA (shRNA) targeting a high homology region between HIF-1α and HIF-2α. It was shown that the expression of the shRNA effectively suppressed the acquisition of radioresistance following the hypoxia mimic. Moreover, it was confirmed that suppression of both subunits resulted in the downregulation of stem cell markers and the suppression of spheroid formation during the hypoxia mimicking-conditions. This shRNA-mediated knockdown method targeting a common region shared by a family of genes may offer a new candidate cancer treatment. - Highlights: • Incubation with CoCl2 confers radioresistance to HeLa cells. • Both HIF-1α and HIF-2α are involved in the acquisition of radioresistance. • An shRNA to a homology region of HIF-1α and HIF-2α suppressed the radioresistance. • The shRNA decreased cells with stem cell markers and a stem cell phenotype

  16. The acquired radioresistance in HeLa cells under conditions mimicking hypoxia was attenuated by a decreased expression of HIF subunit genes induced by RNA interference

    Energy Technology Data Exchange (ETDEWEB)

    Doi, Nobutaka [Department of Radiological Sciences, Graduate School of Medicine and Pharmaceutical Sciences, University of Toyama, Toyama 930-0194 (Japan); New Products Research & Development, Gene Engineering Division, NIPPON GENE Co., Ltd. (Japan); Ogawa, Ryohei, E-mail: ogawa@med.u-toyama.ac.jp [Department of Radiological Sciences, Graduate School of Medicine and Pharmaceutical Sciences, University of Toyama, Toyama 930-0194 (Japan); Cui, Zheng-Guo [Department of Public Health, Graduate School of Medicine and Pharmaceutical Sciences, University of Toyama (Japan); Morii, Akihiro; Watanabe, Akihiko [Department of Urology, Graduate School of Medicine and Pharmaceutical Sciences, University of Toyama (Japan); Kanayama, Shinji; Yoneda, Yuko [New Products Research & Development, Gene Engineering Division, NIPPON GENE Co., Ltd. (Japan); Kondo, Takashi [Department of Radiological Sciences, Graduate School of Medicine and Pharmaceutical Sciences, University of Toyama, Toyama 930-0194 (Japan)

    2015-05-01

    The cancer cells residing in the hypoxic layer are resistant to radiation and these are ones responsible for cancer recurrence after radiation therapy. One of the reasons why hypoxic cancer cells acquire radioresistance may be attributable to changes in the gene expression profile by the activation of hypoxia inducible factors (HIFs). However, the details underlying this process remain unknown. In this study, we investigated the effects of knockdown of HIF subunit genes to elucidate how HIF subunit genes may be involved in the radioresistance acquired by HeLa cells following exposure to a hypoxia mimic. Interestingly, HIF-1α and HIF-2α seemed mutually complementary for each other when either of them was suppressed. We thus suppressed the expression of both genes simultaneously. To do this, we developed a short hairpin RNA (shRNA) targeting a high homology region between HIF-1α and HIF-2α. It was shown that the expression of the shRNA effectively suppressed the acquisition of radioresistance following the hypoxia mimic. Moreover, it was confirmed that suppression of both subunits resulted in the downregulation of stem cell markers and the suppression of spheroid formation during the hypoxia mimicking-conditions. This shRNA-mediated knockdown method targeting a common region shared by a family of genes may offer a new candidate cancer treatment. - Highlights: • Incubation with CoCl{sub 2} confers radioresistance to HeLa cells. • Both HIF-1α and HIF-2α are involved in the acquisition of radioresistance. • An shRNA to a homology region of HIF-1α and HIF-2α suppressed the radioresistance. • The shRNA decreased cells with stem cell markers and a stem cell phenotype.

  17. Selective decrease in cell surface expression and mRNA level of the 55-kDa tumor necrosis factor receptor during differentiation of HL-60 cells into macrophage-like but not granulocyte-like cells

    DEFF Research Database (Denmark)

    Winzen, R; Wallach, D; Engelmann, H;

    1992-01-01

    Expression of the two known receptors for TNF was studied in the promyelocytic leukemia cell line HL-60 before and after differentiation of the cells along the granulocyte lineage (induced by incubation with retinoic acid), or along the macrophage lineage (induced by incubation with the phorbol...

  18. Calpastatin overexpression reduces oxidative stress-induced mitochondrial impairment and cell death in human neuroblastoma SH-SY5Y cells by decreasing calpain and calcineurin activation, induction of mitochondrial fission and destruction of mitochondrial fusion.

    Science.gov (United States)

    Tangmansakulchai, Kulvadee; Abubakar, Zuroida; Kitiyanant, Narisorn; Suwanjang, Wilasinee; Leepiyasakulchai, Chaniya; Govitrapong, Piyarat; Chetsawang, Banthit

    2016-09-01

    Calpain is an intracellular Ca(2+)-dependent protease, and the activation of calpain has been implicated in neurodegenerative diseases. Calpain activity can be regulated by calpastatin, an endogenous specific calpain inhibitor. Several lines of evidence have demonstrated a potential role of calpastatin in preventing calpain-mediated pathogenesis. Additionally, several studies have revealed that calpain activation and mitochondrial damage are involved in the cell death process; however, recent evidence has not clearly indicated a neuroprotective mechanism of calpastatin against calpain-dependent mitochondrial impairment in the process of neuronal cell death. Therefore, the purpose of this study was to investigate the potential ability of calpastatin to inhibit calpain activation and mitochondrial impairment in oxidative stress-induced neuron degeneration. Calpastatin was stably overexpressed in human neuroblastoma SH-SY5Y cells. In non-calpastatin overexpressing SH-SY5Y cells, hydrogen peroxide significantly decreased cell viability, superoxide dismutase activity, mitochondrial membrane potential, ATP production and mitochondrial fusion protein (Opa1) levels in the mitochondrial fraction but increased reactive oxygen species formation, calpain and calcineurin activation, mitochondrial fission protein (Fis1 and Drp1) levels in the mitochondrial fraction and apoptotic cells. Nevertheless, these toxic effects were abolished in hydrogen peroxide-treated calpastatin-overexpressing SH-SY5Y cells. The results of the present study demonstrate the potential ability of calpastatin to diminish calpain and calcineurin activation and mitochondrial impairment in neurons that are affected by oxidative damage. PMID:27453331

  19. 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; Thorsen, Kasper; Fristrup, Niels; Brems-Eskildsen, Anne Sofie; Madsen, Pia Pinholt; Sørensen, Karina Dalsgaard; Borre, Michael; Ørntoft, Torben Falck

    2013-01-01

    cell extracts with an artificial “GAS”-DNA element resulted in an enrichment of the GAS containing DNA-sequence, providing evidence for a potential interaction of TOX3 with the GAS-sequence of STAT1. Conclusions These results provide evidence for an alternative activation of the downstream interferon......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 and...... identification and immunoprecipitation studies were used for DNA binding studies. Results Microarray transcript profiling of 89 bladder biopsies showed a significant upregulation of TOX3 (p< 10-4) in non-muscle invasive (Ta-T1) bladder tumors compared to muscle-invasive (T2-T4) bladder tumors and normal...

  20. Comparison of Monoamine Oxidase Inhibitors in Decreasing Production of the Autotoxic Dopamine Metabolite 3,4-Dihydroxyphenylacetaldehyde in PC12 Cells.

    Science.gov (United States)

    Goldstein, David S; Jinsmaa, Yunden; Sullivan, Patti; Holmes, Courtney; Kopin, Irwin J; Sharabi, Yehonatan

    2016-02-01

    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 Parkinson

  1. [Inhibition of NF-kB Activation Decreases Resistance in Acute Myeloid Leukemia Cells to TRAIL-induced Apoptosis in Multicellular Aggregates].

    Science.gov (United States)

    Fadeev, R S; Solovieva, M E; Slyadovskiy, D A; Zakharov, S G; Fadeeva, I S; Senotov, A S; Golenkov, A K; Akatov, V S

    2015-01-01

    Suppression of resistance in acute myeloid leukemia cells to TRAIL-induced apoptosis in multicellular aggregates, was studied using small molecule inhibitors of the activation of the transcription factor NF-kB - NF-k9 Activation Inhibitor IV and JSH-23 at non-toxic concentrations. NF-kB Activation Inhibitor IV and JSH-23 reduced resistance in the acute myeloid leukemia cells in multicellular aggregates to cytotoxic action of recombinant protein izTRAIL. It is shown that the use of these inhibitors decreased the phosphorylation of the RelA (p65) as a main marker activation of the transcription factor NF-kB. We discuss a possible reason for increasing resistance in acute myeloid leukemia cells to TRAIL-induced apoptosis in multicellular aggregates. PMID:26841509

  2. The novel curcumin analog FLLL32 decreases STAT3 DNA binding activity and expression, and induces apoptosis in osteosarcoma cell lines

    International Nuclear Information System (INIS)

    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

  3. 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.

  4. Deficiency of G1 regulators P53, P21Cip1 and/or pRb decreases hepatocyte sensitivity to TGFβ cell cycle arrest

    International Nuclear Information System (INIS)

    TGFβ is critical to control hepatocyte proliferation by inducing G1-growth arrest through multiple pathways leading to inhibition of E2F transcription activity. The retinoblastoma protein pRb is a key controller of E2F activity and G1/S transition which can be inhibited in viral hepatitis. It is not known whether the impairment of pRb would alter the growth inhibitory potential of TGFβ in disease. We asked how Rb-deficiency would affect responses to TGFβ-induced cell cycle arrest. Primary hepatocytes isolated from Rb-floxed mice were infected with an adenovirus expressing CRE-recombinase to delete the Rb gene. In control cells treatment with TGFβ prevented cells to enter S phase via decreased cMYC activity, activation of P16INK4A and P21Cip and reduction of E2F activity. In Rb-null hepatocytes, cMYC activity decreased slightly but P16INK4A was not activated and the great majority of cells continued cycling. Rb is therefore central to TGFβ-induced cell cycle arrest in hepatocytes. However some Rb-null hepatocytes remained sensitive to TGFβ-induced cell cycle arrest. As these hepatocytes expressed very high levels of P21Cip1 and P53 we investigated whether these proteins regulate pRb-independent signaling to cell cycle arrest by evaluating the consequences of disruption of p53 and p21Cip1. Hepatocytes deficient in p53 or p21Cip1 showed diminished growth inhibition by TGFβ. Double deficiency had a similar impact showing that in cells containing functional pRb; P21Cip and P53 work through the same pathway to regulate G1/S in response to TGFβ. In Rb-deficient cells however, p53 but not p21Cip deficiency had an additive effect highlighting a pRb-independent-P53-dependent effector pathway of inhibition of E2F activity. The present results show that otherwise genetically normal hepatocytes with disabled p53, p21Cip1 or Rb genes respond less well to the antiproliferative effects of TGFβ. As the function of these critical cellular proteins can be impaired by common

  5. Deficiency of G1 regulators P53, P21Cip1 and/or pRb decreases hepatocyte sensitivity to TGFβ cell cycle arrest

    Directory of Open Access Journals (Sweden)

    Harrison David J

    2007-11-01

    Full Text Available Abstract Background TGFβ is critical to control hepatocyte proliferation by inducing G1-growth arrest through multiple pathways leading to inhibition of E2F transcription activity. The retinoblastoma protein pRb is a key controller of E2F activity and G1/S transition which can be inhibited in viral hepatitis. It is not known whether the impairment of pRb would alter the growth inhibitory potential of TGFβ in disease. We asked how Rb-deficiency would affect responses to TGFβ-induced cell cycle arrest. Results Primary hepatocytes isolated from Rb-floxed mice were infected with an adenovirus expressing CRE-recombinase to delete the Rb gene. In control cells treatment with TGFβ prevented cells to enter S phase via decreased cMYC activity, activation of P16INK4A and P21Cip and reduction of E2F activity. In Rb-null hepatocytes, cMYC activity decreased slightly but P16INK4A was not activated and the great majority of cells continued cycling. Rb is therefore central to TGFβ-induced cell cycle arrest in hepatocytes. However some Rb-null hepatocytes remained sensitive to TGFβ-induced cell cycle arrest. As these hepatocytes expressed very high levels of P21Cip1 and P53 we investigated whether these proteins regulate pRb-independent signaling to cell cycle arrest by evaluating the consequences of disruption of p53 and p21Cip1. Hepatocytes deficient in p53 or p21Cip1 showed diminished growth inhibition by TGFβ. Double deficiency had a similar impact showing that in cells containing functional pRb; P21Cip and P53 work through the same pathway to regulate G1/S in response to TGFβ. In Rb-deficient cells however, p53 but not p21Cip deficiency had an additive effect highlighting a pRb-independent-P53-dependent effector pathway of inhibition of E2F activity. Conclusion The present results show that otherwise genetically normal hepatocytes with disabled p53, p21Cip1 or Rb genes respond less well to the antiproliferative effects of TGFβ. As the function of

  6. Interferon induced by UV-irradiated murine cytomegalovirus decreases type C virus expression in BALB/c cells treated by 5-iodo-deoxyuridine or cycloheximide

    International Nuclear Information System (INIS)

    UV-irradiated mouse cytomegalovirus (MCMV) activates at low incidence (2.0 X 10(-4) - 8.0 X 10(-4)) a xenotropic type C virus in Kirsten sarcoma virus-transformed nonproducer BALB 3T3 (KBALB) cells. When KBALB cells are treated with UV-MCMV and subsequently with potent chemical inducers, such as 5-Iododeoxyuridine (IdUrd) or cycloheximide, the activation frequencies of type C virus are significantly lower than in controls exposed to the chemical inducers alone: 60% for IdUrd and 44% for cycloheximide. This decrease of activation appears to be mediated by endogenous interferon (IFN) induced by UV-MCMV in KBALB cells, as could be proven by neutralization of the inhibitory activity. Indeed, after exposure of UV-MCMV-infected cells to anti-IFN gamma globulin, the level of C type virus increases, reaching values obtained with IdUrd or cycloheximide in the absence of UV-MCMV. The inhibitory activity associated with UV-MCMV is dose-dependent and the xenotropic type C virus appears to be more sensitive than the ecotropic (N-tropic) retrovirus of BALB/c cells

  7. An altered gp100 peptide ligand with decreased binding by TCR and CD8alpha dissects T cell cytotoxicity from production of cytokines and activation of NFAT

    Directory of Open Access Journals (Sweden)

    Niels eSchaft

    2013-09-01

    Full Text Available Altered peptide ligands (APLs provide useful tools to study T cell activation and potentially direct immune responses to improve treatment of cancer patients. To better understand and exploit APLs, we studied the relationship between APLs and T cell function in more detail. Here, we tested a broad panel of gp100(280-288 APLs with respect to T cell cytotoxicity, production of cytokines and activation of Nuclear Factor of Activated T cells (NFAT by human T cells gene-engineered with a gp100-HLA-A2-specific TCRalpha/beta. We demonstrated that gp100-specific cytotoxicity, production of cytokines, and activation of NFAT were not affected by APLs with single amino acid substitutions, except for an APL with an amino acid substitution at position 3 (APL A3, which did not elicit any T cell response. A gp100 peptide with a double amino acid mutation (APL S4S6 elicited T cell cytotoxicity and production of IFNgamma, and to a lesser extent TNFalpha, IL-4, and IL-5, but not production of IL-2 and IL-10, or activation of NFAT. Notably, TCR-mediated functions showed decreases in sensitivities for S4S6 versus gp100 wt peptide, which were minor for cytotoxicity but at least a 1000-fold more prominent for the production of cytokines. TCR-engineered T cells did not bind A3-HLA-A2, but did bind S4S6-HLA-A2 although to a lowered extent compared to wt peptide-HLA-A2. Moreover, S4S6-induced T cell function demonstrated an enhanced dependency on CD8alpha. Taken together, most gp100 APLs functioned as agonists, but A3 and S4S6 peptides acted as a null ligand and partial agonist, respectively. Our results further suggest that TCR-mediated cytotoxicity can be dissected from production of cytokines and activation of NFAT, and that the agonist potential of peptide mutants relates to the extent of binding by TCR and CD8alpha. These findings may facilitate the design of APLs to advance the study of T cell activation and their use for therapeutic applications.

  8. Leydig cell micronodules are a common finding in testicular biopsies from men with impaired spermatogenesis and are associated with decreased testosterone/LH ratio

    DEFF Research Database (Denmark)

    Holm, Mette; Rajpert-De Meyts, Ewa; Andersson, Anna-Maria; Skakkebæk, Niels Erik

    2003-01-01

    were examined using a semi-quantitative stereological method. In patients, serum concentrations of testosterone, sex hormone binding globulin (SHBG), luteinizing hormone (LH), follicle stimulating hormone (FSH), oestradiol and inhibin-B were correlated with the findings on histological examination...... hyperstimulated testes, as reflected by an increased LH/testosterone ratio. In conclusion, Leydig cell micronodules were more frequent in biopsies with impaired spermatogenesis and associated with decreased ratios of testicular hormones to gonadotrophins. The presence of micronodules thus seems to be a...

  9. Decreased glutathione content and glutathione S-transferase activity in red blood cells of coal miners with early stages of pneumoconiosis.

    OpenAIRE

    Evelo, C T; Bos, R P; Borm, P J

    1993-01-01

    Blood samples of miners heavily exposed to coal dust were examined for changes in glutathione S-transferase (GST) activity. Decreased GST activity was found in red blood cells of subjects with early stages of coal workers' pneumoconiosis (International Labour Office classification 0/1-1/2) when compared with control miners. At further progression of coal workers' pneumoconiosis (> or = 2/1), the activity of GST was not different from controls. In the same group with moderate coal workers' pne...

  10. Vitamin D binding protein-derived macrophage activating factor inhibits human breast cancer cell proliferation and decreases alpha-N-acetyl galactosaminidase in breast cancer patiens.

    OpenAIRE

    L. Thyer; G.Morucci; J.J.V. Branca; E. Wards; Smith, R; Noakes, D.

    2013-01-01

    Objectives: It was demonstrated that vitamin D binding protein-derived macrophage activating factor (GcMAF) decreases serum alpha-N-acetyl galactosaminidase (nagalase) in breast cancer patients; decrease of nagalase was associated with significant improvement of clinical conditions (Int J Cancer. 2008 Jan 15;122(2):461-7). Since nagalase is both an index of immune suppression and of tumour burden, we studied the direct effects of GcMAF on a human breast cancer cell line (MCF-7), and we com...

  11. Maple polyphenols, ginnalins A-C, induce S- and G2/M-cell cycle arrest in colon and breast cancer cells mediated by decreasing cyclins A and D1 levels.

    Science.gov (United States)

    González-Sarrías, Antonio; Ma, Hang; Edmonds, Maxwell E; Seeram, Navindra P

    2013-01-15

    Polyphenols are bioactive compounds found in plant foods. Ginnalins A-C are polyphenols present in the sap and other parts of the sugar and red maple species which are used to produce maple syrup. Here we evaluated the antiproliferative effects of ginnalins A-C on colon (HCT-116) and breast (MCF-7) tumourigenic and non-tumourigenic colon (CCD-18Co) cells and investigated whether these effects were mediated through cell cycle arrest and/or apoptosis. Ginnalins A-C were twofold more effective against the tumourigenic than non-tumourigenic cells. Among the polyphenols, ginnalin A (84%, HCT-116; 49%, MCF-7) was more effective than ginnalins B and C (50%, HCT-116; 30%, MCF-7) at 50 μM concentrations. Ginnalin A did not induce apoptosis of the cancer cells but arrested cell cycle (in the S- and G(2)/M-phases) and decreased cyclins A and D1 protein levels. These results suggest that maple polyphenols may have potential cancer chemopreventive effects mediated through cell cycle arrest. PMID:23122108

  12. Inhibition of protein kinase C decreases sensitivity of GABA receptor subtype to fipronil insecticide in insect neurosecretory cells.

    Science.gov (United States)

    Murillo, Laurence; Hamon, Alain; Es-Salah-Lamoureux, Zeineb; Itier, Valérie; Quinchard, Sophie; Lapied, Bruno

    2011-12-01

    Phosphorylation by serine/threonine kinases has been described as a new mechanism for regulating the effects of insecticides on insect neuronal receptors and channels. Although insect GABA receptors are commercially important targets for insecticides (e.g. fipronil), their modulation by kinases is poorly understood and the influence of phosphorylation on insecticide sensitivity is unknown. Using the whole-cell patch-clamp technique, we investigated the modulatory effect of PKC and CaMKinase II on GABA receptor subtypes (GABAR1 and GABAR2) in DUM neurons isolated from the terminal abdominal ganglion (TAG) of Periplaneta americana. Chloride currents through GABAR2 were selectively abolished by PMA and PDBu (the PKC activators) and potentiated by Gö6983, an inhibitor of PKC. Furthermore, using KN-62, a specific CaMKinase II inhibitor, we demonstrated that CaMKinase II activation was also involved in the regulation of GABAR2 function. In addition, using CdCl(2) (the calcium channel blocker) and LOE-908, a blocker of TRPγ, we revealed that calcium influx through TRPγ played an important role in kinase activations. Comparative studies performed with CACA, a selective agonist of GABAR1 in DUM neurons confirmed the involvement of these kinases in the specific regulation of GABAR2. Furthermore, our study reported that GABAR1 was less sensitive than GABAR2 to fipronil. This was demonstrated by the biphasic concentration-response curve and the current-voltage relationship established with both GABA and CACA. Finally, we demonstrated that GABAR2 was 10-fold less sensitive to fipronil following inhibition of PKC, whereas inhibition of CaMKinase II did not alter the effect of fipronil. PMID:21684305

  13. Membranous expression of Her3 is associated with a decreased survival in head and neck squamous cell carcinoma

    Directory of Open Access Journals (Sweden)

    Hong Seung-Mo

    2011-07-01

    Full Text Available Abstract Background Head and neck squamous cell carcinoma (HNSCC still remains a lethal malignancy benefiting from the identification of the new target for early detection and/or development of new therapeutic regimens based on a better understanding of the biological mechanism for treatment. The overexpression of Her2 and Her3 receptors have been identified in various solid tumors, but its prognostic relevance in HNSCC remains controversial. Methods Three hundred eighty-seven primary HNSCCs, 20 matching metasis and 17 recurrent HNSCCs were arrayed into tissue microarrays. The relationships between Her2 and Her3 protein expression and clinicopathological parameters/survival of HNSCC patients were analyzed with immunohistochemistry. Results Her3 is detected as either a cytoplasmic or a membranous dominant expression pattern whereas Her2 expression showed uniform membranous form. In primary tumor tissues, high membranous Her2 expression level was found in 104 (26.9% cases while positive membranous and cytoplasmic Her3 expression was observed in 34 (8.8% and 300 (77.5% samples, respectively. Membranous Her2 expression was significantly associated with histological grade (P = 0.021, as grade 2 tumors showed the highest positive expression. Membranous Her3 over-expression was significantly prevalent in metastatic tissues compared to primary tumors (P = 0.003. Survival analysis indicates that membranous Her3 expression is significantly associated with worse overall survival (P = 0.027 and is an independent prognostic factor in multivariate analysis (hazard ratio, 1.51; 95% confidence interval, 1.01-2.23; P = 0.040. Conclusions These results suggest that membranous Her3 expression is strongly associated with poor prognosis of patients with HNSCC and is a potential candidate molecule for targeted therapy.

  14. Magnetofection based on superparamagnetic iron oxide nanoparticle-mediated low lncRNA HOTAIR expression decreases the proliferation and invasion of glioma stem cells.

    Science.gov (United States)

    Fang, Kan; Liu, Peifeng; Dong, Suyan; Guo, Yanjie; Cui, Xinxin; Zhu, Xiaoying; Li, Xuan; Jiang, Lianghan; Liu, Te; Wu, Yuncheng

    2016-08-01

    Glioma stem cells (GSCs) are a special subpopulation of glioma cells that are key to the sensitivity of tumors to treatments and to the possibility of tumor recurrence. Identifying new strategies that inhibit the growth of GSCs are therefore important for developing novel therapies for glioblastoma multiforme (GBM). In this study, CD133+ human glioma stem cells were isolated and cultured. Magnetic nanoparticles were used to mediate the expression of siRNAs targeting the HOTAIR (si-HOTAIR) sequence in human gliomas. Effect of downregulation of HOTAIR expression on proliferation, invasion and in vivo tumorigenicity of human GSCs and underlying molecular mechanisms were further evaluated. The results of the MTT assay and flow cytometric analysis showed that downregulation of HOTAIR expression inhibited cell proliferation and induced cell cycle arrest. Transwell assays demonstrated that downregulation of HOTAIR expression resulted in a decrease in the invasive capability of GSCs. Moreover, magnetic nanoparticle-mediated low expression of HOTAIR effectively reduced the tumorigenic capacity of glioma stem cells in vivo. In addition, the results of qRT-PCR and western blot analysis demonstrated that downregulation of HOTAIR expression significantly increased the expression of PDCD4 in GSCs, in addition to reducing the expression of CCND1 and CDK4. An in-depth mechanistic analysis showed that downregulation of HOTAIR expression reduced the recruitment of downstream molecules, EZH2 and LSD1, thereby activating the expression of PDCD4 at the transcriptional level. In conclusion, downregulation of HOTAIR expression effectively promoted the expression of PDCD4, thereby inhibiting the proliferation, invasion and in vivo tumorigenicity of human GSCs. PMID:27277755

  15. Major Components of Energy Drinks (Caffeine, Taurine, and Guarana Exert Cytotoxic Effects on Human Neuronal SH-SY5Y Cells by Decreasing Reactive Oxygen Species Production

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    Fares Zeidán-Chuliá

    2013-01-01

    Full Text Available Scope. 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. Methods and Results. 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. Conclusion. 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.

  16. Evidence for a calcium-mobilizing receptor triggered by cadmium, or decreasing external pH or sodium in coronary endothelial cells

    International Nuclear Information System (INIS)

    Cadmium (1 μM) or decreasing external pH (pHo) or Na concentration (Nao) transiently increased cytoplasmic free Ca (Cai) by ∼5-fold as measured with fura-2 in endothelial cells grown on cover glasses. Cd or pHo=6 evoked similar peak Cai increases in the presence or absence of external Ca indicating that they trigger the release of stored Ca. Cd or 4 mM Nao, but not pHo 6, caused sustained Cai increases. The sustained phase of the Cai response to Cd required external Ca. Decreasing intracellular pH with propionate had no effect on Cai. Cd and some other divalent metals also stimulated 45Ca efflux. The potency order of the metals that stimulated efflux was Cd > Co > Ni > Fe(II) > Mn. Cd apparently acts at an extracellular site because loading the cells with a heavy metal chelator (TPEN) only slightly affected Cd- or ATP-stimulated 45Ca efflux. Addition of Zn or Cu(II) to the assay solution prevented the stimulation of efflux by Cd and had no effect on ATP-evoked efflux. Decreasing Nao stimulated 45Ca efflux ∼4-fold and slightly increased 45Ca influx. Decreasing Nao to 27 mM or pHo to 6.4 half-maximally increased 45Ca efflux. The stimulation of efflux was independent of external Ca or the osmotic Na substitute. The release of stored Ca, not Ca influx via Na-Ca exchange, causes the increase in Cai and 45Ca efflux evoked by decreasing Nao. Incubating the cells with Zn prior to assaying efflux in the absence of Zn strongly inhibited the stimulation of 45Ca efflux by Cd, pHo 6, and the Nao removal. Desensitization by Zn had no effect on ATP-stimulated efflux. These finding support the hypothesis proposed previously that the trace metals or decreasing external Na or pH release stored Ca by stimulating the same cell surface receptor

  17. Non-thermal plasma inhibits human cervical cancer HeLa cells invasiveness by suppressing the MAPK pathway and decreasing matrix metalloproteinase-9 expression

    Science.gov (United States)

    Li, Wei; Yu, K. N.; Bao, Lingzhi; Shen, Jie; Cheng, Cheng; Han, Wei

    2016-01-01

    Non-thermal plasma (NTP) has been proposed as a novel therapeutic method for anticancer treatment. However, the mechanism underlying its biological effects remains unclear. In this study, we investigated the inhibitory effect of NTP on the invasion of HeLa cells, and explored the possible mechanism. Our results showed that NTP exposure for 20 or 40 s significantly suppressed the migration and invasion of HeLa cells on the basis of matrigel invasion assay and wound healing assay, respectively. Moreover, NTP reduced the activity and protein expression of the matrix metalloproteinase (MMP)-9 enzyme. Western blot analysis indicated that NTP exposure effectively decreased phosphorylation level of both ERK1/2 and JNK, but not p38 MAPK. Furthermore, treatment with MAPK signal pathway inhibitors or NTP all exhibited significant depression of HeLa cells migration and MMP-9 expression. The result showed that NTP synergistically suppressed migration and MMP-9 expression in the presence of ERK1/2 inhibitor and JNK inhibitor, but not p38 MAPK inhibitor. Taken together, these findings suggested that NTP exposure inhibited the migration and invasion of HeLa cells via down-regulating MMP-9 expression in ERK1/2 and JNK signaling pathways dependent manner. These findings provide hints to the potential clinical research and therapy of NTP on cervical cancer metastasis.

  18. Tunable Biodegradable Nanocomposite Hydrogel for Improved Cisplatin Efficacy on HCT-116 Colorectal Cancer Cells and Decreased Toxicity in Rats.

    Science.gov (United States)

    Abdel-Bar, Hend Mohamed; Osman, Rihab; Abdel-Reheem, Amal Youssef; Mortada, Nahed; Awad, Gehanne A S

    2016-02-01

    This work describes the development of a modified nanocomposite thermosensitive hydrogel for controlled cisplatin release and improved cytotoxicity with decreased side effects. The system was characterized in terms of physical properties, morphological architecture and in vitro cisplatin release. Cytotoxicity was tested against human colorectal carcinoma HCT-116. In vivo studies were conducted to evaluate the acute toxicity in terms of rats' survival rate and body weight loss. Nephro and hepatotoxicities were evaluated followed by histopathological alterations of various tissue organs. Nanocomposite thermosensitive hydrogel containing nanosized carrier conferred density and stiffness allowing a zero order drug release for 14 days. Enhanced cytotoxicity with 2-fold decrease in cisplatin IC50 was accomplished. A linear in vivo-in vitro correlation was proved for the system degradation. Higher animal survival rate and lower tissue toxicities proved the decreased toxicity of cisplatin nanocomposite compared to its solution. PMID:26709447

  19. Paullinia cupana Mart. var. sorbilis, guarana, increases survival of Ehrlich ascites carcinoma (EAC) bearing mice by decreasing cyclin-D1 expression and inducing a G0/G1 cell cycle arrest in EAC cells.

    Science.gov (United States)

    Fukumasu, Heidge; Latorre, Andreia Oliveira; Zaidan-Dagli, Maria Lucia

    2011-01-01

    The objective of this work is to report the antiproliferative effect of P. cupana treatment in Ehrlich Ascites Carcinoma (EAC)-bearing animals. Female mice were treated with three doses of powdered P. cupana (100, 1000 and 2000 mg/kg) for 7 days, injected with 10(5) EAC cells and treated up to day 21. In addition, a survival experiment was carried out with the same protocol. P. cupana decreased the ascites volume (p = 0.0120), cell number (p = 0.0004) and hemorrhage (p = 0.0054). This occurred through a G1-phase arrest (p < 0.01) induced by a decreased gene expression of Cyclin D1 in EAC cells. Furthermore, P. cupana significantly increased the survival of EAC-bearing animals (p = 0.0012). In conclusion, the P. cupana growth control effect in this model was correlated with a decreased expression of cyclin D1 and a G1 phase arrest. These results reinforce the cancer therapeutic potential of this Brazilian plant. PMID:20564499

  20. HOXA9 is Underexpressed in Cervical Cancer Cells and its Restoration Decreases Proliferation, Migration and Expression of Epithelial-to-Mesenchymal Transition Genes.

    Science.gov (United States)

    Alvarado-Ruiz, Liliana; Martinez-Silva, Maria Guadalupe; Torres-Reyes, Luis Alberto; Pina-Sanchez, Patricia; Ortiz-Lazareno, Pablo; Bravo-Cuellar, Alejandro; Aguilar-Lemarroy, Adriana; Jave-Suarez, Luis Felipe

    2016-01-01

    HOX transcription factors are evolutionarily conserved in many different species and are involved in important cellular processes such as morphogenesis, differentiation, and proliferation. They have also recently been implicated in carcinogenesis, but their precise role in cancer, especially in cervical cancer (CC), remains unclear. In this work, using microarray assays followed by the quantitative polymerase chain reaction (qPCR), we found that the expression of 25 HOX genes was downregulated in CC derived cell lines compared with nontumorigenic keratinocytes. In particular, the expression of HOXA9 was observed as down-modulated in CCderived cell lines. The expression of HOXA9 has not been previously reported in CC, or in normal keratinocytes of the cervix. We found that normal CC from women without cervical lesions express HOXA9; in contrast, CC cell lines and samples of biopsies from women with CC showed significantly diminished HOXA9 expression. Furthermore, we found that methylation at the first exon of HOXA9 could play an important role in modulating the expression of this gene. Exogenous restoration of HOXA9 expression in CC cell lines decreased cell proliferation and migration, and induced an epithelial-like phenotype. Interestingly, the silencing of human papilloma virus (HPV) E6 and E7 oncogenes induced expression of HOXA9. In conclusion, controlling HOXA9 expression appears to be a necessary step during CC development. Further studies are needed to delineate the role of HOXA9 during malignant progression and to afford more insights into the relationship between downmodulation of HOXA9 and viral HPV oncoprotein expression during cercical cancer development. PMID:27039722

  1. Cobalt chloride decreases fibroblast growth factor-21 expression dependent on oxidative stress but not hypoxia-inducible factor in Caco-2 cells

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Yanlong [School of Pharmacy, Wenzhou Medical College, Wenzhou (China); Department of Medicine, University of Louisville, Louisville, KY (United States); Wang, Chunhong [Second Hospital, Jilin University, Changchun (China); Department of Medicine, University of Louisville, Louisville, KY (United States); Wang, Yuhua [College of Food Science and Engineering, Jilin Agricultural University, Changchun (China); Department of Medicine, University of Louisville, Louisville, KY (United States); Ma, Zhenhua [First Hospital, Xi' an Jiaotong University, Xi' an (China); Department of Medicine, University of Louisville, Louisville, KY (United States); Xiao, Jian [School of Pharmacy, Wenzhou Medical College, Wenzhou (China); McClain, Craig [Department of Medicine, University of Louisville, Louisville, KY (United States); Department of Pharmacology and Toxicology, University of Louisville, Louisville, KY (United States); Alcohol Research Center, University of Louisville, Louisville, KY (United States); Robley Rex Veterans Affairs Medical Center, Louisville, KY (United States); Li, Xiaokun [School of Pharmacy, Wenzhou Medical College, Wenzhou (China); Feng, Wenke, E-mail: wenke.feng@louisville.edu [School of Pharmacy, Wenzhou Medical College, Wenzhou (China); Department of Medicine, University of Louisville, Louisville, KY (United States); Alcohol Research Center, University of Louisville, Louisville, KY (United States)

    2012-10-15

    Fibroblast growth factor-21 (FGF21) is a potential metabolic regulator with multiple beneficial effects on metabolic diseases. FGF21 is mainly expressed in the liver, but is also found in other tissues including the intestine, which expresses β-klotho abundantly. The intestine is a unique organ that operates in a physiologically hypoxic environment, and is responsible for the fat absorption processes including triglyceride breakdown, re-synthesis and absorption into the portal circulation. In the present study, we investigated the effects of hypoxia and the chemical hypoxia inducer, cobalt chloride (CoCl{sub 2}), on FGF21 expression in Caco-2 cells and the consequence of fat accumulation. Physical hypoxia (1% oxygen) and CoCl{sub 2} treatment decreased both FGF21 mRNA and secreted protein levels. Gene silence and inhibition of hypoxia-inducible factor-α (HIFα) did not affect the reduction of FGF21 mRNA and protein levels by hypoxia. However, CoCl{sub 2} administration caused a significant increase in oxidative stress. The addition of n-acetylcysteine (NAC) suppressed CoCl{sub 2}-induced reactive oxygen species (ROS) formation and completely negated CoCl{sub 2}-induced FGF21 loss. mRNA stability analysis demonstrated that the CoCl{sub 2} administration caused a remarkable reduction in FGF21 mRNA stability. Furthermore, CoCl{sub 2} increased intracellular triglyceride (TG) accumulation, along with a reduction in mRNA levels of lipid lipase, hormone sensitive lipase (HSL) and adipose triglyceride lipase (ATGL), and an increase of sterol regulatory element-binding protein-1c (SREBP1c) and stearoyl-coenzyme A (SCD1). Addition of both NAC and recombinant FGF21 significantly attenuated the CoCl{sub 2}-induced TG accumulation. In conclusion, the decrease of FGF21 in Caco-2 cells by chemical hypoxia is independent of HIFα, but dependent on an oxidative stress-mediated mechanism. The regulation of FGF21 by hypoxia may contribute to intestinal lipid metabolism and

  2. Cobalt chloride decreases fibroblast growth factor-21 expression dependent on oxidative stress but not hypoxia-inducible factor in Caco-2 cells

    International Nuclear Information System (INIS)

    Fibroblast growth factor-21 (FGF21) is a potential metabolic regulator with multiple beneficial effects on metabolic diseases. FGF21 is mainly expressed in the liver, but is also found in other tissues including the intestine, which expresses β-klotho abundantly. The intestine is a unique organ that operates in a physiologically hypoxic environment, and is responsible for the fat absorption processes including triglyceride breakdown, re-synthesis and absorption into the portal circulation. In the present study, we investigated the effects of hypoxia and the chemical hypoxia inducer, cobalt chloride (CoCl2), on FGF21 expression in Caco-2 cells and the consequence of fat accumulation. Physical hypoxia (1% oxygen) and CoCl2 treatment decreased both FGF21 mRNA and secreted protein levels. Gene silence and inhibition of hypoxia-inducible factor-α (HIFα) did not affect the reduction of FGF21 mRNA and protein levels by hypoxia. However, CoCl2 administration caused a significant increase in oxidative stress. The addition of n-acetylcysteine (NAC) suppressed CoCl2-induced reactive oxygen species (ROS) formation and completely negated CoCl2-induced FGF21 loss. mRNA stability analysis demonstrated that the CoCl2 administration caused a remarkable reduction in FGF21 mRNA stability. Furthermore, CoCl2 increased intracellular triglyceride (TG) accumulation, along with a reduction in mRNA levels of lipid lipase, hormone sensitive lipase (HSL) and adipose triglyceride lipase (ATGL), and an increase of sterol regulatory element-binding protein-1c (SREBP1c) and stearoyl-coenzyme A (SCD1). Addition of both NAC and recombinant FGF21 significantly attenuated the CoCl2-induced TG accumulation. In conclusion, the decrease of FGF21 in Caco-2 cells by chemical hypoxia is independent of HIFα, but dependent on an oxidative stress-mediated mechanism. The regulation of FGF21 by hypoxia may contribute to intestinal lipid metabolism and absorption. -- Graphical abstract: Physical and

  3. Epoxyeicosatrienoic acid agonist regulates human mesenchymal stem cell-derived adipocytes through activation of HO-1-pAKT signaling and a decrease in PPARγ.

    Science.gov (United States)

    Kim, Dong Hyun; Vanella, Luca; Inoue, Kazuyoshi; Burgess, Angela; Gotlinger, Katherine; Manthati, Vijaya Lingam; Koduru, Sreenivasulu Reddy; Zeldin, Darryl C; Falck, John R; Schwartzman, Michal L; Abraham, Nader G

    2010-12-01

    Human mesenchymal stem cells (MSCs) expressed substantial levels of CYP2J2, a major CYP450 involved in epoxyeicosatrienoic acid (EET) formation. MSCs synthesized significant levels of EETs (65.8 ± 5.8 pg/mg protein) and dihydroxyeicosatrienoic acids (DHETs) (15.83 ± 1.62 pg/mg protein), suggesting the presence of soluble epoxide hydrolase (sEH). The addition of an sEH inhibitor to MSC culture decreased adipogenesis. EETs decreased MSC-derived adipocytes in a concentration-dependent manner, 8,9- and 14,15-EET having the maximum reductive effect on adipogenesis. We examined the effect of 12-(3-hexylureido)dodec-8(Z)-enoic acid, an EET agonist, on MSC-derived adipocytes and demonstrated an increased number of healthy small adipocytes, attenuated fatty acid synthase (FAS) levels (P SnMP) and LY2940002, respectively, reversed EET-induced inhibition of adipogenesis, suggesting that activation of the HO-1-adiponectin axis underlies EET effect in MSCs. These findings indicate that EETs decrease MSC-derived adipocyte stem cell differentiation by upregulation of HO-1-adiponectin-AKT signaling and play essential roles in the regulation of adipocyte differentiation by inhibiting PPARγ, C/EBPα, and FAS and in stem cell development. These novel observations highlight the seminal role of arachidonic acid metabolism in MSCs and suggest that an EET agonist may have potential therapeutic use in the treatment of dyslipidemia, diabetes, and the metabolic syndrome. PMID:20412023

  4. Decreased expression of BRCA1 in SK-BR-3 cells is the result of aberrant activation of the GABP Beta promoter by an NRF-1-containing complex

    Directory of Open Access Journals (Sweden)

    MacDonald Gwen

    2011-05-01

    Full Text Available Abstract Background BRCA1 has recently been identified as a potential regulator of mammary stem/progenitor cell differentiation, and this function may explain the high prevalence of breast cancer in BRCA1 mutation carriers, as well as the downregulation of BRCA1 in a large proportion of sporadic breast cancers. That is, loss of BRCA1 function results in blocked differentiation with expansion of the mammary stem/progenitor cells. Because BRCA1 also maintains genomic integrity, its loss could produce a pool of genetically unstable stem/progenitor cells that are prime targets for further transforming events. Thus, elucidating the regulatory mechanisms of BRCA1 expression is important to our understanding of normal and malignant breast differentiation. Results Loss of BRCA1 expression in the ErbB2-amplified SK-BR-3 cell line was found to be the result of loss of activity of the ets transcription factor GABP, a previously characterized regulator of BRCA1 transcription. The expression of the non-DNA binding GABPβ subunit was shown to be deficient, while the DNA binding subunit, GABPα was rendered unstable by the absence of GABPβ. Deletion analysis of the GABPβ proximal promoter identified a potential NRF-1 binding site as being critical for expression. Supershift analysis, the binding of recombinant protein and chromatin immunoprecipitation confirmed the role of NRF-1 in regulating the expression of GABPβ. The siRNA knockdown of NRF-1 resulted in decreased GABPβ and BRCA1 expression in MCF-7 cells indicating that they form a transcriptional network. NRF-1 levels and activity did not differ between SK-BR-3 and MCF-7 cells, however the NRF-1 containing complex on the GABPβ promoter differed between the two lines and appears to be the result of altered coactivator binding. Conclusions Both NRF-1 and GABP have been linked to the regulation of nuclear-encoded mitochondrial proteins, and the results of this study suggest their expression is

  5. Plasma miR-185 is decreased in patients with esophageal squamous cell carcinoma and might suppress tumor migration and invasion by targeting RAGE.

    Science.gov (United States)

    Jing, Rongrong; Chen, Wen; Wang, Huimin; Ju, Shaoqing; Cong, Hui; Sun, Baolan; Jin, Qin; Chu, Shaopeng; Xu, Lili; Cui, Ming

    2015-11-01

    The receptor for advanced-glycation end products (RAGE) is upregulated in various cancers and has been associated with tumor progression, but little is known about its expression and regulation by microRNAs (miRNAs) in esophageal squamous cell carcinoma (ESCC). Here, we describe miR-185, which represses RAGE expression, and investigate the biological role of miR-185 in ESCC. In this study, we found that the high level of RAGE expression in 29 pairs of paraffin-embedded ESCC tissues was correlated positively with the depth of invasion by immunohistochemistry, suggesting that RAGE was involved in ESCC. We used bioinformatics searches and luciferase reporter assays to investigate the prediction that RAGE was regulated directly by miR-185. Besides, overexpression of miR-185 in ESCC cells was accompanied by 27% (TE-11) and 49% (Eca-109) reduced RAGE expression. The effect was further confirmed in RAGE protein by immunofluorescence in both cell lines. The effects were reversed following cotransfection with miR-185 and high-level expression of the RAGE vector. Furthermore, the biological role of miR-185 in ESCC cell lines was investigated using assays of cell viability, Ki-67 staining, and cell migration and invasion, as well as in a xenograft model. We found that overexpression of miR-185 inhibited migration and invasion by ESCC cells in vitro and reduced their capacity to develop distal pulmonary metastases in vivo partly through the RAGE/heat shock protein 27 pathway. Interestingly, in clinical specimens, the level of plasma miR-185 expression was decreased significantly (P = 0.002) in patients with ESCC [0.500; 95% confidence interval (CI) 0.248-1.676] compared with healthy controls (2.410; 95% CI 0.612-5.671). The value of the area under the receiver-operating characteristic curve was 0.73 (95% CI 0.604-0.855). In conclusion, our findings shed novel light on the role of miR-185/RAGE in ESCC metastasis, and plasma miR-185 has potential as a novel diagnostic biomarker

  6. Ethanol, ethyl and sodium pyruvate decrease the inflammatory responses of human lung epithelial cells via Akt and NF-κB in vitro but have a low impact on hepatocellular cells.

    Science.gov (United States)

    Relja, B; Omid, N; Wagner, N; Mörs, K; Werner, I; Juengel, E; Perl, M; Marzi, I

    2016-02-01

    Increases in pro-inflammatory cytokine levels and tissue-infiltrating leukocytes have been closely linked to increased systemic and local inflammation, which result in organ injury. Previously, we demonstrated the beneficial and hepatoprotective anti-inflammatory effects of acute ethanol (EtOH) ingestion in an in vivo model of acute inflammation. Due to its undesirable side-effects, however, EtOH does not represent a therapeutic option for treatment of acute inflammation. Therefore, in this study, we compared the effects of acute EtOH exposure with ethyl pyruvate (EtP) as an alternative anti-inflammatory drug in an in vitro model of hepatic and pulmonary inflammation. Human hepatocellular carcinoma cells Huh7 and alveolar epithelial cells A549 were stimulated with either interleukin (IL) IL-1β (1 ng/ml, 24 h) or tumor necrosis factor (TNF) (10 ng/ml, 4 h), and then treated with EtP (2.5-10 mM), sodium pyruvate (NaP, 10 mM) or EtOH (85-170 mM) for 1 h. IL-6 or IL-8 release from Huh7 or A549 cells, respectively, was measured by an enzyme‑linked immunosorbent assay. Neutrophil adhesion to cell monolayers and CD54 expression were also analyzed. Bcl-2 and Bax gene expression was determined by RT-qPCR, and western blot analysis was performed to determine the mechanisms involved. Treating A549 cells with either EtOH or EtP significantly reduced the IL-1β- or TNF‑induced IL-8 release, whereas treating Huh7 cells did not significantly alter IL-6 release. Similarly, neutrophil adhesion to stimulated A549 cells was significantly reduced by EtOH or EtP, whereas for Huh7 cells the tendency for reduced neutrophil adhesion rates by EtOH or EtP was not significant. CD54 expression was noticeably reduced in A549 cells, but this was not the case in Huh7 cells after treatment. The Bax/Bcl-2 ratio was dose‑dependently decreased by EtOH and by high-dose EtP in A549 cells, indicating a reduction in apoptosis, whereas this effect was not observed in Huh7 cells. The underlying

  7. Inflammatory disease protective R381Q IL23 receptor polymorphism results in decreased primary CD4+ and CD8+ human T-cell functional responses

    OpenAIRE

    Sarin, Ritu; Wu, Xingxin; Abraham, Clara

    2011-01-01

    The SNP (c.1142G > A;p.R381Q) in the IL-23 receptor (IL23R) confers protection from multiple inflammatory diseases, representing one of the most significant human genetic polymorphisms in autoimmunity. We, therefore, sought to define the functional consequences of this clinically significant variant. We find that CD4+CD45RO+ and CD8+ T cells from healthy R381Q IL23R carriers show decreased IL-23–dependent IL-17 and IL-22 production relative to WT IL23R individuals. This was associated with a ...

  8. Survival of cancer stem cells under hypoxia and serum depletion via decrease in PP2A activity and activation of p38-MAPKAPK2-Hsp27.

    Directory of Open Access Journals (Sweden)

    Shih-Pei Lin

    Full Text Available Hypoxia and serum depletion are common features of solid tumors that occur upon antiangiogenesis, irradiation and chemotherapy across a wide variety of malignancies. Here we show that tumor cells expressing CD133, a marker for colorectal cancer initiating or stem cells, are enriched and survive under hypoxia and serum depletion conditions, whereas CD133- cells undergo apoptosis. CD133+ tumor cells increase cancer stem cell and epithelial-mesenchymal transition properties. Moreover, via screening a panel of tyrosine and serine/threonine kinase pathways, we identified Hsp27 is constitutively activated in CD133+ cells rather than CD133- cell under hypoxia and serum depletion conditions. However, there was no difference in Hsp27 activation between CD133+ and CD133- cells under normal growth condition. Hsp27 activation, which was mediated by the p38MAPK-MAPKAPK2-Hsp27 pathway, is required for CD133+ cells to inhibit caspase 9 and 3 cleavage. In addition, inhibition of Hsp27 signaling sensitizes CD133+ cells to hypoxia and serum depletion -induced apoptosis. Moreover, the antiapoptotic pathway is also activated in spheroid culture-enriched CD133+ cancer stem cells from a variety of solid tumor cells including lung, brain and oral cancer, suggesting it is a common pathway activated in cancer stem cells from multiple tumor types. Thus, activation of PP2A or inactivation of the p38MAPK-MAPKAPK2-Hsp27 pathway may develop new strategies for cancer therapy by suppression of their TIC population.

  9. Rapid dimerization of quercetin through an oxidative mechanism in the presence of serum albumin decreases its ability to induce cytotoxicity in MDA-MB-231 cells

    Energy Technology Data Exchange (ETDEWEB)

    Pham, Anh; Bortolazzo, Anthony [Department of Biological Sciences, San Jose State University, San Jose, CA 95192-0100 (United States); White, J. Brandon, E-mail: Brandon.White@sjsu.edu [Department of Biological Sciences, San Jose State University, San Jose, CA 95192-0100 (United States)

    2012-10-19

    Highlights: Black-Right-Pointing-Pointer Quercetin cannot be detected intracellularly despite killing MDA-MB-231 cells. Black-Right-Pointing-Pointer Quercetin forms a heterodimer through oxidation in media with serum. Black-Right-Pointing-Pointer The quercetin heterodimer does not kill MDA-MB-231 cells. Black-Right-Pointing-Pointer Ascorbic acid stabilizes quercetin increasing cell death in quercetin treated cells. Black-Right-Pointing-Pointer Quercetin, and not a modified form, is responsible for apoptosis and cell death. -- Abstract: Quercetin is a member of the flavonoid family and has been previously shown to have a variety of anti-cancer activities. We and others have reported anti-proliferation, cell cycle arrest, and induction of apoptosis of cancer cells after treatment with quercetin. Quercetin has also been shown to undergo oxidation. However, it is unclear if quercetin or one of its oxidized forms is responsible for cell death. Here we report that quercetin rapidly oxidized in cell culture media to form a dimer. The quercetin dimer is identical to a dimer that is naturally produced by onions. The quercetin dimer and quercetin-3-O-glucopyranoside are unable to cross the cell membrane and do not kill MDA-MB-231 cells. Finally, supplementing the media with ascorbic acid increases quercetin's ability to induce cell death probably by reduction oxidative dimerization. Our results suggest that an unmodified quercetin is the compound that elicits cell death.

  10. Rapid dimerization of quercetin through an oxidative mechanism in the presence of serum albumin decreases its ability to induce cytotoxicity in MDA-MB-231 cells.

    Science.gov (United States)

    Pham, Anh; Bortolazzo, Anthony; White, J Brandon

    2012-10-19

    Quercetin is a member of the flavonoid family and has been previously shown to have a variety of anti-cancer activities. We and others have reported anti-proliferation, cell cycle arrest, and induction of apoptosis of cancer cells after treatment with quercetin. Quercetin has also been shown to undergo oxidation. However, it is unclear if quercetin or one of its oxidized forms is responsible for cell death. Here we report that quercetin rapidly oxidized in cell culture media to form a dimer. The quercetin dimer is identical to a dimer that is naturally produced by onions. The quercetin dimer and quercetin-3-O-glucopyranoside are unable to cross the cell membrane and do not kill MDA-MB-231 cells. Finally, supplementing the media with ascorbic acid increases quercetin's ability to induce cell death probably by reduction oxidative dimerization. Our results suggest that an unmodified quercetin is the compound that elicits cell death. PMID:23000408

  11. Decreased LRIG1 in Human Ovarian Cancer Cell SKOV3 Upregulates MRP-1 and Contributes to the Chemoresistance of VP16.

    Science.gov (United States)

    Yang, Hua; Yao, Jun; Yin, Jiangpin; Wei, Xuan

    2016-05-01

    The leucine-rich repeats and immunoglobulin-like domains (LRIG) are used as tumor suppressors in clinical applications. Although the LRIG has been identified to manipulate the cell proliferation via various oncogenic receptor tyrosine kinases in diverse cancers, its role in multidrug resistance needs to be further elucidated, especially in human ovarian cancer. We herein established that the etoposide (VP16)-resistant SKOV3 human ovarian cancer cell clones (SKOV3/VP16 cells) and mRNA expression of LRIG1 were significantly reduced by the treatment of VP16 in a concentration-dependent manner. Moreover, downregulated LRIG1 in SKOV3 could enhance the colony formation and resist the inhibition of proliferation by VP16, leading to the elevated expression of Bcl-2 and decreased apoptosis of SKOV3. Interestingly, our results uncovered that the multidrug resistance-associated protein 1 (MRP-1) was upregulated for the chemoresistance of VP16. To overcome the chemoresistance of SKOV3, SKOV3/VP16 was ectopically expressed of LRIG1. We found that the inhibition of VP16 on colony formation and proliferation was remarkably enhanced with increased apoptosis in SKOV3/VP16. Furthermore, the expression of MRP-1 and Bcl-2 was also inhibited, suggesting that the LRIG1could negatively control MRP-1 and the apoptosis to improve the sensitivity of VP16-related chemotherapy. PMID:27183435

  12. Decreased pre-surgical CD34+/CD144+ cell number in patients undergoing coronary artery bypass grafting compared to coronary artery disease-free valvular patients

    Directory of Open Access Journals (Sweden)

    Redondo Santiago

    2012-01-01

    Full Text Available Abstract Background Cardiovascular disease has been linked to endothelial progenitor cell (EPC depletion and functional impairment in atherosclerosis and aortic stenosis. EPCs may play a pivotal role in vascular grafting. However, the EPC depletion in coronary artery bypass grafting (CABG patients has not been compared to coronary artery disease-free valvular replacement patients with aortic stenosis. Methods We aimed to assess the basal number of CD34+/KDR+ and CD34+/CD144+ cells in CABG patients, compared to aortic stenosis valvular replacement patients. 100 patients (51 CABG and 49 valvular surgery ones were included in the present study. All CABG or valvular patients had angiographic demonstration of the presence or the absence of coronary artery disease, respectively. Numbers of CD34+/KDR+ and CD34+/CD144+ were assessed by flow cytometry of pre-surgical blood samples. Results We found a lower number of CD34+/CD144+ cells in CABG patients compared to valvular patients (0.21 ± 0.03% vs. 0.47 ± 0.08%, and this difference remained statistically significant after the P was adjusted for multiple comparisons (P = 0.01428. Both groups had more EPCs than healthy controls. Conclusions Pre-surgical CD34+/CD144+ numbers are decreased in CABG patients, compared to valvular patients with absence of coronary disease.

  13. Genetic Inactivation of ATRX Leads to a Decrease in the Amount of Telomeric Cohesin and Level of Telomere Transcription in Human Glioma Cells.

    Science.gov (United States)

    Eid, Rita; Demattei, Marie-Véronique; Episkopou, Harikleia; Augé-Gouillou, Corinne; Decottignies, Anabelle; Grandin, Nathalie; Charbonneau, Michel

    2015-08-01

    Mutations in ATRX (alpha thalassemia/mental retardation syndrome X-linked), a chromatin-remodeling protein, are associated with the telomerase-independent ALT (alternative lengthening of telomeres) pathway of telomere maintenance in several types of cancer, including human gliomas. In telomerase-positive glioma cells, we found by immunofluorescence that ATRX localized not far from the chromosome ends but not exactly at the telomere termini. Chromatin immunoprecipitation (ChIP) experiments confirmed a subtelomeric localization for ATRX, yet short hairpin RNA (shRNA)-mediated genetic inactivation of ATRX failed to trigger the ALT pathway. Cohesin has been recently shown to be part of telomeric chromatin. Here, using ChIP, we showed that genetic inactivation of ATRX provoked diminution in the amount of cohesin in subtelomeric regions of telomerase-positive glioma cells. Inactivation of ATRX also led to diminution in the amount of TERRAs, noncoding RNAs resulting from transcription of telomeric DNA, as well as to a decrease in RNA polymerase II (RNAP II) levels at the telomeres. Our data suggest that ATRX might establish functional interactions with cohesin on telomeric chromatin in order to control TERRA levels and that one or the other or both of these events might be relevant to the triggering of the ALT pathway in cancer cells that exhibit genetic inactivation of ATRX. PMID:26055325

  14. Loss of phosphoinositide 3-kinase γ decreases migration and activation of phagocytes but not T cell activation in antigen-induced arthritis

    Directory of Open Access Journals (Sweden)

    Wetzker Reinhard

    2010-04-01

    Full Text Available Abstract Background Phosphoinositide 3-kinase γ (PI3Kγ has been depicted as a major regulator of inflammatory processes, including leukocyte activation and migration towards several chemokines. This study aims to explore the role of PI3Kγ in the murine model of antigen-induced arthritis (AIA. Methods Development of AIA was investigated in wildtype and PI3Kγ-deficient mice as well as in mice treated with a specific inhibitor of PI3Kγ (AS-605240 in comparison to untreated animals. Inflammatory reactions of leukocytes, including macrophage and T cell activation, and macrophage migration, were studied in vivo and in vitro. Results Genetic deletion or pharmacological inhibition of PI3Kγ induced a marked decrease of clinical symptoms in early AIA, together with a considerably diminished macrophage migration and activation (lower production of NO, IL-1β, IL-6. Also, macrophage and neutrophil infiltration into the knee joint were impaired in vivo. However, T cell functions, measured by cytokine production (TNFα, IFNγ, IL-2, IL-4, IL-5, IL-17 in vitro and DTH reaction in vivo were not altered, and accordingly, disease developed normally at later timepoints Conclusion PI3Kγ specifically affects phagocyte function in the AIA model but has no impact on T cell activation.

  15. Overexpression of stress-inducible OsBURP16, the β subunit of polygalacturonase 1, decreases pectin content and cell adhesion and increases abiotic stress sensitivity in rice.

    Science.gov (United States)

    Liu, Huanhuan; Ma, Yan; Chen, Na; Guo, Siyi; Liu, Huili; Guo, Xiaoyu; Chong, Kang; Xu, Yunyuan

    2014-05-01

    Polygalacturonase (PG), one of the hydrolases responsible for cell wall pectin degradation, is involved in organ consenescence and biotic stress in plants. PG1 is composed of a catalytic subunit, PG2, and a non-catalytic PG1β subunit. OsBURP16 belongs to the PG1β-like subfamily of BURP-family genes and encodes one putative PG1β subunit precursor in rice (Oryza sativa L.). Transcription of OsBURP16 is induced by cold, salinity and drought stresses, as well as by abscisic acid (ABA) treatment. Analysis of plant survival rates, relative ion leakage rates, accumulation levels of H2 O2 and water loss rates of leaves showed that overexpression of OsBURP16 enhanced sensitivity to cold, salinity and drought stresses compared with controls. Young leaves of Ubi::OsBURP16 transgenic plants showed reduced cell adhesion and increased cuticular transpiration rate. Mechanical strength measurement of Ubi::OsBURP16 plants showed that reduced force was required to break leaves as compared with wild type. Transgenic rice showed enhanced PG activity and reduced pectin content. All these results suggested that overexpression of OsBURP16 caused pectin degradation and affected cell wall integrity as well as transpiration rate, which decreased tolerance to abiotic stresses. PMID:24237159

  16. COX-2 inhibition improves immunotherapy and is associated with decreased numbers of myeloid-derived suppressor cells in mesothelioma. Celecoxib influences MDSC function

    OpenAIRE

    Veltman, Joris; Lambers, Margaretha E. H.; Nimwegen, Menno; Hendriks, Rudi; Hoogsteden, Henk; Aerts, Joachim; Hegmans, Joost

    2010-01-01

    textabstractBackground: Myeloid-derived suppressor cells (MDSC) are a heterogeneous population of immature cells that accumulates in tumour-bearing hosts. These cells are induced by tumour-derived factors (e.g. prostaglandins) and have a critical role in immune suppression. MDSC suppress T and NK cell function via increased expression of arginase I and production of reactive oxygen species (ROS) and nitric oxide (NO). Immune suppression by MDSC was found to be one of the main factors for immu...

  17. COX-2 inhibition improves immunotherapy and is associated with decreased numbers of myeloid-derived suppressor cells in mesothelioma. Celecoxib influences MDSC function

    OpenAIRE

    Veltman Joris D; Lambers Margaretha EH; van Nimwegen Menno; Hendriks Rudi W; Hoogsteden Henk C; Aerts Joachim GJV; Hegmans Joost PJJ

    2010-01-01

    Abstract Background Myeloid-derived suppressor cells (MDSC) are a heterogeneous population of immature cells that accumulates in tumour-bearing hosts. These cells are induced by tumour-derived factors (e.g. prostaglandins) and have a critical role in immune suppression. MDSC suppress T and NK cell function via increased expression of arginase I and production of reactive oxygen species (ROS) and nitric oxide (NO). Immune suppression by MDSC was found to be one of the main factors for immunoth...

  18. Attempts to predict the long-term decrease in lung function due to radiotherapy of non-small cell lung cancer

    International Nuclear Information System (INIS)

    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 R2 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 (R2 =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

  19. L655,240, acting as a competitive BACE1 inhibitor,efficiently decreases β-amyloid peptide production in HEK293-APPswe cells

    Institute of Scientific and Technical Information of China (English)

    Qin LU; Wu-yan CHEN; Zhi-yuan ZHU; Jing CHEN; Ye-chun XU; Morakot KAEWPET; Vatcharin RUKACHAISIRIKUL; Li-li CHEN; Xu SHEN

    2012-01-01

    Aim: To identify a small molecule L655,240 as a novel β-secretase (BACE1) inhibitor and to investigate its effects on β-amyloid (Aβ)generation in vitro.Methods: Fluorescence resonance energy transfer (FRET) was used to characterize the inhibitory effect of L655,240 on BACE1.Surface plasmon resonance (SPR) technology-based assay was performed to study the binding affinity of L655,240 for BACE1.The selectivity of L655,240 toward BACE1 over other aspartic proteases was determined with enzymatic assay.The effects of L655,240 on Aβ40,Aβ42,and sAPPβ production were studied in HEK293 cells stably expressing APP695 Swedish mutantK595N/M596L (HEK293-APPswe cells).The activities of BACE1,ν-secretase and α-secretase were assayed,and both the mRNA and protein levels of APP and BACE1 were evaluated using real-time PCR (RT-PCR) and Western blot analysis.Results: L655,240 was determined to be a competitive,selective BACE1 inhibitor (IC50=4.47±1.37 μmol/L),which bound to BACE1 directly (KD=17.9±0.72 μmol/L).L655,240 effectively reduced Aβ40,Aβ42,and sAPPβ production by inhibiting BACE1 without affecting the activities of y-secretase and α-secretase in HEK293-APPswe cells.L655,240 has no effect on APP and BACE1 mRNA or protein levels in HEK293-APPswe cells.Conclusion: The small molecule L655,240 is a novel BACE1 inhibitor that can effectively decreases Aβ production in vitro,thereby highlighting its therapeutic potential for the treatment of Alzheimer's disease.

  20. Involvement of Ca2+ in Vacuole Degradation Caused by a Rapid Temperature Decrease in Saintpaulia Palisade Cells: A Case of Gene Expression Analysis in a Specialized Small Tissue.

    Science.gov (United States)

    Ohnishi, Miwa; Kadohama, Noriaki; Suzuki, Yoshihiro; Kajiyama, Tomoharu; Shichijo, Chizuko; Ishizaki, Kimitsune; Fukaki, Hidehiro; Iida, Hidetoshi; Kambara, Hideki; Mimura, Tetsuro

    2015-07-01

    Saintpaulia (African violet) leaves are known to be damaged by a rapid temperature decrease when cold water is applied to the leaf surface; the injury is ascribed to the chloroplast damage caused by the cytosolic pH decrease following the degradation of the vacuolar membrane in the palisade cells. In this report, we present evidence for the involvement of Ca(2+) in facilitating the collapse of the vacuolar membrane and in turn in the temperature sensitivity of Saintpaulia leaves. In the presence of a Ca(2+) chelator (EGTA) or certain Ca(2+) channel inhibitors (Gd(3+) or La(3+)) but not others (verapamil or nifedipine), the pH of the vacuole, monitored through BCECF (2',7'-bis(carboxyethyl)-4 or 5-carboxyfluorescein) fluorescence, did not increase in response to a rapid temperature drop. These pharmacological observations are consistent with the involvement of mechanosensitive Ca(2+) channels in the collapse of the vacuolar membrane. The high level of expression of an MCA- (Arabidopsis mechanosensitive Ca(2+) channel) like gene, a likely candidate for a mechanosensitive Ca(2+) channel(s) in plant cells, was confirmed in the palisade tissue in Saintpaulia leaves by using a newly developed method of gene expression analysis for the specialized small tissues. PMID:25941231

  1. Carnosine decreased neuronal cell death through targeting glutamate system and astrocyte mitochondrial bioenergetics in cultured neuron/astrocyte exposed to OGD/recovery.

    Science.gov (United States)

    Ouyang, Li; Tian, Yueyang; Bao, Yun; Xu, Huijuan; Cheng, Jiaoyan; Wang, Bingyu; Shen, Yao; Chen, Zhong; Lyu, Jianxin

    2016-06-01

    Previously, we showed that carnosine upregulated the expression level of glutamate transporter 1 (GLT-1), which has been recognized as an important participant in the astrocyte-neuron lactate shuttle (ANLS), with ischemic model in vitro and in vivo. This study was designed to investigate the protective effect of carnosine on neuron/astrocyte co-cultures exposed to OGD/recovery, and to explore whether the ANLS or any other mechanism contributes to carnosine-induced neuroprotection on neuron/astrocyte. Co-cultures were treated with carnosine and exposed to OGD/recovery. Cell death and the extracellular levels of glutamate and GABA were measured. The mitochondrial respiration and glycolysis were detected by Seahorse Bioscience XF96 Extracellular Flux Analyzer. Results showed that carnosine decreased neuronal cell death, increased extracellular GABA level, and abolished the increase in extracellular glutamate and reversed the mitochondrial energy metabolism disorder induced by OGD/recovery. Carnosine also upregulated the mRNA level of neuronal glutamate transporter EAAC1 at 2h after OGD. Dihydrokainate, a specific inhibitor of GLT-1, decreased glycolysis but it did not affect mitochondrial respiration of the cells, and it could not reverse the increase in mitochondrial OXPHOS induced by carnosine in the co-cultures. The levels of mRNAs for monocarboxylate transporter1, 4 (MCT1, 4), which were expressed in astrocytes, and MCT2, the main neuronal MCT, were significantly increased at the early stage of recovery. Carnosine only partly reversed the increased expression of astrocytic MCT1 and MCT4. These results suggest that regulating astrocytic energy metabolism and extracellular glutamate and GABA levels but not the ANLS are involved in the carnosine-induced neuroprotection. PMID:27040711

  2. EPS-SJ Exopolisaccharide Produced by the Strain Lactobacillus paracasei subsp. paracasei BGSJ2-8 Is Involved in Adhesion to Epithelial Intestinal Cells and Decrease on E. coli Association to Caco-2 Cells

    Science.gov (United States)

    Živković, Milica; Miljković, Marija S.; Ruas-Madiedo, Patricia; Markelić, Milica B.; Veljović, Katarina; Tolinački, Maja; Soković, Svetlana; Korać, Aleksandra; Golić, Nataša

    2016-01-01

    The aim of this study was to determine the role of an exopolysaccharide produced by natural dairy isolate Lactobacillus paracasei subsp. paracasei BGSJ2-8, in the adhesion to intestinal epithelial cells and a decrease in Escherichia coli’s association with Caco-2 cells. Annotation of the BGSJ2-8 genome showed the presence of a gene cluster, epsSJ, which encodes the biosynthesis of the strain-specific exopolysaccharide EPS-SJ, detected as two fractions (P1 and P2) by size exclusion chromatography (SEC) coupled with multi-angle laser light scattering (MALLS) detection. SEC-MALLS analysis revealed that an EPS-SJ- mutant (EPS7, obtained by insertion mutagenesis of the glps_2198 gene encoding primary glycosyltransferase) does not produce the P2 fraction of EPS-SJ. Transmission electron microscopy showed that EPS7 mutant has a thinner cell wall compared to the EPS-SJ+ strain BGSJ2-83 (a plasmid free-derivative of BGSJ2-8). Interestingly, strain BGSJ2-83 showed higher adhesion to Caco-2 epithelial intestinal cell line than the EPS7 mutant. Accordingly, BGSJ2-83 effectively reduced E. coli ATCC25922’s association with Caco-2 cells, while EPS7 did not show statistically significant differences. In addition, the effect of EPS-SJ on the proliferation of lymphocytes in gastrointestinal associated lymphoid tissue (GALT) was tested and the results showed that the reduction of GALT lymphocyte proliferation was higher by BGSJ2-83 than by the mutant. To the best of our knowledge this is the first report indicating that the presence of EPS (EPS-SJ) on the surface of lactobacilli can improve communication between bacteria and intestinal epithelium, implying its possible role in gut colonization. PMID:27014210

  3. Nogo-B regulates migration and contraction of airway smooth muscle cells by decreasing ARPC 2/3 and increasing MYL-9 expression

    Directory of Open Access Journals (Sweden)

    Cai Zailong

    2011-01-01

    Full Text Available Abstract Background Abnormal proliferation, apoptosis, migration and contraction of airway smooth muscle (ASM cells in airway remodeling in asthma are basically excessive repair responses to a network of inflammatory mediators such as PDGF, but the mechanisms of such responses remain unclear. Nogo-B, a member of the reticulum family 4(RTN4, is known to play a key role in arteriogenesis and tissue repair. Further studies are needed to elucidate the role of Nogo-B in airway smooth muscle abnormalities. Methods A mouse model of chronic asthma was established by repeated OVA inhalation and subjected to Nogo-B expression analysis using immunohistochemistry and Western Blotting. Then, primary human bronchial smooth muscle cells (HBSMCs were cultured in vitro and a siRNA interference was performed to knockdown the expression of Nogo-B in the cells. The effects of Nogo-B inhibition on PDGF-induced HBSMCs proliferation, migration and contraction were evaluated. Finally, a proteomic analysis was conducted to unveil the underlying mechanisms responsible for the function of Nogo-B. Results Total Nogo-B expression was approximately 3.08-fold lower in chronic asthmatic mice compared to naïve mice, which was obvious in the smooth muscle layer of the airways. Interference of Nogo-B expression by siRNA resulted nearly 96% reduction in mRNA in cultured HBSMCs. In addition, knockdown of Nogo-B using specific siRNA significantly decreased PDGF-induced migration of HBSMCs by 2.3-fold, and increased the cellular contraction by 16% compared to negative controls, but had limited effects on PDGF-induced proliferation. Furthermore, using proteomic analysis, we demonstrate that the expression of actin related protein 2/3 complex subunit 5 (ARPC 2/3 decreased and, myosin regulatory light chain 9 isoform a (MYL-9 increased after Nogo-B knockdown. Conclusions These data define a novel role for Nogo-B in airway remodeling in chronic asthma. Endogenous Nogo-B, which may exert

  4. Decreased expression levels of PIWIL1, PIWIL2, and PIWIL4 are associated with worse survival in renal cell carcinoma patients

    Directory of Open Access Journals (Sweden)

    Iliev R

    2016-01-01

    Full Text Available Robert Iliev,1,2 Michal Stanik,3 Michal Fedorko,4 Alexandr Poprach,1 Petra Vychytilova-Faltejskova,1,2 Katerina Slaba,2 Marek Svoboda,1 Pavel Fabian,5 Dalibor Pacik,1 Jan Dolezel,2 Ondrej Slaby3,4 1Department of Comprehensive Cancer Care, Masaryk Memorial Cancer Institute, 2Central European Institute of Technology, Masaryk University, 3Department of Urologic Oncology, Masaryk Memorial Cancer Institute, 4Department of Urology, University Hospital Brno, Masaryk University Brno, 5Department of Diagnostic and Experimental Pathology, Masaryk Memorial Cancer Institute, Brno, Czech RepublicAbstract: Piwi-interacting RNAs (piRNAs are a newly discovered class of small non-coding RNAs involved in silencing of transposable elements and in sequence-specific chromatin modifications. PIWI proteins (PIWIL belong to the family of Argonaute genes/proteins, bind piRNAs and their functioning have been described mainly in germ-line cells and more recently also in stem cells and cancer cells. There are four PIWI proteins PIWIL1, PIWIL2, PIWIL3, and PIWIL4 discovered in human till now. Recent studies suggested that deregulated the expression of Piwi proteins and selected piRNAs is common to many types of cancers. We found significantly lower expression of PIWIL1 (P<0.0001 and piR-823 (P=0.0001 in tumor tissue in comparison to paired renal parenchyma. Further, we observed progressive decrease in PIWIL1 (P=0.0228, PIWIL2 (P=0.0015, and PIWIL4 (P=0.0028 expression levels together with increasing clinical stage. PIWIL2 (P=0.0073 and PIWIL4 (P=0.0001 expression progressively decreased also with increasing Fuhrman grade. Most importantly, low-expression levels of PIWIL1 (P=0.009, PIWIL2 (P<0.0001, and PIWIL4 (P=0.0065 were significantly associated with worse overall survival in renal cell carcinoma (RCC patients. Our results suggest the involvement of PIWIL genes and piR-823 in RCC pathogenesis, and indicate PIWIL1, PIWIL2, and PIWIL4 as potential prognostic biomarkers in

  5. Isolation of cell lines with decreased or deficient nitrate-reductase activity from cell-suspension culture of mono- (2n=x=12) and dihaploid (2n=2x=24) Solanum tuberosum plants

    International Nuclear Information System (INIS)

    In order to obtain in vitro cell lines resistant to high chlorate concentrations, suspension-incubated cells of mono- (2n = x = 12) and dihaploid (2n = 2x = 24) Solanum tuberosum plants were exposed to the action of gamma-rays (500 r/min) doses of 4 and 5 kR. These doses had an inhibiting effect on the plating efficiency and cell survival in the control medium. An increase in the plating efficiency of cells radiated in the selection medium may indicate induced mutagenesis consisting in the loss of nitrate reductase activity in these cells. Ten chlorate-resistant cell lines, selected from radiated cells, successfully developed for 3 years on the selection medium with a high chlorate concentration and when pasaged on the control medium for over a year, have not restored the properties of wild type cells, thus becoming ''stable variants''

  6. Deficiency of Kruppel-like factor KLF4 in myeloid-derived suppressor cells inhibits tumor pulmonary metastasis in mice accompanied by decreased fibrocytes.

    Science.gov (United States)

    Shi, Y; Ou, L; Han, S; Li, M; Pena, M M O; Pena, E A; Liu, C; Nagarkatti, M; Fan, D; Ai, W

    2014-01-01

    The importance of immunosuppressive myeloid-derived suppressor cells (MDSCs) bearing monocyte markers in tumor metastasis has been well established. Recently, it was reported that these cells possess phenotypic plasticity and differentiate into fibrocytes, very distinct cells that are precursors of tumorigenic myofibroblasts. However, the importance of this transdifferentiation in tumor metastasis has not been explored. Here, we describe the role of MDSC-derived fibrocytes in tumor metastasis that is regulated by Kruppel-like factor 4 (KLF4), a transcription factor that is critical to monocyte differentiation and to promotion of cancer development. Using mouse metastasis models of melanoma and breast cancer, we found that KLF4 knockout was associated with significantly reduced pulmonary metastasis, which was accompanied by decreased populations of MDSCs, fibrocytes and myofibroblasts in the lung. Cause-effect studies by adoptive transfer revealed that KLF4 deficiency in MDSCs led to significantly reduced lung metastasis that was associated with fewer MDSC-derived fibrocytes and myofibroblasts. Mechanistically, KLF4 deficiency significantly compromised the generation of fibrocytes from MDSCs in vitro. During this process, KLF4 expression levels were tightly linked with those of fibroblast-specific protein-1 (FSP-1), deficiency of which resulted in no metastasis in mice as has been previously reported. In addition, KLF4 bound directly to the FSP-1 promoter as determined by chromatin immunoprecipitation and overexpression of KLF4 increased the FSP-1 promoter activities. Taken together, our results suggest that MDSCs not only execute their immunosuppressive function to promote metastatic seeding as reported before, but also boost metastatic tumor growth after they adopt a fibrocyte fate. Therefore, KLF4-mediated fibrocyte generation from MDSCs may represent a novel mechanism of MDSCs contributing to tumor metastasis and supports the feasibility of inhibiting KLF4 or

  7. The fibrate decreases radiation sensitivity via peroxisome proliferator-activated receptor {alpha}-mediated superoxide dismutase induction in HeLa cells

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Xianguang; An, Zhengzhe; Song, Hye Jin; Kim, Won Dong; Park, Woo Yoon [Chungbuk National University College of Medicine, Cheongju (Korea, Republic of); Jang, Seong Soon [The Catholic University of Korea College of Medicine, Seoul (Korea, Republic of); Yu, Jae Ran [Konkuk University College of Medicine, Chungju (Korea, Republic of)

    2012-06-15

    The fibrates are ligands for peroxisome proliferator-activated receptor (PPAR) {alpha} and used clinically as hypolipidemic drugs. The fibrates are known to cause peroxisome proliferation, enhance superoxide dismutase (SOD) expression and catalase activity. The antioxidant actions of the fibrates may modify radiation sensitivity. Here, we investigated the change of the radiation sensitivity in two cervix cancer cell lines in combination with fenofi brate (FF). Activity and protein expression of SOD were measured according to the concentration of FF. The mRNA expressions were measured by using real time reverse-transcription polymerase chain reaction. Combined cytotoxic effect of FF and radiation was measured by using clonogenic assay. In HeLa cells total SOD activity was increased with increasing FF doses up to 30 {mu}M. In the other hand, the catalase activity was increased a little. As with activity the protein expression of SOD1 and SOD2 was increased with increasing doses of FF. The mRNAs of SOD1, SOD2, PPAR{alpha} and PPAR{gamma} were increased with increasing doses of FF. The reactive oxygen species (ROS) produced by radiation was decreased by preincubation with FF. The surviving fractions (SF) by combining FF and radiation was higher than those of radiation alone. In Me180 cells SOD and catalase activity were not increased with FF. Also, the mRNAs of SOD1, SOD2, and PPAR{alpha} were not increased with FF. However, the mRNA of PPAR{gamma} was increased with FF. FF can reduce radiation sensitivity by ROS scavenging via SOD induction in HeLa. SOD induction by FF is related with PPAR{alpha}.

  8. Hesperetin and its sulfate and glucuronide metabolites inhibit TNF-α induced human aortic endothelial cell migration and decrease plasminogen activator inhibitor-1 (PAI-1) levels.

    Science.gov (United States)

    Giménez-Bastida, Juan Antonio; González-Sarrías, Antonio; Vallejo, Fernando; Espín, Juan Carlos; Tomás-Barberán, Francisco A

    2016-01-01

    Epidemiological, clinical and preclinical studies have reported the protection offered by citrus consumption, mainly orange, against cardiovascular diseases, which is primarily mediated by the antiatherogenic and vasculoprotective effects of the flavanone hesperetin-7-O-rutinoside (hesperidin). However, flavanone aglycones or glycosides are not present in the bloodstream but their derived phase-II metabolites could be the actual bioactive molecules. To date, only a few studies have explored the effects of circulating hesperetin-derived metabolites (glucuronides and sulfates) on endothelial cells. Herein, we describe for the first time the effects of hesperetin 3'-O-glucuronide, hesperetin 7-O-glucuronide, hesperetin 3'-O-sulfate, hesperetin 7-O-sulfate and hesperetin on human aortic endothelial cell (HAEC) migration upon pro-inflammatory stimuli as an essential step to angiogenesis. Hesperetin and its derived metabolites, at physiologically relevant concentrations (1-10 μM), significantly attenuated cell migration in the presence of the pro-inflammatory cytokine TNF-α (50 ng mL(-1)), which was accompanied and perhaps mediated by a significant decrease in the levels of the thrombogenic plasminogen activator inhibitor-1 (PAI-1). However, hesperetin metabolites did not counteract the TNF-α-induced production of pro-inflammatory interleukin-6 (IL-6) and IL-8. We also study here for the first time, the metabolism of hesperetin and its derived metabolites by HAEC with and without a pro-inflammatory stimulus. All these results reinforce the concept according to which circulating phase-II hesperetin metabolites are critical molecules contributing to the cardioprotective effects upon consumption of citrus fruits such as orange. PMID:26456097

  9. Acetylated H4K16 by MYST1 protects UROtsa cells from arsenic toxicity and is decreased following chronic arsenic exposure

    International Nuclear Information System (INIS)

    Arsenic, a human carcinogen that is associated with an increased risk of bladder cancer, is commonly found in drinking water. An important mechanism by which arsenic is thought to be carcinogenic is through the induction of epigenetic changes that lead to aberrant gene expression. Previously, we reported that the SAS2 gene is required for optimal growth of yeast in the presence of arsenite (AsIII). Yeast Sas2p is orthologous to human MYST1, a histone 4 lysine 16 (H4K16) acetyltransferase. Here, we show that H4K16 acetylation is necessary for the resistance of yeast to AsIII through the modulation of chromatin state. We further explored the role of MYST1 and H4K16 acetylation in arsenic toxicity and carcinogenesis in human bladder epithelial cells. The expression of MYST1 was knocked down in UROtsa cells, a model of bladder epithelium that has been used to study arsenic-induced carcinogenesis. Silencing of MYST1 reduced acetylation of H4K16 and induced sensitivity to AsIII and to its more toxic metabolite monomethylarsonous acid (MMAIII) at doses relevant to high environmental human exposures. In addition, both AsIII and MMAIII treatments decreased global H4K16 acetylation levels in a dose- and time-dependent manner. This indicates that acetylated H4K16 is required for resistance to arsenic and that a reduction in its levels as a consequence of arsenic exposure may contribute to toxicity in UROtsa cells. Based on these findings, we propose a novel role for the MYST1 gene in human sensitivity to arsenic.

  10. Secondhand smoke in combination with ambient air pollution exposure is associated with increasedx CpG methylation and decreased expression of IFN-γ in T effector cells and Foxp3 in T regulatory cells in children

    Directory of Open Access Journals (Sweden)

    Kohli Arunima

    2012-09-01

    Full Text Available Abstract Background Secondhand smoke (SHS and ambient air pollution (AAP exposures have been associated with increased prevalence and severity of asthma and DNA modifications of immune cells. In the current study, we examined the association between SHS and AAP with DNA methylation and expression of interferon-gamma (IFN-γ and forkhead box protein 3 (Foxp3 in T cell populations. Methods Subjects 7–18 years old were recruited from Fresno (high AAP; n = 62 and Stanford, CA (low AAP; n = 40 and divided into SHS-exposed (Fresno: n = 31, Stanford: n = 6 and non-SHS-exposed (nSHS; Fresno: n = 31, Stanford: n = 34 groups. T cells purified from peripheral blood were assessed for levels of DNA methylation and expression of IFN-γ (in effector T cells or Foxp3 (in regulatory T cells. Results Analysis showed a significant increase in mean % CpG methylation of IFN-γ and Foxp3 associated with SHS exposure (IFN-γ: FSHS 62.10%, FnSHS 41.29%, p p Foxp3: FSHS 74.60%, FnSHS 54.44%, p p IFN-γ: FSHS 0.75, FnSHS 1.52, p p Foxp3: FSHS 0.75, FnSHS 3.29, p p IFN-γ: FSHS vs. SSHS, p p Foxp3: FSHS vs. SSHS, p p IFN-γ: FSHS vs. SSHS, p p Foxp3: FSHS vs. SSHS, p p IFN-γ: p = 0.15; Foxp3: p = 0.27, nor was Foxp3 expression (p = 0.08; IFN-γ expression was significantly decreased in AAP-only subjects (p  Conclusions Exposures to SHS and AAP are associated with significant hypermethylation and decreased expression of IFN-γ in Teffs and Foxp3 in Tregs. Relative contributions of each exposure to DNA modification and asthma pathogenesis warrant further investigation.

  11. MALDI Mass Spectrometry Imaging Reveals Decreased CK5 Levels in Vulvar Squamous Cell Carcinomas Compared to the Precursor Lesion Differentiated Vulvar Intraepithelial Neoplasia.

    Science.gov (United States)

    Zhang, Chao; Arentz, Georgia; Winderbaum, Lyron; Lokman, Noor A; Klingler-Hoffmann, Manuela; Mittal, Parul; Carter, Christopher; Oehler, Martin K; Hoffmann, Peter

    2016-01-01

    Vulvar cancer is the fourth most common gynecological cancer worldwide. However, limited studies have been completed on the molecular characterization of vulvar squamous cell carcinoma resulting in a poor understanding of the disease initiation and progression. Analysis and early detection of the precursor lesion of HPV-independent vulvar squamous cell carcinoma (VSCC), differentiated vulvar intraepithelial neoplasia (dVIN), is of great importance given dVIN lesions have a high level of malignant potential. Here we present an examination of adjacent normal vulvar epithelium, dVIN, and VSCC from six patients by peptide Matrix-assisted laser desorption/ionization Mass Spectrometry Imaging (MALDI-MSI). The results reveal the differential expression of multiple peptides from the protein cytokeratin 5 (CK5) across the three vulvar tissue types. The difference observed in the relative abundance of CK5 by MALDI-MSI between the healthy epithelium, dVIN, and VSCC was further analyzed by immunohistochemistry (IHC) in tissue from eight VSCC patients. A decrease in CK5 immunostaining was observed in the VSCC compared to the healthy epithelium and dVIN. These results provide an insight into the molecular fingerprint of the vulvar intraepithelial neoplasia that appears to be more closely related to the healthy epithelium than the VSCC. PMID:27399691

  12. Metformin Decreases Reactive Oxygen Species, Enhances Osteogenic Properties of Adipose-Derived Multipotent Mesenchymal Stem Cells In Vitro, and Increases Bone Density In Vivo.

    Science.gov (United States)

    Marycz, Krzysztof; Tomaszewski, Krzysztof A; Kornicka, Katarzyna; Henry, Brandon Michael; Wroński, Sebastian; Tarasiuk, Jacek; Maredziak, Monika

    2016-01-01

    Due to its pleiotropic effects, the commonly used drug metformin has gained renewed interest among medical researchers. While metformin is mainly used for the treatment of diabetes, recent studies suggest that it may have further application in anticancer and antiaging therapies. In this study, we investigated the proliferative potential, accumulation of oxidative stress factors, and osteogenic and adipogenic differentiation potential of mouse adipose-derived stem cells (MuASCs) isolated from mice treated with metformin for 8 weeks. Moreover, we investigated the influence of metformin supplementation on mice bone density and bone element composition. The ASCs isolated from mice who were treated with metformin for 8 weeks showed highest proliferative potential, generated a robust net of cytoskeletal projections, had reduced expression of markers associated with cellular senescence, and decreased amount of reactive oxygen species in comparison to control group. Furthermore, we demonstrated that these cells possessed greatest osteogenic differentiation potential, while their adipogenic differentiation ability was reduced. We also demonstrated that metformin supplementation increases bone density in vivo. Our result stands as a valuable source of data regarding the in vivo influence of metformin on ASCs and bone density and supports a role for metformin in regenerative medicine. PMID:27195075

  13. Metformin Decreases Reactive Oxygen Species, Enhances Osteogenic Properties of Adipose-Derived Multipotent Mesenchymal Stem Cells In Vitro, and Increases Bone Density In Vivo

    Directory of Open Access Journals (Sweden)

    Krzysztof Marycz

    2016-01-01

    Full Text Available Due to its pleiotropic effects, the commonly used drug metformin has gained renewed interest among medical researchers. While metformin is mainly used for the treatment of diabetes, recent studies suggest that it may have further application in anticancer and antiaging therapies. In this study, we investigated the proliferative potential, accumulation of oxidative stress factors, and osteogenic and adipogenic differentiation potential of mouse adipose-derived stem cells (MuASCs isolated from mice treated with metformin for 8 weeks. Moreover, we investigated the influence of metformin supplementation on mice bone density and bone element composition. The ASCs isolated from mice who were treated with metformin for 8 weeks showed highest proliferative potential, generated a robust net of cytoskeletal projections, had reduced expression of markers associated with cellular senescence, and decreased amount of reactive oxygen species in comparison to control group. Furthermore, we demonstrated that these cells possessed greatest osteogenic differentiation potential, while their adipogenic differentiation ability was reduced. We also demonstrated that metformin supplementation increases bone density in vivo. Our result stands as a valuable source of data regarding the in vivo influence of metformin on ASCs and bone density and supports a role for metformin in regenerative medicine.

  14. Metformin Decreases Reactive Oxygen Species, Enhances Osteogenic Properties of Adipose-Derived Multipotent Mesenchymal Stem Cells In Vitro, and Increases Bone Density In Vivo

    Science.gov (United States)

    Marycz, Krzysztof; Tomaszewski, Krzysztof A.; Kornicka, Katarzyna; Henry, Brandon Michael; Wroński, Sebastian; Tarasiuk, Jacek; Maredziak, Monika

    2016-01-01

    Due to its pleiotropic effects, the commonly used drug metformin has gained renewed interest among medical researchers. While metformin is mainly used for the treatment of diabetes, recent studies suggest that it may have further application in anticancer and antiaging therapies. In this study, we investigated the proliferative potential, accumulation of oxidative stress factors, and osteogenic and adipogenic differentiation potential of mouse adipose-derived stem cells (MuASCs) isolated from mice treated with metformin for 8 weeks. Moreover, we investigated the influence of metformin supplementation on mice bone density and bone element composition. The ASCs isolated from mice who were treated with metformin for 8 weeks showed highest proliferative potential, generated a robust net of cytoskeletal projections, had reduced expression of markers associated with cellular senescence, and decreased amount of reactive oxygen species in comparison to control group. Furthermore, we demonstrated that these cells possessed greatest osteogenic differentiation potential, while their adipogenic differentiation ability was reduced. We also demonstrated that metformin supplementation increases bone density in vivo. Our result stands as a valuable source of data regarding the in vivo influence of metformin on ASCs and bone density and supports a role for metformin in regenerative medicine. PMID:27195075

  15. MALDI Mass Spectrometry Imaging Reveals Decreased CK5 Levels in Vulvar Squamous Cell Carcinomas Compared to the Precursor Lesion Differentiated Vulvar Intraepithelial Neoplasia

    Directory of Open Access Journals (Sweden)

    Chao Zhang

    2016-07-01

    Full Text Available Vulvar cancer is the fourth most common gynecological cancer worldwide. However, limited studies have been completed on the molecular characterization of vulvar squamous cell carcinoma resulting in a poor understanding of the disease initiation and progression. Analysis and early detection of the precursor lesion of HPV-independent vulvar squamous cell carcinoma (VSCC, differentiated vulvar intraepithelial neoplasia (dVIN, is of great importance given dVIN lesions have a high level of malignant potential. Here we present an examination of adjacent normal vulvar epithelium, dVIN, and VSCC from six patients by peptide Matrix-assisted laser desorption/ionization Mass Spectrometry Imaging (MALDI-MSI. The results reveal the differential expression of multiple peptides from the protein cytokeratin 5 (CK5 across the three vulvar tissue types. The difference observed in the relative abundance of CK5 by MALDI-MSI between the healthy epithelium, dVIN, and VSCC was further analyzed by immunohistochemistry (IHC in tissue from eight VSCC patients. A decrease in CK5 immunostaining was observed in the VSCC compared to the healthy epithelium and dVIN. These results provide an insight into the molecular fingerprint of the vulvar intraepithelial neoplasia that appears to be more closely related to the healthy epithelium than the VSCC.

  16. Regulation of Cell Migration by Sphingomyelin Synthases: Sphingomyelin in Lipid Rafts Decreases Responsiveness to Signaling by the CXCL12/CXCR4 Pathway

    OpenAIRE

    Asano, Satoshi; Kitatani, Kazuyuki; Taniguchi, Makoto; Hashimoto, Mayumi; Zama, Kota; Mitsutake, Susumu; Igarashi, Yasuyuki; Takeya, Hiroyuki; KIGAWA, JUNZO; Hayashi, Akira; Umehara, Hisanori; Okazaki, Toshiro

    2012-01-01

    Sphingomyelin synthase (SMS) catalyzes the formation of sphingomyelin, a major component of the plasma membrane and lipid rafts. To investigate the role of SMS in cell signaling and migration induced by binding of the chemokine CXCL12 to CXCR4, we used mouse embryonic fibroblasts deficient in SMS1 and/or SMS2 and examined the effects of SMS deficiency on cell migration. SMS deficiency promoted cell migration through a CXCL12/CXCR4-dependent signaling pathway involving extracellular signal-reg...

  17. Beta-adrenergic receptors on murine lymphocytes: density varies with cell maturity and lymphocyte subtype and is decreased after antigen administration

    International Nuclear Information System (INIS)

    beta-Adrenergic receptors were assayed on intact, viable, murine splenocytes and thymocytes using the labeled adrenergic antagonists [3H]-dihydroalprenolol l-[ring propyl-3H(N)] ([3H]DHA) and 4-(3-t-butylamino-2-hydroxypropoxy)-[5,7-3H]benzimidazol-2-one ([3H]CGP 12177). The sites detected by [3H]DHA did not always possess the characteristics of beta-adrenergic receptors and were demonstrated to be stereospecific only after the addition of the binding assay. Populations of cells from C57Bl/6 inbred and CF1 outbred mice were compared. Purified T cells from C57Bl/6 mice had fewer receptors than did either whole spleen or B cells. Thymocytes from either strain had significantly fewer receptors than did the other lymphocyte populations. However, mature medullary thymocytes purified from C57Bl/6 mice had higher numbers of receptors per cell which were comparable to those of the splenic T cell. Radiation-resistant splenocytes recovered from CF1 mice 24 hr after 700 rad of irradiation possessed greatly increased numbers of receptors per cell. Immunization with sheep red blood cells caused a significant reduction in the density of receptors on splenocytes from C57Bl/6 mice. The wide variations observed in the density of beta-adrenergic receptors, possibly related to cell maturity or state of activation, seem to provide opportunities for differential modulation of cell functions by either endogenous or exogenous adrenergic agents

  18. Decreased percentage of CD4+Foxp3+TGF-β+ and increased percentage of CD4+IL-17+ cells in bronchoalveolar lavage of asthmatics

    OpenAIRE

    Barczyk, Adam; Pierzchala, Wladyslaw; Caramori, Gaetano; Wiaderkiewicz, Ryszard; Kaminski, Marcin; Barnes, Peter J; Adcock, Ian M.

    2014-01-01

    Background Asthma is a chronic inflammatory disorder of the airways with the proven role of Th2 cells in its pathogenesis. The role and characteristic of different subsets of CD4+ cells is much less known. Aim The aim of the study was to analyze the incidence of different subsets of CD4+ T cells, in particular different subsets of CD4+ cells with the co-expression of different cytokines. Methods Twenty five stable asthmatic and twelve age-matched control subjects were recruited to the study. ...

  19. The anti-proliferative effect of L-carnosine correlates with a decreased expression of hypoxia inducible factor 1 alpha in human colon cancer cells.

    Directory of Open Access Journals (Sweden)

    Barbara Iovine

    Full Text Available In recent years considerable attention has been given to the use of natural substances as anticancer drugs. The natural antioxidant dipeptide L-carnosine belongs to this class of molecules because it has been proved to have a significant anticancer activity both in vitro and in vivo. Previous studies have shown that L-carnosine inhibits the proliferation of human colorectal carcinoma cells by affecting the ATP and Reactive Oxygen Species (ROS production. In the present study we identified the Hypoxia-Inducible Factor 1α (HIF-1α as a possible target of L-carnosine in HCT-116 cell line. HIF-1α protein is over-expressed in multiple types of human cancer and is the major cause of resistance to drugs and radiation in solid tumours. Of particular interest are experimental data supporting the concept that generation of ROS provides a redox signal for HIF-1α induction, and it is known that some antioxidants are able to suppress tumorigenesis by inhibiting HIF-1α. In the current study we found that L-carnosine reduces the HIF-1α protein level affecting its stability and decreases the HIF-1 transcriptional activity. In addition, we demonstrated that L-carnosine is involved in ubiquitin-proteasome system promoting HIF-1α degradation. Finally, we compared the antioxidant activity of L-carnosine with that of two synthetic anti-oxidant bis-diaminotriazoles (namely 1 and 2, respectively. Despite these three compounds have the same ability in reducing intracellular ROS, 1 and 2 are more potent scavengers and have no effect on HIF-1α expression and cancer cell proliferation. These findings suggest that an analysis of L-carnosine antioxidant pathway will clarify the mechanism underlying the anti-proliferative effects of this dipeptide on colon cancer cells. However, although the molecular mechanism by which L-carnosine down regulates or inhibits the HIF-1α activity has not been yet elucidated, this ability may be promising in treating hypoxia

  20. The anti-proliferative effect of L-carnosine correlates with a decreased expression of hypoxia inducible factor 1 alpha in human colon cancer cells.

    Science.gov (United States)

    Iovine, Barbara; Oliviero, Giorgia; Garofalo, Mariangela; Orefice, Maria; Nocella, Francesca; Borbone, Nicola; Piccialli, Vincenzo; Centore, Roberto; Mazzone, Massimiliano; Piccialli, Gennaro; Bevilacqua, Maria Assunta

    2014-01-01

    In recent years considerable attention has been given to the use of natural substances as anticancer drugs. The natural antioxidant dipeptide L-carnosine belongs to this class of molecules because it has been proved to have a significant anticancer activity both in vitro and in vivo. Previous studies have shown that L-carnosine inhibits the proliferation of human colorectal carcinoma cells by affecting the ATP and Reactive Oxygen Species (ROS) production. In the present study we identified the Hypoxia-Inducible Factor 1α (HIF-1α) as a possible target of L-carnosine in HCT-116 cell line. HIF-1α protein is over-expressed in multiple types of human cancer and is the major cause of resistance to drugs and radiation in solid tumours. Of particular interest are experimental data supporting the concept that generation of ROS provides a redox signal for HIF-1α induction, and it is known that some antioxidants are able to suppress tumorigenesis by inhibiting HIF-1α. In the current study we found that L-carnosine reduces the HIF-1α protein level affecting its stability and decreases the HIF-1 transcriptional activity. In addition, we demonstrated that L-carnosine is involved in ubiquitin-proteasome system promoting HIF-1α degradation. Finally, we compared the antioxidant activity of L-carnosine with that of two synthetic anti-oxidant bis-diaminotriazoles (namely 1 and 2, respectively). Despite these three compounds have the same ability in reducing intracellular ROS, 1 and 2 are more potent scavengers and have no effect on HIF-1α expression and cancer cell proliferation. These findings suggest that an analysis of L-carnosine antioxidant pathway will clarify the mechanism underlying the anti-proliferative effects of this dipeptide on colon cancer cells. However, although the molecular mechanism by which L-carnosine down regulates or inhibits the HIF-1α activity has not been yet elucidated, this ability may be promising in treating hypoxia-related diseases. PMID

  1. Warburg effect increases steady-state ROS condition in cancer cells through decreasing their antioxidant capacities (anticancer effects of 3-bromopyruvate through antagonizing Warburg effect).

    Science.gov (United States)

    El Sayed, Salah Mohamed; Mahmoud, Ahmed Alamir; El Sawy, Samer Ahmed; Abdelaal, Esam Abdelrahim; Fouad, Amira Murad; Yousif, Reda Salah; Hashim, Marwa Shaban; Hemdan, Shima Badawy; Kadry, Zainab Mahmoud; Abdelmoaty, Mohamed Ahmed; Gabr, Adel Gomaa; Omran, Faten M; Nabo, Manal Mohamed Helmy; Ahmed, Nagwa Sayed

    2013-11-01

    Cancer cells undergo an increased steady-state ROS condition compared to normal cells. Among the major metabolic differences between cancer cells and normal cells is the dependence of cancer cells on glycolysis as a major source of energy even in the presence of oxygen (Warburg effect). In Warburg effect, glucose is catabolized to lactate that is extruded through monocarboxylate transporters to the microenvironment of cancer cells, while in normal cells, glucose is metabolized into pyruvate that is not extruded. Pyruvate is a potent antioxidant, while lactate has no antioxidant effect. Pyruvate in normal cells may be further metabolized to acetyl CoA and then through Krebs cycle with production of antioxidant intermediates e.g. citrate, malate and oxaloacetate together with the reducing equivalents (NADH.H+). Through activity of mitochondrial transhydrogenase, NADH.H+ replenishes NADPH.H+, coenzyme of glutathione reductase which replenishes reduced form of glutathione (potent antioxidant). This enhances antioxidant capacities of normal cells, while cancer cells exhibiting Warburg effect may be deprived of all that antioxidant capabilities due to loss of extruded lactate (substrate for Krebs cycle). Although intrinsic oxidative stress in cancer cells is high, it may be prevented from reaching progressively increasing levels that are cytotoxic to cancer cells. This may be due to some antioxidant effects exerted by hexokinase II (HK II) and NADPH.H+ produced through HMP shunt. Glycolytic phenotype in cancer cells maintains a high non-toxic oxidative stress in cancer cells and may be responsible for their malignant behavior. Through HK II, glycolysis fuels the energetic arm of malignancy, the mitotic arm of malignancy (DNA synthesis through HMP shunt pathway) and the metastatic arm of malignancy (hyaluronan synthesis through uronic acid pathway) in addition to the role of phosphohexose isomerase (autocrine motility factor). All those critical three arms start with the

  2. A decrease in the addition of new cells in the nucleus accumbens and prefrontal cortex between puberty and adulthood in male rats.

    Science.gov (United States)

    Staffend, Nancy A; Mohr, Margaret A; DonCarlos, Lydia L; Sisk, Cheryl L

    2014-06-01

    Adolescence involves shifts in social behaviors, behavioral flexibility, and adaptive risk-taking that coincide with structural remodeling of the brain. We previously showed that new cells are added to brain regions associated with sexual behaviors, suggesting that cytogenesis may be a mechanism for acquiring adult-typical behaviors during adolescence. Whether pubertal cell addition occurs in brain regions associated with behavioral flexibility or motivation and whether these patterns differ between pubertal and adult animals had not been determined. Therefore, we assessed patterns of cell proliferation or survival in the prefrontal cortex and nucleus accumbens. Pubertal and adult male rats were given injections of bromo-deoxyuridine (BrdU). To assess cell proliferation, half of the animals from each group were sacrificed 24 h following the last injection. The remaining animals were sacrificed at Day 30 following the last injection to evaluate cell survival. Adult animals had significantly lower densities of BrdU-immunoreactive (ir) cells in the prefrontal cortex, irrespective of post-BrdU survival time, whereas in the nucleus accumbens, adult animals had a lower density of BrdU-ir cells at the short survival time; however, the density of BrdU-ir cells was equivalent in pubertal and adult animals at the longer survival time. These data provide evidence that cell addition during puberty may contribute to the remodeling of brain regions associated with behavioral flexibility and motivation, and this cell addition continues into adulthood, albeit at lower levels. Higher levels of cell proliferation or survival in younger animals may reflect a higher level of plasticity, possibly contributing to the dynamic remodeling of the pubertal brain. PMID:24339170

  3. Sickle Cell Disease

    Science.gov (United States)

    ... sickle cell disease? Sickle cell disease, also called sickle cell anemia, is a hereditary condition (which means it runs ... or blocks blood and oxygen reaching nearby tissues. Sickle cell disease ... the whites of the eyes) Anemia (the decreased ability of the blood to carry ...

  4. T Cells

    Science.gov (United States)

    T Cells - National Multiple Sclerosis Society Skip to navigation Skip to content Menu Navigation National Multiple Sclerosis Society Sign ... Is MS? Definition of MS T Cells T Cells Share Smaller Text Larger Text Print In this ...

  5. Cell counting.

    Science.gov (United States)

    Phelan, M C; Lawler, G

    2001-05-01

    This unit presents protocols for counting cells using either a hemacytometer or electronically using a Coulter counter. Cell counting with a hemacytometer permits effective discrimination of live from dead cells using trypan blue exclusion. In addition, the procedure is less subject to errors arising from cell clumping or size heterogeneity. Counting cells is more quickly and easily performed using an electronic counter, but live-dead discrimination is unreliable. Cell populations containing large numbers of dead cells and/or cell clumps are difficult to count accurately. In addition, electronic counting requires resetting of the instrument for cell populations of different sizes; heterogeneous populations can give rise to inaccurate counts, and resting and activated cells may require counting at separate settings. In general, electronic cell counting is best performed on fresh peripheral blood cells. PMID:18770655

  6. Activation of chloride current and decrease of cell volume by ATP in nasopharyngeal carcinoma cells%ATP激活鼻咽癌细胞氯电流并减小细胞容积

    Institute of Scientific and Technical Information of China (English)

    何庆丰; 王立伟; 毛建文; 孙雪荣; 李攀; 钟平; 聂思槐; Tim JACOB; 陈丽新

    2004-01-01

    Whole-cell patch clamp and cell volume measurement techniques were used to investigate the ATP-activated chloride current and the ATP effect on cell volume in nasopharyngeal carcinoma cells. Extracellular application of ATP in micromolar concentrations activated a current with the properties of modest outward rectification and negligible time-dependent inactivation in a dose-dependent manner. The current reversed at a potential [(-0.05±0.03) mV] close to the Cl equilibrium potential (-0.9 mV). Substitution of Cl- with gluconate in the extracellular solution decreased the ATP-activated current and shifted the reversal potential positively. NPPB, one of the chloride channel blockers,inhibited the current by (81.03±9.36)%. The current was also depressed by the P2Y purinoceptor antagonist, reactive blue 2, by (67.39±5.06)%.ATP (50 μmol/L) decreased the cell volume under the isotonic condition. Depletion of extracellular and intracellular Cl- abolished the ATP effect on cell volume. The results suggest that extracellular ATP of micromolar scales can induce a chloride current associated with cell volume regulation by activation of chloride channel through binding to purinoceptor P2Y.%采用全细胞膜片钳技术和细胞容积测量技术,在低分化鼻咽癌细胞株CNE-2Z上观察ATP诱导的Cl-电流的特性及其对细胞容积的影响.细胞外微摩尔水平的ATP以剂量依赖性的方式激活一个具有弱外向整流特性,没有时间依赖性失活的电流,此电流的反转电位[(0.05±0.03)mV]接近Cl-的平衡电位(-0.9mV).用葡萄糖酸置换细胞外液Cl-后,ATP激活的电流明显减小并且反转电位发生改变.氯通道抑制剂NPPB(200 μmol/L)可以抑制这一电流[(81.03±9.3)%].此电流亦可被嘌呤受体(P2Y)拮抗剂反应蓝2抑制[(67.39±5.06)%].50 μmol/L的ATP使在等渗状态下的细胞容积缩小,替代和耗竭细胞外、内的Cl-后,ATP的这一作用消失.这些结果提示细胞外微摩

  7. Leydig cell micronodules are a common finding in testicular biopsies from men with impaired spermatogenesis and are associated with decreased testosterone/LH ratio

    DEFF Research Database (Denmark)

    Holm, Mette; Rajpert-De Meyts, Ewa; Andersson, Anna-Maria;

    2003-01-01

    To assess the biological significance of Leydig cell 'hyperplasia' in man, Leydig cell distribution, volume, and function were studied in patients with infertility or testicular cancer and in suddenly deceased controls. A total of 156 biopsies from 95 patients and 18 necropsies from 13 controls w...

  8. Decrease in the production of beta-amyloid by berberine inhibition of the expression of beta-secretase in HEK293 cells

    Directory of Open Access Journals (Sweden)

    Zhu Feiqi

    2011-12-01

    Full Text Available Abstract Background Berberine (BER, the major alkaloidal component of Rhizoma coptidis, has multiple pharmacological effects including inhibition of acetylcholinesterase, reduction of cholesterol and glucose levels, anti-inflammatory, neuroprotective and neurotrophic effects. It has also been demonstrated that BER can reduce the production of beta-amyloid40/42, which plays a critical and primary role in the pathogenesis of Alzheimer's disease. However, the mechanism by which it accomplishes this remains unclear. Results Here, we report that BER could not only significantly decrease the production of beta-amyloid40/42 and the expression of beta-secretase (BACE, but was also able to activate the extracellular signal-regulated kinase1/2 (ERK1/2 pathway in a dose- and time-dependent manner in HEK293 cells stably transfected with APP695 containing the Swedish mutation. We also find that U0126, an antagonist of the ERK1/2 pathway, could abolish (1 the activation activity of BER on the ERK1/2 pathway and (2 the inhibition activity of BER on the production of beta-amyloid40/42 and the expression of BACE. Conclusion Our data indicate that BER decreases the production of beta-amyloid40/42 by inhibiting the expression of BACE via activation of the ERK1/2 pathway.

  9. Deleted in malignant brain tumors 1 (DMBT1) elicits increased VEGF and decreased IL-6 production in type II lung epithelial cells

    DEFF Research Database (Denmark)

    Müller, Hanna; Nagel, Christian; Weiss, Christel;

    2015-01-01

    between VEGF and IL-6 levels to DMBT1 expression in the lungs of preterm and term infants and in lung epithelial cells in vitro. METHODS: We examined by ELISA VEGF levels in 120 tracheal aspirates of 57 preterm and term infants and tested for correlation with different perinatal factors as well as with...... DMBT1 levels. To examine the effect of DMBT1 on VEGF and IL-6 expression we compared type II lung epithelial A549 cells stably transfected with a DMBT1 expression plasmid (DMBT1+ cells) to A549 cells stably transfected with an empty expression plasmid (DMBT1- cells). The concentrations of VEGF and IL-6...... that DMBT1 promotes VEGF and suppresses IL-6 production in alveolar tissues, which could point to DMBT1 having a possible role in the transition from inflammation to regeneration and being a potentially useful clinical marker....

  10. Temporal profile of magnetic resonance angiography and decreased ratio of regulatory T cells after immunological adjuvant administration to mice lacking RNF213, a susceptibility gene for moyamoya disease.

    Science.gov (United States)

    Kanoke, Atsushi; Fujimura, Miki; Niizuma, Kuniyasu; Fujimura, Taku; Kakizaki, Aya; Ito, Akira; Sakata, Hiroyuki; Sato-Maeda, Mika; Kure, Shigeo; Tominaga, Teiji

    2016-07-01

    Moyamoya disease (MMD) is a chronic, occlusive cerebrovascular disease with an unknown etiology and is characterized by an abnormal vascular network at the base of the brain. Recent studies identified the RNF213 gene (RNF213) as an important susceptibility gene for MMD; however, the mechanisms underlying the RNF213 abnormality related to MMD have not yet been elucidated. We previously reported that Rnf213-deficient mice and Rnf213 p. R4828K knock-in mice did not spontaneously develop MMD, indicating the importance of secondary insults in addition to genetic factors in the pathogenesis of MMD. The most influential secondary insult is considered to be an immunological reaction because RNF213 is predominantly expressed in immunological tissues. Therefore, we herein attempted to evaluate the role of an immunological stimulation as a supplementary insult to the target disruption of RNF213 in the pathophysiology of MMD. Rnf213-deficient mice were treated with strong immunological adjuvants including muramyl dipeptide (MDP)-Lys (L18), and then underwent time-sequential magnetic resonance angiography (MRA) up to 40 weeks of age. The results obtained did not reveal any characteristic finding of MMD, and no significant difference was observed in MRA findings or the anatomy of the circle of Willis between Rnf213-deficient mice and wild-type mice after the administration of MDP-Lys (L18). The ratio of regulatory T cells after the administration of MDP-Lys (L18) was significantly decreased in Rnf213-deficient mice (p<0.01), suggesting the potential role of the RNF213 abnormality in the differentiation of regulatory T cells. Although the mechanisms underlying the development of MMD currently remain unclear, the RNF213 abnormality may compromise immunological self-tolerance, thereby contributing to the development of MMD. PMID:26972532

  11. Heterologous expression of C. elegans fat-1 decreases the n-6/n-3 fatty acid ratio and inhibits adipogenesis in 3T3-L1 cells

    Energy Technology Data Exchange (ETDEWEB)

    An, Lei, E-mail: anleim@yahoo.com.cn [Ministry of Agriculture Key Laboratory of Animal Genetics, Breeding and Reproduction, National Engineering Laboratory for Animal Breeding, College of Animal Science and Technology, China Agricultural University, Beijing 100193 (China); Pang, Yun-Wei, E-mail: yunweipang@126.com [Ministry of Agriculture Key Laboratory of Animal Genetics, Breeding and Reproduction, National Engineering Laboratory for Animal Breeding, College of Animal Science and Technology, China Agricultural University, Beijing 100193 (China); Gao, Hong-Mei, E-mail: Gaohongmei_123@yahoo.cn [Ministry of Agriculture Key Laboratory of Animal Genetics, Breeding and Reproduction, National Engineering Laboratory for Animal Breeding, College of Animal Science and Technology, China Agricultural University, Beijing 100193 (China); Research Unit for Animal Life Sciences, Animal Resource Science Center, Graduate School of Agricultural and Life Sciences, The University of Tokyo, Ibaraki-Iwama 319-0206 (Japan); Tao, Li, E-mail: Eunice8023@yahoo.cn [Ministry of Agriculture Key Laboratory of Animal Genetics, Breeding and Reproduction, National Engineering Laboratory for Animal Breeding, College of Animal Science and Technology, China Agricultural University, Beijing 100193 (China); College of Animal Science and Technology, Jilin Agricultural University, Changchun, Jilin 130118 (China); Miao, Kai, E-mail: miaokai7@163.com [Ministry of Agriculture Key Laboratory of Animal Genetics, Breeding and Reproduction, National Engineering Laboratory for Animal Breeding, College of Animal Science and Technology, China Agricultural University, Beijing 100193 (China); Wu, Zhong-Hong, E-mail: wuzhh@cau.edu.cn [Ministry of Agriculture Key Laboratory of Animal Genetics, Breeding and Reproduction, National Engineering Laboratory for Animal Breeding, College of Animal Science and Technology, China Agricultural University, Beijing 100193 (China); and others

    2012-11-23

    Highlights: Black-Right-Pointing-Pointer Expression of C. elegans fat-1 reduces the n-6/n-3 PUFA ratio in 3T3-L1 cells. Black-Right-Pointing-Pointer fat-1 inhibits the proliferation and differentiation of 3T3-L1 preadipocytes. Black-Right-Pointing-Pointer fat-1 reduces lipid deposition in 3T3-L1 adipocytes. Black-Right-Pointing-Pointer The lower n-6/n-3 ratio induces apoptosis in 3T3-L1 adipocytes. -- Abstract: In general, a diet enriched in polyunsaturated fatty acids (PUFAs) inhibits the development of obesity and decreases adipose tissue. The specific impacts of n-3 and n-6 PUFAs on adipogenesis, however, have not been definitively determined. Traditional in vivo and in vitro supplementation studies have yielded inconsistent or even contradictory results, which likely reflect insufficiently controlled experimental systems. Caenorhabditiselegans fat-1 gene encodes an n-3 fatty acid desaturase, and its heterologous expression represents an effective method both for altering the n-6/n-3 PUFA ratio and for evaluating the biological effects of n-3 and n-6 PUFAs. We sought to determine whether a reduced n-6/n-3 ratio could influence adipogenesis in 3T3-L1 cells. Lentivirus-mediated introduction of the fat-1 gene into 3T3-L1 preadipocytes significantly reduced the n-6/n-3 ratio and inhibited preadipocyte proliferation and differentiation. In mature adipocytes, fat-1 expression reduced lipid deposition, as measured by Oil Red O staining, and induced apoptosis. Our results indicate that a reduced n-6/n-3 ratio inhibits adipogenesis through several mechanisms and that n-3 PUFAs more effectively inhibit adipogenesis (but not lipogenesis) than do n-6 PUFAs.

  12. Inhibition of p38 mitogen-activated protein kinase may decrease intestinal epithelial cell apoptosis and improve intestinal epithelial barrier function after ischemia- reperfusion injury

    Institute of Scientific and Technical Information of China (English)

    Shu-Yun Zheng; Xiao-Bing Fu; Jian-Guo Xu; Jing-Yu Zhao; Tong-Zhu Sun; Wei Chen

    2005-01-01

    AIM: To investigate the role of p38 mitogen-activated protein kinase in rat small intestine after ischemia-reperfusion (I/R)insult and the relationship between activation of p38 MAPK and apoptotic cell death of intestine.METHODS: Ninety Wistar rats were divided randomly into three groups, namely sham-operated group (C), I/R vehicle group (R) and SB203580 pre-treated group(S).In groups R and S, the superior mesenteric artery(SMA)was separated and occluded for 45 min, then released for reperfusion for0.25, 0.5, 1, 2, 6, 12 and 24 h. In group C, SMA was separated without occlusion. Plasma D-lactate levels were examined and histological changes were observed under a light microscope. The activity of p38 MAPK was determined by Western immunoblotting and apoptotic cells were detected by the terminal deoxynucleotidyl transferase (TdT)-mediated dUDP-biotin nick end labeling (TUNEL).RESULTS: Intestinal ischemia followed by reperfusion activated p38 MAPK, and the maximal level of activation (7.3-fold vs sham-operated group) was reached 30 min after I/R. Treatment with SB 203580, a p38 MAPK inhibitor,reduced intestinal apoptosis (26.72±3.39% vs62.50±3.08%in I/R vehicle, P<0.01) and decreased plasma D-lactate level (0.78±0.15 mmol/L in I/R vehicle vs0.42±0.17 mmol/L in SB-treated group) and improved post-ischemic intestinal histological damage.CONCLUSION: p38 MAPK plays a crucial role in the signal transduction pathway mediating post-ischemic intestinal apoptosis, and inhibition of p38 MAPK may attenuate ischemia-reperfusion injury.

  13. Pioglitazone significantly prevented decreased rate of neural differentiation of mouse embryonic stem cells which was reduced by Pex11β knock-down.

    Science.gov (United States)

    Esmaeili, M; Ghaedi, K; Shoaraye Nejati, A; Nematollahi, M; Shiralyian, H; Nasr-Esfahani, M H

    2016-01-15

    Peroxisomes constitute special cellular organelles which display a variety of metabolic functions including fatty acid oxidation and free radical elimination. Abundance of these flexible organelles varies in response to different environmental stimuli. It has been demonstrated that PEX11β, a peroxisomal membrane elongation factor, is involved in the regulation of size, shape and number of peroxisomes. To investigate the role of PEX11β in neural differentiation of mouse embryonic stem cells (mESCs), we generated a stably transduced mESCs line that derives the expression of a short hairpin RNA against Pex11β gene following doxycycline (Dox) induction. Knock-down of Pex11β, during neural differentiation, significantly reduced the expression of neural progenitor cells and mature neuronal markers (p<0.05) indicating that decreased expression of PEX11β suppresses neuronal maturation. Additionally, mRNA levels of other peroxisome-related genes such as PMP70, Pex11α, Catalase, Pex19 and Pex5 were also significantly reduced by Pex11β knock-down (p<0.05). Interestingly, pretreatment of transduced mESCs with peroxisome proliferator-activated receptor γ agonist (pioglitazone (Pio)) ameliorated the inhibitory effects of Pex11β knock down on neural differentiation. Pio also significantly (p<0.05) increased the expression of neural progenitor and mature neuronal markers besides the expression of peroxisomal genes in transduced mESC. Results elucidated the importance of Pex11β expression in neural differentiation of mESCs, thereby highlighting the essential role of peroxisomes in mammalian neural differentiation. The observation that Pio recovered peroxisomal function and improved neural differentiation of Pex11β knocked-down mESCs, proposes a potential new pharmacological implication of Pio for neurogenesis in patients with peroxisomal defects. PMID:26562432

  14. Melatonin decreases estrogen receptor binding to estrogen response elements sites on the OCT4 gene in human breast cancer stem cells

    Science.gov (United States)

    Lopes, Juliana; Arnosti, David; Trosko, James E.; Tai, Mei-Hui; Zuccari, Debora

    2016-01-01

    Cancer stem cells (CSCs) pose a challenge in cancer treatment, as these cells can drive tumor growth and are resistant to chemotherapy. Melatonin exerts its oncostatic effects through the estrogen receptor (ER) pathway in cancer cells, however its action in CSCs is unclear. Here, we evaluated the effect of melatonin on the regulation of the transcription factor OCT4 (Octamer Binding 4) by estrogen receptor alpha (ERα) in breast cancer stem cells (BCSCs). The cells were grown as a cell suspension or as anchorage independent growth, for the mammospheres growth, representing the CSCs population and treated with 10 nM estrogen (E2) or 10 μM of the environmental estrogen Bisphenol A (BPA) and 1 mM of melatonin. At the end, the cell growth as well as OCT4 and ERα expression and the binding activity of ERα to the OCT4 was assessed. The increase in number and size of mammospheres induced by E2 or BPA was reduced by melatonin treatment. Furthermore, binding of the ERα to OCT4 was reduced, accompanied by a reduction of OCT4 and ERα expression. Thus, melatonin treatment is effective against proliferation of BCSCs in vitro and impacts the ER pathway, demonstrating its potential therapeutic use in breast cancer. PMID:27551335

  15. Treatment with granulocyte colony-stimulating factor decreases the capacity of hematopoietic progenitor cells for generation of lymphocytes in human immunodeficiency virus-infected persons

    DEFF Research Database (Denmark)

    Nielsen, Susanne Dam; Clark, D R; Hutchings, M;

    1999-01-01

    An obstacle to stem cell gene therapy for AIDS is the limited numbers of hematopoietic progenitors available. Granulocyte colony-stimulating factor (G-CSF) is used for mobilization of progenitors, but little is known about the functional characteristics of mobilized progenitors, and immature and T...... cell progenitors may not be mobilized. This study examined the effect of G-CSF on the function of progenitors. Ten human immunodeficiency virus-infected patients received G-CSF (filgrastim, 300 microgram/day) for 5 days. Absolute numbers of immature and T cell progenitors did not increase. The ability...

  16. Hydrogen sulfide decreases high glucose/palmitate-induced autophagy in endothelial cells by the Nrf2-ROS-AMPK signaling pathway

    OpenAIRE

    Liu, Jiaqi; Wu, Jichao; Sun, Aili; Sun, Yu; Yu, Xiangjing; Liu, Ning; Dong, Shiyun; Yang, Fan; Zhang, Linxue; Zhong, Xin; Xu, Changqing; Lu, Fanghao; Zhang, Weihua

    2016-01-01

    Background Excessive autophagy induced by extravagant oxidative stress is the main reason for diabetes-induced vascular endothelial cells dysfunction. Hydrogen sulfide (H2S) has anti-oxidative effects but its regulation on excessive autophagy of vascular endothelial cells is unclear. Methods In this study, aorta of db/db mice (28 weeks old) and rat aortic endothelial cells (RAECs) treated with 40 mM glucose and 500 μM palmitate acted as type II diabetic animal and cellular models, respectivel...

  17. Molecular events associated with increased regenerative capacity of the goldfish retinal ganglion cells following X-irradiation: decreased level of axonal growth inhibitors

    International Nuclear Information System (INIS)

    In our previous work we established conditions to study the contribution of non-neuronal cells to the process of goldfish optic nerve regeneration. This issue has been studied successfully by adapting the use of X-irradiation to manipulate division of non-neuronal cells associated with the injured nerve. The regenerative capacity of the goldfish retinal ganglion cells was determined subsequent to the X-ray treatment. The authors present an analysis of the molecular events associated with regeneration and enhanced regenerative capacity which follows X-irradiation. (Auth.)

  18. Tanshinone IIA inhibits human breast cancer MDA-MB-231 cells by decreasing LC3-II, Erb-B2 and NF-κBp65

    OpenAIRE

    Su, Chin-Cheng; Chien, Su-Yu; Kuo, Shou-Jen; Chen, Yao-Li; CHENG, CHUN-YUAN; Chen, Dar-Ren

    2012-01-01

    The ability of tanshinone IIA (Tan-IIA) to inhibit the proliferation of human breast cancer cell lines in vitro and in vivo is well documented. However, the molecular mechanisms have not been fully elucidated. In the present study, MDA-MB-231 cells were treated with different concentrations of Tan-IIA for 48 h, followed by protein extraction for western blotting. For an in vivo study, MDA-MB-231 cells were implanted directly into female SCID mice which were divided randomly into three groups ...

  19. R-CHOP regimen can significantly decrease the risk of disease relapse and progression in patients with non-germinal center B-cell subtype diffuse large B-cell lymphoma

    Institute of Scientific and Technical Information of China (English)

    Xiao-Hui He; Mei Dong; Sheng-Yu Zhou; Jian-Liang Yang; Peng Liu; Chang-Gong Zhang; Yan Qin; Feng-Yi Feng; Yuan-Kai Shi; Bo Li; Sheng Yang; Ning Lu; Xun Zhang; Shuang-Mei Zou; Ye-Xiong Li; Yong-Wen Song; Shan Zheng

    2012-01-01

    To further explore the role of rituximab when added to the CHOP-like regimen in the treatment of immunohistochemically defined non-germinal center B-cell subtype (non-GCB) diffuse large B-cell lymphoma (DLBCL),159 newly diagnosed DLBCL patients were studied retrospectively based on the immunohistochemical evaluation of CD10,Bcl-6,MUM-1,and Bcl-2.Altogether,110 patients underwent the CHOP-like regimen,and rituximab was added for the other 49 patients.Cox regression analysis showed that compared with the CHOP-like regimen,the rituximab-based regimen (R-CHOP regimen)significantly decreased the risk of disease relapse and progression in CD10-negative patients (P =0.001),Bcl-6-negative patients (P =0.01),and MUM-1-positive patients (P =0.003).The risk of disease relapse in patients with non-GCB subtype (P =0.002) also decreased.In contrast,patients with the opposite immunohistochemical marker expression profile and GCB subtype did not benefit from treatment with the R-CHOP regimen.In addition,non-GCB subtype patients had a significantly higher expression rate of Bcl-2than GCB subtype patients (P =0.042).Although univariate analysis found that both Bcl-2-positive and -negative patients had significantly higher event-free survival rates with the R-CHOP regimen,only Bcl-2positivity (P =0.004) maintained significance in the Cox regression analysis.We conclude that the addition of rituximab can significantly improve the prognosis of patients with non-GCB subtype DLBCL,which is closely related to the expression of CD10,Bcl-6,MUM-1,and Bcl-2.

  20. Galvanic Cells

    Science.gov (United States)

    Young, I. G.

    1973-01-01

    Many standard physical chemistry textbooks contain ambiguities which lead to confusion about standard electrode potentials, calculating cell voltages, and writing reactions for galvanic cells. This article shows how standard electrode potentials can be used to calculate cell voltages and deduce cell reactions. (Author/RH)

  1. Abrupt decrease of c—myc expression by antisense transcripts induses terminal differentiation and apoptosis in human promyelocytic leukemia HL—60 cells

    Institute of Scientific and Technical Information of China (English)

    HAOXIUJUAN; PEIHSIENTANG; 等

    1996-01-01

    This study was designed using c-myc antisense transcripts to evaluate how alteration of c-myc expression in human myeloid leukemic HL-60 cells could influence the myelomonocytic differentiation and induction of apoptosis.The recombinant plasmid pDACx expressing antisense transcripts to c-myc fragment containing a part of intron 1 and 137 nt exon 2 was constructed.pDACx was transfected into HL-60 cell line by lipofectin reagent.Cytochemical stainings including NBT reduction,peroxidase and α-NAE as well as detection of CD13 and CD33 antigens by flow cytometric analysis indicated occurrence of myelomonocytic differentiation in cells expressing antisense transcripts to c-myc.DNA degradation measured by DNA gel electrophoresis and typical morphological changes observed under electron microscope proved the swith-on of apoptosis in terminally differentiating HL-60 cells.

  2. Luteolin decreases IGF-II production and downregulates insulin-like growth factor-I receptor signaling in HT-29 human colon cancer cells

    Directory of Open Access Journals (Sweden)

    Lim Do

    2012-01-01

    Full Text Available Abstract Background Luteolin is a 3',4',5,7-tetrahydroxyflavone found in various fruits and vegetables. We have shown previously that luteolin reduces HT-29 cell growth by inducing apoptosis and cell cycle arrest. The objective of this study was to examine whether luteolin downregulates the insulin-like growth factor-I receptor (IGF-IR signaling pathway in HT-29 cells. Methods In order to assess the effects of luteolin and/or IGF-I on the IGF-IR signaling pathway, cells were cultured with or without 60 μmol/L luteolin and/or 10 nmol/L IGF-I. Cell proliferation, DNA synthesis, and IGF-IR mRNA levels were evaluated by a cell viability assay, [3H]thymidine incorporation assays, and real-time polymerase chain reaction, respectively. Western blot analyses, immunoprecipitation, and in vitro kinase assays were conducted to evaluate the secretion of IGF-II, the protein expression and activation of IGF-IR, and the association of the p85 subunit of phophatidylinositol-3 kinase (PI3K with IGF-IR, the phosphorylation of Akt and extracellular signal-regulated kinase (ERK1/2, and cell division cycle 25c (CDC25c, and PI3K activity. Results Luteolin (0 - 60 μmol/L dose-dependently reduced the IGF-II secretion of HT-29 cells. IGF-I stimulated HT-29 cell growth but did not abrogate luteolin-induced growth inhibition. Luteolin reduced the levels of the IGF-IR precursor protein and IGF-IR transcripts. Luteolin reduced the IGF-I-induced tyrosine phosphorylation of IGF-IR and the association of p85 with IGF-IR. Additionally, luteolin inhibited the activity of PI3K activity as well as the phosphorylation of Akt, ERK1/2, and CDC25c in the presence and absence of IGF-I stimulation. Conclusions The present results demonstrate that luteolin downregulates the activation of the PI3K/Akt and ERK1/2 pathways via a reduction in IGF-IR signaling in HT-29 cells; this may be one of the mechanisms responsible for the observed luteolin-induced apoptosis and cell cycle arrest.

  3. LITHOCHOLIC ACID DECREASES EXPRESSION OF UGT2B7 IN CACO-2 CELLS: A POTENTIAL ROLE FOR A NEGATIVE FARNESOID X RECEPTOR RESPONSE ELEMENT

    OpenAIRE

    Lu, Yuan; Heydel, Jean-Marie; LI, XIN; Bratton, Stacie; Lindblom, Tim; Radominska-Pandya, Anna

    2005-01-01

    Human UDP-glucuronosyltransferase (UGT) 2B7 is the major isoform catalyzing the glucuronidation of a variety of endogenous compounds including bile acids. To determine the role of bile acids in the regulation of UGT2B7 expression, Caco-2 cells were incubated with the natural human farnesoid X receptor (hFXR) ligand, chenodeoxycholic acid, as well as the secondary bile acid, lithocholic acid, derived from chenodeoxycholic acid. Incubation of Caco-2 cells with lithocholic acid in the absence of...

  4. Porphyromonas gingivalis Gingipain-R Enhances Interleukin-8 but Decreases Gamma Interferon-Inducible Protein 10 Production by Human Gingival Fibroblasts in Response to T-Cell Contact

    Science.gov (United States)

    Oido-Mori, Mari; Rezzonico, Roger; Wang, Pao-Li; Kowashi, Yusuke; Dayer, Jean-Michel; Baehni, Pierre C.; Chizzolini, Carlo

    2001-01-01

    Proteases produced by Porphyromonas gingivalis, an oral pathogen, are considered important virulence factors and may affect the responses of cells equipped with proteinase-activated receptors. The aim of this study was to investigate the effect of the arginine-specific cysteine protease gingipain-R produced by P. gingivalis on chemokine production by human gingival fibroblasts (HGF) and the effect of gingipain-R treatment on the subsequent contact-dependent activation of HGF by T cells. HGF incubated in the presence of purified 47-kDa gingipain-R showed increased levels of interleukin-8 (IL-8) mRNA. Cyclooxygenase-2 (COX-2) mRNA was also induced. Further exposure of HGF to activated T cells resulted in the dose- and time-dependent enhancement of IL-8 transcription and release. T-cell membrane-bound tumor necrosis factor (TNF) was the ligand inducing IL-8 production by HGF, since TNF neutralization abrogated HGF responses to T-cell contact. The enhanced IL-8 release was due, at least in part, to prostaglandin-E2 production, which was mostly blocked by indomethacin. Gingipain-R proteolytic activity was required since heat inactivation, specific synthetic protease inhibitors, and the natural substrate competitor histatin 5 abrogated its effects. The enhanced production of IL-8 in response to T-cell contact was specific since monocyte chemotactic protein-1 (MCP-1) production was unaffected while interferon-gamma-inducible protein-10 (IP-10) was inhibited. The sum of these activities may result in the recruitment of differential cell types to sites of inflammation since IL-8 preferentially recruits neutrophils and IP-10 attracts activated T cells and may be relevant to the pathogenesis of periodontitis. PMID:11401991

  5. COX-2 inhibition improves immunotherapy and is associated with decreased numbers of myeloid-derived suppressor cells in mesothelioma. Celecoxib influences MDSC function

    Directory of Open Access Journals (Sweden)

    Veltman Joris D

    2010-08-01

    Full Text Available Abstract Background Myeloid-derived suppressor cells (MDSC are a heterogeneous population of immature cells that accumulates in tumour-bearing hosts. These cells are induced by tumour-derived factors (e.g. prostaglandins and have a critical role in immune suppression. MDSC suppress T and NK cell function via increased expression of arginase I and production of reactive oxygen species (ROS and nitric oxide (NO. Immune suppression by MDSC was found to be one of the main factors for immunotherapy insufficiency. Here we investigate if the in vivo immunoregulatory function of MDSC can be reversed by inhibiting prostaglandin synthesis by specific COX-2 inhibition focussing on ROS production by MDSC subtypes. In addition, we determined if dietary celecoxib treatment leads to refinement of immunotherapeutic strategies. Methods MDSC numbers and function were analysed during tumour progression in a murine model for mesothelioma. Mice were inoculated with mesothelioma tumour cells and treated with cyclooxygenase-2 (COX-2 inhibitor celecoxib, either as single agent or in combination with dendritic cell-based immunotherapy. Results We found that large numbers of infiltrating MDSC co-localise with COX-2 expression in those areas where tumour growth takes place. Celecoxib reduced prostaglandin E2 levels in vitro and in vivo. Treatment of tumour-bearing mice with dietary celecoxib prevented the local and systemic expansion of all MDSC subtypes. The function of MDSC was impaired as was noticed by reduced levels of ROS and NO and reversal of T cell tolerance; resulting in refinement of immunotherapy. Conclusions We conclude that celecoxib is a powerful tool to improve dendritic cell-based immunotherapy and is associated with a reduction in the numbers and suppressive function of MDSC. These data suggest that immunotherapy approaches benefit from simultaneously blocking cyclooxygenase-2 activity.

  6. COX-2 inhibition improves immunotherapy and is associated with decreased numbers of myeloid-derived suppressor cells in mesothelioma. Celecoxib influences MDSC function

    International Nuclear Information System (INIS)

    Myeloid-derived suppressor cells (MDSC) are a heterogeneous population of immature cells that accumulates in tumour-bearing hosts. These cells are induced by tumour-derived factors (e.g. prostaglandins) and have a critical role in immune suppression. MDSC suppress T and NK cell function via increased expression of arginase I and production of reactive oxygen species (ROS) and nitric oxide (NO). Immune suppression by MDSC was found to be one of the main factors for immunotherapy insufficiency. Here we investigate if the in vivo immunoregulatory function of MDSC can be reversed by inhibiting prostaglandin synthesis by specific COX-2 inhibition focussing on ROS production by MDSC subtypes. In addition, we determined if dietary celecoxib treatment leads to refinement of immunotherapeutic strategies. MDSC numbers and function were analysed during tumour progression in a murine model for mesothelioma. Mice were inoculated with mesothelioma tumour cells and treated with cyclooxygenase-2 (COX-2) inhibitor celecoxib, either as single agent or in combination with dendritic cell-based immunotherapy. We found that large numbers of infiltrating MDSC co-localise with COX-2 expression in those areas where tumour growth takes place. Celecoxib reduced prostaglandin E2 levels in vitro and in vivo. Treatment of tumour-bearing mice with dietary celecoxib prevented the local and systemic expansion of all MDSC subtypes. The function of MDSC was impaired as was noticed by reduced levels of ROS and NO and reversal of T cell tolerance; resulting in refinement of immunotherapy. We conclude that celecoxib is a powerful tool to improve dendritic cell-based immunotherapy and is associated with a reduction in the numbers and suppressive function of MDSC. These data suggest that immunotherapy approaches benefit from simultaneously blocking cyclooxygenase-2 activity

  7. Decreased intratumoral Foxp3 Tregs and increased dendritic cell density by neoadjuvant chemotherapy associated with favorable prognosis in advanced gastric cancer

    OpenAIRE

    Hu, Min; Li, Kai; Maskey, Ninu; Xu, Zhigao; Peng, Chunwei; Wang, Bicheng; Li, Yan; Yang, Guifang

    2014-01-01

    Although neoadjuvant chemotherapy (NACT) has been increasingly used to improve the outcome of advanced gastric cancer (GC) for decades, its precise efficacy has been difficult to evaluate yet. Abundant studies have investigated the predictive factors that represent the effect of NACT on advanced GC. In the present study, the intratumoral infiltration of regulatory T cells (Tregs) and dendritic cells (DCs) response to NACT in advanced GC and their correlation with prognosis were evaluated. Inf...

  8. Stem Cells

    OpenAIRE

    Madhukar Thakur

    2009-01-01

    Objective: The objective of this presentation is to create awareness of stem cell applications in the ISORBE community and to foster a strategy of how the ISORBE community can disseminate information and promote the use of radiolabeled stem cells in biomedical applications. Methods: The continued excitement in Stem Cells, in many branches of basic and applied biomedical science, stems from the remarkable ability of stem cells to divide and develop into different types of cells in ...

  9. Cell Wall

    OpenAIRE

    Jamet, Elisabeth; Canut, Hervé; Boudart, Georges; Albenne, Cécile; Pont-Lezica, Rafael F

    2008-01-01

    This chapter covers our present knowledge of cell wall proteomics highlighting the distinctive features of cell walls and cell wall proteins in relation to problems encountered for protein extraction, separation and identification. It provides clues to design strategies for efficient cell wall proteomic studies. It gives an overview of the kinds of proteins that have yet been identified: the expected proteins vs the identified proteins. Finally, the new vision of the cell wall proteome, and t...

  10. Increased levels of soluble CD226 in sera accompanied by decreased membrane CD226 expression on peripheral blood mononuclear cells from cancer patients

    Directory of Open Access Journals (Sweden)

    Xu Zhuwei

    2009-06-01

    Full Text Available Abstract Background As a cellular membrane triggering receptor, CD226 is involved in the NK cell- or CTL-mediated lysis of tumor cells of different origin, including freshly isolated tumor cells and tumor cell lines. Here, we evaluated soluble CD226 (sCD226 levels in sera, and membrane CD226 (mCD226 expression on peripheral blood mononuclear cells (PBMC from cancer patients as well as normal subjects, and demonstrated the possible function and origin of the altered sCD226, which may provide useful information for understanding the mechanisms of tumor escape and for immunodiagnosis and immunotherapy. Results Soluble CD226 levels in serum samples from cancer patients were significantly higher than those in healthy individuals (P P Conclusion These findings suggest that sCD226 might be shed from cell membranes by certain proteases, and, further, sCD226 may be used as a predictor for monitoring cancer, and more important, a possible immunotherapy target, which may be useful in clinical application.

  11. Triptolide inhibits B7-H1 expression on proinflammatory factor activated renal tubular epithelial cells by decreasing NF-kappaB transcription.

    Science.gov (United States)

    Chen, Yongwen; Zhang, Jingbo; Li, Jingyi; Zhao, Tingting; Zou, Liyun; Tang, Yan; Zhang, Xiaoping; Wu, Yuzhang

    2006-03-01

    Triptolide has been used extensively in China for the treatment of autoimmune diseases and tumor for many centuries. Nevertheless, little is known about its exact immunosuppressive and anti-inflammatory properties. Increasing recognition of the importance of renal tubular epithelial cells (TECs) in renal diseases raises the question whether triptolide can regulate TEC activity. In this study, various cultured human and murine TECs were exposed to tumor necrotic factor-alpha (TNF-alpha) and triptolide, followed to examine the expression of B7-H1 and B7-DC. Flow cytometric analysis revealed that B7-H1 but not B7-DC constitutively expresses on TECs, and the B7-H1 protein expression was profoundly up-regulated by the stimulation of TNF-alpha with a dose-dependent manner. However, triptolide under non-cytotoxic concentration could down-regulate B7-H1 expression on activated TECs at both mRNA and protein level. This effect was transcription factor NF-kappaB dependent. Interestingly, the significant damping effect of triptolide on B7-H1 signal could promote interleukin-2 production by T cell hybridoma (C10) after antigen presentation and enhance cytokine (IFN-gamma and IL-2) secretion by anti-CD3 activated T cells. Our results indicated that triptolide could regulate TEC activity via B7-H1, in addition to previously reported it directly affects the production of some inflammatory factors by T cells, tumor cells and peripheral blood mononuclear cells. PMID:16129490

  12. PINK1 Deficiency Decreases Expression Levels of mir-326, mir-330, and mir-3099 during Brain Development and Neural Stem Cell Differentiation.

    Science.gov (United States)

    Choi, Insup; Woo, Joo Hong; Jou, Ilo; Joe, Eun-Hye

    2016-02-01

    PTEN-induced putative kinase 1 (PINK1) is a Parkinson's disease (PD) gene. We examined miRNAs regulated by PINK1 during brain development and neural stem cell (NSC) differentiation, and found that lvels of miRNAs related to tumors and inflammation were different between 1-day-old-wild type (WT) and PINK1-knockout (KO) mouse brains. Notably, levels of miR-326, miR-330 and miR-3099, which are related to astroglioma, increased during brain development and NSC differentiation, and were significantly reduced in the absence of PINK1. Interestingly, in the presence of ciliary neurotrophic factor (CNTF), which pushes differentiation of NSCs into astrocytes, miR-326, miR-330, and miR-3099 levels in KO NSCs were also lower than those in WT NSCs. Furthermore, mimics of all three miRNAs increased expression of the astrocytic marker glial fibrillary acidic protein (GFAP) during differentiation of KO NSCs, but inhibitors of these miRNAs decreased GFAP expression in WT NSCs. Moreover, these miRNAs increased the translational efficacy of GFAP through the 3'-UTR of GFAP mRNA. Taken together, these results suggest that PINK1 deficiency reduce expression levels of miR-326, miR-330 and miR-3099, which may regulate GFAP expression during NSC differentiation and brain development. PMID:26924929

  13. OTUB1 overexpression in mesangial cells is a novel regulator in the pathogenesis of glomerulonephritis through the decrease of DCN level.

    Directory of Open Access Journals (Sweden)

    Yan Zhang

    Full Text Available BACKGROUND: OTUB1 is a member of OTUs (Ovarian-tumor-domain-containing proteases, a deubiquitinating enzymes family (DUBs, which was shown as a proteasome-associated DUB to be involved in the proteins Ub-dependent degradation. It has been reported that OTUB1 was expressed in kidney tissue. But its concrete cellular location and function in the kidney remain unclear. Decorin (DCN in mesangial cells (MC is considered to be a potentially important factor for antagonizing glomerulonephritides, and its degradation is mediated by ubiquitination. The aim of this study is to investigate the role of OTUB1 expression in MC and its relationship with DCN during glomerulonephritis. METHODOLOGY/PRINCIPAL FINDINGS: Using quantitative RT-PCR and Western blot, we demonstrated that OTUB1 mRNA and protein were constitutively expressed in cultured rat MC and found to be upregulated by the stimulation of IL-1β or ATS. OTUB1 overexpression was detected in the mesangial area of glomeruli in some immunocomplex mediated nephritides such as IgA nephropathy, acute diffuse proliferative glomerulonephritis and lupus nephritis by immunohistochemistry. The immunoprecipitation assay demonstrated that OTUB1 interacted with DCN. The overexpression of OTUB1 enhanced the ubiquitination and degradation of DCN in MC. CONCLUSION/SIGNIFICANCE: These data showed the inflammatory injury could up-regulate OTUB1 expression in MC, which might attribute the promoting effect of OTUB1 on glomerulonephritides to the decrease of DCN level.

  14. Inactivation of the srtA Gene in Streptococcus gordonii Inhibits Cell Wall Anchoring of Surface Proteins and Decreases In Vitro and In Vivo Adhesion

    Science.gov (United States)

    Bolken, Tové C.; Franke, Christine A.; Jones, Kevin F.; Zeller, Gloria O.; Jones, C. Hal; Dutton, Emma K.; Hruby, Dennis E.

    2001-01-01

    The srtA gene product, SrtA, has been shown to be required for cell wall anchoring of protein A as well as virulence in the pathogenic bacterium Staphylococcus aureus. There are five major mechanisms for displaying proteins at the surface of gram-positive bacteria (P. Cossart and R. Jonquieres, Proc. Natl. Acad. Sci. USA 97:5013–5015, 2000). However, since many of the known surface proteins of gram-positive bacteria are believed to be exported and anchored via the sortase pathway, it was of interest to determine if srtA plays a similar role in other gram-positive bacteria. To that end, the srtA gene in the human oral commensal organism Streptococcus gordonii was insertionally inactivated. The srtA mutant S. gordonii exhibited a marked reduction in quantity of a specific anchored surface protein. Furthermore, the srtA mutant had reduced binding to immobilized human fibronectin and had a decreased ability to colonize the oral mucosa of mice. Taken together, these results suggest that the activity of SrtA plays an important role in the biology of nonpathogenic as well as pathogenic gram-positive cocci. PMID:11119491

  15. Cell Membrane Softening in Cancer Cells

    Science.gov (United States)

    Schmidt, Sebastian; Händel, Chris; Käs, Josef

    Biomechanical properties are useful characteristics and regulators of the cell's state. Current research connects mechanical properties of the cytoskeleton to many cellular processes but does not investigate the biomechanics of the plasma membrane. We evaluated thermal fluctuations of giant plasma membrane vesicles, directly derived from the plasma membranes of primary breast and cervical cells and observed a lowered rigidity in the plasma membrane of malignant cells compared to non-malignant cells. To investigate the specific role of membrane rigidity changes, we treated two cell lines with the Acetyl-CoA carboxylase inhibitor Soraphen A. It changed the lipidome of cells and drastically increased membrane stiffness by up regulating short chained membrane lipids. These altered cells had a decreased motility in Boyden chamber assays. Our results indicate that the thermal fluctuations of the membrane, which are much smaller than the fluctuations driven by the cytoskeleton, can be modulated by the cell and have an impact on adhesion and motility.

  16. Snail modulates cell metabolism in MDCK cells

    International Nuclear Information System (INIS)

    Highlights: ► MDCK/snail cells were more sensitive to glucose deprivation than MDCK/neo cells. ► MDCK/snail cells had decreased oxidative phosphorylation, O2 consumption and ATP content. ► TCA cycle enzyme activity, but not expression, was lower in MDCK/snail cells. ► MDCK/snail cells showed reduced PDH activity and increased PDK1 expression. ► MDCK/snail cells showed reduced expression of GLS2 and ACLY. -- Abstract: Snail, a repressor of E-cadherin gene transcription, induces epithelial-to-mesenchymal transition and is involved in tumor progression. Snail also mediates resistance to cell death induced by serum depletion. By contrast, we observed that snail-expressing MDCK (MDCK/snail) cells undergo cell death at a higher rate than control (MDCK/neo) cells in low-glucose medium. Therefore, we investigated whether snail expression influences cell metabolism in MDCK cells. Although gylcolysis was not affected in MDCK/snail cells, they did exhibit reduced pyruvate dehydrogenase (PDH) activity, which controls pyruvate entry into the tricarboxylic acid (TCA) cycle. Indeed, the activity of multiple enzymes involved in the TCA cycle was decreased in MDCK/snail cells, including that of mitochondrial NADP+-dependent isocitrate dehydrogenase (IDH2), succinate dehydrogenase (SDH), and electron transport Complex II and Complex IV. Consequently, lower ATP content, lower oxygen consumption and increased survival under hypoxic conditions was also observed in MDCK/snail cells compared to MDCK/neo cells. In addition, the expression and promoter activity of pyruvate dehydrogenase kinase 1 (PDK1), which phosphorylates and inhibits the activity of PDH, was increased in MDCK/snail cells, while expression levels of glutaminase 2 (GLS2) and ATP-citrate lyase (ACLY), which are involved in glutaminolysis and fatty acid synthesis, were decreased in MDCK/snail cells. These results suggest that snail modulates cell metabolism by altering the expression and activity of key enzymes

  17. Snail modulates cell metabolism in MDCK cells

    Energy Technology Data Exchange (ETDEWEB)

    Haraguchi, Misako, E-mail: haraguci@m3.kufm.kagoshima-u.ac.jp [Department of Biochemistry and Molecular Biology, Graduate School of Medical and Dental Sciences, Kagoshima University, Kagoshima 890-8544 (Japan); Indo, Hiroko P. [Department of Maxillofacial Radiology, Graduate School of Medical and Dental Sciences, Kagoshima University, Kagoshima 890-8544 (Japan); Iwasaki, Yasumasa [Health Care Center, Kochi University, Kochi 780-8520 (Japan); Iwashita, Yoichiro [Department of Maxillofacial Radiology, Graduate School of Medical and Dental Sciences, Kagoshima University, Kagoshima 890-8544 (Japan); Fukushige, Tomoko [Department of Dermatology, Graduate School of Medical and Dental Sciences, Kagoshima University, Kagoshima 890-8544 (Japan); Majima, Hideyuki J. [Department of Maxillofacial Radiology, Graduate School of Medical and Dental Sciences, Kagoshima University, Kagoshima 890-8544 (Japan); Izumo, Kimiko; Horiuchi, Masahisa [Department of Environmental Medicine, Graduate School of Medical and Dental Sciences, Kagoshima University, Kagoshima 890-8544 (Japan); Kanekura, Takuro [Department of Dermatology, Graduate School of Medical and Dental Sciences, Kagoshima University, Kagoshima 890-8544 (Japan); Furukawa, Tatsuhiko [Department of Molecular Oncology, Graduate School of Medical and Dental Sciences, Kagoshima University, Kagoshima 890-8544 (Japan); Ozawa, Masayuki [Department of Biochemistry and Molecular Biology, Graduate School of Medical and Dental Sciences, Kagoshima University, Kagoshima 890-8544 (Japan)

    2013-03-22

    Highlights: ► MDCK/snail cells were more sensitive to glucose deprivation than MDCK/neo cells. ► MDCK/snail cells had decreased oxidative phosphorylation, O{sub 2} consumption and ATP content. ► TCA cycle enzyme activity, but not expression, was lower in MDCK/snail cells. ► MDCK/snail cells showed reduced PDH activity and increased PDK1 expression. ► MDCK/snail cells showed reduced expression of GLS2 and ACLY. -- Abstract: Snail, a repressor of E-cadherin gene transcription, induces epithelial-to-mesenchymal transition and is involved in tumor progression. Snail also mediates resistance to cell death induced by serum depletion. By contrast, we observed that snail-expressing MDCK (MDCK/snail) cells undergo cell death at a higher rate than control (MDCK/neo) cells in low-glucose medium. Therefore, we investigated whether snail expression influences cell metabolism in MDCK cells. Although gylcolysis was not affected in MDCK/snail cells, they did exhibit reduced pyruvate dehydrogenase (PDH) activity, which controls pyruvate entry into the tricarboxylic acid (TCA) cycle. Indeed, the activity of multiple enzymes involved in the TCA cycle was decreased in MDCK/snail cells, including that of mitochondrial NADP{sup +}-dependent isocitrate dehydrogenase (IDH2), succinate dehydrogenase (SDH), and electron transport Complex II and Complex IV. Consequently, lower ATP content, lower oxygen consumption and increased survival under hypoxic conditions was also observed in MDCK/snail cells compared to MDCK/neo cells. In addition, the expression and promoter activity of pyruvate dehydrogenase kinase 1 (PDK1), which phosphorylates and inhibits the activity of PDH, was increased in MDCK/snail cells, while expression levels of glutaminase 2 (GLS2) and ATP-citrate lyase (ACLY), which are involved in glutaminolysis and fatty acid synthesis, were decreased in MDCK/snail cells. These results suggest that snail modulates cell metabolism by altering the expression and activity of

  18. Combination of lithium chloride and pEGFP-N1-BmK CT effectively decreases proliferation and migration of C6 glioma cells.

    Science.gov (United States)

    Fu, Yuejun; Jiao, Yanmei; Zheng, Shuhua; Liang, Aihua; Hu, Fengyun

    2016-03-01

    Deleterious invasiveness of glioma cells into the normal brain tissue is endorsed by its inherent ability to regulate the receptor-mediated adhesive properties, extracellular matrix degradation and remodeling and elevated secretory ability of metalloproteinase (MMPs) such as MMP-2. By doing so, it will create an intercellular space for the invasion of glioma cells. Here, we reported that combination of gene therapy Buthus martensii Karsch (BmK) CT, a type of scorpion toxin peptide, with lithium chloride (LiCl), clinically used as mood stabilizer, could inhibit the migration and invasion of C6 glioma cells. The results showed that concomitant administration of LiCl and pEGFP-N1-BmK CT on glioma cells would hamper pro-MMP2 secretion and in the meantime, inhibited its proliferation in a synergistic manner. These results try to extrapolate the potential interplay between the combined treatment of LiCl and BmK CT with signaling pathways β-catenin, MMP, GSK-3 in C6 glioma cells. This strategy can stand for a novel approach designated for the development of a new method for glioma therapy. PMID:25286828

  19. Super pharmacological levels of calcitriol (1,25-(OH)2D3) inhibits mineral deposition and decreases cell proliferation in a strain dependent manner in chicken mesenchymal stem cells undergoing osteogenic differentiation in vitro

    Science.gov (United States)

    Pande, Vivek V.; Chousalkar, Kapil C.; Bhanugopan, Marie S.; Quinn, Jane C.

    2015-01-01

    The biologically active form of vitamin D3, calcitriol (1,25-(OH)2D3), plays a key role in mineral homeostasis and bone formation and dietary vitamin D3 deficiency is a major cause of bone disorders in poultry. Supplementary dietary cholecalciferol (25-hydroxyvitamin D, 25-OH), the precursor of calcitriol, is commonly employed to combat this problem; however, dosage must be carefully determined as excess dietary vitamin D can cause toxicity resulting in a decrease in bone calcification, hypercalcinemia and renal failure. Despite much research on the therapeutic administration of dietary vitamin D in humans, the relative sensitivity of avian species to exogenous vitamin D has not been well defined. In order to determine the effects of exogenous 1,25-(OH)2D3 during avian osteogenesis, chicken bone marrow-derived mesenchymal stem cells (BM-MSCs) were exposed to varying doses of 1,25-(OH)2D3 during in vitro osteogenic differentiation and examined for markers of early proliferation and osteogenic induction. Similar to humans and other mammals, poultry BM-MSCs were found to be highly sensitive to exogenous 1,25-(OH)2D3 with super pharmacological levels exerting significant inhibition of mineralization and loss of cell proliferation in vitro. Strain related differences were apparent, with BM-MCSs derived from layers strains showing a higher level of sensitivity to 1,25-(OH)2D3 than those from broilers. These data suggest that understanding species and strain specific sensitivities to 1,25-(OH)2D3 is important for optimizing bone health in the poultry industry and that use of avian BM-MSCs are a useful tool for examining underlying effects of genetic variation in poultry. PMID:26500277

  20. A prebiotic, Celmanax™, decreases Escherichia coli O157:H7 colonization of bovine cells and feed-associated cytotoxicity in vitro

    Directory of Open Access Journals (Sweden)

    Juba Jean

    2011-04-01

    expressed Stx1 and Stx2, but more significantly, Stxs were also present in the blood clot blocking the jejunum. Mycotoxin analysis of the corn crop confirmed the presence of fumonisin, NIV, ZEAR, DON, 15-ADON, 3-ADON, NEO, DAS, HT-2 and T-2. Feed extracts were toxic to enterocytes and 0.1% Celmanax™ removed the cytotoxicity in vitro. There was no effect of Dairyman's Choice™ paste on feed-extract activity in vitro. Fumonisin, T-2, ZEAR and DON were toxic to bovine cells and 0.1% Celmanax™ removed the cytotoxicity in vitro. Celmanax™ also directly decreased E. coli O157:H7 colonization of mucosal explants and a colonic cell line in a dose-dependent manner. There was no effect of Dairyman's Choice™ paste on E. coli O157:H7 colonization in vitro. The inclusion of the prebiotic and probiotic in the feed was associated with a decline in disease. Conclusion The current study confirmed an association between mycotoxigenic fungi in the feed and the development of JHS in cattle. This association was further expanded to include mycotoxins in the feed and mixtures of STECs colonizing the severely hemorrhaged tissues. Future studies should examine the extent of involvement of the different STEC in the infection process. The prebiotic, Celmanax™, acted as an anti-adhesive for STEC colonization and a mycotoxin binder in vitro. Future studies should determine the extent of involvement of the prebiotic in altering disease.

  1. Expression of the sFLT1 gene in cord blood cells is associated to maternal arsenic exposure and decreased birth weight.

    Directory of Open Access Journals (Sweden)

    Sylvie Remy

    Full Text Available There is increasing epidemiologic evidence that arsenic exposure in utero is associated with adverse pregnancy outcomes and may contribute to long-term health effects. These effects may occur at low environmental exposures but the underlying molecular mechanism is not clear. We collected cord blood samples of 183 newborns to identify associations between arsenic levels and birth anthropometric parameters in an area with very low arsenic exposure. Our core research aim was to screen for transcriptional marks that mechanistically explain these associations. Multiple regression analyses showed that birth weight decreased with 47 g (95% CI: 16-78 g for an interquartile range increase of 0.99 μg/L arsenic. The model was adjusted for child's sex, maternal smoking during pregnancy, gestational age, and parity. Higher arsenic concentrations and reduced birth weight were positively associated with changes in expression of the sFLT1 (soluble fms-like tyrosine kinase-1 gene in cord blood cells in girls. The protein product of sFLT1 is a scavenger of vascular endothelial growth factor (VEGF in the extracellular environment and plays a key role in the inhibition of placental angiogenesis. In terms of fetal development, inhibition of placental angiogenesis leads to impaired nutrition and hence to growth retardation. Various genes related to DNA methylation and oxidative stress showed also changed expression in relation to arsenic exposure but were not related to birth outcome parameters. In conclusion, this study suggests that increased expression of sFLT1 is an intermediate marker that points to placental angiogenesis as a pathway linking prenatal arsenic exposure to reduced birth weight.

  2. Biotin Supplementation Decreases the Expression of the SERCA3 Gene (ATP2A3) in Jurkat Cells, thus Triggering Unfolded Protein Response*

    OpenAIRE

    Griffin, Jacob B.; Rodriguez-Melendez, Rocio; Dode, Leonard; Wuytack, Frank; Zempleni, Janos

    2005-01-01

    Protein folding in the endoplasmic reticulum (ER) depends on Ca2+; uptake of Ca2+ into the ER is mediated by SERCA3. The 5′-flanking region of the SERCA3 gene (ATP2A3) contains numerous binding sites for the transcription factors Sp1 and Sp3. Biotin affects the nuclear abundance of Sp1 and Sp3, which may act as transcriptional activators or repressors. Here we determined whether biotin affects the expression of the SERCA3 gene and, thus, protein folding in human lymphoid cells. Jurkat cells w...

  3. Cell Motility

    CERN Document Server

    Lenz, Peter

    2008-01-01

    Cell motility is a fascinating example of cell behavior which is fundamentally important to a number of biological and pathological processes. It is based on a complex self-organized mechano-chemical machine consisting of cytoskeletal filaments and molecular motors. In general, the cytoskeleton is responsible for the movement of the entire cell and for movements within the cell. The main challenge in the field of cell motility is to develop a complete physical description on how and why cells move. For this purpose new ways of modeling the properties of biological cells have to be found. This long term goal can only be achieved if new experimental techniques are developed to extract physical information from these living systems and if theoretical models are found which bridge the gap between molecular and mesoscopic length scales. Cell Motility gives an authoritative overview of the fundamental biological facts, theoretical models, and current experimental developments in this fascinating area.

  4. Secondhand smoke in combination with ambient air pollution exposure is associated with increased CpG methylation and decreased expression of IFN-gamma in T effector cells and Foxp3 in T regulatory cells in children

    OpenAIRE

    Kohli, Arunima; Garcia, Marco A; Miller, Rachel L.; Maher, Christina; Humblet, Olivier; Hammond, S; Nadeau, Kari

    2012-01-01

    Abstract Background Secondhand smoke (SHS) and ambient air pollution (AAP) exposures have been associated with increased prevalence and severity of asthma and DNA modifications of immune cells. In the current study, we examined the association between SHS and AAP with DNA methylation and expression of interferon-gamma (IFN-γ) and forkhead box protein 3 (Foxp3) in T cell populations. Methods Subjects 7–18 years old were recruited from Fresno (high AAP; n ...

  5. Retinoid- and sodium-butyrate– induced decrease in heat shock protein 70 membrane-positive tumor cells is associated with reduced sensitivity to natural killer cell lysis, growth delay, and altered growth morphology

    OpenAIRE

    Gehrmann, Mathias; Schönberger, Johann; Zilch, Tanja; Rossbacher, Lydia; Thonigs, Gerald; Eilles, Christoph; Multhoff, Gabriele

    2005-01-01

    Human tumors frequently present heat shock protein 70 (Hsp70) on their cell membranes, whereas corresponding normal tissues fail to do so. Therefore, an Hsp70 membrane-positive phenotype provided a tumor-specific marker. Moreover, membrane-bound Hsp70 provides a target structure for the cytolytic attack mediated by natural killer (NK) cells. Vitamin A derivatives 13-cis retinoic acid (13-RA) and all-trans retinoic acid (ATRA) and sodium-butyrate (SBU) are known for their redifferentiating cap...

  6. Human bronchial epithelial cells exposed in vitro to diesel exhaust particles exhibit alterations in cell rheology and cytotoxicity associated with decrease in antioxidant defenses and imbalance in pro- and anti-apoptotic gene expression.

    Science.gov (United States)

    Seriani, Robson; de Souza, Claudia Emanuele Carvalho; Krempel, Paloma Gava; Frias, Daniela Perroni; Matsuda, Monique; Correia, Aristides Tadeu; Ferreira, Márcia Zotti Justo; Alencar, Adriano Mesquita; Negri, Elnara Marcia; Saldiva, Paulo Hilário Nascimento; Mauad, Thais; Macchione, Mariangela

    2016-05-01

    Diesel exhaust particles (DEPs) from diesel engines produce adverse alterations in cells of the airways by activating intracellular signaling pathways and apoptotic gene overexpression, and also by influencing metabolism and cytoskeleton changes. This study used human bronchial epithelium cells (BEAS-2B) in culture and evaluates their exposure to DEPs (15ug/mL for 1 and 2 h) in order to determine changes to cell rheology (viscoelasticity) and gene expression of the enzymes involved in oxidative stress, apoptosis, and cytotoxicity. BEAS-2B cells exposed to DEPs were found to have a significant loss in stiffness, membrane stability, and mitochondrial activity. The genes involved in apoptosis [B cell lymphoma 2 (BCL-2 and caspase-3)] presented inversely proportional expressions (p = 0.05, p = 0.01, respectively), low expression of the genes involved in antioxidant responses [SOD1 (superoxide dismutase 1); SOD2 (superoxide dismutase 2), and GPx (glutathione peroxidase) (p = 0.01)], along with an increase in cytochrome P450, family 1, subfamily A, polypeptide 1 (CYP1A1) (p = 0.01). These results suggest that alterations in cell rheology and cytotoxicity could be associated with oxidative stress and imbalance between pro- and anti-apoptotic genes. PMID:26856867

  7. Holy Basil Leaf Extract Decreases Tumorigenicity and Metastasis of Aggressive Human Pancreatic Cancer Cells in vitro and in vivo: Potential Role in Therapy

    Science.gov (United States)

    Shimizu, Tomohiro; Torres, María P.; Chakraborty, Subhankar; Souchek, Joshua J.; Rachagani, Satyanarayana; Kaur, Sukhwinder; Macha, Muzafar; Ganti, Apar K.; Hauke, Ralph J; Batra, Surinder K.

    2013-01-01

    There is an urgent need to develop alternative therapies against lethal pancreatic cancer (PC). Ocimum sanctum (“Holy Basil”) has been used for thousands of years in traditional Indian medicine, but its anti-tumorigenic effect remains largely unexplored. Here, we show that extracts of O. sanctum leaves inhibit the proliferation, migration, invasion, and induce apoptosis of PC cells in vitro. The expression of genes that promote the proliferation, migration and invasion of PC cells including activated ERK-1/2, FAK, and p65 (subunit of NF-κB), was downregulated in PC cells after O. sanctum treatment. Intraperitoneal injections of the aqueous extract significantly inhibited the growth of orthotopically transplanted PC cells in vivo (p<0.05). Genes that inhibit metastasis (E-cadherin) and induce apoptosis (BAD) were significantly upregulated in tumors isolated from mice treated with O. sanctum extracts, while genes that promote survival (Bcl-2 and Bcl-xL) and chemo/radiation resistance (AURKA, Chk1 and Survivin) were downregulated. Overall, our study suggests that leaves of O. sanctum could be a potential source of novel anticancer compounds in the future. PMID:23523869

  8. Inhibitory effect of berberine on the invasion of human lung cancer cells via decreased productions of urokinase-plasminogen activator and matrix metalloproteinase-2

    International Nuclear Information System (INIS)

    Berberine, a compound isolated from medicinal herbs, has been reported with many pharmacological effects related to anti-cancer and anti-inflammation capabilities. In this study, we observed that berberine exerted a dose- and time-dependent inhibitory effect on the motility and invasion ability of a highly metastatic A549 cells under non-cytotoxic concentrations. In cancer cell migration and invasion process, matrix-degrading proteinases are required. A549 cell treated with berberine at various concentrations showed reduced ECM proteinases including matrix metalloproteinase-2 (MMP2) and urokinase-plasminogen activator (u-PA) by gelatin and casein zymography analysis. The inhibitory effect is likely to be at the transcriptional level, since the reduction in the transcripts levels was corresponding to the proteins. Moreover, berberine also exerted its action via regulating tissue inhibitor of metalloproteinase-2 (TIMP-2) and urokinase-plasminogen activator inhibitor (PAI). The upstream mediators of the effect involved c-jun, c-fos and NF-κB, as evidenced by reduced phosphorylation of the proteins. These findings suggest that berberine possesses an anti-metastatic effect in non-small lung cancer cell and may, therefore, be helpful in clinical treatment

  9. Interference RNA (RNAi)-based silencing of endogenous thrombopoietin receptor (Mpl) in Dami cells resulted in decreased hNUDC-mediated megakaryocyte proliferation and differentiation

    International Nuclear Information System (INIS)

    Recently our laboratory reported evidence showing that hNUDC acts as an additional cytokine for thrombopoietin receptor (Mpl). Previously known as the human homolog of a fungal nuclear migration protein, hNUDC plays a critical role in megakaryocyte differentiation and maturation. Here we sought to further clarify the hNUDC-Mpl ligand-receptor relationship by utilizing interference RNA (RNAi) to knockdown Mpl expression in a megakaryocyte cell line. We created U6 promoter driven constructs to express short hairpin RNAs (shRNA) with affinity for different sites on Mpl mRNA. By including Mpl-EGFP fusion protein in these constructs, we were able to effectively screen the shRNA that was most efficient in inhibiting Mpl mRNA expression. This shRNA was subsequently transferred into a lentivirus vector and transduced into Dami cells, a cell line which constitutively expresses endogenous Mpl. This lentiviral vector was also designed to simultaneously express EGFP to monitor transfection efficiency. Our results show that lentivirus can be used to effectively deliver shRNAs into Dami cells and cause specific inhibition of Mpl protein expression after transduction. Furthermore, we show the functional effects of shRNA-mediated Mpl silencing by demonstrating reduced hNUDC stimulated megakaryocyte proliferation and differentiation. Thus, the use of a RNAi knockdown strategy has allowed us to pinpoint the connection of hNUDC with Mpl in the regulation of megakaryocyte maturation.

  10. Bovine viral diarrhea virus type 2 in vivo infection modulates TLR4 responsiveness in differentiated Myeloid cells which is associated with decreased MyD88 expression

    Science.gov (United States)

    Bovine viral diarrhea virus (BVDV) causes clinical signs in cattle ranging from mild to severe acute infection which can lead to increased susceptibility to secondary bacteria. In this study we examined the effects of BVDV genotype 2 (BVDV2) infection on the ability of myeloid lineage cells derived...

  11. Mechanical stimulation of cyclic tensile strain induces reduction of pluripotent related gene expressions via activation of Rho/ROCK and subsequent decreasing of AKT phosphorylation in human induced pluripotent stem cells

    Energy Technology Data Exchange (ETDEWEB)

    Teramura, Takeshi, E-mail: teramura@med.kindai.ac.jp [Institute of Advanced Clinical Medicine, Kinki University, Faculty of Medicine, Osaka (Japan); Takehara, Toshiyuki; Onodera, Yuta [Institute of Advanced Clinical Medicine, Kinki University, Faculty of Medicine, Osaka (Japan); Nakagawa, Koichi; Hamanishi, Chiaki [Department of Orthopaedic Surgery, Kinki University, Faculty of Medicine, Osaka (Japan); Fukuda, Kanji [Institute of Advanced Clinical Medicine, Kinki University, Faculty of Medicine, Osaka (Japan); Department of Orthopaedic Surgery, Kinki University, Faculty of Medicine, Osaka (Japan)

    2012-01-13

    Highlights: Black-Right-Pointing-Pointer Mechanical stimulation is an important factor for regulation of stem cell fate. Black-Right-Pointing-Pointer Cyclic stretch to human induced pluripotent stem cells activated small GTPase Rho. Black-Right-Pointing-Pointer Rho-kinase activation attenuated pluripotency via inhibition of AKT activation. Black-Right-Pointing-Pointer This reaction could be reproduced only by transfection of dominant active Rho. Black-Right-Pointing-Pointer Rho/ROCK are important molecules in mechanotransduction and control of stemness. -- Abstract: Mechanical stimulation has been shown to regulate the proliferation and differentiation of stem cells. However, the effects of the mechanical stress on the stemness or related molecular mechanisms have not been well determined. Pluripotent stem cells such as embryonic stem (ES) cells and induced pluripotent stem (iPS) cells are used as good materials for cell transplantation therapy and research of mammalian development, since they can self-renew infinitely and differentiate into various cell lineages. Here we demonstrated that the mechanical stimulation to human iPS cells altered alignment of actin fibers and expressions of the pluripotent related genes Nanog, POU5f1 and Sox2. In the mechanically stimulated iPS cells, small GTPase Rho was activated and interestingly, AKT phosphorylation was decreased. Inhibition of Rho-associated kinase ROCK recovered the AKT phosphorylation and the gene expressions. These results clearly suggested that the Rho/ROCK is a potent primary effector of mechanical stress in the pluripotent stem cells and it participates to pluripotency-related signaling cascades as an upper stream regulator.

  12. Live-cell luciferase assay of drug resistant cells

    OpenAIRE

    sprotocols

    2015-01-01

    To date, multiplexing cell-based assay is essential for high-throughput screening of molecular targets. Measuring multiple parameters of a single sample increases consistency and decrease time and cost of assay. Functional assay of living cell is useful as a first step of multiplexing assay, because live-cell assay allows following second assay using cell lysate or stained cell. However, live-cell assay of drug resistant cells that are highly activated of drug efflux mechanisms is sometimes u...

  13. Decreased survival of mosquito cells after stable transfection with a Drosophila ecdysteroid response element: Possible involvement of a 40 kDa DNA binding protein

    Directory of Open Access Journals (Sweden)

    Gitanjali Jayachandran

    2002-11-01

    Full Text Available Homologous transfection systems provide a useful tool for characterizing promoters and other regulatory elements from cloned genes. We have used cultured Aedes albopictus C7-10 mosquito cells to evaluate expression of 20-hydroxyecdysone-inducible genes. Although this cell line has previously been shown to synthesize components of the ecdysteroid receptor and ecdysone-inducible proteins, the well-characterized ecdysteroid response element (EcRE from the Drosophila hsp27 promoter failed to confer a substantial 20-hydroxyecdysone mediated induction in transfected mosquito cells. Recovery of stably transformed clones was also reduced in a DNA dependent manner when the EcREs were in the sense orientation, relative to control plasmids lacking the EcREs or containing an antisense construct. Finally, when tandem EcREs were placed within the hsp70 promoter, CAT activity was detected only after prolonged enzyme incubation, suggesting that the DNA interfered with cellular metabolism. In these constructs, we noted that the promoter DNA contained several potential binding sites for the activator protein-1 (AP-1 transcription factor, one of which lay between the tandem EcREs. On southwestern blots, a 40 kDa nuclear protein from C7-10 cells bound to DNA containing AP-1 sites. A DNA affinity column was used to partially purify the 40 kDa protein, and western analysis showed that the mosquito protein cross-reacted with a heterologous antibody to JUN. Likewise, mRNA from C7-10 cells cross-hybridized with the jun cDNA from Drosophila. These results suggest that like estrogen, 20-hydroxyecdysone interfaces with AP-1 as a co-activator protein that modulates the overall hormone response.

  14. Solar cells

    Science.gov (United States)

    Cuquel, A.; Roussel, M.

    The physical and electronic characteristics of solar cells are discussed in terms of space applications. The principles underlying the photovoltaic effect are reviewed, including an analytic model for predicting the performance of individual cells and arrays of cells. Attention is given to the effects of electromagnetic and ionizing radiation, micrometeors, thermal and mechanical stresses, pollution and degassing encountered in space. The responses of different types of solar cells to the various performance-degrading agents are examined, with emphasis on techniques for quality assurance in the manufacture and mounting of Si cells.

  15. Nestin(+) cells direct inflammatory cell migration in atherosclerosis.

    Science.gov (United States)

    Del Toro, Raquel; Chèvre, Raphael; Rodríguez, Cristina; Ordóñez, Antonio; Martínez-González, José; Andrés, Vicente; Méndez-Ferrer, Simón

    2016-01-01

    Atherosclerosis is a leading death cause. Endothelial and smooth muscle cells participate in atherogenesis, but it is unclear whether other mesenchymal cells contribute to this process. Bone marrow (BM) nestin(+) cells cooperate with endothelial cells in directing monocyte egress to bloodstream in response to infections. However, it remains unknown whether nestin(+) cells regulate