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Sample records for hexarelin increases cell

  1. Hexarelin Protects Rodent Pancreatic Β-Cells Function from Cytotoxic Effects of Streptozotocin Involving Mitochondrial Signalling Pathways In Vivo and In Vitro.

    Directory of Open Access Journals (Sweden)

    Yan Zhao

    Full Text Available Mitochondrial functions are crucial for pancreatic β-cell survival and glucose-induced insulin secretion. Hexarelin (Hex is a synthetic small peptide ghrelin analogue, which has been shown to protect cardiomyocytes from the ischemia-reperfusion process. In this study, we used in vitro and in vivo models of streptozotocin (STZ-induced β-cell damage to study the protective effect of Hex and the associated mechanisms. We found that STZ produced a cytotoxic effect in a dose- and time-dependent manner in MIN6 cells (a mouse β-cell line. Hex (1.0 μM decreased the STZ-induced damage in β-cells. Rhodamine 123 assay and superoxide DHE production assay revealed that Hex ameliorated STZ-induced mitochondrial damage and excessive superoxide activity in β-cells. In addition, Hex significantly reduced STZ-induced expression of cleaved Caspases-3, Caspases-9 and the ratio of pro-apoptotic protein Bax to anti-apoptotic protein Bcl-2 in MIN6 cells. We further examined the in vivo effect of Hex in a rat model of type 1 diabetes induced by STZ injection. Hex ameliorated STZ-induced decrease in plasma insulin and protected the structure of islets from STZ-induced disruption. Hex also ameliorated STZ-induced expression of cleaved Caspase-9 and the Bax in β-cells. In conclusion, our data indicate that Hex is able to protects β-cell mass from STZ-caused cytotoxic effects involving mitochondrial pathways in vitro and in vivo. Hex may serve as a potential protective agent for the management of diabetes.

  2. Increased voltage photovoltaic cell

    Science.gov (United States)

    Ross, B.; Bickler, D. B.; Gallagher, B. D. (Inventor)

    1985-01-01

    A photovoltaic cell, such as a solar cell, is provided which has a higher output voltage than prior cells. The improved cell includes a substrate of doped silicon, a first layer of silicon disposed on the substrate and having opposite doping, and a second layer of silicon carbide disposed on the first layer. The silicon carbide preferably has the same type of doping as the first layer.

  3. Increasing the lifetime of fuel cell catalysts

    NARCIS (Netherlands)

    Latsuzbaia, R.

    2015-01-01

    In this thesis, I discuss a novel idea of fuel cell catalyst regeneration to increase lifetime of the PEM fuel cell electrode/catalyst operation and, therefore, reduce the catalyst costs. As many of the catalyst degradation mechanisms are difficult to avoid, the regeneration is alternative option to

  4. Nanostructured magnesium increases bone cell density.

    Science.gov (United States)

    Weng, Lucy; Webster, Thomas J

    2012-12-07

    Magnesium has attracted some attention in orthopedics due to its biodegradability and mechanical properties. Since magnesium is an essential natural mineral for bone growth, it can be expected that as a biomaterial, it would support bone formation. However, upon degradation in the body, magnesium releases OH(-) which results in an alkaline pH that can be detrimental to cell density (for example, osteoblasts or bone forming cells). For this reason, modification of magnesium may be necessary to compensate for such detrimental effects to cells. This study created biologically inspired nanoscale surface features on magnesium by soaking magnesium in various concentrations of NaOH (from 1 to 10 N) and for various periods of time (from 10 to 30 min). The results provided the first evidence of increased roughness, surface energy, and consequently greater osteoblast adhesion, after 4 h as well as density up to 7 days on magnesium treated with any concentration of NaOH for any length of time compared to untreated controls. For these reasons, this study suggests that soaking magnesium in NaOH could be an inexpensive, simple and effective manner to promote osteoblast functions for numerous orthopedic applications and, thus, should be further studied.

  5. Nanostructured magnesium increases bone cell density

    International Nuclear Information System (INIS)

    Weng, Lucy; Webster, Thomas J

    2012-01-01

    Magnesium has attracted some attention in orthopedics due to its biodegradability and mechanical properties. Since magnesium is an essential natural mineral for bone growth, it can be expected that as a biomaterial, it would support bone formation. However, upon degradation in the body, magnesium releases OH − which results in an alkaline pH that can be detrimental to cell density (for example, osteoblasts or bone forming cells). For this reason, modification of magnesium may be necessary to compensate for such detrimental effects to cells. This study created biologically inspired nanoscale surface features on magnesium by soaking magnesium in various concentrations of NaOH (from 1 to 10 N) and for various periods of time (from 10 to 30 min). The results provided the first evidence of increased roughness, surface energy, and consequently greater osteoblast adhesion, after 4 h as well as density up to 7 days on magnesium treated with any concentration of NaOH for any length of time compared to untreated controls. For these reasons, this study suggests that soaking magnesium in NaOH could be an inexpensive, simple and effective manner to promote osteoblast functions for numerous orthopedic applications and, thus, should be further studied. (paper)

  6. Cigarette smoking increases white blood cell aggregation in whole blood.

    OpenAIRE

    Bridges, A B; Hill, A; Belch, J J

    1993-01-01

    We studied the effect of chronic cigarette smoking on white blood cell aggregation, increased aggregation predisposes to microvascular occlusion and damage. Current smokers had significantly increased white blood cell aggregation when compared with non smokers. The presence of chronically activated white blood cells in current smokers may be relevant in the pathogenesis of ischaemic vascular disease.

  7. Increasing Stem Cell Dose Promotes Posttransplant Immune Reconstitution.

    Science.gov (United States)

    Xu, Ning; Shen, Sylvie; Dolnikov, Alla

    2017-04-01

    Umbilical cord blood (UCB) transplantation can provide a successful therapeutic option for patients that have no suitable related donor. UCB transplantation is often limited by the relatively small hematopoietic stem cell (HSC) numbers in UCB especially for adult recipients. Early neutrophil and platelet engraftment correlates with the stem cell numbers in UCB transplant. Compared to other HSC sources, immune reconstitution following UCB transplant is slower and complicated by increased frequency of opportunistic infections. The effect of HSC numbers in UCB transplant on immune reconstitution was not thoroughly examined. Using immunocompromised mice transplanted with purified UCB CD34+ stem cells, we have demonstrated that increasing the numbers of CD34+ cells in the transplant promotes hematopoietic and immune reconstitution. At early stages posttransplant, high stem cell dose generated relatively more B cells, while lower dose generated more myeloid and T cells. Thus, the size of the stem cell graft appears to modulate the differentiation potential of infused stem cells. In addition, increasing stem cell dose in the transplant improved CD8+ T cell development and delayed late memory T cell skewing in expense of naive T cells highlighting the importance of HSC dose to maintain the pool of naive T cells able to develop strong immune responses. Transplantation of ex vivo expanded CD34+ cells did not promote, but rather delayed immune reconstitution suggesting the loss of primitive lymphoid precursor cells during ex vivo expansion.

  8. Invasive Glioblastoma Cells Acquire Stemness and Increased Akt Activation

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    Jennifer R. Molina

    2010-06-01

    Full Text Available Glioblastoma multiforme (GBM is the most frequent and most aggressive brain tumor in adults. The dismal prognosis is due to postsurgery recurrences arising from escaped invasive tumor cells. The signaling pathways activated in invasive cells are under investigation, and models are currently designed in search for therapeutic targets. We developed here an in vivo model of human invasive GBM in mouse brain from a GBM cell line with moderate tumorigenicity that allowed simultaneous primary tumor growth and dispersal of tumor cells in the brain parenchyma. This strategy allowed for the first time the isolation and characterization of matched sets of tumor mass (Core and invasive (Inv cells. Both cell populations, but more markedly Inv cells, acquired stem cell markers, neurosphere renewal ability, and resistance to rapamycin-induced apoptosis relative to parental cells. The comparative phenotypic analysis between Inv and Core cells showed significantly increased tumorigenicity in vivo and increased invasion with decreased proliferation in vitro for Inv cells. Examination of a large array of signaling pathways revealed extracellular signal-regulated kinase (Erk down-modulation and Akt activation in Inv cells and an opposite profile in Core cells. Akt activation correlated with the increased tumorigenicity, stemness, and invasiveness, whereas Erk activation correlated with the proliferation of the cells. These results underscore complementary roles of the Erk and Akt pathways for GBM proliferation and dispersal and raise important implications for a concurrent inhibitory therapy.

  9. Increased DNA-repair in spleen cells of M. Hodgkin

    International Nuclear Information System (INIS)

    Frischauf, H.; Neumann, E.; Howanietz, L.; Dolejs, I.; Tuschl, H.; Altmann, H.

    1974-11-01

    In spleen cells of control patients and cells of Morbus Hodgkin, DNA-repair after gamma- and UV-irradiation was determined measuring the incorporated 3H-thymidine activity in the DNA. Additionally, the ratio of labeled cells compared to non-labeled cells and the grains per cell were evaluated by autoradiographic investigations. DNA-content per cell was measured using pulsecytophotometry. A significant increase of DNA-repair capacity after gamma-irradiation was found by density gradient centrifugation in alkaline sucrose. The same trend could be shown by investigations of unscheduled DNA-synthesis using autoradiographic method. (author)

  10. Increased radiation resistance in lithium-counterdoped silicon solar cells

    Science.gov (United States)

    Weinberg, I.; Swartz, C. K.; Mehta, S.

    1984-01-01

    Lithium-counterdoped n(+)p silicon solar cells are found to exhibit significantly increased radiation resistance to 1-MeV electron irradiation when compared to boron-doped n(+)p silicon solar cells. In addition to improved radiation resistance, considerable damage recovery by annealing is observed in the counterdoped cells at T less than or equal to 100 C. Deep level transient spectroscopy measurements are used to identify the defect whose removal results in the low-temperature aneal. It is suggested that the increased radiation resistance of the counterdoped cells is primarily due to interaction of the lithium with interstitial oxygen.

  11. Ouabain Increases Gap Junctional Communication in Epithelial Cells

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    Arturo Ponce

    2014-11-01

    Full Text Available Background/Aims: The finding that endogenous ouabain acts as a hormone prompted efforts to elucidate its physiological function. In previous studies, we have shown that 10 nM ouabain (i.e., a concentration within the physiological range modulates cell-cell contacts such as tight junctions and apical/basolateral polarity. In this study, we examined whether 10 nM ouabain affects another important cell-cell feature: gap junction communication (GJC. Methods: We employed two different approaches: 1 analysis of the cell-to-cell diffusion of neurobiotin injected into a particular MDCK cell (epithelial cells from dog kidneys in a confluent monolayer by counting the number of neighboring cells reached by the probe and 2 measurement of the electrical capacitance. Results: We found that 10 nM ouabain increase GJC by 475% within 1 hour. The Na+-K+-ATPase acts as a receptor of ouabain. In previous works we have shown that ouabain activates c-Src and ERK1/2 in 1 hour; in the present study we show that the inhibition of these proteins block the effect of ouabain on GJC. This increase in GJC does not require synthesis of new protein components, because the inhibitors cycloheximide and actinomycin D did not affect this phenomenon. Using silencing assays we also demonstrate that this ouabain-induced enhancement of GJC involves connexins 32 and 43. Conclusion: Ouabain 10 nM increases GJC in MDCK cells.

  12. DA-6034 Induces [Ca(2+)]i Increase in Epithelial Cells.

    Science.gov (United States)

    Yang, Yu-Mi; Park, Soonhong; Ji, Hyewon; Kim, Tae-Im; Kim, Eung Kweon; Kang, Kyung Koo; Shin, Dong Min

    2014-04-01

    DA-6034, a eupatilin derivative of flavonoid, has shown potent effects on the protection of gastric mucosa and induced the increases in fluid and glycoprotein secretion in human and rat corneal and conjunctival cells, suggesting that it might be considered as a drug for the treatment of dry eye. However, whether DA-6034 induces Ca(2+) signaling and its underlying mechanism in epithelial cells are not known. In the present study, we investigated the mechanism for actions of DA-6034 in Ca(2+) signaling pathways of the epithelial cells (conjunctival and corneal cells) from human donor eyes and mouse salivary gland epithelial cells. DA-6034 activated Ca(2+)-activated Cl(-) channels (CaCCs) and increased intracellular calcium concentrations ([Ca(2+)]i) in primary cultured human conjunctival cells. DA-6034 also increased [Ca(2+)]i in mouse salivary gland cells and human corneal epithelial cells. [Ca(2+)]i increase of DA-6034 was dependent on the Ca(2+) entry from extracellular and Ca(2+) release from internal Ca(2+) stores. Interestingly, these effects of DA-6034 were related to ryanodine receptors (RyRs) but not phospholipase C/inositol 1,4,5-triphosphate (IP3) pathway and lysosomal Ca(2+) stores. These results suggest that DA-6034 induces Ca(2+) signaling via extracellular Ca(2+) entry and RyRs-sensitive Ca(2+) release from internal Ca(2+) stores in epithelial cells.

  13. Increased hydrostatic pressure enhances motility of lung cancer cells.

    Science.gov (United States)

    Kao, Yu-Chiu; Lee, Chau-Hwang; Kuo, Po-Ling

    2014-01-01

    Interstitial fluid pressures within most solid tumors are significantly higher than that in the surrounding normal tissues. Therefore, cancer cells must proliferate and migrate under the influence of elevated hydrostatic pressure while a tumor grows. In this study, we developed a pressurized cell culture device and investigated the influence of hydrostatic pressure on the migration speeds of lung cancer cells (CL1-5 and A549). The migration speeds of lung cancer cells were increased by 50-60% under a 20 mmHg hydrostatic pressure. We also observed that the expressions of aquaporin in CL1-5 and A549 cells were increased under the hydrostatic pressure. Our preliminary results indicate that increased hydrostatic pressure plays an important role in tumor metastasis.

  14. Targeting development of incretin-producing cells increases insulin secretion

    DEFF Research Database (Denmark)

    Petersen, Natalia; Reimann, Frank; van Es, Johan H

    2015-01-01

    the number of intestinal L cells, which produce GLP-1, is an alternative strategy to augment insulin responses and improve glucose tolerance. Blocking the NOTCH signaling pathway with the γ-secretase inhibitor dibenzazepine increased the number of L cells in intestinal organoid-based mouse and human culture...... of the development of incretin-producing cells in the intestine has potential as a therapeutic strategy to improve glycemic control....

  15. Tandem photovoltaic solar cells and increased solar energy conversion efficiency

    Science.gov (United States)

    Loferski, J. J.

    1976-01-01

    Tandem photovoltaic cells, as proposed by Jackson (1955) to increase the efficiency of solar energy conversion, involve the construction of a system of stacked p/n homojunction photovoltaic cells composed of different semiconductors. It had been pointed out by critics, however, that the total power which could be extracted from the cells in the stack placed side by side was substantially greater than the power obtained from the stacked cells. A reexamination of the tandem cell concept in view of the development of the past few years is conducted. It is concluded that the use of tandem cell systems in flat plate collectors, as originally envisioned by Jackson, may yet become feasible as a result of the development of economically acceptable solar cells for large scale terrestrial power generation.

  16. Transgenic cells with increased plastoquinone levels and methods of use

    Energy Technology Data Exchange (ETDEWEB)

    Sayre, Richard T.; Subramanian, Sowmya; Cahoon, Edgar

    2016-12-27

    Disclosed herein are transgenic cells expressing a heterologous nucleic acid encoding a prephenate dehydrogenase (PDH) protein, a heterologous nucleic acid encoding a homogentisate solanesyl transferase (HST) protein, a heterologous nucleic acid encoding a deoxyxylulose phosphate synthase (DXS) protein, or a combination of two or more thereof. In particular examples, the disclosed transgenic cells have increased plastoquinone levels. Also disclosed are methods of increasing cell growth rates or production of biomass by cultivating transgenic cells expressing a heterologous nucleic acid encoding a PDH protein, a heterologous nucleic acid encoding an HST protein, a heterologous nucleic acid encoding a DXS protein, or a combination of two or more thereof under conditions sufficient to produce cell growth or biomass.

  17. Defective TFH Cell Function and Increased TFR Cells Contribute to Defective Antibody Production in Aging.

    Science.gov (United States)

    Sage, Peter T; Tan, Catherine L; Freeman, Gordon J; Haigis, Marcia; Sharpe, Arlene H

    2015-07-14

    Defective antibody production in aging is broadly attributed to immunosenescence. However, the precise immunological mechanisms remain unclear. Here, we demonstrate an increase in the ratio of inhibitory T follicular regulatory (TFR) cells to stimulatory T follicular helper (TFH) cells in aged mice. Aged TFH and TFR cells are phenotypically distinct from those in young mice, exhibiting increased programmed cell death protein-1 expression but decreased ICOS expression. Aged TFH cells exhibit defective antigen-specific responses, and programmed cell death protein-ligand 1 blockade can partially rescue TFH cell function. In contrast, young and aged TFR cells have similar suppressive capacity on a per-cell basis in vitro and in vivo. Together, these studies reveal mechanisms contributing to defective humoral immunity in aging: an increase in suppressive TFR cells combined with impaired function of aged TFH cells results in reduced T-cell-dependent antibody responses in aged mice. Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.

  18. Increasing the efficiency of polymer solar cells by silicon nanowires

    Energy Technology Data Exchange (ETDEWEB)

    Eisenhawer, B; Sivakov, V; Pietsch, M; Andrae, G; Falk, F [Institute of Photonic Technology, Albert-Einstein-Strasse 9, 07743 Jena (Germany); Sensfuss, S, E-mail: bjoern.eisenhawer@ipht-jena.de [Thuringian Institute for Textile and Plastics Research, Breitscheidstrasse 97, 07407 Rudolstadt (Germany)

    2011-08-05

    Silicon nanowires have been introduced into P3HT:[60]PCBM solar cells, resulting in hybrid organic/inorganic solar cells. A cell efficiency of 4.2% has been achieved, which is a relative improvement of 10% compared to a reference cell produced without nanowires. This increase in cell performance is possibly due to an enhancement of the electron transport properties imposed by the silicon nanowires. In this paper, we present a novel approach for introducing the nanowires by mixing them into the polymer blend and subsequently coating the polymer/nanowire blend onto a substrate. This new onset may represent a viable pathway to producing nanowire-enhanced polymer solar cells in a reel to reel process.

  19. Increasing the efficiency of polymer solar cells by silicon nanowires

    International Nuclear Information System (INIS)

    Eisenhawer, B; Sivakov, V; Pietsch, M; Andrae, G; Falk, F; Sensfuss, S

    2011-01-01

    Silicon nanowires have been introduced into P3HT:[60]PCBM solar cells, resulting in hybrid organic/inorganic solar cells. A cell efficiency of 4.2% has been achieved, which is a relative improvement of 10% compared to a reference cell produced without nanowires. This increase in cell performance is possibly due to an enhancement of the electron transport properties imposed by the silicon nanowires. In this paper, we present a novel approach for introducing the nanowires by mixing them into the polymer blend and subsequently coating the polymer/nanowire blend onto a substrate. This new onset may represent a viable pathway to producing nanowire-enhanced polymer solar cells in a reel to reel process.

  20. Autophagy involved in resveratrol increased radiosensitivity in glioma stem cells

    International Nuclear Information System (INIS)

    Long Linmei; Zhang Qingqing; Yang Neng; Ji Wenjun; Song Yunzhen; Zhao Jianghu; Liang Zhongqin

    2012-01-01

    Objective: To investigate the effect of Resveratrol combined with X-ray on radiosensitivity in glioma stem cells. Methods: The proliferation inhibition of glioma stem cells induced by X-rays and Resveratrol was assessed with MTT assay. The activation of proapoptotic effect was characterized by Hoechst 33258 stain. MDC stain and Western blot analysis were used to analyze the autophagy mechanism in X-rays-induced death of glioma stem cells. Results: MTT assay indicated that X-rays and Resveratrol decreased the viability of glioma stem cells (P<0.05); we found the proliferative inhibition of glioma stem cells was declined when we used 3-MA to inhibit autophagy(P<0.05). When the cells were treated by the Resveratrol and x-rays, their spherical shape were changed. Apoptosis was induced in glioma stem cells by combined X-rays and Resveratrol as detected by Hoechst 33258 staining. In addition, autophagy was induced in glioma stem cells in the combined treatment group as detected by MDC staining. Western blotting showed that Bcl-2 expression was decreased. in the combined treatment group (P<0.01), and the LC3-Ⅱ expression was increased in the combined treatment group (P<0.01). Conclusion: Resveratrol can increased the radiation sensitivity of glioma stem cells, the apoptosis and autophagy was induced in the glioma stem cells in the combined treatment X-rays and Resveratrol. Our results suggest that autophagy plays an essential role in the regulation of radiosensitization of glioma stem cells. (authors)

  1. UV-enhanced reactivation in mammalian cells: increase by caffeine

    International Nuclear Information System (INIS)

    Lytle, C.D.; Iacangelo, A.L.; Lin, C.H.; Goddard, J.G.

    1981-01-01

    It has been reported that caffeine decreases UV-enhanced reactivation of UV-irradiated Herpes simplex virus in CV-l monkey kidney cells. That occurred when there was no delay between cell irradiation and virus infection. In the present study, virus infection was delayed following cell irradiation to allow an 'induction' period separate from the 'expression' period which occurs during the virus infection. Thus, the effects of caffeine on 'induction' and 'expression' could be determined separately. Caffeine increased the expression of UV-enhanced reactivation, while causing a small decrease in the 'induction' of enhanced reactivation. (author)

  2. EDA-containing fibronectin increases proliferation of embryonic stem cells.

    Directory of Open Access Journals (Sweden)

    Noelia Losino

    Full Text Available Embryonic stem cells (ESC need a set of specific factors to be propagated. They can also grow in conditioned medium (CM derived from a bovine granulosa cell line BGC (BGC-CM, a medium that not only preserves their main features but also increases ESC´s proliferation rate. The mitogenic properties of this medium were previously reported, ascribing this effect to an alternative spliced generated fibronectin isoform that contains the extra domain A (FN EDA(+. Here, we investigated if the FN EDA(+ isoform increased proliferation of mouse and human ES cells. We analyzed cell proliferation using conditioned media produced by different mouse embryonic fibroblast (MEF lines genetically engineered to express FN constitutively including or excluding the EDA domain (FN EDA(-, and in media supplemented with recombinant peptides containing or not the EDA. We found that the presence of EDA in the medium increased mouse and human ESC's proliferation rate. Here we showed for the first time that this FN isoform enhances ESC's proliferation. These findings suggest a possible conserved behavior for regulation of ES cells proliferation by this FN isoform and could contribute to improve their culturing conditions both for research and cell therapy.

  3. Bactericidal Antibiotics Increase Hydroxyphenyl Fluorescein Signal by Altering Cell Morphology

    DEFF Research Database (Denmark)

    Paulander, Wilhelm; Wang, Ying; Folkesson, Sven Anders

    2014-01-01

    It was recently proposed that for bactericidal antibiotics a common killing mechanism contributes to lethality involving indirect stimulation of hydroxyl radical (OH center dot) formation. Flow cytometric detection of OH center dot by hydroxyphenyl fluorescein (HPF) probe oxidation was used...... to support this hypothesis. Here we show that increased HPF signals in antibiotics-exposed bacterial cells are explained by fluorescence associated with increased cell size, and do not reflect reactive oxygen species (ROS) concentration. Independently of antibiotics, increased fluorescence was seen...... for elongated cells expressing the oxidative insensitive green fluorescent protein (GFP). Although our data question the role of ROS in lethality of antibiotics other research approaches point to important interplays between basic bacterial metabolism and antibiotic susceptibility. To underpin...

  4. Veratridine increases the survival of retinal ganglion cells in vitro

    Directory of Open Access Journals (Sweden)

    S.P.F. Pereira

    1997-12-01

    Full Text Available Neuronal cell death is an important phenomenon involving many biochemical pathways. This degenerative event has been studied to understand how the cells activate the mechanisms that lead to self-destruction. Target cells and afferent cells play a relevant role in the regulation of natural cell death. We studied the effect of veratridine (1.5, 3.0, 4.5 and 6.0 µM on the survival of neonatal rat retinal ganglion cells in vitro. Veratridine (3.0 µM, a well-known depolarizing agent that opens the Na+ channel, promoted a two-fold increase in the survival of retinal ganglion cells kept in culture for 48 h. This effect was dose-dependent and was blocked by 1.0 µM tetrodotoxin (a classical voltage-dependent Na+ channel blocker and 30.0 µM flunarizine (a Na+ and Ca2+ channel blocker. These results indicate that electrical activity is also important for the maintenance of retinal ganglion cell survival in vitro

  5. Adipose-Derived Stem Cells Respond to Increased Osmolarities.

    Directory of Open Access Journals (Sweden)

    Urška Potočar

    Full Text Available Cell therapies present a feasible option for the treatment of degenerated cartilaginous and intervertebral disc (IVD tissues. Microenvironments of these tissues are specific and often differ from the microenvironment of cells that, could be potentially used for therapy, e.g. human adipose-derived stem cells (hASC. To ensure safe and efficient implantation of hASC, it is important to evaluate how microenvironmental conditions at the site of implantation affect the implanted cells. This study has demonstrated that cartilaginous tissue-specific osmolarities ranging from 400-600 mOsm/L affected hASC in a dose- and time-dependent fashion in comparison to 300 mOsm/L. Increased osmolarities resulted in transient (nuclear DNA and actin reorganisation and non-transient, long-term morphological changes (vesicle formation, increase in cell area, and culture morphology, as well as reduced proliferation in monolayer cultures. Increased osmolarities diminished acid proteoglycan production and compactness of chondrogenically induced pellet cultures, indicating decreased chondrogenic potential. Viability of hASC was strongly dependent on the type of culture, with hASC in monolayer culture being more tolerant to increased osmolarity compared to hASC in suspension, alginate-agarose hydrogel, and pellet cultures, thus emphasizing the importance of choosing relevant in vitro conditions according to the specifics of clinical application.

  6. Hyaluronan in aged collagen matrix increases prostate epithelial cell proliferation

    Science.gov (United States)

    Damodarasamy, Mamatha; Vernon, Robert B.; Chan, Christina K.; Plymate, Stephen R.; Wight, Thomas N.

    2015-01-01

    The extracellular matrix (ECM) of the prostate, which is comprised primarily of collagen, becomes increasingly disorganized with age, a property that may influence the development of hyperplasia and cancer. Collageous ECM extracted from the tails of aged mice exhibits many characteristics of collagen in aged tissues, including the prostate. When polymerized into a 3-dimensional (3D) gel, these collagen extracts can serve as models for the study of specific cell-ECM interactions. In the present study, we examined the behaviors of human prostatic epithelial cell lines representing normal prostate epithelial cells (PEC), benign prostatic hyperplasia (BPH-1), and adenocarcinoma (LNCaP) cultured in contact with 3D gels made from collagen extracts of young and aged mice. We found that proliferation of PEC, BPH-1, and LNCaP cells were all increased by culture on aged collagen gels relative to young collagen gels. In examining age-associated differences in the composition of the collagen extracts, we found that aged and young collagen had a similar amount of several collagen-associated ECM components, but aged collagen had a much greater content of the glycosaminoglycan hyaluronan (HA) than young collagen. The addition of HA (of similar size and concentration to that found in aged collagen extracts) to cells placed in young collagen elicited significantly increased proliferation in BPH-1 cells, but not in PEC or LNCaP cells, relative to controls not exposed to HA. Of note, histochemical analyses of human prostatic tissues showed significantly higher expression of HA in BPH and prostate cancer stroma relative to stroma of normal prostate. Collectively, these results suggest that changes in ECM involving increased levels of HA contribute to the growth of prostatic epithelium with aging. PMID:25124870

  7. TSA increases C/EBP‑α expression by increasing its lysine acetylation in hepatic stellate cells.

    Science.gov (United States)

    Tao, Li-Li; Ding, Di; Yin, Wei-Hua; Peng, Ji-Ying; Hou, Chen-Jian; Liu, Xiu-Ping; Chen, Yao-Li

    2017-11-01

    CCAAT enhancer binding protein‑α (C/EBP‑α) is a transcription factor expressed only in certain tissues, including the liver. It has been previously demonstrated that C/EBP‑α may induce apoptosis in hepatic stellate cells (HSCs), raising the question of whether acetylation of C/EBP‑α is associated with HSCs, and the potential associated mechanism. A total of three histone deacetylase inhibitors (HDACIs), including trichostatin A (TSA), suberoylanilide hydroxamic acid and nicotinamide, were selected to determine whether acetylation affects C/EBP‑α expression. A Cell Counting Kit‑8 assay was used to determine the rate of proliferation inhibition following treatment with varying doses of the three HDACIs in HSC‑T6 and BRL‑3A cells. Western blot analysis was used to examine Caspase‑3, ‑8, ‑9, and ‑12 levels in HSC‑T6 cells treated with adenoviral‑C/EBP‑α and/or TSA. Following treatment with TSA, a combination of reverse transcription‑quantitative polymerase chain reaction and western blot analyses was used to determine the inherent C/EBP‑α mRNA and protein levels in HSC‑T6 cells at 0, 1, 2, 4, 8, 12, 24, 36 and 48 h. Nuclear and cytoplasmic proteins were extracted to examine C/EBP‑α distribution. Co‑immunoprecipitation analysis was used to examine the lysine acetylation of C/EBP‑α. It was observed that TSA inhibited the proliferation of HSC‑T6 cells to a greater extent compared with BRL‑3A cells, following treatment with the three HDACIs. TSA induced apoptosis in HSC‑T6 cells and enhanced the expression of C/EBP‑α. Following treatment of HSC‑T6 cells with TSA, inherent C/EBP‑α expression increased in a time‑dependent manner, and its lysine acetylation simultaneously increased. Therefore, the results of the present study suggested that TSA may increase C/EBP‑α expression by increasing its lysine acetylation in HSCs.

  8. Light-induced performance increase of silicon heterojunction solar cells

    KAUST Repository

    Kobayashi, Eiji; De Wolf, Stefaan; Levrat, Jacques; Christmann, Gabriel; Descoeudres, Antoine; Nicolay, Sylvain; Despeisse, Matthieu; Watabe, Yoshimi; Ballif, Christophe

    2016-01-01

    Silicon heterojunction solar cells consist of crystalline silicon (c-Si) wafers coated with doped/intrinsic hydrogenated amorphous silicon (a-Si:H) bilayers for passivating-contact formation. Here, we unambiguously demonstrate that carrier injection either due to light soaking or (dark) forward-voltage bias increases the open circuit voltage and fill factor of finished cells, leading to a conversion efficiency gain of up to 0.3% absolute. This phenomenon contrasts markedly with the light-induced degradation known for thin-film a-Si:H solar cells. We associate our performance gain with an increase in surface passivation, which we find is specific to doped a-Si:H/c-Si structures. Our experiments suggest that this improvement originates from a reduced density of recombination-active interface states. To understand the time dependence of the observed phenomena, a kinetic model is presented.

  9. Light-induced performance increase of silicon heterojunction solar cells

    KAUST Repository

    Kobayashi, Eiji

    2016-10-11

    Silicon heterojunction solar cells consist of crystalline silicon (c-Si) wafers coated with doped/intrinsic hydrogenated amorphous silicon (a-Si:H) bilayers for passivating-contact formation. Here, we unambiguously demonstrate that carrier injection either due to light soaking or (dark) forward-voltage bias increases the open circuit voltage and fill factor of finished cells, leading to a conversion efficiency gain of up to 0.3% absolute. This phenomenon contrasts markedly with the light-induced degradation known for thin-film a-Si:H solar cells. We associate our performance gain with an increase in surface passivation, which we find is specific to doped a-Si:H/c-Si structures. Our experiments suggest that this improvement originates from a reduced density of recombination-active interface states. To understand the time dependence of the observed phenomena, a kinetic model is presented.

  10. Progranulin increases phagocytosis by retinal pigment epithelial cells in culture.

    Science.gov (United States)

    Murase, Hiromi; Tsuruma, Kazuhiro; Kuse, Yoshiki; Shimazawa, Masamitsu; Hara, Hideaki

    2017-12-01

    Retinal pigment epithelium (RPE) cells take part in retinal preservation, such as phagocytizing the shed photoreceptor outer segments (POS), every day. The incomplete phagocytic function accelerates RPE degeneration and formation of the toxic by-product lipofuscin. Excessive lipofuscin accumulation is characteristic of various blinding diseases in the human eye. Progranulin is a cysteine-rich protein that has multiple biological activities, and it has a high presence in the retina. Progranulin has been recognized to be involved in macrophage phagocytosis in the brain. The purpose of this study is to determine whether progranulin influences phagocytosis by RPE cells. All experiments were performed on primary human RPE (hRPE) cells in culture. pHrodo was used to label the isolated porcine POS, and quantification of pHrodo fluorescence was used to determine the degree of phagocytosis. Western blotting and immunohistochemistry of key proteins involved in phagocytosis were used to clarify the mechanism of progranulin. Progranulin increased RPE phagocytosis in hydrogen peroxide-treated and nontreated RPE cells. The phosphorylated form of Mer tyrosine kinase, which is important for POS internalization, was significantly increased in the progranulin-exposed cells. This increase was attenuated by SU11274, an inhibitor of hepatic growth factor receptor. Under the oxidative stress condition, exposure to progranulin led to an approximately twofold increase in integrin alpha-v, which is associated with the first step in recognition of POS by RPE cells. These results suggest that progranulin could be an effective stimulator for RPE phagocytosis and could repair RPE function. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

  11. KCl stimulation increases norepinephrine transporter function in PC12 cells.

    Science.gov (United States)

    Mandela, Prashant; Ordway, Gregory A

    2006-09-01

    The norepinephrine transporter (NET) plays a pivotal role in terminating noradrenergic signaling and conserving norepinephrine (NE) through the process of re-uptake. Recent evidence suggests a close association between NE release and regulation of NET function. The present study evaluated the relationship between release and uptake, and the cellular mechanisms that govern these processes. KCl stimulation of PC12 cells robustly increased [3H]NE uptake via the NET and simultaneously increased [3H]NE release. KCl-stimulated increases in uptake and release were dependent on Ca2+. Treatment of cells with phorbol-12-myristate-13-acetate (PMA) or okadaic acid decreased [3H]NE uptake but did not block KCl-stimulated increases in [3H]NE uptake. In contrast, PMA increased [3H]NE release and augmented KCl-stimulated release, while okadaic acid had no effects on release. Inhibition of Ca2+-activated signaling cascades with KN93 (a Ca2+ calmodulin-dependent kinase inhibitor), or ML7 and ML9 (myosin light chain kinase inhibitors), reduced [3H]NE uptake and blocked KCl-stimulated increases in uptake. In contrast, KN93, ML7 and ML9 had no effect on KCl-stimulated [3H]NE release. KCl-stimulated increases in [3H]NE uptake were independent of transporter trafficking to the plasma membrane. While increases in both NE release and uptake mediated by KCl stimulation require Ca2+, different intracellular mechanisms mediate these two events.

  12. Immobilisation increases yeast cells' resistance to dehydration-rehydration treatment.

    Science.gov (United States)

    Borovikova, Diana; Rozenfelde, Linda; Pavlovska, Ilona; Rapoport, Alexander

    2014-08-20

    This study was performed with the goal of revealing if the dehydration procedure used in our new immobilisation method noticeably decreases the viability of yeast cells in immobilised preparations. Various yeasts were used in this research: Saccharomyces cerevisiae cells that were rather sensitive to dehydration and had been aerobically grown in an ethanol-containing medium, a recombinant strain of S. cerevisiae grown in aerobic conditions which were completely non-resistant to dehydration and an anaerobically grown bakers' yeast strain S. cerevisiae, as well as a fairly resistant Pichia pastoris strain. Experiments performed showed that immobilisation of all these strains essentially increased their resistance to a dehydration-rehydration treatment. The increase of cells' viability (compared with control cells dehydrated in similar conditions) was from 30 to 60%. It is concluded that a new immobilisation method, which includes a dehydration stage, does not lead to an essential loss of yeast cell viability. Correspondingly, there is no risk of losing the biotechnological activities of immobilised preparations. The possibility of producing dry, active yeast preparations is shown, for those strains that are very sensitive to dehydration and which can be used in biotechnology in an immobilised form. Finally, the immobilisation approach can be used for the development of efficient methods for the storage of recombinant yeast strains. Copyright © 2014 Elsevier B.V. All rights reserved.

  13. Anoxia increases potassium conductance in hippocampal nerve cells.

    Science.gov (United States)

    Hansen, A J; Hounsgaard, J; Jahnsen, H

    1982-07-01

    The effect of anoxia on nerve cell function was studied by intra- and extracellular microelectrode recordings from the CA1 and CA3 region in guinea pig hippocampal slices. Hyperpolarization and concomitant reduction of the nerve cell input resistance was observed early during anoxia. During this period the spontaneous activity first disappeared, then the evoked activity gradually disappeared. The hyperpolarization was followed by depolarization and an absence of a measurable input resistance. All the induced changes were reversed when the slice was reoxygenated. Reversal of the electro-chemical gradient for Cl- across the nerve cell membrane did not affect the course of events during anoxia. Aminopyridines blocked the anoxic hyperpolarization and attenuated the decrease of membrane resistance, but had no effect on the later depolarization. Blockers of synaptic transmission. Mn++, Mg++ and of Na+-channels (TTX) were without effect on the nerve cell changes during anoxia. It is suggested that the reduction of nerve cell excitability in anoxia is primarily due to increased K+-conductance. Thus, the nerve cells are hyperpolarized and the input resistance reduced, causing higher threshold and reduction of synaptic potentials. The mechanism of the K+-conductance activation is unknown at present.

  14. Mesenchymal Stem Cells Respond to Hypoxia by Increasing Diacylglycerols.

    Science.gov (United States)

    Lakatos, Kinga; Kalomoiris, Stefanos; Merkely, Béla; Nolta, Jan A; Fierro, Fernando A

    2016-02-01

    Mesenchymal stem cells (MSC) are currently being tested clinically for a plethora of conditions, with most approaches relying on the secretion of paracrine signals by MSC to modulate the immune system, promote wound healing, and induce angiogenesis. Hypoxia has been shown to affect MSC proliferation, differentiation, survival and secretory profile. Here, we investigate changes in the lipid composition of human bone marrow-derived MSC after exposure to hypoxia. Using mass spectrometry, we compared the lipid profiles of MSC derived from five different donors, cultured for two days in either normoxia (control) or hypoxia (1% oxygen). Hypoxia induced a significant increase of total triglycerides, fatty acids and diacylglycerols (DG). Remarkably, reduction of DG levels using the phosphatidylcholine-specific phospholipase C inhibitor D609 inhibited the secretion of VEGF and Angiopoietin-2, but increased the secretion of interleukin-8, without affecting significantly their respective mRNA levels. Functionally, incubation of MSC in hypoxia with D609 inhibited the potential of the cells to promote migration of human endothelial cells in a wound/scratch assay. Hence, we show that hypoxia induces in MSC an increase of DG that may affect the angiogenic potential of these cells. © 2015 Wiley Periodicals, Inc.

  15. Growth hormone increases vascular cell adhesion molecule 1 expression

    DEFF Research Database (Denmark)

    Hansen, Troels Krarup; Fisker, Sanne; Dall, Rolf

    2004-01-01

    We investigated the impact of GH administration on endothelial adhesion molecules, vascular cell adhesion molecule-1 (VCAM-1) and E-selectin, in vivo and in vitro. Soluble VCAM-1, E-selectin, and C-reactive protein concentrations were measured before and after treatment in 25 healthy subjects...... and 25 adult GH-deficient (GHD) patients randomized to GH treatment or placebo. Furthermore, we studied the direct effect of GH and IGF-I and serum from GH-treated subjects on basal and TNF alpha-stimulated expression of VCAM-1 and E-selectin on cultured human umbilical vein endothelial cells. Baseline......% confidence interval: 95.0-208.7 microg/liter); P cells, there was no direct stimulatory effect of either GH or IGF-I on the expression of VCAM-1 and E-selectin, but serum from GH-treated healthy subjects significantly increased the expression of VCAM-1 (P

  16. Increased mast cell numbers in a calcaneal tendon overuse model.

    Science.gov (United States)

    Pingel, J; Wienecke, J; Kongsgaard, M; Behzad, H; Abraham, T; Langberg, H; Scott, A

    2013-12-01

    Tendinopathy is often discovered late because the initial development of tendon pathology is asymptomatic. The aim of this study was to examine the potential role of mast cell involvement in early tendinopathy using a high-intensity uphill running (HIUR) exercise model. Twenty-four male Wistar rats were divided in two groups: running group (n = 12); sedentary control group (n = 12). The running-group was exposed to the HIUR exercise protocol for 7 weeks. The calcaneal tendons of both hind limbs were dissected. The right tendon was used for histologic analysis using Bonar score, immunohistochemistry, and second harmonic generation microscopy (SHGM). The left tendon was used for quantitative polymerase chain reaction (qPCR) analysis. An increased tendon cell density in the runners were observed compared to the controls (P = 0.05). Further, the intensity of immunostaining of protein kinase B, P = 0.03; 2.75 ± 0.54 vs 1.17 ± 0.53, was increased in the runners. The Bonar score (P = 0.05), and the number of mast cells (P = 0.02) were significantly higher in the runners compared to the controls. Furthermore, SHGM showed focal collagen disorganization in the runners, and reduced collagen density (P = 0.03). IL-3 mRNA levels were correlated with mast cell number in sedentary animals. The qPCR analysis showed no significant differences between the groups in the other analyzed targets. The current study demonstrates that 7-week HIUR causes structural changes in the calcaneal tendon, and further that these changes are associated with an increased mast cell density. © 2013 The Authors. Scand J Med Sci Sports published by John Wiley & Sons Ltd.

  17. Mutations That Alter the Bacterial Cell Envelope Increase Lipid Production

    Energy Technology Data Exchange (ETDEWEB)

    Lemmer, Kimberly C.; Zhang, Weiping; Langer, Samantha J.; Dohnalkova, Alice; Hu, Dehong; Lemke, Rachelle A.; Piotrowski, Jeff S.; Orr, Galya; Noguera, Daniel R.; Donohue, Timothy J.

    2017-05-23

    ABSTRACT

    Lipids from microbes offer a promising source of renewable alternatives to petroleum-derived compounds. In particular, oleaginous microbes are of interest because they accumulate a large fraction of their biomass as lipids. In this study, we analyzed genetic changes that alter lipid accumulation inRhodobacter sphaeroides. By screening anR. sphaeroidesTn5mutant library for insertions that increased fatty acid content, we identified 10 high-lipid (HL) mutants for further characterization. These HL mutants exhibited increased sensitivity to drugs that target the bacterial cell envelope and changes in shape, and some had the ability to secrete lipids, with two HL mutants accumulating ~60% of their total lipids extracellularly. When one of the highest-lipid-secreting strains was grown in a fed-batch bioreactor, its lipid content was comparable to that of oleaginous microbes, with the majority of the lipids secreted into the medium. Based on the properties of these HL mutants, we conclude that alterations of the cell envelope are a previously unreported approach to increase microbial lipid production. We also propose that this approach may be combined with knowledge about biosynthetic pathways, in this or other microbes, to increase production of lipids and other chemicals.

    IMPORTANCEThis paper reports on experiments to understand how to increase microbial lipid production. Microbial lipids are often cited as one renewable replacement for petroleum-based fuels and chemicals, but strategies to increase the yield of these compounds are needed to achieve this goal. While lipid biosynthesis is often well understood, increasing yields of these compounds to industrially relevant levels is a challenge, especially since genetic, synthetic biology, or engineering approaches are not feasible in many microbes. We show that altering the bacterial cell envelope can be used to increase

  18. Increased regulatory T cells in acute lymphoblastic leukemia patients.

    Science.gov (United States)

    Idris, Siti-Zuleha; Hassan, Norfarazieda; Lee, Le-Jie; Md Noor, Sabariah; Osman, Raudhawati; Abdul-Jalil, Marsitah; Nordin, Abdul-Jalil; Abdullah, Maha

    2015-10-01

    Regulation in adaptive immune response balances a fine line that prevents instigation of self-damage or fall into unresponsiveness permitting abnormal cell growth. Mechanisms that keep this balance in check include regulatory T cells (Tregs). Tregs consist of a small but heterogeneous population which may be identified by the phenotype, CD3+CD4+CD25+CD127-. Role of Tregs in pathogenesis of cancers is thus far supported by evidence of increased Tregs in various cancers and may contribute to poorer prognosis. Tregs may also be important in acute leukemias. A review of the literature on Tregs in acute leukemias was conducted and Tregs were determined in B-cell acute lymphoblastic leukemias (ALLs). Studies on Tregs in B-cell ALL are few and controversial. We observed a significantly increased percentage of Tregs (mean ± SD, 9.72 ± 3.79% vs. 7.05 ± 1.74%; P = 0.047) in the bone marrow/peripheral blood of ALL (n = 17) compared to peripheral blood of normal controls (n = 35). A positive trend between Tregs and age (R = 0.474, P = 0.055, n = 17) implicates this factor of poor prognosis in B-cell ALL. Tregs in cancer are particularly significant in immunotherapy. The manipulation of the immune system to treat cancer has for a long time ignored regulatory mechanisms inducible or in place. In lymphoma studies tumor-specific mechanisms that are unlike conventional methods in the induction of Tregs have been hypothesized. In addition, tumor-infiltrating Tregs may present different profiles from peripheral blood pictures. Tregs will continue to be dissected to reveal their mysteries and their impact on clinical significance.

  19. Microparticle Shedding from Neural Progenitor Cells and Vascular Compartment Cells Is Increased in Ischemic Stroke.

    Science.gov (United States)

    Chiva-Blanch, Gemma; Suades, Rosa; Crespo, Javier; Peña, Esther; Padró, Teresa; Jiménez-Xarrié, Elena; Martí-Fàbregas, Joan; Badimon, Lina

    2016-01-01

    Ischemic stroke has shown to induce platelet and endothelial microparticle shedding, but whether stroke induces microparticle shedding from additional blood and vascular compartment cells is unclear. Neural precursor cells have been shown to replace dying neurons at sites of brain injury; however, if neural precursor cell activation is associated to microparticle shedding, and whether this activation is maintained at long term and associates to stroke type and severity remains unknown. We analyzed neural precursor cells and blood and vascular compartment cells microparticle shedding after an acute ischemic stroke. Forty-four patients were included in the study within the first 48h after the onset of stroke. The cerebral lesion size was evaluated at 3-7 days of the stroke. Circulating microparticles from neural precursor cells and blood and vascular compartment cells (platelets, endothelial cells, erythrocytes, leukocytes, lymphocytes, monocytes and smooth muscle cells) were analyzed by flow cytometry at the onset of stroke and at 7 and 90 days. Forty-four age-matched high cardiovascular risk subjects without documented vascular disease were used as controls. Compared to high cardiovascular risk controls, patients showed higher number of neural precursor cell- and all blood and vascular compartment cell-derived microparticles at the onset of stroke, and after 7 and 90 days. At 90 days, neural precursor cell-derived microparticles decreased and smooth muscle cell-derived microparticles increased compared to levels at the onset of stroke, but only in those patients with the highest stroke-induced cerebral lesions. Stroke increases blood and vascular compartment cell and neural precursor cell microparticle shedding, an effect that is chronically maintained up to 90 days after the ischemic event. These results show that stroke induces a generalized blood and vascular cell activation and the initiation of neuronal cell repair process after stroke. Larger cerebral lesions

  20. Microparticle Shedding from Neural Progenitor Cells and Vascular Compartment Cells Is Increased in Ischemic Stroke.

    Directory of Open Access Journals (Sweden)

    Gemma Chiva-Blanch

    Full Text Available Ischemic stroke has shown to induce platelet and endothelial microparticle shedding, but whether stroke induces microparticle shedding from additional blood and vascular compartment cells is unclear. Neural precursor cells have been shown to replace dying neurons at sites of brain injury; however, if neural precursor cell activation is associated to microparticle shedding, and whether this activation is maintained at long term and associates to stroke type and severity remains unknown. We analyzed neural precursor cells and blood and vascular compartment cells microparticle shedding after an acute ischemic stroke.Forty-four patients were included in the study within the first 48h after the onset of stroke. The cerebral lesion size was evaluated at 3-7 days of the stroke. Circulating microparticles from neural precursor cells and blood and vascular compartment cells (platelets, endothelial cells, erythrocytes, leukocytes, lymphocytes, monocytes and smooth muscle cells were analyzed by flow cytometry at the onset of stroke and at 7 and 90 days. Forty-four age-matched high cardiovascular risk subjects without documented vascular disease were used as controls.Compared to high cardiovascular risk controls, patients showed higher number of neural precursor cell- and all blood and vascular compartment cell-derived microparticles at the onset of stroke, and after 7 and 90 days. At 90 days, neural precursor cell-derived microparticles decreased and smooth muscle cell-derived microparticles increased compared to levels at the onset of stroke, but only in those patients with the highest stroke-induced cerebral lesions.Stroke increases blood and vascular compartment cell and neural precursor cell microparticle shedding, an effect that is chronically maintained up to 90 days after the ischemic event. These results show that stroke induces a generalized blood and vascular cell activation and the initiation of neuronal cell repair process after stroke. Larger

  1. Inhibition of autophagy induced by proteasome inhibition increases cell death in human SHG-44 glioma cells.

    Science.gov (United States)

    Ge, Peng-Fei; Zhang, Ji-Zhou; Wang, Xiao-Fei; Meng, Fan-Kai; Li, Wen-Chen; Luan, Yong-Xin; Ling, Feng; Luo, Yi-Nan

    2009-07-01

    The ubiquitin-proteasome system (UPS) and lysosome-dependent macroautophagy (autophagy) are two major intracellular pathways for protein degradation. Recent studies suggest that proteasome inhibitors may reduce tumor growth and activate autophagy. Due to the dual roles of autophagy in tumor cell survival and death, the effect of autophagy on the destiny of glioma cells remains unclear. In this study, we sought to investigate whether inhibition of the proteasome can induce autophagy and the effects of autophagy on the fate of human SHG-44 glioma cells. The proteasome inhibitor MG-132 was used to induce autophagy in SHG-44 glioma cells, and the effect of autophagy on the survival of SHG-44 glioma cells was investigated using an autophagy inhibitor 3-MA. Cell viability was measured by MTT assay. Apoptosis and cell cycle were detected by flow cytometry. The expression of autophagy related proteins was determined by Western blot. MG-132 inhibited cell proliferation, induced cell death and cell cycle arrest at G(2)/M phase, and activated autophagy in SHG-44 glioma cells. The expression of autophagy-related Beclin-1 and LC3-I was significantly up-regulated and part of LC3-I was converted into LC3-II. However, when SHG-44 glioma cells were co-treated with MG-132 and 3-MA, the cells became less viable, but cell death and cell numbers at G(2)/M phase increased. Moreover, the accumulation of acidic vesicular organelles was decreased, the expression of Beclin-1 and LC3 was significantly down-regulated and the conversion of LC3-II from LC3-I was also inhibited. Inhibition of the proteasome can induce autophagy in human SHG-44 glioma cells, and inhibition of autophagy increases cell death. This discovery may shed new light on the effect of autophagy on modulating the fate of SHG-44 glioma cells.Acta Pharmacologica Sinica (2009) 30: 1046-1052; doi: 10.1038/aps.2009.71.

  2. CDTE alloys and their application for increasing solar cell performance

    Science.gov (United States)

    Swanson, Drew E.

    Cadmium Telluride (CdTe) thin film solar is the largest manufactured solar cell technology in the United States and is responsible for one of the lowest costs of utility scale solar electricity at a purchase agreement of $0.0387/kWh. However, this cost could be further reduced by increasing the cell efficiency. To bridge the gap between the high efficiency technology and low cost manufacturing, a research and development tool and process was built and tested. This fully automated single vacuum PV manufacturing tool utilizes multiple inline close space sublimation (CSS) sources with automated substrate control. This maintains the proven scalability of the CSS technology and CSS source design but with the added versatility of independent substrate motion. This combination of a scalable deposition technology with increased cell fabrication flexibility has allowed for high efficiency cells to be manufactured and studied. The record efficiency of CdTe solar cells is lower than fundamental limitations due to a significant deficit in voltage. It has been modeled that there are two potential methods of decreasing this voltage deficiency. The first method is the incorporation of a high band gap film at the back contact to induce a conduction-band barrier that can reduce recombination by reflecting electrons from the back surface. The addition of a Cd1-x MgxTe (CMT) layer at the back of a CdTe solar cell should induce this desired offset and reflect both photoelectrons and forward-current electrons away from the rear surface. Higher collection of photoelectrons will increase the cells current and the reduction of forward current will increase the cells voltage. To have the optimal effect, CdTe must have reasonable carrier lifetimes and be fully depleted. To achieve this experimentally, CdTe layers have been grown sufficiently thin to help produce a fully depleted cell. A variety of measurements including performance curves, transmission electron microscopy, x

  3. Aging increases cell-to-cell transcriptional variability upon immune stimulation.

    Science.gov (United States)

    Martinez-Jimenez, Celia Pilar; Eling, Nils; Chen, Hung-Chang; Vallejos, Catalina A; Kolodziejczyk, Aleksandra A; Connor, Frances; Stojic, Lovorka; Rayner, Timothy F; Stubbington, Michael J T; Teichmann, Sarah A; de la Roche, Maike; Marioni, John C; Odom, Duncan T

    2017-03-31

    Aging is characterized by progressive loss of physiological and cellular functions, but the molecular basis of this decline remains unclear. We explored how aging affects transcriptional dynamics using single-cell RNA sequencing of unstimulated and stimulated naïve and effector memory CD4 + T cells from young and old mice from two divergent species. In young animals, immunological activation drives a conserved transcriptomic switch, resulting in tightly controlled gene expression characterized by a strong up-regulation of a core activation program, coupled with a decrease in cell-to-cell variability. Aging perturbed the activation of this core program and increased expression heterogeneity across populations of cells in both species. These discoveries suggest that increased cell-to-cell transcriptional variability will be a hallmark feature of aging across most, if not all, mammalian tissues. Copyright © 2017, American Association for the Advancement of Science.

  4. SATB2 expression increased anchorage-independent growth and cell migration in human bronchial epithelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Wu, Feng; Jordan, Ashley; Kluz, Thomas [Department of Environmental Medicine, New York University School of Medicine, 57 Old Forge Road, Tuxedo, NY 10987 (United States); Shen, Steven [Center for Health Informatics and Bioinformatics, New York University Langone Medical Center, New York, NY 10016 (United States); Sun, Hong; Cartularo, Laura A. [Department of Environmental Medicine, New York University School of Medicine, 57 Old Forge Road, Tuxedo, NY 10987 (United States); Costa, Max, E-mail: Max.Costa@nyumc.org [Department of Environmental Medicine, New York University School of Medicine, 57 Old Forge Road, Tuxedo, NY 10987 (United States)

    2016-02-15

    The special AT-rich sequence-binding protein 2 (SATB2) is a protein that binds to the nuclear matrix attachment region of the cell and regulates gene expression by altering chromatin structure. In our previous study, we reported that SATB2 gene expression was induced in human bronchial epithelial BEAS-2B cells transformed by arsenic, chromium, nickel and vanadium. In this study, we show that ectopic expression of SATB2 in the normal human bronchial epithelial cell-line BEAS-2B increased anchorage-independent growth and cell migration, meanwhile, shRNA-mediated knockdown of SATB2 significantly decreased anchorage-independent growth in Ni transformed BEAS-2B cells. RNA sequencing analyses of SATB2 regulated genes revealed the enrichment of those involved in cytoskeleton, cell adhesion and cell-movement pathways. Our evidence supports the hypothesis that SATB2 plays an important role in BEAS-2B cell transformation. - Highlights: • We performed SATB2 overexpression in the BEAS-2B cell line. • We performed SATB2 knockdown in a Ni transformed BEAS-2B cell line. • SATB2 induced anchorage-independent growth and increased cell migration. • SATB2 knockdown significantly decreased anchorage-independent growth. • We identified alterations in gene involved in cytoskeleton, cell adhesion.

  5. SATB2 expression increased anchorage-independent growth and cell migration in human bronchial epithelial cells

    International Nuclear Information System (INIS)

    Wu, Feng; Jordan, Ashley; Kluz, Thomas; Shen, Steven; Sun, Hong; Cartularo, Laura A.; Costa, Max

    2016-01-01

    The special AT-rich sequence-binding protein 2 (SATB2) is a protein that binds to the nuclear matrix attachment region of the cell and regulates gene expression by altering chromatin structure. In our previous study, we reported that SATB2 gene expression was induced in human bronchial epithelial BEAS-2B cells transformed by arsenic, chromium, nickel and vanadium. In this study, we show that ectopic expression of SATB2 in the normal human bronchial epithelial cell-line BEAS-2B increased anchorage-independent growth and cell migration, meanwhile, shRNA-mediated knockdown of SATB2 significantly decreased anchorage-independent growth in Ni transformed BEAS-2B cells. RNA sequencing analyses of SATB2 regulated genes revealed the enrichment of those involved in cytoskeleton, cell adhesion and cell-movement pathways. Our evidence supports the hypothesis that SATB2 plays an important role in BEAS-2B cell transformation. - Highlights: • We performed SATB2 overexpression in the BEAS-2B cell line. • We performed SATB2 knockdown in a Ni transformed BEAS-2B cell line. • SATB2 induced anchorage-independent growth and increased cell migration. • SATB2 knockdown significantly decreased anchorage-independent growth. • We identified alterations in gene involved in cytoskeleton, cell adhesion.

  6. Inflammation increases cells expressing ZSCAN4 and progenitor cell markers in the adult pancreas

    Science.gov (United States)

    Azuma, Sakiko; Yokoyama, Yukihiro; Yamamoto, Akiko; Kyokane, Kazuhiro; Niida, Shumpei; Ishiguro, Hiroshi; Ko, Minoru S. H.

    2013-01-01

    We have recently identified the zinc finger and SCAN domain containing 4 (Zscan4), which is transiently expressed and regulates telomere elongation and genome stability in mouse embryonic stem (ES) cells. The aim of this study was to examine the expression of ZSCAN4 in the adult pancreas and elucidate the role of ZSCAN4 in tissue inflammation and subsequent regeneration. The expression of ZSCAN4 and other progenitor or differentiated cell markers in the human pancreas was immunohistochemically examined. Pancreas sections of alcoholic or autoimmune pancreatitis patients before and under maintenance corticosteroid treatment were used in this study. In the adult human pancreas a small number of ZSCAN4-positive (ZSCAN4+) cells are present among cells located in the islets of Langerhans, acini, ducts, and oval-shaped cells. These cells not only express differentiated cell markers for each compartment of the pancreas but also express other tissue stem/progenitor cell markers. Furthermore, the number of ZSCAN4+ cells dramatically increased in patients with chronic pancreatitis, especially in the pancreatic tissues of autoimmune pancreatitis actively regenerating under corticosteroid treatment. Interestingly, a number of ZSCAN4+ cells in the pancreas of autoimmune pancreatitis returned to the basal level after 1 yr of maintenance corticosteroid treatment. In conclusion, coexpression of progenitor cell markers and differentiated cell markers with ZSCAN4 in each compartment of the pancreas may indicate the presence of facultative progenitors for both exocrine and endocrine cells in the adult pancreas. PMID:23599043

  7. Increased mast cell numbers in a calcaneal tendon overuse model

    DEFF Research Database (Denmark)

    Pingel, Jessica; Wienecke, Jacob; Kongsgaard Madsen, Mads

    2013-01-01

    Tendinopathy is often discovered late because the initial development of tendon pathology is asymptomatic. The aim of this study was to examine the potential role of mast cell involvement in early tendinopathy using a high-intensity uphill running (HIUR) exercise model. Twenty-four male Wistar rats...... = 0.03; 2.75 ± 0.54 vs 1.17 ± 0.53, was increased in the runners. The Bonar score (P = 0.05), and the number of mast cells (P = 0.02) were significantly higher in the runners compared to the controls. Furthermore, SHGM showed focal collagen disorganization in the runners, and reduced collagen density...... (P = 0.03). IL-3 mRNA levels were correlated with mast cell number in sedentary animals. The qPCR analysis showed no significant differences between the groups in the other analyzed targets. The current study demonstrates that 7-week HIUR causes structural changes in the calcaneal tendon, and further...

  8. Hakai reduces cell-substratum adhesion and increases epithelial cell invasion

    International Nuclear Information System (INIS)

    Rodríguez-Rigueiro, Teresa; Valladares-Ayerbes, Manuel; Haz-Conde, Mar; Aparicio, Luis A; Figueroa, Angélica

    2011-01-01

    The dynamic regulation of cell-cell adhesions is crucial for developmental processes, including tissue formation, differentiation and motility. Adherens junctions are important components of the junctional complex between cells and are necessary for maintaining cell homeostasis and normal tissue architecture. E-cadherin is the prototype and best-characterized protein member of adherens junctions in mammalian epithelial cells. Regarded as a tumour suppressor, E-cadherin loss is associated with poor prognosis in carcinoma. The E3 ubiquitin-ligase Hakai was the first reported posttranslational regulator of the E-cadherin complex. Hakai specifically targetted E-cadherin for internalization and degradation and thereby lowered epithelial cell-cell contact. Hakai was also implicated in controlling proliferation, and promoted cancer-related gene expression by increasing the binding of RNA-binding protein PSF to RNAs encoding oncogenic proteins. We sought to investigate the possible implication of Hakai in cell-substratum adhesions and invasion in epithelial cells. Parental MDCK cells and MDCK cells stably overexpressing Hakai were used to analyse cell-substratum adhesion and invasion capabilities. Western blot and immunofluoresecence analyses were performed to assess the roles of Paxillin, FAK and Vinculin in cell-substratum adhesion. The role of the proteasome in controlling cell-substratum adhesion was studied using two proteasome inhibitors, lactacystin and MG132. To study the molecular mechanisms controlling Paxillin expression, MDCK cells expressing E-cadherin shRNA in a tetracycline-inducible manner was employed. Here, we present evidence that implicate Hakai in reducing cell-substratum adhesion and increasing epithelial cell invasion, two hallmark features of cancer progression and metastasis. Paxillin, an important protein component of the cell-matrix adhesion, was completely absent from focal adhesions and focal contacts in Hakai-overexpressing MDCK cells. The

  9. Solanine induced apoptosis and increased chemosensitivity to Adriamycin in T-cell acute lymphoblastic leukemia cells.

    Science.gov (United States)

    Yi, Ying-Jie; Jia, Xiu-Hong; Wang, Jian-Yong; Chen, Jie-Ru; Wang, Hong; Li, You-Jie

    2018-05-01

    Solanine is an alkaloid and is the main extract of the traditional Chinese herb, Solanum nigrum Linn . It has been reported that Solanine has anti-inflammatory and antitumor properties. The present study aimed to investigate the antitumor effect of Solanine in Jurkat cells and demonstrate the molecular mechanism of antitumor activity of Solanine. A Cell Counting Kit-8 assay demonstrated that Solanine inhibited the proliferation of Jurkat cells in a dose-and time-dependent manner. Cell apoptosis was measured by flow cytometry. Flow cytometry revealed that Solanine induced apoptosis in a dose-dependent manner in Jurkat cells. Reverse transcription-quantitative polymerase chain reaction demonstrated that Solanine modulated the mRNA levels of B-cell lymphoma-2 (Bcl-2) and Bcl-2-associated X protein (Bax). Additionally, Bcl-2 and Bax expression was measured using western blot analysis. Western blot analysis revealed a significant increase in the expression of Bax and decrease in the expression of Bcl-2. Solanine increased the chemosensitivity of Jurkat cells to Adriamycin. In summary, the present results indicated that the antitumor activity of Solanine was associated with inhibition of cell proliferation, induction of apoptosis and increasing cytotoxicity of Adriamycin. Therefore, Solanine may have potential as a novel agent for the treatment of acute lymphocytic leukemia.

  10. Lactic Acid Bacteria from Kefir Increase Cytotoxicity of Natural Killer Cells to Tumor Cells.

    Science.gov (United States)

    Yamane, Takuya; Sakamoto, Tatsuji; Nakagaki, Takenori; Nakano, Yoshihisa

    2018-03-27

    The Japanese fermented beverage, homemade kefir, contains six lactic acid bacteria: Lactococcus. lactis subsp. Lactis , Lactococcus . lactis subsp. Cremoris , Lactococcus. Lactis subsp. Lactis biovar diacetylactis , Lactobacillus plantarum , Leuconostoc meseuteroides subsp. Cremoris and Lactobacillus casei . In this study, we found that a mixture of the six lactic acid bacteria from kefir increased the cytotoxicity of human natural killer KHYG-1 cells to human chronic myelogenous leukemia K562 cells and colorectal tumor HCT116 cells. Furthermore, levels of mRNA expression and secretion of IFN-γ (interferon gamma) increased in KHYG-1 cells that had been treated with the six lactic acid bacteria mixture from kefir. The results suggest that the six lactic acid bacteria mixture from kefir has strong effects on natural immunity and tumor cell cytotoxicity.

  11. Lactic Acid Bacteria from Kefir Increase Cytotoxicity of Natural Killer Cells to Tumor Cells

    Directory of Open Access Journals (Sweden)

    Takuya Yamane

    2018-03-01

    Full Text Available The Japanese fermented beverage, homemade kefir, contains six lactic acid bacteria: Lactococcus. lactis subsp. Lactis, Lactococcus. lactis subsp. Cremoris, Lactococcus. Lactis subsp. Lactis biovar diacetylactis, Lactobacillus plantarum, Leuconostoc meseuteroides subsp. Cremoris and Lactobacillus casei. In this study, we found that a mixture of the six lactic acid bacteria from kefir increased the cytotoxicity of human natural killer KHYG-1 cells to human chronic myelogenous leukemia K562 cells and colorectal tumor HCT116 cells. Furthermore, levels of mRNA expression and secretion of IFN-γ (interferon gamma increased in KHYG-1 cells that had been treated with the six lactic acid bacteria mixture from kefir. The results suggest that the six lactic acid bacteria mixture from kefir has strong effects on natural immunity and tumor cell cytotoxicity.

  12. Human lactoferrin increases Helicobacter pylori internalisation into AGS cells.

    Science.gov (United States)

    Coray, Dorien S; Heinemann, Jack A; Tyrer, Peter C; Keenan, Jacqueline I

    2012-05-01

    Helicobacter pylori has high global infection rates and can cause other undesirable clinical manifestations such as duodenal ulcer (DU) and gastric cancer (GC). Frequencies of re-infection after therapeutic clearance and rates of DU versus GC vary geographically and differ markedly between developed and developing countries, which suggests additional factors may be involved. The possibility that, in vivo, lactoferrin (Lf) may play a subtle role in modulating micronutrient availability or bacterial internalisation with implications for disease etiology is considered. Lf is an iron binding protein produced in mammals that has antimicrobial and immunomodulatory properties. Some bacteria that regularly colonise mammalian hosts have adapted to living in high Lf environments and we investigated if this included the gastric pathogen H. pylori. We found that H. pylori was able to use iron from fully iron-saturated human Lf (hLf) whereas partially iron-saturated hLf (apo) did not increase H. pylori growth. Instead, apo-hLf increased adherence to and internalisation of bacteria into cultured epithelial cells. By increasing internalisation, we speculate that apo-human lactoferrin may contribute to H. pylori's ability to persistence in the human stomach, an observation that potentially has implications for the risk of H. pylori-associated disease.

  13. Increase of corneal epithelium cell radioresistance during regeneration

    International Nuclear Information System (INIS)

    Popova, M.F.; Bulyakova, N.V.; Azarova, V.S.

    1985-01-01

    A comparative study of the radiosensitivity of the normal and regenerating cornea epithelium of C 57 Bl mice was performed on the cellular level, the duration of the cell cycle being taken into account. Criteria of radiation injuries were the number of chromosome aberrations, mitotic index and duration of mitotic block. The anterior part of the head was irradiated singly with 1.75, 3.5 or 7.0 Gy and also repeatedly 3.5 + 3.5 at a 24-hours interval. The corneas were fixed 2, 4, 6, 12, 24, 48, 72 and 96 hours after irradiation. In all cases of irradiated mice the regenerating epithelium showed a shorter mitotic block and significantly lower cytogenetic injury as compared with the controls. Effects of fractionated irradiation were only shown in the regenerating epithelium. The results obtained indicate that regenerating epithelium cells of the cornea are significantly more radioresistant than normal epithelium due to activation of post-radiation recovery, and also, possibly, due to an increase in the content of endogenous radioprotectors. (author)

  14. Increased theta band EEG power in sickle cell disease patients

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    Case M

    2017-12-01

    Full Text Available Michelle Case,1 Sina Shirinpour,1 Huishi Zhang,1 Yvonne H Datta,2 Stephen C Nelson,3 Karim T Sadak,4 Kalpna Gupta,2 Bin He1,5 1Department of Biomedical Engineering, 2Department of Medicine, University of Minnesota, 3Pediatric Hematology-Oncology, Children’s Hospitals and Clinics of Minnesota, 4Pediatric Hematology-Oncology, University of Minnesota Masonic Children’s Hospital, 5Institute for Engineering in Medicine, University of Minnesota, Minneapolis, MN, USA Objective: Pain is a major issue in the care of patients with sickle cell disease (SCD. The mechanisms behind pain and the best way to treat it are not well understood. We studied how electroencephalography (EEG is altered in SCD patients. Methods: We recruited 20 SCD patients and compared their resting state EEG to that of 14 healthy controls. EEG power was found across frequency bands using Welch’s method. Electrophysiological source imaging was assessed for each frequency band using the eLORETA algorithm. Results: SCD patients had increased theta power and decreased beta2 power compared to controls. Source localization revealed that areas of greater theta band activity were in areas related to pain processing. Imaging parameters were significantly correlated to emergency department visits, which indicate disease severity and chronic pain intensity. Conclusion: The present results support the pain mechanism referred to as thalamocortical dysrhythmia. This mechanism causes increased theta power in patients. Significance: Our findings show that EEG can be used to quantitatively evaluate differences between controls and SCD patients. Our results show the potential of EEG to differentiate between different levels of pain in an unbiased setting, where specific frequency bands could be used as biomarkers for chronic pain. Keywords: sickle cell disease, electroencephalography, chronic pain, electrophysiological source imaging, thalamocortical dysrhythmia

  15. Increased cell surface metallopeptidase activity in cells undergoing UV-induced apoptosis

    International Nuclear Information System (INIS)

    Piva, T.J.; Davern, C.M.; Ellem, K.A.O.

    1999-01-01

    Full text: We have previously shown that UVC irradiation activated a range of cell surface peptidases (CSP) in HeLa cell monolayer cultures 20 h post-irradiation (1). In cells undergoing apoptosis there is an increase in CSP activity compared to control viable cells in cultures which have been treated by a wide range of agents including UV-irradiation (2). In order to further understand the mechanism involved in this process, we induced apoptosis in HeLa cells using 500 Jm -2 UVB. The separation of viable, apoptotic and necrotic cells of irradiated HeLa cell cultures was made by FACS analysis and sorting. The three populations were distinguished by their staining with PI and Hoechst 33342 dyes. CSP activity was measured using the P9 assay developed in this laboratory (1-3). The viable fraction of the irradiated cells had a higher level of CSP activity compared to unirradiated controls. The level of CSP activity in the apoptotic fraction was higher than that of the viable fraction, however that of the necrotic fraction was significantly lower. This finding agreed with that seen in UVC-irradiated (50 Jm -2 ) cultures (2). In order to elucidate the mechanism by which CSP activity was increased in UVB-irradiated cells undergoing apoptosis, the cultures were treated with the following agents: bestatin, aminopeptidase inhibitor, DEVD, caspase 3 inhibitor, and 3-aminobenzamide (3AB), PARP activation inhibitor. Bestatin and DEVD did not affect the level of CSP activity in the different cell subpopulations following UVB-irradiation. Treatment with 3AB abolished the increased CSP activity seen in the viable and apoptotic fraction following UVB-irradiation. All treated cells had the same morphology as observed under EM. The degree of phosphatidylserine eversion on the cell membrane was similar as were the cleavage profiles of PARP and actin. Only DEVD-treated cells had reduced caspase 3 activity which confirmed that the activation of CSP activity in apoptotic cells is

  16. Centrosome Amplification Increases Single-Cell Branching in Post-mitotic Cells.

    Science.gov (United States)

    Ricolo, Delia; Deligiannaki, Myrto; Casanova, Jordi; Araújo, Sofia J

    2016-10-24

    Centrosome amplification is a hallmark of cancer, although we are still far from understanding how this process affects tumorigenesis [1, 2]. Besides the contribution of supernumerary centrosomes to mitotic defects, their biological effects in the post-mitotic cell are not well known. Here, we exploit the effects of centrosome amplification in post-mitotic cells during single-cell branching. We show that Drosophila tracheal cells with extra centrosomes branch more than wild-type cells. We found that mutations in Rca1 and CycA affect subcellular branching, causing tracheal tip cells to form more than one subcellular lumen. We show that Rca1 and CycA post-mitotic cells have supernumerary centrosomes and that other mutant conditions that increase centrosome number also show excess of subcellular lumen branching. Furthermore, we show that de novo lumen formation is impaired in mutant embryos with fewer centrioles. The data presented here define a requirement for the centrosome as a microtubule-organizing center (MTOC) for the initiation of subcellular lumen formation. We propose that centrosomes are necessary to drive subcellular lumen formation. In addition, centrosome amplification increases single-cell branching, a process parallel to capillary sprouting in blood vessels [3]. These results shed new light on how centrosomes can contribute to pathology independently of mitotic defects. Copyright © 2016 Elsevier Ltd. All rights reserved.

  17. Protease-activated receptor 2 agonist increases cell proliferation and invasion of human pancreatic cancer cells

    Science.gov (United States)

    XIE, LIQUN; DUAN, ZEXING; LIU, CAIJU; ZHENG, YANMIN; ZHOU, JING

    2015-01-01

    The aim of this study was to determine the expression of protease-activated receptor 2 (PAR-2) in the human pancreatic cancer cell line SW1990, and to evaluate its effect on cell proliferation and invasion. The expression of PAR-2 protein and mRNA in SW1990 cells was determined by immunocytochemistry and reverse transcription polymerase chain reaction (PCR), respectively. MTT and cell invasion and migration assays, as well as semi-quantitative PCR and zymography analysis, were additionally performed. PAR-2 mRNA was significantly upregulated in the cells treated with trypsin or the PAR-2 activating peptide Ser-Leu-Ile-Gly-Lys-Val (SLIGKV) (P0.05). Trypsin and SLIGKV significantly promoted SW1990 cell proliferation in a dose- and time-dependent manner (P<0.05). Compared with the control group, trypsin and SLIGKV significantly increased the mRNA expression (P<0.01) and gelatinolytic activity (P<0.01) of matrix metalloproteinase (MMP)-2. In conclusion, PAR-2 is expressed in SW1990 cells. PAR-2 activation may promote the invasion and migration of human pancreatic cancer cells by increasing MMP-2 expression. PMID:25452809

  18. PCM1 Depletion Inhibits Glioblastoma Cell Ciliogenesis and Increases Cell Death and Sensitivity to Temozolomide

    Directory of Open Access Journals (Sweden)

    Lan B. Hoang-Minh

    2016-10-01

    Full Text Available A better understanding of the molecules implicated in the growth and survival of glioblastoma (GBM cells and their response to temozolomide (TMZ, the standard-of-care chemotherapeutic agent, is necessary for the development of new therapies that would improve the outcome of current GBM treatments. In this study, we characterize the role of pericentriolar material 1 (PCM1, a component of centriolar satellites surrounding centrosomes, in GBM cell proliferation and sensitivity to genotoxic agents such as TMZ. We show that PCM1 is expressed around centrioles and ciliary basal bodies in patient GBM biopsies and derived cell lines and that its localization is dynamic throughout the cell cycle. To test whether PCM1 mediates GBM cell proliferation and/or response to TMZ, we used CRISPR/Cas9 genome editing to generate primary GBM cell lines depleted of PCM1. These PCM1-depleted cells displayed reduced AZI1 satellite protein localization and significantly decreased proliferation, which was attributable to increased apoptotic cell death. Furthermore, PCM1-depleted lines were more sensitive to TMZ toxicity than control lines. The increase in TMZ sensitivity may be partly due to the reduced ability of PCM1-depleted cells to form primary cilia, as depletion of KIF3A also ablated GBM cells' ciliogenesis and increased their sensitivity to TMZ while preserving PCM1 localization. In addition, the co-depletion of KIF3A and PCM1 did not have any additive effect on TMZ sensitivity. Together, our data suggest that PCM1 plays multiple roles in GBM pathogenesis and that associated pathways could be targeted to augment current or future anti-GBM therapies.

  19. Brain Cell Swelling During Hypocapnia Increases with Hyperglycemia or Ketosis

    Science.gov (United States)

    Glaser, Nicole; Bundros, Angeliki; Anderson, Steve; Tancredi, Daniel; Lo, Weei; Orgain, Myra; O'Donnell, Martha

    2014-01-01

    Severe hypocapnia increases the risk of DKA-related cerebral injury in children, but the reason for this association is unclear. To determine whether the effects of hypocapnia on the brain are altered during hyperglycemia or ketosis, we induced hypocapnia (pCO2 20 ± 3 mmHg) via mechanical ventilation in three groups of juvenile rats: 25 controls, 22 hyperglycemic rats (serum glucose 451± 78 mg/dL) and 15 ketotic rats (beta-hydroxy butyrate 3.0 ± 1.0 mmol/L). We used magnetic resonance imaging to measure cerebral blood flow (CBF) and apparent diffusion coefficient (ADC) values in these groups and in 17 ventilated rats with normal pCO2 (40±3 mmHg). In a subset (n=35), after 2 hrs of hypocapnia, pCO2 levels were normalized (40±3 mmHg) and ADC and CBF measurements repeated. Declines in CBF with hypocapnia occurred in all groups. Normalization of pCO2 after hypocapnia resulted in striatal hyperemia. These effects were not substantially altered by hyperglycemia or ketosis, however, declines in ADC during hypocapnia were greater during both hyperglycemia and ketosis. We conclude that brain cell swelling associated with hypocapnia is increased by both hyperglycemia and ketosis, suggesting that these metabolic conditions may make the brain more vulnerable to injury during hypocapnia. PMID:24443981

  20. Y-27632 Increases Sensitivity of PANC-1 Cells to EGCG in Regulating Cell Proliferation and Migration.

    Science.gov (United States)

    Liu, Xing; Bi, Yongyi

    2016-10-03

    BACKGROUND The study aimed to investigate the inhibitory effect of (1R,4r)-4-((R)-1-aminoethyl)-N-(pyridin-4-yl) cyclohexanecarboxamide (Y-27632) and (-)-epigallocatechin-3-gallate (EGCG) on the proliferation and migration of PANC-1 cells. EGCG, found in green tea, has been previously shown to be one of the most abundant and powerful catechins in cancer prevention and treatment. Y-27632, a selective inhibitor of rho-associated protein kinase 1, is widely used in treating cardiovascular disease, inflammation, and cancer. MATERIAL AND METHODS PANC-1 cells, maintained in Dulbecco's Modified Eagle's Medium, were treated with dimethyl sulfoxide (control) as well as different concentrations (20, 40, 60, and 80 μg/mL) of EGCG for 48 h. In addition, PANC-1 cells were treated separately with 60 μg/mL EGCG, 20 μM Y-27632, and EGCG combined with Y-27632 (60 μg/mL EGCG + 20 μM Y-27632) for 48 h. The effect of EGCG and Y-27632 on the proliferation and migration of PANC-1 cells was evaluated using Cell Counting Kit-8 and transwell migration assays. The expression of peroxisome proliferator-activated receptor alpha (PPARα) and Caspase-3 mRNA was determined by Quantitative real-time polymerase chain reaction (RT-qPCR). RESULTS EGCG (20-80 μg/mL) inhibited cell viability in a dose-dependent manner. Y-27632 enhanced the sensitivity of PANC-1 cells to EGCG (by increasing the expression of PPARa and Caspase-3 mRNA) and suppressed cell proliferation. PANC-1 cell migration was inhibited by treatment with a combination of EGCG and Y-27632. CONCLUSIONS Y-27632 increases the sensitivity of PANC-1 cells to EGCG in regulating cell proliferation and migration, which is likely to be related to the expression of PPARa mRNA and Caspase-3 mRNA.

  1. Lactic Acid Bacteria from Kefir Increase Cytotoxicity of Natural Killer Cells to Tumor Cells

    OpenAIRE

    Takuya Yamane; Tatsuji Sakamoto; Takenori Nakagaki; Yoshihisa Nakano

    2018-01-01

    The Japanese fermented beverage, homemade kefir, contains six lactic acid bacteria: Lactococcus. lactis subsp. Lactis, Lactococcus. lactis subsp. Cremoris, Lactococcus. Lactis subsp. Lactis biovar diacetylactis, Lactobacillus plantarum, Leuconostoc meseuteroides subsp. Cremoris and Lactobacillus casei. In this study, we found that a mixture of the six lactic acid bacteria from kefir increased the cytotoxicity of human natural killer KHYG-1 cells to human chronic myelogenous leukemia K562 cell...

  2. IGFBP1 increases β-cell regeneration by promoting α- to β-cell transdifferentiation.

    Science.gov (United States)

    Lu, Jing; Liu, Ka-Cheuk; Schulz, Nadja; Karampelias, Christos; Charbord, Jérémie; Hilding, Agneta; Rautio, Linn; Bertolino, Philippe; Östenson, Claes-Göran; Brismar, Kerstin; Andersson, Olov

    2016-09-15

    There is great interest in therapeutically harnessing endogenous regenerative mechanisms to increase the number of β cells in people with diabetes. By performing whole-genome expression profiling of zebrafish islets, we identified 11 secreted proteins that are upregulated during β-cell regeneration. We then tested the proteins' ability to potentiate β-cell regeneration in zebrafish at supraphysiological levels. One protein, insulin-like growth factor (Igf) binding-protein 1 (Igfbp1), potently promoted β-cell regeneration by potentiating α- to β-cell transdifferentiation. Using various inhibitors and activators of the Igf pathway, we show that Igfbp1 exerts its regenerative effect, at least partly, by inhibiting Igf signaling. Igfbp1's effect on transdifferentiation appears conserved across species: Treating mouse and human islets with recombinant IGFBP1 in vitro increased the number of cells co-expressing insulin and glucagon threefold. Moreover, a prospective human study showed that having high IGFBP1 levels reduces the risk of developing type-2 diabetes by more than 85%. Thus, we identify IGFBP1 as an endogenous promoter of β-cell regeneration and highlight its clinical importance in diabetes. © 2016 The Authors. Published under the terms of the CC BY NC ND 4.0 license.

  3. Separator Characteristics for Increasing Performance of Microbial Fuel Cells

    KAUST Repository

    Zhang, Xiaoyuan

    2009-11-01

    Two challenges for improving the performance of air cathode, single-chamber microbial fuel cells (MFCs) include increasing Coulombic efficiency (CE) and decreasing internal resistance. Nonbiodegradable glass fiber separators between the two electrodes were shown to increase power and CE, compared to cloth separators (J-cloth) that were degraded over time. MFCtestswereconductedusing glass fibermatswith thicknesses of 1.0mm (GF1) or 0.4 mm (GF0.4), a cation exchange membrane (CEM), and a J-cloth (JC), using reactors with different configurations. Higher power densities were obtained with either GF1 (46 ± 4 W/m3) or JC (46 ± 1 W/m3) in MFCs with a 2 cm electrode spacing, when the separator was placed against the cathode (S-configuration), rather than MFCs with GF0.4 (36 ± 1 W/m3) or CEM (14 ± 1 W/m3). Power was increased to 70 ± 2 W/m3 by placing the electrodes on either side of the GF1 separator (single separator electrode assembly, SSEA) and further to 150 ± 6 W/m3 using two sets of electrodes spaced 2 cm a part (double separator electrode assembly, DSEA). Reducing the DSEA electrode spacing to 0.3 cm increased power to 696 ± 26 W/m3 as a result of a decrease in the ohmic resistance from 5.9 to 2.2 Ω. The main advantages of a GF1 separator compared to JC were an improvement in the CE from 40% to 81% (S-configuration), compared to only 20-40% for JC under similar conditions, and the fact that GF1 was not biodegradable. The high CE for the GF1 separator was attributed to a low oxygen mass transfer coefficient (ko ) 5.0 x 10-5 cm/s). The GF1 andJCmaterials differed in the amount of biomass that accumulated on the separator and its biodegradability, which affected long-term power production and oxygen transport. These results show that materials and mass transfer properties of separators are important factors for improving power densities, CE, and long-term performance of MFCs. © 2009 American Chemical Society.

  4. Th17 Cells and Activated Dendritic Cells Are Increased in Vitiligo Lesions

    Science.gov (United States)

    Fuentes-Duculan, Judilyn; Moussai, Dariush; Gulati, Nicholas; Sullivan-Whalen, Mary; Gilleaudeau, Patricia; Cohen, Jules A.; Krueger, James G.

    2011-01-01

    Background Vitiligo is a common skin disorder, characterized by progressive skin de-pigmentation due to the loss of cutaneous melanocytes. The exact cause of melanocyte loss remains unclear, but a large number of observations have pointed to the important role of cellular immunity in vitiligo pathogenesis. Methodology/Principal Findings In this study, we characterized T cell and inflammation-related dermal dendritic cell (DC) subsets in pigmented non-lesional, leading edge and depigmented lesional vitiligo skin. By immunohistochemistry staining, we observed enhanced populations of CD11c+ myeloid dermal DCs and CD207+ Langerhans cells in leading edge vitiligo biopsies. DC-LAMP+ and CD1c+ sub-populations of dermal DCs expanded significantly in leading edge and lesional vitiligo skin. We also detected elevated tissue mRNA levels of IL-17A in leading edge skin biopsies of vitiligo patients, as well as IL-17A positive T cells by immunohistochemistry and immunofluorescence. Langerhans cells with activated inflammasomes were also noted in lesional vitiligo skin, along with increased IL-1ß mRNA, which suggest the potential of Langerhans cells to drive Th17 activation in vitiligo. Conclusions/Significance These studies provided direct tissue evidence that implicates active Th17 cells in vitiligo skin lesions. We characterized new cellular immune elements, in the active margins of vitiligo lesions (e.g. populations of epidermal and dermal dendritic cells subsets), which could potentially drive the inflammatory responses. PMID:21541348

  5. Increased FDG bone marrow uptake after intracoronary progenitor cell therapy

    Energy Technology Data Exchange (ETDEWEB)

    Doebert, N.; Menzel, C.; Diehl, M.; Hamscho, N.; Zaplatnikov, K.; Gruenwald, F. [Dept. of Nuclear Medicine, Univ. of Frankfurt (Germany)

    2005-02-01

    Patients with coronary artery disease who undergo FDG PET for therapy monitoring after intracoronary progenitor cell infusion (PCT) show an increased bone marrow uptake in some cases. Aim of the study was to evaluate the systemic bone marrow glucose metabolism in this patient group after PCT. Patients, methods: FDG bone marrow uptake (BMU), measured as standardized uptake value (SUVmax) in the thoracic spine, was retrospectively evaluated in 23 control patients who did not receive PCT and in 75 patients who received PCT 3{+-}2.2 days before PET scanning. Five out of them were pretreated with granulocyte colony-stimulating factor (G-CSF) 5 days prior to PCT and 10{+-}1.2 days before PET scanning. In 39 patients who received only PCT without G-CSF and underwent PET therapy monitoring 4 months later, baseline and follow up bone marrow uptake were measured. Leucocytes, C-reactive protein (CRP) levels and the influence of nicotine consumption were compared with the BMU. Results: In patients (n=70) who received PCT without G-CSF, BMU media (1.3) was slightly, but significantly higher than in the controls (1.0) (p=0.02) regardless nicotine consumption. BMU did not change significantly 4 months later (1.2) (p=0.41, n.s.). After G-CSF pretreatment, patients showed a significantly higher bone marrow uptake (3.7) compared to patients only treated with PCT (1.3) (p=0.023). Leucocyte blood levels were significantly higher in patients with a BMU {>=}2.5 compared to patients with a bone marrow SUVmax<2.5 (p<0.001). CRP values did not correlate with the BMU (rho -0.02, p=0.38). Conclusion: Monitoring PCT patients, a slightly increased FDG BMU may be observed which remains unchanged for several months. Unspecific bone marrow reactions after PCT may be associated with increased leucocyte blood levels and play a role in the changed systemic glucose BMU. In addition, pretreatment with G-CSF shows an intense amplitifcation of BMU. (orig.)

  6. High epitope expression levels increase competition between T cells.

    Directory of Open Access Journals (Sweden)

    Almut Scherer

    2006-08-01

    Full Text Available Both theoretical predictions and experimental findings suggest that T cell populations can compete with each other. There is some debate on whether T cells compete for aspecific stimuli, such as access to the surface on antigen-presenting cells (APCs or for specific stimuli, such as their cognate epitope ligand. We have developed an individual-based computer simulation model to study T cell competition. Our model shows that the expression level of foreign epitopes per APC determines whether T cell competition is mainly for specific or aspecific stimuli. Under low epitope expression, competition is mainly for the specific epitope stimuli, and, hence, different epitope-specific T cell populations coexist readily. However, if epitope expression levels are high, aspecific competition becomes more important. Such between-specificity competition can lead to competitive exclusion between different epitope-specific T cell populations. Our model allows us to delineate the circumstances that facilitate coexistence of T cells of different epitope specificity. Understanding mechanisms of T cell coexistence has important practical implications for immune therapies that require a broad immune response.

  7. β-Cell Replacement Strategies: The Increasing Need for a "β-Cell Dogma".

    Science.gov (United States)

    Vieira, Andhira; Druelle, Noémie; Avolio, Fabio; Napolitano, Tiziana; Navarro-Sanz, Sergi; Silvano, Serena; Collombat, Patrick

    2017-01-01

    Type 1 diabetes is an auto-immune disease resulting in the loss of pancreatic β-cells and, consequently, in chronic hyperglycemia. Insulin supplementation allows diabetic patients to control their glycaemia quite efficiently, but treated patients still display an overall shortened life expectancy and an altered quality of life as compared to their healthy counterparts. In this context and due to the ever increasing number of diabetics, establishing alternative therapies has become a crucial research goal. Most current efforts therefore aim at generating fully functional insulin-secreting β-like cells using multiple approaches. In this review, we screened the literature published since 2011 and inventoried the selected markers used to characterize insulin-secreting cells generated by in vitro differentiation of stem/precursor cells or by means of in vivo transdifferentiation. By listing these features, we noted important discrepancies when comparing the different approaches for the initial characterization of insulin-producing cells as true β-cells. Considering the recent advances achieved in this field of research, the necessity to establish strict guidelines has become a subject of crucial importance, especially should one contemplate the next step, which is the transplantation of in vitro or ex vivo generated insulin-secreting cells in type 1 diabetic patients.

  8. Glucocorticoid receptor beta increases migration of human bladder cancer cells.

    Science.gov (United States)

    McBeth, Lucien; Nwaneri, Assumpta C; Grabnar, Maria; Demeter, Jonathan; Nestor-Kalinoski, Andrea; Hinds, Terry D

    2016-05-10

    Bladder cancer is observed worldwide having been associated with a host of environmental and lifestyle risk factors. Recent investigations on anti-inflammatory glucocorticoid signaling point to a pathway that may impact bladder cancer. Here we show an inverse effect on the glucocorticoid receptor (GR) isoform signaling that may lead to bladder cancer. We found similar GRα expression levels in the transitional uroepithelial cancer cell lines T24 and UMUC-3. However, the T24 cells showed a significant (p bladder cancer cells. Therefore, GRβ may have a significant role in bladder cancer, and possibly serve as a therapeutic target for the disease.

  9. Medullospheres from DAOY, UW228 and ONS-76 cells: increased stem cell population and proteomic modifications.

    Science.gov (United States)

    Zanini, Cristina; Ercole, Elisabetta; Mandili, Giorgia; Salaroli, Roberta; Poli, Alice; Renna, Cristiano; Papa, Valentina; Cenacchi, Giovanna; Forni, Marco

    2013-01-01

    Medulloblastoma (MB) is an aggressive pediatric tumor of the Central Nervous System (CNS) usually treated according to a refined risk stratification. The study of cancer stem cells (CSC) in MB is a promising approach aimed at finding new treatment strategies. The CSC compartment was studied in three characterized MB cell lines (DAOY, UW228 and ONS-76) grown in standard adhesion as well as being grown as spheres, which enables expansion of the CSC population. MB cell lines, grown in adherence and as spheres, were subjected to morphologic analysis at the light and electron microscopic level, as well as cytofluorimetric determinations. Medullospheres (MBS) were shown to express increasingly immature features, along with the stem cells markers: CD133, Nestin and β-catenin. Proteomic analysis highlighted the differences between MB cell lines, demonstrating a unique protein profile for each cell line, and minor differences when grown as spheres. In MBS, MALDI-TOF also identified some proteins, that have been linked to tumor progression and resistance, such as Nucleophosmin (NPM). In addition, immunocytochemistry detected Sox-2 as a stemness marker of MBS, as well as confirming high NPM expression. Culture conditioning based on low attachment flasks and specialized medium may provide new data on the staminal compartment of CNS tumors, although a proteomic profile of CSC is still elusive for MB.

  10. Medullospheres from DAOY, UW228 and ONS-76 cells: increased stem cell population and proteomic modifications.

    Directory of Open Access Journals (Sweden)

    Cristina Zanini

    Full Text Available BACKGROUND: Medulloblastoma (MB is an aggressive pediatric tumor of the Central Nervous System (CNS usually treated according to a refined risk stratification. The study of cancer stem cells (CSC in MB is a promising approach aimed at finding new treatment strategies. METHODOLOGY/PRINCIPAL FINDINGS: The CSC compartment was studied in three characterized MB cell lines (DAOY, UW228 and ONS-76 grown in standard adhesion as well as being grown as spheres, which enables expansion of the CSC population. MB cell lines, grown in adherence and as spheres, were subjected to morphologic analysis at the light and electron microscopic level, as well as cytofluorimetric determinations. Medullospheres (MBS were shown to express increasingly immature features, along with the stem cells markers: CD133, Nestin and β-catenin. Proteomic analysis highlighted the differences between MB cell lines, demonstrating a unique protein profile for each cell line, and minor differences when grown as spheres. In MBS, MALDI-TOF also identified some proteins, that have been linked to tumor progression and resistance, such as Nucleophosmin (NPM. In addition, immunocytochemistry detected Sox-2 as a stemness marker of MBS, as well as confirming high NPM expression. CONCLUSIONS/SIGNIFICANCE: Culture conditioning based on low attachment flasks and specialized medium may provide new data on the staminal compartment of CNS tumors, although a proteomic profile of CSC is still elusive for MB.

  11. Increasing cell culture population doublings for long-term growth of finite life span human cell cultures

    Science.gov (United States)

    Stampfer, Martha R; Garbe, James C

    2015-02-24

    Cell culture media formulations for culturing human epithelial cells are herein described. Also described are methods of increasing population doublings in a cell culture of finite life span human epithelial cells and prolonging the life span of human cell cultures. Using the cell culture media disclosed alone and in combination with addition to the cell culture of a compound associated with anti-stress activity achieves extended growth of pre-stasis cells and increased population doublings and life span in human epithelial cell cultures.

  12. Increased somatic cell mutant frequency in atomic bomb survivors

    International Nuclear Information System (INIS)

    Hakoda, Masayuki; Akiyama, Mitoshi; Kyoizumi, Seishi; Awa, A.A.; Yamakido, Michio; Otake, Masanori.

    1988-05-01

    Frequencies of mutant T-cells in peripheral blood, which are deficient in the activity of hypoxanthine guanine phosphoribosyltransferase (HPRT) were determined for atomic bomb survivors by direct clonal assay using a previously reported method. Results from 30 exposed survivors (exposed to more than 1 rad) and 17 age- and sex-matched controls (exposed to less than 1 rad) were analyzed. The mean mutant frequency (Mf) in the exposed (5.2 x 10 -6 ; range 0.8 - 14.4 x 10 -6 ) was significantly higher than in controls (3.4 x 10 -6 ; range 1.3 - 9.3 x 10 -6 ), a fact not attributable to lower nonmutant cell cloning efficiencies in the exposed group since cell cloning efficiencies were virtually identical in both groups. An initial analysis of the data did not reveal a significant correlation between individual Mfs and individual radiation dose estimates when the latter were defined by the original, tentative estimates (T65D), even though there was a significant positive correlation of Mfs with individual frequency of lymphocytes bearing chromosome aberration. However, reanalysis using the newer revised individual dose estimates (DS86) for 27 exposed survivors and 17 controls did reveal a significant but shallow positive correlation between T-cell Mf values and individual exposure doses. These results indicate that HPRT mutation in vivo in human T-cells could be detected in these survivors 40 years after the presumed mutational event. (author)

  13. An Aminopropyl Carbazole Derivative Induces Neurogenesis by Increasing Final Cell Division in Neural Stem Cells.

    Science.gov (United States)

    Shin, Jae-Yeon; Kong, Sun-Young; Yoon, Hye Jin; Ann, Jihyae; Lee, Jeewoo; Kim, Hyun-Jung

    2015-07-01

    P7C3 and its derivatives, 1-(3,6-dibromo-9H-carbazol-9-yl)-3-(p-tolylamino)propan-2-ol (1) and N-(3-(3,6-dibromo-9H-carbazol-9-yl)-2-hydroxypropyl)-N-(3-methoxyphenyl)-4-methylbenzenesulfonamide (2), were previously reported to increase neurogenesis in rat neural stem cells (NSCs). Although P7C3 is known to increase neurogenesis by protecting newborn neurons, it is not known whether its derivatives also have protective effects to increase neurogenesis. In the current study, we examined how 1 induces neurogenesis. The treatment of 1 in NSCs increased numbers of cells in the absence of epidermal growth factor (EGF) and fibroblast growth factor 2 (FGF2), while not affecting those in the presence of growth factors. Compound 1 did not induce astrocytogenesis during NSC differentiation. 5-Bromo-2'-deoxyuridine (BrdU) pulsing experiments showed that 1 significantly enhanced BrdU-positive neurons. Taken together, our data suggest that 1 promotes neurogenesis by the induction of final cell division during NSC differentiation.

  14. MAPK inhibitors, particularly the JNK inhibitor, increase cell death effects in H2O2-treated lung cancer cells via increased superoxide anion and glutathione depletion.

    Science.gov (United States)

    Park, Woo Hyun

    2018-02-01

    Reactive oxygen species (ROS), especially hydrogen peroxide (H2O2), induce apoptosis in cancer cells by regulating mitogen-activated protein kinase (MAPK) signaling pathways. The present study investigated the effects of MAPK inhibitors on cell growth and death as well as changes in ROS and glutathione (GSH) levels in H2O2-treated Calu-6 and A549 lung cancer cells. H2O2 inhibited growth and induced death of Calu-6 and A549 lung cancer cells. All MAPK inhibitors appeared to enhance growth inhibition in H2O2-treated Calu-6 and A549 lung cancer cells and increased the percentage of Annexin V-FITC-positive cells in these cancer cells. Among the MAPK inhibitors, a JNK inhibitor significantly augmented the loss of mitochondrial membrane potential (MMP; ΔΨm) in H2O2-treated Calu-6 and A549 lung cancer cells. Intracellular ROS levels were significantly increased in the H2O2-treated cells at 1 and 24 h. Only the JNK inhibitor increased ROS levels in the H2O2-treated cells at 1 h and all MAPK inhibitors raised superoxide anion levels in these cells at 24 h. In addition, H2O2 induced GSH depletion in Calu-6 and A549 cells and the JNK inhibitor significantly enhanced GSH depletion in H2O2‑treated cells. Each of the MAPK inhibitors altered ROS and GSH levels differently in the Calu-6 and A549 control cells. In conclusion, H2O2 induced growth inhibition and death in lung cancer cells through oxidative stress and depletion of GSH. The enhanced effect of MAPK inhibitors, especially the JNK inhibitor, on cell death in H2O2-treated lung cancer cells was correlated with increased O2•- levels and GSH depletion.

  15. Passivation Using Molecular Halides Increases Quantum Dot Solar Cell Performance

    KAUST Repository

    Lan, Xinzheng; Voznyy, Oleksandr; Kiani, Amirreza; Garcí a de Arquer, F. Pelayo; Abbas, Abdullah Saud; Kim, Gi-Hwan; Liu, Mengxia; Yang, Zhenyu; Walters, Grant; Xu, Jixian; Yuan, Mingjian; Ning, Zhijun; Fan, Fengjia; Kanjanaboos, Pongsakorn; Kramer, Illan; Zhitomirsky, David; Lee, Philip; Perelgut, Alexander; Hoogland, Sjoerd; Sargent, Edward H.

    2015-01-01

    © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim. Here we report a solution-based passivation scheme is developed featuring the use of molecular iodine and PbS colloidal quantum dots (CQDs). The improved passivation translates into a longer carrier diffusion length in the solid film. This allows thicker solar-cell devices to be built while preserving efficient charge collection, leading to a certified power conversion efficiency of 9.9%, which is a new record in CQD solar cells.

  16. Passivation Using Molecular Halides Increases Quantum Dot Solar Cell Performance.

    Science.gov (United States)

    Lan, Xinzheng; Voznyy, Oleksandr; Kiani, Amirreza; García de Arquer, F Pelayo; Abbas, Abdullah Saud; Kim, Gi-Hwan; Liu, Mengxia; Yang, Zhenyu; Walters, Grant; Xu, Jixian; Yuan, Mingjian; Ning, Zhijun; Fan, Fengjia; Kanjanaboos, Pongsakorn; Kramer, Illan; Zhitomirsky, David; Lee, Philip; Perelgut, Alexander; Hoogland, Sjoerd; Sargent, Edward H

    2016-01-13

    A solution-based passivation scheme is developed featuring the use of molecular iodine and PbS colloidal quantum dots (CQDs). The improved passivation translates into a longer carrier diffusion length in the solid film. This allows thicker solar-cell devices to be built while preserving efficient charge collection, leading to a certified power conversion efficiency of 9.9%, which is a new record in CQD solar cells. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  17. Passivation Using Molecular Halides Increases Quantum Dot Solar Cell Performance

    KAUST Repository

    Lan, Xinzheng

    2015-11-18

    © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim. Here we report a solution-based passivation scheme is developed featuring the use of molecular iodine and PbS colloidal quantum dots (CQDs). The improved passivation translates into a longer carrier diffusion length in the solid film. This allows thicker solar-cell devices to be built while preserving efficient charge collection, leading to a certified power conversion efficiency of 9.9%, which is a new record in CQD solar cells.

  18. Increased expression of T-helper cell activation markers in ...

    African Journals Online (AJOL)

    Ehab

    expression of these activation markers would be of value in monitoring asthma severity and the response to ... Key words: Children, atopic asthma, T-helper cell subsets, glucocorticoid inhalation, lower respiratory infections, CD45RO ...... budesonide, and placebo on mucosal inflammation and clinical indices in mild asthma.

  19. Anticoagulant drugs increase natural killer cell activity in lung cancer

    Czech Academy of Sciences Publication Activity Database

    Bobek, M.; Boubelík, Michael; Fišerová, Anna; Luptovcová, Martina; Vannucci, Luca; Kacprzak, G.; Kolodzej, J.; Majewski, A.M.; Hoffman, R. M.

    2005-01-01

    Roč. 47, č. 2 (2005), s. 215-223 ISSN 0169-5002 Institutional research plan: CEZ:AV0Z5052915 Keywords : anticoagulant drugs * lung cancer * NK cells Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 3.172, year: 2005

  20. Astaxanthin increases progesterone production in cultured bovine luteal cells.

    Science.gov (United States)

    Kamada, Hachiro; Akagi, Satoshi; Watanabe, Shinya

    2017-06-29

    Although astaxanthin (AST) is known to be a strong antioxidant, its effects on reproductive function in domestic animals have not yet been elucidated in detail. Therefore, we investigated the effects of AST on luteal cells, which produce progesterone (P4), an important hormone for maintaining pregnancy. Luteal cells were prepared by collagenase dispersion of the corpus luteum (CL). The addition of racemic AST at a low concentration (production than RR-AST. When 1 mg/kg·body weight of SS-AST derived from green algae was fed to cows for 2 weeks, its concentration in blood plasma was 10.9 nM on average, which was sufficient to expect an in vitro effect on the production of P4 in cows. These results suggested the potential of SS-AST supplements for cows to elevate luteal function.

  1. Increased T cell trafficking as adjunct therapy for HIV-1

    Science.gov (United States)

    Wolinsky, Steven M.; McLean, Angela R.

    2018-01-01

    Although antiretroviral drug therapy suppresses human immunodeficiency virus-type 1 (HIV-1) to undetectable levels in the blood of treated individuals, reservoirs of replication competent HIV-1 endure. Upon cessation of antiretroviral therapy, the reservoir usually allows outgrowth of virus and approaches to targeting the reservoir have had limited success. Ongoing cycles of viral replication in regions with low drug penetration contribute to this persistence. Here, we use a mathematical model to illustrate a new approach to eliminating the part of the reservoir attributable to persistent replication in drug sanctuaries. Reducing the residency time of CD4 T cells in drug sanctuaries renders ongoing replication unsustainable in those sanctuaries. We hypothesize that, in combination with antiretroviral drugs, a strategy to orchestrate CD4 T cell trafficking could contribute to a functional cure for HIV-1 infection. PMID:29499057

  2. Growth hormone increases vascular cell adhesion molecule 1 expression

    DEFF Research Database (Denmark)

    Hansen, Troels Krarup; Fisker, Sanne; Dall, Rolf

    2004-01-01

    and 25 adult GH-deficient (GHD) patients randomized to GH treatment or placebo. Furthermore, we studied the direct effect of GH and IGF-I and serum from GH-treated subjects on basal and TNF alpha-stimulated expression of VCAM-1 and E-selectin on cultured human umbilical vein endothelial cells. Baseline...... levels of VCAM-1, but not E-selectin, were significantly lower in GHD patients than in healthy subjects (362 +/- 15 microg/liter vs. 516 +/- 21 microg/liter, P liter (95......% confidence interval: 95.0-208.7 microg/liter); P

  3. Mesenchymal stem cells increase T-regulatory cells and improve healing following trauma and hemorrhagic shock.

    Science.gov (United States)

    Gore, Amy V; Bible, Letitia E; Song, Kimberly; Livingston, David H; Mohr, Alicia M; Sifri, Ziad C

    2015-07-01

    Rodent lungs undergo full histologic recovery within 1 week following unilateral lung contusion (LC). However, when LC is followed by hemorrhagic shock (HS), healing is impaired. We hypothesize that the intravenous administration of mesenchymal stem cells (MSCs) in animals undergoing combined LC followed by HS (LCHS) will improve wound healing. Male Sprague-Dawley rats (n = 5-6 per group) were subjected to LCHS with or without the injection of a single intravenous dose of 5 × 10 MSCs following return of shed blood after HS. Rats were sacrificed 7 days following injury. Flow cytometry was used to determine the T-regulatory cell (Treg) population in peripheral blood. Lung histology was graded using a well-established lung injury score (LIS). Components of the LIS include average inflammatory cells per high-power field over 30 fields, interstitial edema, pulmonary edema, and alveolar integrity, with total scores ranging from 0 to 11. Data were analyzed by analysis of variance followed by Tukey's multiple comparison test, expressed as mean (SD). p healing with an LIS unchanged from naive. The addition of HS resulted in a persistently elevated LIS score, whereas the addition of MSCs to LCHS decreased the LIS score back to naive levels. The change in LIS was driven by a significant decrease in edema scores. In rats undergoing LC alone, 10.5% (3.3%) of CD4 cells were Tregs. The addition of HS caused no significant change in Treg population (9.3% [0.7%]), whereas LCHS + MSC significantly increased the population to 18.2% (6.8%) in peripheral blood (p healing following trauma and HS is improved by a single dose of MSCs given immediately after injury. This enhanced healing is associated with an increase in the Treg population and a significant decrease in lung edema score as compared with animals undergoing LCHS. Further study into the role of Tregs in MSC-mediated wound healing is warranted.

  4. Cancer-initiating cells derived from established cervical cell lines exhibit stem-cell markers and increased radioresistance

    International Nuclear Information System (INIS)

    López, Jacqueline; Poitevin, Adela; Mendoza-Martínez, Veverly; Pérez-Plasencia, Carlos; García-Carrancá, Alejandro

    2012-01-01

    Cancer-initiating cells (CICs) are proposed to be responsible for the generation of metastasis and resistance to therapy. Accumulating evidences indicates CICs are found among different human cancers and cell lines derived from them. Few studies address the characteristics of CICs in cervical cancer. We identify biological features of CICs from four of the best-know human cell lines from uterine cervix tumors. (HeLa, SiHa, Ca Ski, C-4 I). Cells were cultured as spheres under stem-cell conditions. Flow cytometry was used to detect expression of CD34, CD49f and CD133 antigens and Hoechst 33342 staining to identify side population (SP). Magnetic and fluorescence-activated cell sorting was applied to enrich and purify populations used to evaluate tumorigenicity in nude mice. cDNA microarray analysis and in vitro radioresistance assay were carried out under standard conditions. CICs, enriched as spheroids, were capable to generate reproducible tumor phenotypes in nu-nu mice and serial propagation. Injection of 1 × 10 3 dissociated spheroid cells induced tumors in the majority of animals, whereas injection of 1 × 10 5 monolayer cells remained nontumorigenic. Sphere-derived CICs expressed CD49f surface marker. Gene profiling analysis of HeLa and SiHa spheroid cells showed up-regulation of CICs markers characteristic of the female reproductive system. Importantly, epithelial to mesenchymal (EMT) transition-associated markers were found highly expressed in spheroid cells. More importantly, gene expression analysis indicated that genes required for radioresistance were also up-regulated, including components of the double-strand break (DSB) DNA repair machinery and the metabolism of reactive oxygen species (ROS). Dose-dependent radiation assay indicated indeed that CICs-enriched populations exhibit an increased resistance to ionizing radiation (IR). We characterized a self-renewing subpopulation of CICs found among four well known human cancer-derived cell lines (HeLa, Si

  5. Using Fuel Cells to Increase the Range of Battery Electric Vehicles | News

    Science.gov (United States)

    | NREL Using Fuel Cells to Increase the Range of Battery Electric Vehicles Using Fuel Cells to significantly lower capital and lifecycle costs than additional battery capacity alone. And while fuel-cell -Duty Battery Electric Vehicles through the Use of Hydrogen Fuel Cells"-presented at the Society of

  6. Cancer cells recovering from damage exhibit mitochondrial restructuring and increased aerobic glycolysis

    Energy Technology Data Exchange (ETDEWEB)

    Akakura, Shin; Ostrakhovitch, Elena; Sanokawa-Akakura, Reiko [Frontiers in Bioscience Research Institute in Aging and Cancer, University of California, Irvine, CA (United States); Tabibzadeh, Siamak, E-mail: fbs@bioscience.org [Frontiers in Bioscience Research Institute in Aging and Cancer, University of California, Irvine, CA (United States); Dept of Oncologic Radiology, University of California, Irvine, CA (United States)

    2014-06-13

    Highlights: • Some cancer cells recover from severe damage that causes cell death in majority of cells. • Damage-Recovered (DR) cancer cells show reduced mitochondria, mDNA and mitochondrial enzymes. • DR cells show increased aerobic glycolysis, ATP, cell proliferation, and resistance to damage. • DR cells recovered from in vivo damage also show increased glycolysis and proliferation rate. - Abstract: Instead of relying on mitochondrial oxidative phosphorylation, most cancer cells rely heavily on aerobic glycolysis, a phenomenon termed as “the Warburg effect”. We considered that this effect is a direct consequence of damage which persists in cancer cells that recover from damage. To this end, we studied glycolysis and rate of cell proliferation in cancer cells that recovered from severe damage. We show that in vitro Damage-Recovered (DR) cells exhibit mitochondrial structural remodeling, display Warburg effect, and show increased in vitro and in vivo proliferation and tolerance to damage. To test whether cancer cells derived from tumor microenvironment can show similar properties, we isolated Damage-Recovered (T{sup DR}) cells from tumors. We demonstrate that T{sup DR} cells also show increased aerobic glycolysis and a high proliferation rate. These findings show that Warburg effect and its consequences are induced in cancer cells that survive severe damage.

  7. Increasing RpoS expression causes cell death in Borrelia burgdorferi.

    Directory of Open Access Journals (Sweden)

    Linxu Chen

    Full Text Available RpoS, one of the two alternative σ factors in Borrelia burgdorferi, is tightly controlled by multiple regulators and, in turn, determines expression of many critical virulence factors. Here we show that increasing RpoS expression causes cell death. The immediate effect of increasing RpoS expression was to promote bacterial division and as a consequence result in a rapid increase in cell number before causing bacterial death. No DNA fragmentation or degradation was observed during this induced cell death. Cryo-electron microscopy showed induced cells first formed blebs, which were eventually released from dying cells. Apparently blebbing initiated cell disintegration leading to cell death. These findings led us to hypothesize that increasing RpoS expression triggers intracellular programs and/or pathways that cause spirochete death. The potential biological significance of induced cell death may help B. burgdorferi regulate its population to maintain its life cycle in nature.

  8. Maximal exercise increases mucosal associated invariant T cell frequency and number in healthy young men.

    Science.gov (United States)

    Hanson, Erik D; Danson, Eli; Nguyen-Robertson, Catriona V; Fyfe, Jackson J; Stepto, Nigel K; Bartlett, David B; Sakkal, Samy

    2017-11-01

    Mucosal associated invariant T (MAIT) cells have properties of the innate and acquired immune systems. While the response to vigorous exercise has been established for most leukocytes, MAIT cells have not been investigated. Therefore, the purpose was to determine if MAIT cell lymphocytosis occurs with acute maximal aerobic exercise and if this response is influenced by exercise duration, cardiovascular fitness, or body composition. Twenty healthy young males with moderate fitness levels performed an extended graded exercise test until volitional fatigue. Peripheral blood mononuclear cells were isolated from venous blood obtained prior and immediately after exercise and were labeled to identify specific T cell populations using flow cytometry. The percentage of MAIT cells relative to total T cells significantly increased from 3.0 to 3.8% and absolute MAIT cell counts increased by 2.2-fold following maximal exercise. MAIT cell subpopulation proportions were unchanged with exercise. Within cytotoxic T lymphocytes (CTL), MAIT cells consisted of 8% of these cells and this remained constant after exercise. MAIT cell counts and changes with exercise were not affected by body composition, VO 2peak , or exercise duration. Maximal exercise doubled MAIT cell numbers and showed preferential mobilization within total T cells but the response was not influenced by fitness levels, exercise duration, or body composition. These results suggest that acute exercise could be used to offset MAIT cell deficiencies observed with certain pathologies. MAIT cells also make up a substantial proportion of CTLs, which may have implications for cytotoxicity assays using these cells.

  9. Increasing Human Neural Stem Cell Transplantation Dose Alters Oligodendroglial and Neuronal Differentiation after Spinal Cord Injury

    Directory of Open Access Journals (Sweden)

    Katja M. Piltti

    2017-06-01

    Full Text Available Multipotent human central nervous system-derived neural stem cells transplanted at doses ranging from 10,000 (low to 500,000 (very high cells differentiated predominantly into the oligodendroglial lineage. However, while the number of engrafted cells increased linearly in relationship to increasing dose, the proportion of oligodendrocytic cells declined. Increasing dose resulted in a plateau of engraftment, enhanced neuronal differentiation, and increased distal migration caudal to the transplantation sites. Dose had no effect on terminal sensory recovery or open-field locomotor scores. However, total human cell number and decreased oligodendroglial proportion were correlated with hindlimb girdle coupling errors. Conversely, greater oligodendroglial proportion was correlated with increased Ab step pattern, decreased swing speed, and increased paw intensity, consistent with improved recovery. These data suggest that transplant dose, and/or target niche parameters can regulate donor cell engraftment, differentiation/maturation, and lineage-specific migration profiles.

  10. Treatment of HeLa cells with Giloe (Tinospora cordifolia meirs) increases the radiosensitivity by increasing DNA damage

    International Nuclear Information System (INIS)

    Varma, Hari Krishna; Jagetia, Ganesh Chandra; Nayak, Vijayashree

    2014-01-01

    Radiotherapy is an important treatment modality and screening of phytoceuticals may enhance the clinical outcome of radiotherapy, therefore radiosensitizing activity of various guduchi (Tinospora cordifolia) extracts was studied in HeLa cells. Chromosomal aberrations were scored in HeLa cells treated with 10 μg/ml of aqueous, methanol, or methylene chloride guduchi extracts or doxorubicin before exposure to 0, 0.5, 1, 2 or 3 Gy of γ-radiation at 12, 24, 36 or 48 h post-irradiation. Irradiation of HeLa cells caused a dose dependent rise in the chromatid breaks, chromosome breaks, dicentric, centric rings, acentric fragments and total aberrations at all post-irradiation times and the dose response was linear quadratic for all types of aberrations scored. Chromatid breaks increased up to 12 h post-irradiation and declined steadily up to 48 h post-irradiation, whereas chromosome breaks, dicentric, acentric fragments and total aberrations elevated up to 24 h post-irradiation and declined thereafter. However, centric rings continued to rise steadily up to 48 h post-irradiation. Treatment of HeLa cells with aqueous, methanol or methylene chloride guduchi extract or doxorubicin before irradiation significantly enhanced various types of chromosomal aberrations and a maximum rise in the chromosome aberrations was observed in the HeLa cells treated with methylene chloride extract before irradiation when compared to other groups. Various guduchi extracts enhanced the effect of radiation in HeLa cells by increasing the molecular damage to cellular genome and their effect was similar to or even greater than doxorubicin (positive control) pretreatment, depending on the type of guduchi extract used. (author)

  11. Follicle stimulating hormone increases spermatogonial stem cell colonization during in vitro co-culture.

    Science.gov (United States)

    Narenji Sani, Reza; Tajik, Parviz; Yousefi, Mohammad Hassan; Movahedin, Mansoureh; Qasemi-Panahi, Babak; Shafiei, Shiva; Ahmadi Hamedani, Mahmood

    2013-01-01

    The complex process of spermatogenesis is regulated by various factors. Studies on spermatogonial stem cells (SCCs) have provided very important tool to improve herd genetic and different field. 0.2 to 0.3 percent of total cells of seminiferous tubules is consist of spermatogonial stem cells. To investigate and biomanipulation of these cells, proliferation and viability rate of cells should be increased in vitro, at first. Follicle stimulating hormone (FSH) has been suggested to play a determinant role in the survival of germ cells in addition to increasing spermatogonial proliferation. In this study, the in vitro effects of FSH on spermatogonial cell colony formation were investigated. Sertoli and spermatogonial cells were isolated from 3-5 months old calves. The identity of the Sertoli cells and spermatogonial stem cells were confirmed through immunocytochemistry and colony morphology, respectively. Co-cultured Sertoli and spermatogonial cells were treated with FSH in different dose of 10, 20 and 40 IU mL(-1) FSH, before colony assay. Results indicated that, FSH increased in vitro colonization of spermatogonial cells in comparison with control group. In conclusion, using FSH provided proper bovine spermatogonial stem cell culture medium for in vitro study of these cells.

  12. Tumor cell adhesion to endothelial cells is increased by endotoxin via an upregulation of beta-1 integrin expression.

    LENUS (Irish Health Repository)

    Andrews, E J

    2012-02-03

    BACKGROUND: Recent studies have demonstrated that metastatic disease develops from tumor cells that adhere to endothelial cells and proliferate intravascularly. The beta-1 integrin family and its ligand laminin have been shown to be important in tumor-to-endothelial cell adhesion. Lipopolysaccharide (LPS) has been implicated in the increased metastatic tumor growth that is seen postoperatively. We postulated that LPS increases tumor cell expression of beta-1 integrins and that this leads to increased adhesion. METHODS: The human metastatic colon cancer cell line LS174T was labeled with an enhanced green fluorescent protein (eGFP) using retroviral transfection. Cell cultures were treated with LPS for 1, 2, and 4 h (n = 6 each) and were subsequently cocultured for 30 or 120 min with confluent human umbilical vein endothelial cells (HUVECs), to allow adherence. Adherent tumor cells were counted using fluorescence microscopy. These experiments were carried out in the presence or absence of a functional blocking beta-1 integrin monoclonal antibody (4B4). Expression of beta-1 integrin and laminin on tumor and HUVECs was assessed using flow cytometric analysis. Tumor cell NF-kappaB activation after incubation with LPS was measured. RESULTS: Tumor cell and HUVEC beta-1 integrin expression and HUVEC expression of laminin were significantly (P < 0.05) enhanced after incubation with LPS. Tumor cell adhesion to HUVECs was significantly increased. Addition of the beta-1 integrin blocking antibody reduced tumor cell adhesion to control levels. LPS increased tumor cell NF-kappaB activation. CONCLUSIONS: Exposure to LPS increases tumor cell adhesion to the endothelium through a beta-1 integrin-mediated pathway that is NF-kappaB dependent. This may provide a target for immunotherapy directed at reducing postoperative metastatic tumor growth.

  13. Increased dermal mast cell prevalence and susceptibility to development of basal cell carcinoma in humans

    DEFF Research Database (Denmark)

    Grimbaldeston, Michele A; Skov, Lone; Finlay-Jones, John J

    2002-01-01

    eliminate them. Studies in a range of inbred mouse strains as well as mast cell-depleted mice reconstituted with mast cell precursors support a functional link between histamine-staining dermal mast cells and the extent of susceptibility to UVB-induced systemic immunomodulation. Humans, like mouse strains......, display variations in dermal mast cell prevalence. In a study of Danish and South Australian BCC patients and control subjects, one 4-mm punch biopsy of non-sun-exposed buttock skin was sampled from each participant. This skin site was investigated to avoid any changes in mast cell prevalence caused...... by sun exposure. Two sections (4 microm) per biopsy were immunohistochemically stained for detection of histamine-containing dermal mast cells. Computer-generated image analysis evaluated dermal mast cell prevalence in both sections by quantifying the total number of mast cells according to the total...

  14. Human umbilical cord mesenchymal stem cells increase interleukin-9 production of CD4+ T cells

    Science.gov (United States)

    Yang, Zhou Xin; Chi, Ying; Ji, Yue Ru; Wang, You Wei; Zhang, Jing; Luo, Wei Feng; Li, Li Na; Hu, Cai Dong; Zhuo, Guang Sheng; Wang, Li Fang; Han, Zhi-Bo; Han, Zhong Chao

    2017-01-01

    Mesenchymal stem cells (MSC) are able to differentiate into cells of multiple lineage, and additionally act to modulate the immune response. Interleukin (IL)-9 is primarily produced by cluster of differentiation (CD)4+ T cells to regulate the immune response. The present study aimed to investigate the effect of human umbilical cord derived-MSC (UC-MSC) on IL-9 production of human CD4+ T cells. It was demonstrated that the addition of UC-MSC to the culture of CD4+ T cells significantly enhanced IL-9 production by CD4+ T cells. Transwell experiments suggested that UC-MSC promotion of IL-9 production by CD4+ T cells was dependent on cell-cell contact. Upregulated expression of CD106 was observed in UC-MSC co-cultured with CD4+ T cells, and the addition of a blocking antibody of CD106 significantly impaired the ability of UC-MSC to promote IL-9 production by CD4+ T cells. Therefore, the results of the present study demonstrated that UC-MSC promoted the generation of IL-9 producing cells, which may be mediated, in part by CD106. The findings may act to expand understanding and knowledge of the immune modulatory role of UC-MSC. PMID:29042945

  15. Increased osmolarity and cell clustering preserve canine notochordal cell phenotype in culture

    NARCIS (Netherlands)

    Spillekom, S.; Smolders, L.A.; Grinwis, G.C.M.; Arkesteijn, I.T.M.; Ito, K.; Meij, B.P.; Tryfonidou, M.A.

    2014-01-01

    Degeneration of the intervertebral disc (IVD) is associated with a loss of notochordal cells (NCs) from the nucleus pulposus (NP) and their replacement by chondrocyte-like cells. NCs are known to maintain extracellular matrix quality and stimulate the chondrocyte-like NP cells, making NCs attractive

  16. Increasing ethanol productivity during xylose fermentation by cell recycling of recombinant Saccharomyces cerevisiae

    DEFF Research Database (Denmark)

    Roca, Christophe Francois Aime; Olsson, Lisbeth

    2003-01-01

    The influence of cell recycling of xylose-fermenting Saccharomyces cerevisiae TMB3001 was investigated during continuous cultivation on a xylose-glucose mixture. By using cell recycling at the dilution rate (D) of 0.05 h(-1), the cell-mass concentration could be increased from 2.2 g l(-1) to 22 g l...... ethanol productivity was in the range of 0.23-0.26 g g(-1) h(-1) with or without cell recycling, showing that an increased cell-mass concentration did not influence the efficiency of the yeast....

  17. Human umbilical cord mesenchymal stem cells increase interleukin-9 production of CD4+ T cells

    OpenAIRE

    Yang, Zhou Xin; Chi, Ying; Ji, Yue Ru; Wang, You Wei; Zhang, Jing; Luo, Wei Feng; Li, Li Na; Hu, Cai Dong; Zhuo, Guang Sheng; Wang, Li Fang; Han, Zhi-Bo; Han, Zhong Chao

    2017-01-01

    Mesenchymal stem cells (MSC) are able to differentiate into cells of multiple lineage, and additionally act to modulate the immune response. Interleukin (IL)-9 is primarily produced by cluster of differentiation (CD)4+ T cells to regulate the immune response. The present study aimed to investigate the effect of human umbilical cord derived-MSC (UC-MSC) on IL-9 production of human CD4+ T cells. It was demonstrated that the addition of UC-MSC to the culture of CD4+ T cells significantly enhance...

  18. Adrenaline promotes cell proliferation and increases chemoresistance in colon cancer HT29 cells through induction of miR-155

    International Nuclear Information System (INIS)

    Pu, Jun; Bai, Danna; Yang, Xia; Lu, Xiaozhao; Xu, Lijuan; Lu, Jianguo

    2012-01-01

    Highlights: ► Adrenaline increases colon cancer cell proliferation and its resistance to cisplatin. ► Adrenaline activates NFκB in a dose dependent manner. ► NFκB–miR-155 pathway contributes to cell proliferation and resistance to cisplatin. -- Abstract: Recently, catecholamines have been described as being involved in the regulation of cancer genesis and progression. Here, we reported that adrenaline increased the cell proliferation and decreased the cisplatin induced apoptosis in HT29 cells. Further study found that adrenaline increased miR-155 expression in an NFκB dependent manner. HT29 cells overexpressing miR-155 had a higher cell growth rate and more resistance to cisplatin induced apoptosis. In contrast, HT29 cells overexpressing miR-155 inhibitor displayed decreased cell proliferation and sensitivity to cisplatin induced cell death. In summary, our study here revealed that adrenaline–NFκB–miR-155 pathway at least partially contributes to the psychological stress induced proliferation and chemoresistance in HT29 cells, shedding light on increasing the therapeutic strategies of cancer chemotherapy.

  19. Adrenaline promotes cell proliferation and increases chemoresistance in colon cancer HT29 cells through induction of miR-155

    Energy Technology Data Exchange (ETDEWEB)

    Pu, Jun [Department of General Surgery, Tangdu Hospital of the Fourth Military Medical University, Xi' an 710038 (China); Bai, Danna [Department of Cardiology, 323 Hospital of PLA, Xi' an 710054 (China); Yang, Xia [Department of Teaching and Medical Administration, Tangdu Hospital of the Fourth Military Medical University, Xi' an 710038 (China); Lu, Xiaozhao [Department of Nephrology, The 323 Hospital of PLA, Xi' an 710054 (China); Xu, Lijuan, E-mail: 13609296272@163.com [Department of Nephrology, The 323 Hospital of PLA, Xi' an 710054 (China); Lu, Jianguo, E-mail: lujianguo029@yahoo.com.cn [Department of General Surgery, Tangdu Hospital of the Fourth Military Medical University, Xi' an 710038 (China)

    2012-11-16

    Highlights: Black-Right-Pointing-Pointer Adrenaline increases colon cancer cell proliferation and its resistance to cisplatin. Black-Right-Pointing-Pointer Adrenaline activates NF{kappa}B in a dose dependent manner. Black-Right-Pointing-Pointer NF{kappa}B-miR-155 pathway contributes to cell proliferation and resistance to cisplatin. -- Abstract: Recently, catecholamines have been described as being involved in the regulation of cancer genesis and progression. Here, we reported that adrenaline increased the cell proliferation and decreased the cisplatin induced apoptosis in HT29 cells. Further study found that adrenaline increased miR-155 expression in an NF{kappa}B dependent manner. HT29 cells overexpressing miR-155 had a higher cell growth rate and more resistance to cisplatin induced apoptosis. In contrast, HT29 cells overexpressing miR-155 inhibitor displayed decreased cell proliferation and sensitivity to cisplatin induced cell death. In summary, our study here revealed that adrenaline-NF{kappa}B-miR-155 pathway at least partially contributes to the psychological stress induced proliferation and chemoresistance in HT29 cells, shedding light on increasing the therapeutic strategies of cancer chemotherapy.

  20. Increasing procaspase 8 expression using repurposed drugs to induce HIV infected cell death in ex vivo patient cells.

    Directory of Open Access Journals (Sweden)

    Rahul Sampath

    Full Text Available HIV persists because a reservoir of latently infected CD4 T cells do not express viral proteins and are indistinguishable from uninfected cells. One approach to HIV cure suggests that reactivating HIV will activate cytotoxic pathways; yet when tested in vivo, reactivating cells do not die sufficiently to reduce cell-associated HIV DNA levels. We recently showed that following reactivation from latency, HIV infected cells generate the HIV specific cytotoxic protein Casp8p41 which is produced by HIV protease cleaving procaspase 8. However, cell death is prevented, possibly due to low procaspase 8 expression. Here, we tested whether increasing procaspase 8 levels in CD4 T cells will produce more Casp8p41 following HIV reactivation, causing more reactivated cells to die. Screening 1277 FDA approved drugs identified 168 that increased procaspase 8 expression by at least 1.7-fold. Of these 30 were tested for anti-HIV effects in an acute HIVIIIb infection model, and 9 drugs at physiologic relevant levels significantly reduced cell-associated HIV DNA. Primary CD4 T cells from ART suppressed HIV patients were treated with one of these 9 drugs and reactivated with αCD3/αCD28. Four drugs significantly increased Casp8p41 levels following HIV reactivation, and decreased total cell associated HIV DNA levels (flurbiprofen: p = 0.014; doxycycline: p = 0.044; indomethacin: p = 0.025; bezafibrate: P = 0.018 without effecting the viability of uninfected cells. Thus procaspase 8 levels can be increased pharmacologically and, in the context of HIV reactivation, increase Casp8p41 causing death of reactivating cells and decreased HIV DNA levels. Future studies will be required to define the clinical utility of this or similar approaches.

  1. Increasing cell-device adherence using cultured insect cells for receptor-based biosensors

    Science.gov (United States)

    Terutsuki, Daigo; Mitsuno, Hidefumi; Sakurai, Takeshi; Okamoto, Yuki; Tixier-Mita, Agnès; Toshiyoshi, Hiroshi; Mita, Yoshio; Kanzaki, Ryohei

    2018-03-01

    Field-effect transistor (FET)-based biosensors have a wide range of applications, and a bio-FET odorant sensor, based on insect (Sf21) cells expressing insect odorant receptors (ORs) with sensitivity and selectivity, has emerged. To fully realize the practical application of bio-FET odorant sensors, knowledge of the cell-device interface for efficient signal transfer, and a reliable and low-cost measurement system using the commercial complementary metal-oxide semiconductor (CMOS) foundry process, will be indispensable. However, the interfaces between Sf21 cells and sensor devices are largely unknown, and electrode materials used in the commercial CMOS foundry process are generally limited to aluminium, which is reportedly toxic to cells. In this study, we investigated Sf21 cell-device interfaces by developing cross-sectional specimens. Calcium imaging of Sf21 cells expressing insect ORs was used to verify the functions of Sf21 cells as odorant sensor elements on the electrode materials. We found that the cell-device interface was approximately 10 nm wide on average, suggesting that the adhesion mechanism of Sf21 cells may differ from that of other cells. These results will help to construct accurate signal detection from expressed insect ORs using FETs.

  2. [Astragalus polysaccharide may increase sensitivity of cervical cancer HeLa cells to cisplatin by regulating cell autophagy].

    Science.gov (United States)

    Zhai, Qiu-Li; Hu, Xiang-Dan; Xiao, Jing; Yu, Dong-Qing

    2018-02-01

    This study aimed to investigate the possible sensitivity of Astragalus polysaccharides, in order to improve the chemosensitivity of cervical cancer HeLa cells to cisplatin by regulating the cell autophagy, and explore its possible mechanism. In this study, HeLa cells were divided into control group, cisplatin group, Astragalus polysaccharide group, and Astragalus polysaccharide combined with cisplatin group. MTT assay was used to detect the proliferation of cervical cancer HeLa cells. Flow cytometry was used to detect the apoptosis and cycle of HeLa cells in each experimental group. RT-PCR was used to detect the mRNA expression of autophagy-related proteins beclin1, LC3Ⅱ and p62. The expression levels of autophagy-related proteins beclin1, LC3Ⅱ, LC3Ⅰ and p62 were detected by WB method. MTT results showed that compared with the control group, the proliferation of HeLa cells was significantly inhibited in each administration group( P HeLa cells was significantly increased( P HeLa cells to cisplatin by regulating the cell autophagy. Its possible mechanism of action is correlated with the up-regulation of autophagy-related proteins beclin1, the promote the conversion from LC3Ⅰ to LC3Ⅱ, the down-regulation of labeled protein p62, and the enhancement of HeLa cell autophagic activity, thereby increasing the sensitivity of HeLa cells to cisplatin chemotherapy. Copyright© by the Chinese Pharmaceutical Association.

  3. Membrane fluidity increases during apoptosis of sheep ileal Peyer's patch B cells

    International Nuclear Information System (INIS)

    Jourd'heuil, D.; Aspinall, A.; Reynolds, J.D.; Meddings, J.B.

    1996-01-01

    To investigate specific plasma membrane structural changes associated with apoptosis, whole cells and purified plasma membranes of apoptotic B cells from the ileal Peyer's patch of sheep were analyzed for their 'membrane fluidity'. The ileal Peyer's patch of sheep provided a large number of B cells required for plasma membrane isolation (>5 x 10 9 ). As the incidence of apoptosis increased with time of culture, the fluidity of purified plasma membranes, as measured with the fluorophore DPH (diphenylhexatriene), increased. To evaluate this phenomenon with intact cells, B cells at different apoptotic stages were fractionated on discontinuous Percoll gradients. Similar results were obtained using the fluorophore TMA-DPH (trimethylammoniumdiphenylhexatriene), which has been shown to localize specifically to the plasma membrane. Functionally, the increase in plasma membrane fluidity associated with apoptosis may represent either a mechanism to cycle phosphatidylserine to the outer leaflet, mediating phagocytic recognition of apoptotic cells, or a consequence of this event. (author). 20 refs., 1 tab., 4 figs

  4. β-cell dysfunction due to increased ER stress in a stem cell model of Wolfram syndrome.

    Science.gov (United States)

    Shang, Linshan; Hua, Haiqing; Foo, Kylie; Martinez, Hector; Watanabe, Kazuhisa; Zimmer, Matthew; Kahler, David J; Freeby, Matthew; Chung, Wendy; LeDuc, Charles; Goland, Robin; Leibel, Rudolph L; Egli, Dieter

    2014-03-01

    Wolfram syndrome is an autosomal recessive disorder caused by mutations in WFS1 and is characterized by insulin-dependent diabetes mellitus, optic atrophy, and deafness. To investigate the cause of β-cell failure, we used induced pluripotent stem cells to create insulin-producing cells from individuals with Wolfram syndrome. WFS1-deficient β-cells showed increased levels of endoplasmic reticulum (ER) stress molecules and decreased insulin content. Upon exposure to experimental ER stress, Wolfram β-cells showed impaired insulin processing and failed to increase insulin secretion in response to glucose and other secretagogues. Importantly, 4-phenyl butyric acid, a chemical protein folding and trafficking chaperone, restored normal insulin synthesis and the ability to upregulate insulin secretion. These studies show that ER stress plays a central role in β-cell failure in Wolfram syndrome and indicate that chemical chaperones might have therapeutic relevance under conditions of ER stress in Wolfram syndrome and other forms of diabetes.

  5. Increased centrosome amplification in aged stem cells of the Drosophila midgut

    International Nuclear Information System (INIS)

    Park, Joung-Sun; Pyo, Jung-Hoon; Na, Hyun-Jin; Jeon, Ho-Jun; Kim, Young-Shin; Arking, Robert; Yoo, Mi-Ae

    2014-01-01

    Highlights: • Increased centrosome amplification in ISCs of aged Drosophila midguts. • Increased centrosome amplification in ISCs of oxidative stressed Drosophila midguts. • Increased centrosome amplification in ISCs by overexpression of PVR, EGFR, and AKT. • Supernumerary centrosomes can be responsible for abnormal ISC polyploid cells. • Supernumerary centrosomes can be a useful marker for aging stem cells. - Abstract: Age-related changes in long-lived tissue-resident stem cells may be tightly linked to aging and age-related diseases such as cancer. Centrosomes play key roles in cell proliferation, differentiation and migration. Supernumerary centrosomes are known to be an early event in tumorigenesis and senescence. However, the age-related changes of centrosome duplication in tissue-resident stem cells in vivo remain unknown. Here, using anti-γ-tubulin and anti-PH3, we analyzed mitotic intestinal stem cells with supernumerary centrosomes in the adult Drosophila midgut, which may be a versatile model system for stem cell biology. The results showed increased centrosome amplification in intestinal stem cells of aged and oxidatively stressed Drosophila midguts. Increased centrosome amplification was detected by overexpression of PVR, EGFR, and AKT in intestinal stem cells/enteroblasts, known to mimic age-related changes including hyperproliferation of intestinal stem cells and hyperplasia in the midgut. Our data show the first direct evidence for the age-related increase of centrosome amplification in intestinal stem cells and suggest that the Drosophila midgut is an excellent model for studying molecular mechanisms underlying centrosome amplification in aging adult stem cells in vivo

  6. Increased centrosome amplification in aged stem cells of the Drosophila midgut

    Energy Technology Data Exchange (ETDEWEB)

    Park, Joung-Sun; Pyo, Jung-Hoon; Na, Hyun-Jin; Jeon, Ho-Jun; Kim, Young-Shin [Department of Molecular Biology, Pusan National University, Busan 609-735 (Korea, Republic of); Arking, Robert, E-mail: aa2210@wayne.edu [Department of Biological Sciences, Wayne State University, Detroit, MI 48202 (United States); Yoo, Mi-Ae, E-mail: mayoo@pusan.ac.kr [Department of Molecular Biology, Pusan National University, Busan 609-735 (Korea, Republic of)

    2014-07-25

    Highlights: • Increased centrosome amplification in ISCs of aged Drosophila midguts. • Increased centrosome amplification in ISCs of oxidative stressed Drosophila midguts. • Increased centrosome amplification in ISCs by overexpression of PVR, EGFR, and AKT. • Supernumerary centrosomes can be responsible for abnormal ISC polyploid cells. • Supernumerary centrosomes can be a useful marker for aging stem cells. - Abstract: Age-related changes in long-lived tissue-resident stem cells may be tightly linked to aging and age-related diseases such as cancer. Centrosomes play key roles in cell proliferation, differentiation and migration. Supernumerary centrosomes are known to be an early event in tumorigenesis and senescence. However, the age-related changes of centrosome duplication in tissue-resident stem cells in vivo remain unknown. Here, using anti-γ-tubulin and anti-PH3, we analyzed mitotic intestinal stem cells with supernumerary centrosomes in the adult Drosophila midgut, which may be a versatile model system for stem cell biology. The results showed increased centrosome amplification in intestinal stem cells of aged and oxidatively stressed Drosophila midguts. Increased centrosome amplification was detected by overexpression of PVR, EGFR, and AKT in intestinal stem cells/enteroblasts, known to mimic age-related changes including hyperproliferation of intestinal stem cells and hyperplasia in the midgut. Our data show the first direct evidence for the age-related increase of centrosome amplification in intestinal stem cells and suggest that the Drosophila midgut is an excellent model for studying molecular mechanisms underlying centrosome amplification in aging adult stem cells in vivo.

  7. Autophagy inhibitor chloroquine increases sensitivity to cisplatin in QBC939 cholangiocarcinoma cells by mitochondrial ROS.

    Directory of Open Access Journals (Sweden)

    Xianzhi Qu

    Full Text Available The tumor cells have some metabolic characteristics of the original tissues, and the metabolism of the tumor cells is closely related to autophagy. However, the mechanism of autophagy and metabolism in chemotherapeutic drug resistance is still poorly understood. In this study, we investigated the role and mechanism of autophagy and glucose metabolism in chemotherapeutic drug resistance by using cholangiocarcinoma QBC939 cells with primary cisplatin resistance and hepatocellular carcinoma HepG2 cells. We found that QBC939 cells with cisplatin resistance had a higher capacity for glucose uptake, consumption, and lactic acid generation, and higher activity of the pentose phosphate pathway compared with HepG2 cells, and the activity of PPP was further increased after cisplatin treatment in QBC939 cells. It is suggested that there are some differences in the metabolism of glucose in hepatocellular carcinoma and cholangiocarcinoma cells, and the activation of PPP pathway may be related to the drug resistance. Through the detection of autophagy substrates p62 and LC3, found that QBC939 cells have a higher flow of autophagy, autophagy inhibitor chloroquine can significantly increase the sensitivity of cisplatin in cholangiocarcinoma cells compared with hepatocellular carcinoma HepG2 cells. The mechanism may be related to the inhibition of QBC939 cells with higher activity of the PPP, the key enzyme G6PDH, which reduces the antioxidant capacity of cells and increases intracellular ROS, especially mitochondrial ROS. Therefore, we hypothesized that autophagy and the oxidative stress resistance mediated by glucose metabolism may be one of the causes of cisplatin resistance in cholangiocarcinoma cells. It is suggested that according to the metabolism characteristics of tumor cells, inhibition of autophagy lysosome pathway with chloroquine may be a new route for therapeutic agents against cholangiocarcinoma.

  8. Simvastatin and metformin inhibit cell growth in hepatitis C virus infected cells via mTOR increasing PTEN and autophagy.

    Directory of Open Access Journals (Sweden)

    José A Del Campo

    Full Text Available Hepatitis C virus (HCV infection has been related to increased risk of development of hepatocellular carcinoma (HCC while metformin (M and statins treatment seemed to protect against HCC development. In this work, we aim to identify the mechanisms by which metformin and simvastatin (S could protect from liver cancer. Huh7.5 cells were infected with HCV particles and treated with M+S. Human primary hepatocytes were treated with M+S. Treatment with both drugs inhibited Huh7.5 cell growth and HCV infection. In non-infected cells S increased translational controlled tumor protein (TCTP and phosphatase and tensin homolog (PTEN proteins while M inhibited mammalian target of rapamycin (mTOR and TCTP. Simvastatin and metformin co-administered down-regulated mTOR and TCTP, while PTEN was increased. In cells infected by HCV, mTOR, TCTP, p62 and light chain 3B II (LC3BII were increased and PTEN was decreased. S+M treatment increased PTEN, p62 and LC3BII in Huh7.5 cells. In human primary hepatocytes, metformin treatment inhibited mTOR and PTEN, but up-regulated p62, LC3BII and Caspase 3. In conclusion, simvastatin and metformin inhibited cell growth and HCV infection in vitro. In human hepatocytes, metformin increased cell-death markers. These findings suggest that M+S treatment could be useful in therapeutic prevention of HCV-related hepatocellular carcinoma.

  9. Vorinostat increases carboplatin and paclitaxel activity in non-small cell lung cancer cells

    OpenAIRE

    Owonikoko, Taofeek K.; Ramalingam, Suresh S.; Kanterewicz, Beatriz; Balius, Trent; Belani, Chandra P.; Hershberger, Pamela A.

    2010-01-01

    We observed a 53% response rate in non-small cell lung cancer (NSCLC) patients treated with vorinostat plus paclitaxel/carboplatin in a Phase I trial. Studies were undertaken to investigate the mechanism (s) underlying this activity. Growth inhibition was assessed in NSCLC cells by MTT assay after 72 h of continuous drug exposure. Vorinostat (1 µM) inhibited growth by: 17±7% in A549, 28±6% in 128-88T, 39±8% in Calu1, and 41±7% in 201T cells. Vorinostat addition to carboplatin or paclitaxel le...

  10. Curcumin Inhibits Growth of Human NCI-H292 Lung Squamous Cell Carcinoma Cells by Increasing FOXA2 Expression

    Directory of Open Access Journals (Sweden)

    Lingling Tang

    2018-02-01

    Full Text Available Lung squamous cell carcinoma (LSCC is a common histological lung cancer subtype, but unlike lung adenocarcinoma, limited therapeutic options are available for treatment. Curcumin, a natural compound, may have anticancer effects in various cancer cells, but how it may be used to treat LSCC has not been well studied. Here, we applied curcumin to a human NCI-H292 LSCC cell line to test anticancer effects and explored underlying potential mechanisms of action. Curcumin treatment inhibited NCI-H292 cell growth and increased FOXA2 expression in a time-dependent manner. FOXA2 expression was decreased in LSCC tissues compared with adjacent normal tissues and knockdown of FOXA2 increased NCI-H292 cells proliferation. Inhibition of cell proliferation by curcumin was attenuated by FOXA2 knockdown. Moreover inhibition of STAT3 pathways by curcumin increased FOXA2 expression in NCI-H292 cells whereas a STAT3 activator (IL-6 significantly inhibited curcumin-induced FOXA2 expression. Also, SOCS1 and SOCS3, negative regulators of STAT3 activity, were upregulated by curcumin treatment. Thus, curcumin inhibited human NCI-H292 cells growth by increasing FOXA2 expression via regulation of STAT3 signaling pathways.

  11. Altered features and increased chemosensitivity of human breast cancer cells mediated by adipose tissue-derived mesenchymal stromal cells

    International Nuclear Information System (INIS)

    Kucerova, Lucia; Skolekova, Svetlana; Matuskova, Miroslava; Bohac, Martin; Kozovska, Zuzana

    2013-01-01

    Mesenchymal stromal cells (MSCs) represent heterogeneous cell population suitable for cell therapies in regenerative medicine. MSCs can also substantially affect tumor biology due to their ability to be recruited to the tumor stroma and interact with malignant cells via direct contacts and paracrine signaling. The aim of our study was to characterize molecular changes dictated by adipose tissue-derived mesenchymal stromal cells (AT-MSCs) and the effects on drug responses in human breast cancer cells SKBR3. The tumor cells were either directly cocultured with AT-MSCs or exposed to MSCs-conditioned medium (MSC-CM). Changes in cell biology were evaluated by kinetic live cell imaging, fluorescent microscopy, scratch wound assay, expression analysis, cytokine secretion profiling, ATP-based viability and apoptosis assays. The efficiency of cytotoxic treatment in the presence of AT-MSCs or MSCs-CM was analyzed. The AT-MSCs altered tumor cell morphology, induced epithelial-to-mesenchymal transition, increased mammosphere formation, cell confluence and migration of SKBR3. These features were attributed to molecular changes induced by MSCs-secreted cytokines and chemokines in breast cancer cells. AT-MSCs significantly inhibited the proliferation of SKBR3 cells in direct cocultures which was shown to be dependent on the SDF-1α/CXCR4 signaling axis. MSC-CM-exposed SKBR3 or SKBR3 in direct coculture with AT-MSCs exhibited increased chemosensitivity and induction of apoptosis in response to doxorubicin and 5-fluorouracil. Our work further highlights the multi-level nature of tumor-stromal cell interplay and demonstrates the capability of AT-MSCs and MSC-secreted factors to alter the anti-tumor drug responses

  12. Follicle stimulating hormone increases spermatogonial stem cell colonization during in vitro co - culture

    Directory of Open Access Journals (Sweden)

    Reza Narenji Sani

    2013-03-01

    Full Text Available The complex process of spermatogenesis is regulated by various factors. Studies onspermatogonial stem cells(SCCshave provided very important tool to improve herd geneticand different field. 0.2 to 0.3 percent of total cells of seminiferous tubules is consist ofspermatogonial stem cells. To investigate and biomanipulation of these cells, proliferationand viability rate of cells should be increasedin vitro, at first. Follicle stimulating hormone(FSH has been suggested to play a determinant role in the survival of germ cells in additionto increasing spermatogonial proliferation. In this study, thein vitroeffects ofFSHonspermatogonial cell colony formation were investigated. Sertoli and spermatogonial cellswere isolated from 3-5 months old calves. The identity of theSertoli cells and spermatogonialstem cells were confirmed through immunocytochemistry and colony morphology,respectively. Co-cultured Sertoli and spermatogonial cells were treatedwithFSHin differentdose of10, 20 and 40 IU mL-1FSH, before colony assay.Results indicated that,FSHincreasedin vitrocolonization of spermatogonial cells in comparison with control group. In conclusion,usingFSHprovided proper bovine spermatogonial stem cell culture medium forin vitrostudy of these cells.

  13. Retinoic acid-treated pluripotent stem cells undergoing neurogenesis present increased aneuploidy and micronuclei formation.

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    Rafaela C Sartore

    Full Text Available The existence of loss and gain of chromosomes, known as aneuploidy, has been previously described within the central nervous system. During development, at least one-third of neural progenitor cells (NPCs are aneuploid. Notably, aneuploid NPCs may survive and functionally integrate into the mature neural circuitry. Given the unanswered significance of this phenomenon, we tested the hypothesis that neural differentiation induced by all-trans retinoic acid (RA in pluripotent stem cells is accompanied by increased levels of aneuploidy, as previously described for cortical NPCs in vivo. In this work we used embryonal carcinoma (EC cells, embryonic stem (ES cells and induced pluripotent stem (iPS cells undergoing differentiation into NPCs. Ploidy analysis revealed a 2-fold increase in the rate of aneuploidy, with the prevalence of chromosome loss in RA primed stem cells when compared to naïve cells. In an attempt to understand the basis of neurogenic aneuploidy, micronuclei formation and survivin expression was assessed in pluripotent stem cells exposed to RA. RA increased micronuclei occurrence by almost 2-fold while decreased survivin expression by 50%, indicating possible mechanisms by which stem cells lose their chromosomes during neural differentiation. DNA fragmentation analysis demonstrated no increase in apoptosis on embryoid bodies treated with RA, indicating that cell death is not the mandatory fate of aneuploid NPCs derived from pluripotent cells. In order to exclude that the increase in aneuploidy was a spurious consequence of RA treatment, not related to neurogenesis, mouse embryonic fibroblasts were treated with RA under the same conditions and no alterations in chromosome gain or loss were observed. These findings indicate a correlation amongst neural differentiation, aneuploidy, micronuclei formation and survivin downregulation in pluripotent stem cells exposed to RA, providing evidence that somatically generated chromosomal

  14. Inhibition of thromboxane synthase induces lung cancer cell death via increasing the nuclear p27

    International Nuclear Information System (INIS)

    Leung, Kin Chung; Hsin, Michael K.Y.; Chan, Joey S.Y.; Yip, Johnson H.Y.; Li, Mingyue; Leung, Billy C.S.; Mok, Tony S.K.; Warner, Timothy D.; Underwood, Malcolm J.; Chen, George G.

    2009-01-01

    The role of thromboxane in lung carcinogenesis is not clearly known, though thromboxane B2 (TXB 2 ) level is increased and antagonists of thromboxane receptors or TXA2 can induce apoptosis of lung cancer cells. p27, an atypical tumor suppressor, is normally sequestered in the nucleus. The increased nuclear p27 may result in apoptosis of tumor cells. We hypothesize that the inhibition of thromboxane synthase (TXS) induces the death of lung cancer cells and that such inhibition is associated with the nuclear p27 level. Our experiment showed that the inhibition of TXS significantly induced the death or apoptosis in lung cancer cells. The activity of TXS was increased in lung cancer. The nuclear p27 was remarkably reduced in lung cancer tissues. The inhibition of TXS caused the cell death and apoptosis of lung cancer cells, likely via the elevation of the nuclear p27 since the TXS inhibition promoted the nuclear p27 level and the inhibition of p27 by its siRNA recovered the cell death induced by TXS inhibition. Collectively, lung cancer cells produce high levels of TXB 2 but their nuclear p27 is markedly reduced. The inhibition of TXS results in the p27-related induction of cell death in lung cancer cells.

  15. Inhibition of thromboxane synthase induces lung cancer cell death via increasing the nuclear p27

    Energy Technology Data Exchange (ETDEWEB)

    Leung, Kin Chung; Hsin, Michael K.Y.; Chan, Joey S.Y.; Yip, Johnson H.Y.; Li, Mingyue; Leung, Billy C.S. [Department of Surgery, The Chinese University of Hong Kong, Shatin, New Territories (Hong Kong); Mok, Tony S.K. [Department of Clinical Oncology, The Chinese University of Hong Kong, Shatin, New Territories (Hong Kong); Warner, Timothy D. [The William Harvey Research Institute, Queen Mary University of London, London (United Kingdom); Underwood, Malcolm J. [Department of Surgery, The Chinese University of Hong Kong, Shatin, New Territories (Hong Kong); Chen, George G., E-mail: gchen@cuhk.edu.hk [Department of Surgery, The Chinese University of Hong Kong, Shatin, New Territories (Hong Kong)

    2009-10-15

    The role of thromboxane in lung carcinogenesis is not clearly known, though thromboxane B2 (TXB{sub 2}) level is increased and antagonists of thromboxane receptors or TXA2 can induce apoptosis of lung cancer cells. p27, an atypical tumor suppressor, is normally sequestered in the nucleus. The increased nuclear p27 may result in apoptosis of tumor cells. We hypothesize that the inhibition of thromboxane synthase (TXS) induces the death of lung cancer cells and that such inhibition is associated with the nuclear p27 level. Our experiment showed that the inhibition of TXS significantly induced the death or apoptosis in lung cancer cells. The activity of TXS was increased in lung cancer. The nuclear p27 was remarkably reduced in lung cancer tissues. The inhibition of TXS caused the cell death and apoptosis of lung cancer cells, likely via the elevation of the nuclear p27 since the TXS inhibition promoted the nuclear p27 level and the inhibition of p27 by its siRNA recovered the cell death induced by TXS inhibition. Collectively, lung cancer cells produce high levels of TXB{sub 2} but their nuclear p27 is markedly reduced. The inhibition of TXS results in the p27-related induction of cell death in lung cancer cells.

  16. Increased Toxicity of Chemotherapeutic Drugs by All-Trans Retinoic Acid in Cd44 Cells

    Directory of Open Access Journals (Sweden)

    A Abbasi

    2016-03-01

    Full Text Available BACKGROUND AND OBJECTIVE: In recent studies, undifferentiated CD44 cells have been introduced as the major cause of chemotherapeutic drug resistance in esophageal cancer. In this study, we aimed to evaluate the effects of all-trans retinoic acid on reducing chemotherapeutic drug resistance and improving the associated toxic effects. METHODS: In this clinical study, CD44+ and CD44- cells were separated from KYSE-30 cell line, using magnetic-activated cell sorting (MACS method. The cytotoxic effects of retinoic acid treatment, combined with cisplatin and 5-fluorouracil, were separately evaluated in two cell groups, i.e., CD44+ and CD44-. Cytotoxicity was determined by identifying cellular metabolic activity, acridine orange/ethidium bromide staining, and flow cytometry. FINDINGS: In this study, CD44 marker was expressed in 6.25% of the cell population in KYSE-30 cell line. The results of flow cytometry revealed that treatment with a combination of retinoic acid and chemotherapeutic drugs could improve cell cycle arrest in CD44+ cells (p<0.05, unlike CD44- cells. Determination of cellular metabolic activity, increased cell apoptosis along with decreased half maximal inhibitory concentration (IC50, and acridine orange/ethidium bromide staining were indicative of the increased percentage of primary and secondary apoptotic CD44+ cells. However, in CD44- cells, these effects were only observed by using a combination of retinoic acid and cisplatin (p<0.05. CONCLUSION: The present results showed that all-trans retinoic acid could increase the toxicity of cisplatin and 5-fluorouracil in CD44+ cells.

  17. Self-reactive CD4+ T cells and B cells in the blood in health and autoimmune disease: increased frequency of thyroglobulin-reactive cells in Graves' disease

    DEFF Research Database (Denmark)

    Nielsen, Claus H; Moeller, Ane Christine; Hegedüs, Laszlo

    2006-01-01

    The mechanisms underlying activation of potentially self-reactive circulating B cells and T cells remain unclear. We measured the uptake of a self-antigen, thyroglobulin, by antigen presenting cells, and the subsequent proliferation of CD4(+) T cells and B cells from healthy controls and patients...... with autoimmune thyroiditis. In Hashimoto's thyroiditis, B cells bound increased amounts of thyroglobulin in a complement- and autoantibody-dependent manner, and the thyroglobulin-elicited proliferation of CD4(+) T cells and B cells was complement dependent. Increased proportions of Tg-responsive CD4(+) T cells...... and B cells were found in patients with Graves' disease. Notably, both patient groups and healthy controls exhibited higher proliferative responses to thyroglobulin than to a foreign recall antigen, tetanus toxoid. Our results suggest that self-tolerance can be broken by exposure of circulating...

  18. Lamotrigine increases the number of BrdU-labeled cells in the rat hippocampus

    DEFF Research Database (Denmark)

    Kondziella, Daniel; Strandberg, Joakim; Lindquist, Catarina

    2011-01-01

    Antidepressant medication and electroconvulsive therapy stabilize mood symptoms and increase hippocampal neurogenesis. We examined whether lamotrigine, suggested to give rise to mood-stabilizing and antidepressant effects in addition to its antiepileptic properties, also increases the number of n...... in the granule cell layer of the dentate gyrus showed an increased number of newborn cells in rats receiving lamotrigine (42.6 ± 3.5 cells/slice) compared with valproate (31.6 ± 2.8) and controls (32.2 ± 3.1; P...

  19. Enzalutamide inhibits proliferation of gemcitabine-resistant bladder cancer cells with increased androgen receptor expression.

    Science.gov (United States)

    Kameyama, Koji; Horie, Kengo; Mizutani, Kosuke; Kato, Taku; Fujita, Yasunori; Kawakami, Kyojiro; Kojima, Toshio; Miyazaki, Tatsuhiko; Deguchi, Takashi; Ito, Masafumi

    2017-01-01

    Advanced bladder cancer is treated mainly with gemcitabine and cisplatin, but most patients eventually become resistance. Androgen receptor (AR) signaling has been implicated in bladder cancer as well as other types of cancer including prostate cancer. In this study, we investigated the expression and role of AR in gemcitabine-resistant bladder cancer cells and also the potential of enzalutamide, an AR inhibitor, as a therapeutic for the chemoresistance. First of all, we established gemcitabine-resistant T24 cells (T24GR) from T24 bladder cancer cells and performed gene expression profiling. Microarray analysis revealed upregulation of AR expression in T24GR cells compared with T24 cells. AR mRNA and protein expression was confirmed to be increased in T24GR cells, respectively, by quantitative RT-PCR and western blot analysis, which was associated with more potent AR transcriptional activity as measured by luciferase reporter assay. The copy number of AR gene in T24GR cells determined by PCR was twice as many as that of T24 cells. AR silencing by siRNA transfection resulted in inhibition of proliferation of T24GR cells. Cell culture in charcoal-stripped serum and treatment with enzalutamide inhibited growth of T24GR cells, which was accompanied by cell cycle arrest. AR transcriptional activity was found to be reduced in T24GR cells by enzalutamide treatment. Lastly, enzalutamide also inhibited cell proliferation of HTB5 bladder cancer cells that express AR and possess intrinsic resistance to gemcitabine. Our results suggest that enzalutamide may have the potential to treat patients with advanced gemcitabine-resistant bladder cancer with increased AR expression.

  20. Epidermal growth factor increases LRF/Pokemon expression in human prostate cancer cells.

    Science.gov (United States)

    Aggarwal, Himanshu; Aggarwal, Anshu; Agrawal, Devendra K

    2011-10-01

    Leukemia/lymphoma related factor/POK erythroid myeloid ontogenic factor (LRF/Pokemon) is a member of the POK family of proteins that promotes oncogenesis in several forms of cancer. Recently, we found higher LRF expression in human breast and prostate carcinomas compared to the corresponding normal tissues. The aim of this study was to examine the regulation of LRF expression in human prostate cells. Epidermal growth factor (EGF) and its receptors mediate several tumorigenic cascades that regulate cell differentiation, proliferation, migration and survival of prostate cancer cells. There was significantly higher level of LRF expression in the nucleus of LNCaP and PC-3 cells than RWPE-1 cells. A significant increase in LRF expression was observed with increasing doses of EGF in more aggressive and androgen-sensitive prostate cancer cells suggesting that EGF signaling pathway is critical in upregulating the expression of LRF/Pokemon to promote oncogenesis. Copyright © 2011 Elsevier Inc. All rights reserved.

  1. Increased T-regulatory cells within lymphocyte follicles in moderate COPD

    DEFF Research Database (Denmark)

    Plumb, J; Smyth, L J C; Adams, H R

    2009-01-01

    Lymphoid follicles in the lung parenchyma are a characteristic feature of chronic obstructive pulmonary disease (COPD). There are reports of altered CD4 T-regulatory cell numbers in COPD lungs, but the location of these cells within COPD lung tissue specific follicles has not been investigated......, as well as lymphoid clusters lacking organisation. The percentage of CD4 cells that were T-regulatory cells were significantly increased (p = 0.02) within COPD (16%) follicles compared with smokers (10%) and nonsmokers (8%). In contrast, there was no change (p>0.05) in the percentage of T-regulatory cells...... in clusters or the subepithelium between groups. Lymphoid follicles in COPD patients have increased T-regulatory cells. Therefore, T-regulatory activity may be altered within COPD lymphoid follicles....

  2. Increasing the Energy Efficiency of Aluminum-Reduction Cells Using Modified Cathodes

    Science.gov (United States)

    Jianping, Peng; Yang, Song; Yuezhong, Di; Yaowu, Wang; Naixiang, Feng

    2017-10-01

    A cathode with an inclined surface (5°) and increased bar collector height (230 mm high) was incorporated into two 300-kA industrial aluminum-reduction cells. The voltage of the cells with the modified cathode was reduced by approximately 200 mV when compared with that of a conventional cell with a flat cathode. Through the use of simulations, the reduction in the cell voltage was attributed to the cathode modification (40 mV) and a reduced electrolyte level of 0.5 cm (160 mV). As a result of reduced anode cathode distance (ACD), the ledge toe was extended to the anode shadow by 12 cm. This caused a large inverted horizontal current and a velocity increase. The ledge profile returned to the desired position when the cells were insulated more effectively, and the metal velocity and metal crest in the modified cells were reduced accordingly.

  3. Increased Incidence of T-Cell Malignancies in Patients with Chronic Lymphocytic Leukemia

    OpenAIRE

    Choi, Goda; van den Broek, Esther C; Stam, Olga CG; van Noesel, C.J.M.; Tonino, Sanne H.; Kater, Armon P.

    2015-01-01

    We present a patient with chemotherapy-refractory Chronic Lymphocytic Leukemia (CLL) in whom postmortem examination showed hepatosplenomegaly, with both multiple small-cellular CLL lesions and large-cellular, monoclonal T-cell infiltrates. Following this case, the co-incidence of T-cell malignancies and CLL was studied using Dutch and American cancer registry databases. Analysis showed an excess risk for T-cell malignancies in CLL patients, with increased standardized incidence ratios compare...

  4. The increased susceptibility of hematopoietic stem cells to Friend leukemia virus in the repopulating period

    International Nuclear Information System (INIS)

    Hirashima, Kunitake; Kumatori, Toshiyuki

    1977-01-01

    The present study considers two fundamental problems of leukemia and stem cells: (1) whether the target cells for malignant transformation of Friend leukemia virus (FV) are the pluripotential stem cells; (2) whether the susceptibility of the target cell is changeable depending on the proliferating state of the cell. In the experiments, inbred 8- to 12-week-old C3H/He and BC3Fl hybrid mice were used. As target cells for FV, two candidate cell populations should be considered. These are the pluripotential stem cells expressed as CFUs and the committed stem cells expressed as erythropoietin-responsive cells (ERC). According to our preliminary experiments on the recovery patterns of CFUs or ERC, after a single 150 rads X-irradiation (X) or administration of 215 mg/kg of cyclophosphamide (CY), it was clear that the repopulation of CFUs was quite different from that of ERC. The target cells for FV-induced transformation are the stem cells expressed as CFUs and the susceptibility of these increases under conditions of active repopulation after X-irradiation and cyclophosphamide administration. After the irradiation especially, this effect is dose-dependent for doses over 25 rads at 2 weeks after irradiation. (Auth.)

  5. TNFRSF14 aberrations in follicular lymphoma increase clinically significant allogeneic T-cell responses.

    Science.gov (United States)

    Kotsiou, Eleni; Okosun, Jessica; Besley, Caroline; Iqbal, Sameena; Matthews, Janet; Fitzgibbon, Jude; Gribben, John G; Davies, Jeffrey K

    2016-07-07

    Donor T-cell immune responses can eradicate lymphomas after allogeneic hematopoietic stem cell transplantation (AHSCT), but can also damage healthy tissues resulting in harmful graft-versus-host disease (GVHD). Next-generation sequencing has recently identified many new genetic lesions in follicular lymphoma (FL). One such gene, tumor necrosis factor receptor superfamily 14 (TNFRSF14), abnormal in 40% of FL patients, encodes the herpes virus entry mediator (HVEM) which limits T-cell activation via ligation of the B- and T-lymphocyte attenuator. As lymphoma B cells can act as antigen-presenting cells, we hypothesized that TNFRSF14 aberrations that reduce HVEM expression could alter the capacity of FL B cells to stimulate allogeneic T-cell responses and impact the outcome of AHSCT. In an in vitro model of alloreactivity, human lymphoma B cells with TNFRSF14 aberrations had reduced HVEM expression and greater alloantigen-presenting capacity than wild-type lymphoma B cells. The increased immune-stimulatory capacity of lymphoma B cells with TNFRSF14 aberrations had clinical relevance, associating with higher incidence of acute GVHD in patients undergoing AHSCT. FL patients with TNFRSF14 aberrations may benefit from more aggressive immunosuppression to reduce harmful GVHD after transplantation. Importantly, this study is the first to demonstrate the impact of an acquired genetic lesion on the capacity of tumor cells to stimulate allogeneic T-cell immune responses which may have wider consequences for adoptive immunotherapy strategies. © 2016 by The American Society of Hematology.

  6. Sexual activity increases the number of newborn cells in the accessory olfactory bulb of male rats.

    Directory of Open Access Journals (Sweden)

    Wendy ePortillo

    2012-07-01

    Full Text Available In rodents, sexual behavior depends on the adequate detection of sexually relevant stimuli. The olfactory bulb (OB is a region of the adult mammalian brain undergoing constant cell renewal by continuous integration of new granular and periglomerular neurons in the accessory (AOB and main (MOB olfactory bulbs. The proliferation, migration, survival, maturation, and integration of these new cells to the OB depend on the stimulus that the subjects received. We have previously shown that 15 days after females control (paced the sexual interaction an increase in the number of cells is observed in the AOB. No changes are observed in the number of cells when females are not allowed to control the sexual interaction. In the present study we investigated if in male rats sexual behavior increases the number of new cells in the OB. Male rats were divided in five groups: 1 males that did not receive any sexual stimulation, 2 males that were exposed to female odors, 3 males that mated for 1 h and could not pace their sexual interaction, 4 males that paced their sexual interaction and ejaculated 1 time and 5 males that paced their sexual interaction and ejaculated 3 times. All males received three injections of the DNA synthesis marker bromodeoxyuridine at 1h intervals, starting 1h before the beginning of the behavioral test. Fifteen days later, males were sacrificed and the brains were processed to identify new cells and to evaluate if they differentiated into neurons. The number of newborn cells increased in the granular cell layer (also known as the internal cell layer of the AOB in males that ejaculated one or three times controlling (paced the rate of the sexual interaction. Some of these new cells were identified as neurons. In contrast, no significant differences were found in the mitral cell layer (also known as the external cell layer and glomerular cell layer of the AOB. In addition, no significant differences were found between groups in the MOB in

  7. Increasing the safety and efficacy of chimeric antigen receptor T cell therapy

    Directory of Open Access Journals (Sweden)

    Hua Li

    2017-04-01

    Full Text Available Abstract Chimeric antigen receptor (CAR T cell therapy is a promising cancer treatment that has recently been undergoing rapid development. However, there are still some major challenges, including precise tumor targeting to avoid off-target or “on-target/off-tumor” toxicity, adequate T cell infiltration and migration to solid tumors and T cell proliferation and persistence across the physical and biochemical barriers of solid tumors. In this review, we focus on the primary challenges and strategies to design safe and effective CAR T cells, including using novel cutting-edge technologies for CAR and vector designs to increase both the safety and efficacy, further T cell modification to overcome the tumor-associated immune suppression, and using gene editing technologies to generate universal CAR T cells. All these efforts promote the development and evolution of CAR T cell therapy and move toward our ultimate goal—curing cancer with high safety, high efficacy, and low cost.

  8. CNPY2 promoted the proliferation of renal cell carcinoma cells and increased the expression of TP53

    International Nuclear Information System (INIS)

    Taniguchi, Hidefumi; Ito, Saya; Ueda, Takashi; Morioka, Yukako; Kayukawa, Naruhiro; Ueno, Akihisa; Nakagawa, Hideo; Fujihara, Atsuko; Ushijima, So; Kanazawa, Motohiro; Hongo, Fumiya; Ukimura, Osamu

    2017-01-01

    Renal cell carcinoma (RCC) is the most common type of kidney cancer. However, the mechanisms underlying the progression of the disease are not well understood. The data in this report suggest that canopy FGF signaling regulator 2 (CNPY2) is a promoter of RCC progression. We found that CNPY2 significantly promoted growth of RCC cells and upregulated TP53 gene expression. Although TP53 is widely known as a tumor suppressor, in RCC TP53 promoted tumor cell growth. A typical p53 target gene, CDKN1A, was upregulated by both p53 and CNPY2 in RCC cells, suggesting that CNPY2 increased the expression level of TP53. Consistent with these results, CNPY2 and TP53 expression levels were positively correlated in RCC patients. These findings suggested that CNPY2 promoted cancer cell growth in RCC through regulating TP53 gene expression. - Highlights: • CNPY2 promoted growth of renal cell carcinoma (RCC) cells. • TP53 expression levels were increased by CNPY2 in RCC cells. • Growth of RCC cells was promoted by TP53. • CNPY2 expression positively correlated with TP53 expression in RCC patients.

  9. Ibrutinib Therapy Increases T Cell Repertoire Diversity in Patients with Chronic Lymphocytic Leukemia.

    Science.gov (United States)

    Yin, Qingsong; Sivina, Mariela; Robins, Harlan; Yusko, Erik; Vignali, Marissa; O'Brien, Susan; Keating, Michael J; Ferrajoli, Alessandra; Estrov, Zeev; Jain, Nitin; Wierda, William G; Burger, Jan A

    2017-02-15

    The Bruton's tyrosine kinase inhibitor ibrutinib is a highly effective, new targeted therapy for chronic lymphocytic leukemia (CLL) that thwarts leukemia cell survival, growth, and tissue homing. The effects of ibrutinib treatment on the T cell compartment, which is clonally expanded and thought to support the growth of malignant B cells in CLL, are not fully characterized. Using next-generation sequencing technology, we characterized the diversity of TCRβ-chains in peripheral blood T cells from 15 CLL patients before and after 1 y of ibrutinib therapy. We noted elevated CD4 + and CD8 + T cell numbers and a restricted TCRβ repertoire in all pretreatment samples. After 1 y of ibrutinib therapy, elevated peripheral blood T cell numbers and T cell-related cytokine levels had normalized, and T cell repertoire diversity increased significantly. Dominant TCRβ clones in pretreatment samples declined or became undetectable, and the number of productive unique clones increased significantly during ibrutinib therapy, with the emergence of large numbers of low-frequency TCRβ clones. Importantly, broader TCR repertoire diversity was associated with clinical efficacy and lower rates of infections during ibrutinib therapy. These data demonstrate that ibrutinib therapy increases diversification of the T cell compartment in CLL patients, which contributes to cellular immune reconstitution. Copyright © 2017 by The American Association of Immunologists, Inc.

  10. Increased frequencies of IL-31-producing T cells are found in chronic atopic dermatitis skin

    NARCIS (Netherlands)

    Szegedi, Krisztina; Kremer, Andreas E.; Kezic, Sanja; Teunissen, Marcel B. M.; Bos, Jan D.; Luiten, Rosalie M.; Res, Pieter C.; Middelkamp-Hup, Maritza A.

    2012-01-01

    Interleukin (IL)-31 has been associated with pruritus, a characteristic feature of atopic dermatitis (AD). Local T cell responses may be responsible for the increased level of IL-31 mRNA observed in AD. We investigated the frequency of IL-31-producing T cells in AD lesions, as well as their cytokine

  11. 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…

  12. Staurosporine Increases Lentiviral Vector Transduction Efficiency of Human Hematopoietic Stem and Progenitor Cells

    Directory of Open Access Journals (Sweden)

    Gretchen Lewis

    2018-06-01

    Full Text Available Lentiviral vector (LVV-mediated transduction of human CD34+ hematopoietic stem and progenitor cells (HSPCs holds tremendous promise for the treatment of monogenic hematological diseases. This approach requires the generation of a sufficient proportion of gene-modified cells. We identified staurosporine, a serine/threonine kinase inhibitor, as a small molecule that could be added to the transduction process to increase the proportion of genetically modified HSPCs by overcoming a LVV entry barrier. Staurosporine increased vector copy number (VCN approximately 2-fold when added to mobilized peripheral blood (mPB CD34+ cells prior to transduction. Limited staurosporine treatment did not affect viability of cells post-transduction, and there was no difference in in vitro colony formation compared to vehicle-treated cells. Xenotransplantation studies identified a statistically significant increase in VCN in engrafted human cells in mouse bone marrow at 4 months post-transplantation compared to vehicle-treated cells. Prostaglandin E2 (PGE2 is known to increase transduction efficiency of HSPCs through a different mechanism. Combining staurosporine and PGE2 resulted in further enhancement of transduction efficiency, particularly in short-term HSPCs. The combinatorial use of small molecules, such as staurosporine and PGE2, to enhance LVV transduction of human CD34+ cells is a promising method to improve transduction efficiency and subsequent potential therapeutic benefit of gene therapy drug products. Keywords: lentiviral, HSPC, transduction

  13. Elevated levels of homocysteine increase IL-6 production in monocytic Mono Mac 6 cells

    NARCIS (Netherlands)

    van Aken, B. E.; Jansen, J.; van Deventer, S. J.; Reitsma, P. H.

    2000-01-01

    Hyperhomocysteinemia is a risk factor for atherosclerosis and thrombosis. The aim of this study was to analyze if exposure of monocytic cells to increased levels of homocysteine (HCY) induces the accumulation of inflammatory mediators. Interleukin (IL)-6 production by monocytic cell line Mono Mac 6

  14. Mammary stem cells: Novel markers and novel approaches to increase lactation efficiency

    Science.gov (United States)

    Mammary stem cells (MaSC) provide for net growth, renewal and turnover of mammary epithelial cells, and are therefore potential targets for strategies to increase production efficiency. Appropriate regulation of MaSC can potentially benefit milk yield, persistency, dry period management and tissue r...

  15. EGFR inhibitor C225 increases the radiosensitivity of human lung squamous cancer cells

    Directory of Open Access Journals (Sweden)

    Yang Ruijie

    2010-10-01

    Full Text Available Abstract Background The purpose of the present study is to investigate the direct biological effects of the epidermal growth factor receptor (EGFR inhibitor C225 on the radiosensitivity of human lung squamous cancer cell-H520. H520 cells were treated with different dosage of 60Co γ ray irradiation (1.953 Gy/min in the presence or absence of C225. The cellular proliferation, colony forming capacity, apoptosis, the cell cycle distribution as well as caspase-3 were analyzed in vitro. Results We found that C225 treatment significantly increased radiosensitivity of H-520 cells to irradiation, and led to cell cycle arrest in G1 phase, whereas 60Co γ ray irradiation mainly caused G2 phase arrest. H-520 cells thus displayed both the G1 and G2 phase arrest upon treatment with C225 in combination with 60Co γ ray irradiation. Moreover, C225 treatment significantly increased the apoptosis percentage of H-520 cells (13.91% ± 1.88% compared with the control group (5.75% ± 0.64%, P Conclusion In this regard, C225 treatment may make H-520 cells more sensitive to irradiation through the enhancement of caspase-3 mediated tumor cell apoptosis and cell cycle arrest.

  16. Diabetes increases susceptibility of primary cultures of rat proximal tubular cells to chemically induced injury

    International Nuclear Information System (INIS)

    Zhong Qing; Terlecky, Stanley R.; Lash, Lawrence H.

    2009-01-01

    Diabetic nephropathy is characterized by increased oxidative stress and mitochondrial dysfunction. In the present study, we prepared primary cultures of proximal tubular (PT) cells from diabetic rats 30 days after an ip injection of streptozotocin and compared their susceptibility to oxidants (tert-butyl hydroperoxide, methyl vinyl ketone) and a mitochondrial toxicant (antimycin A) with that of PT cells isolated from age-matched control rats, to test the hypothesis that PT cells from diabetic rats exhibit more cellular and mitochondrial injury than those from control rats when exposed to these toxicants. PT cells from diabetic rats exhibited higher basal levels of reactive oxygen species (ROS) and higher mitochondrial membrane potential, demonstrating that the PT cells maintain the diabetic phenotype in primary culture. Incubation with either the oxidants or mitochondrial toxicant resulted in greater necrotic and apoptotic cell death, greater evidence of morphological damage, greater increases in ROS, and greater decreases in mitochondrial membrane potential in PT cells from diabetic rats than in those from control rats. Pretreatment with either the antioxidant N-acetyl-L-cysteine or a catalase mimetic provided equivalent protection of PT cells from both diabetic and control rats. Despite the greater susceptibility to oxidative and mitochondrial injury, both cytoplasmic and mitochondrial glutathione concentrations were markedly higher in PT cells from diabetic rats, suggesting an upregulation of antioxidant processes in diabetic kidney. These results support the hypothesis that primary cultures of PT cells from diabetic rats are a valid model in which to study renal cellular function in the diabetic state.

  17. Increased endothelial cell-leukocyte interaction in murine schistosomiasis: possible priming of endothelial cells by the disease.

    Directory of Open Access Journals (Sweden)

    Suellen D S Oliveira

    Full Text Available BACKGROUND AND AIMS: Schistosomiasis is an intravascular parasitic disease associated with inflammation. Endothelial cells control leukocyte transmigration and vascular permeability being modulated by pro-inflammatory mediators. Recent data have shown that endothelial cells primed in vivo in the course of a disease keep the information in culture. Herein, we evaluated the impact of schistosomiasis on endothelial cell-regulated events in vivo and in vitro. METHODOLOGY AND PRINCIPAL FINDINGS: The experimental groups consisted of Schistosoma mansoni-infected and age-matched control mice. In vivo infection caused a marked influx of leukocytes and an increased protein leakage in the peritoneal cavity, characterizing an inflamed vascular and cellular profile. In vitro leukocyte-mesenteric endothelial cell adhesion was higher in cultured cells from infected mice as compared to controls, either in the basal condition or after treatment with the pro-inflammatory cytokine tumor necrosis factor (TNF. Nitric oxide (NO donation reduced leukocyte adhesion to endothelial cells from control and infected groups; however, in the later group the effect was more pronounced, probably due to a reduced NO production. Inhibition of control endothelial NO synthase (eNOS increased leukocyte adhesion to a level similar to the one observed in the infected group. Besides, the adhesion of control leukocytes to endothelial cells from infected animals is similar to the result of infected animals, confirming that schistosomiasis alters endothelial cells function. Furthermore, NO production as well as the expression of eNOS were reduced in cultured endothelial cells from infected animals. On the other hand, the expression of its repressor protein, namely caveolin-1, was similar in both control and infected groups. CONCLUSION/SIGNIFICANCE: Schistosomiasis increases vascular permeability and endothelial cell-leukocyte interaction in vivo and in vitro. These effects are partially

  18. Increased p21ras activity in human fibroblasts transduced with survivin enhances cell proliferation

    International Nuclear Information System (INIS)

    Temme, Achim; Diestelkoetter-Bachert, Petra; Schmitz, Marc; Morgenroth, Agnieszka; Weigle, Bernd; Rieger, Michael A.; Kiessling, Andrea; Rieber, E. Peter

    2005-01-01

    Survivin is critically involved in mitosis and when overexpressed enhances the activity of the Aurora B kinase, a serine-threonine kinase belonging to the family of oncogenic Aurora/IpI1p-related kinases. Both proteins interact with Ras GTPase-activating protein suggesting an impact on the Ras pathway. This study aimed at defining the role of survivin in proliferation and potential transformation of cells. When survivin was overexpressed in normal human lung fibroblasts, the characteristic track lanes of fibroblasts were disturbed and the rate of cell proliferation was increased. An enhanced level of p21 ras mRNA and protein expression and concomitant rise in levels of activated p21 ras were observed. Despite increased proliferation cell survival remained dependent on serum and cells were not able to form colonies in soft agar assays. These data suggest that overexpression of survivin increases cell growth but, despite the increase in active p21 ras , is not sufficient to transform primary cells. Yet, in addition to its anti-apoptotic function it might contribute to the accelerated growth of tumour cells by increasing p21 ras activity

  19. Thiamine and benfotiamine prevent increased apoptosis in endothelial cells and pericytes cultured in high glucose.

    Science.gov (United States)

    Beltramo, E; Berrone, E; Buttiglieri, S; Porta, M

    2004-01-01

    High glucose induces pathological alterations in small and large vessels, possibly through increased formation of AGE, activation of aldose reductase and protein kinase C, and increased flux through the hexosamine pathway. We showed previously that thiamine and benfotiamine correct delayed replication and increase lactate production in endothelial cells subjected to high glucose. We now aim at verifying the effects of thiamine and benfotiamine on cell cycle, apoptosis, and expression of adhesion molecules in endothelial cells and pericytes, under high ambient glucose. Human umbilical vein endothelial cells and bovine retinal pericytes were cultured in normal (5.6 mmol/L) or high (28 mmol/L) glucose, with or without thiamine or benfotiamine, 50 or 100 micro mol/L. Apoptosis was determined by two separate ELISA methods, measuring DNA fragmentation and caspase-3 activity, respectively. Cell cycle and integrin subunits alpha3, alpha5, and beta1 concentration were measured by flow cytometry. Apoptosis was increased in high glucose after 3 days of culture, both in endothelium and pericytes. Thiamine and benfotiamine reversed such effects. Neither cell cycle traversal nor integrin concentrations were modified in these experimental conditions. Thiamine and benfotiamine correct increased apoptosis due to high glucose in cultured vascular cells. Further elucidations of the mechanisms through which they work could help set the basis for clinical use of this vitamin in the prevention and/or treatment of diabetic microangiopathy. Copyright 2004 John Wiley & Sons, Ltd.

  20. 2-methoxyestradiol-mediated anti-tumor effect increases osteoprotegerin expression in osteosarcoma cells.

    Science.gov (United States)

    Benedikt, Michaela B; Mahlum, Eric W; Shogren, Kristen L; Subramaniam, Malayannan; Spelsberg, Thomas C; Yaszemski, Michael J; Maran, Avudaiappan

    2010-04-01

    Osteosarcoma is a bone tumor that frequently develops during adolescence. 2-Methoxyestradiol (2-ME), a naturally occurring metabolite of 17beta-estradiol, induces cell cycle arrest and cell death in human osteosarcoma cells. To investigate whether the osteoprotegrin (OPG) protein plays a role in 2-ME actions, we studied the effect of 2-ME treatment on OPG gene expression in human osteosarcoma cells. 2-ME treatment induced OPG gene promoter activity and mRNA levels. Also, Western blot analysis showed that 2-ME treatment increased OPG protein levels in MG63, KHOS, 143B and LM7 osteosarcoma cells by 3-, 1.9-, 2.8-, and 2.5-fold, respectively, but did not affect OPG expression in normal bone cells. In addition, increases in OPG protein levels were observed in osteosarcoma cell culture media after 3 days of 2-ME treatment. The effect of 2-ME on osteosarcoma cells was ligand-specific as parent estrogen, 17beta-estradiol and a tumorigenic estrogen metabolite, 16alpha-hydroxyestradiol, which do not affect osteosarcoma cell cycle and cell death, had no effect on OPG protein expression. Furthermore, co-treating osteosarcoma cells with OPG protein did not further enhance 2-ME-mediated anti-tumor effects. OPG-released in 2-ME-treated cultures led to an increase in osteoblastic activity and a decrease in osteoclast number, respectively. These findings suggest that OPG is not directly involved in 2-ME-mediated anti-proliferative effects in osteosarcoma cells, but rather participates in anti-resorptive functions of 2-ME in bone tumor environment. Copyright 2010 Wiley-Liss, Inc.

  1. Natural killer cells in obesity: impaired function and increased susceptibility to the effects of cigarette smoke.

    LENUS (Irish Health Repository)

    O'Shea, Donal

    2012-02-01

    BACKGROUND: Obese individuals who smoke have a 14 year reduction in life expectancy. Both obesity and smoking are independently associated with increased risk of malignancy. Natural killer cells (NK) are critical mediators of anti-tumour immunity and are compromised in obese patients and smokers. We examined whether NK cell function was differentially affected by cigarette smoke in obese and lean subjects. METHODOLOGY AND PRINCIPAL FINDINGS: Clinical data and blood were collected from 40 severely obese subjects (BMI>40 kg\\/m(2)) and 20 lean healthy subjects. NK cell levels and function were assessed using flow cytometry and cytotoxicity assays. The effect of cigarette smoke on NK cell ability to kill K562 tumour cells was assessed in the presence or absence of the adipokines leptin and adiponectin. NK cell levels were significantly decreased in obese subjects compared to lean controls (7.6 vs 16.6%, p = 0.0008). NK function was also significantly compromised in obese patients (30% +\\/- 13% vs 42% +\\/-12%, p = 0.04). Cigarette smoke inhibited NK cell ability to kill tumour cell lines (p<0.0001). NK cells from obese subjects were even more susceptible to the inhibitory effects of smoke compared to lean subjects (33% vs 28%, p = 0.01). Cigarette smoke prevented NK cell activation, as well as perforin and interferon-gamma secretion upon tumour challenge. Adiponectin but not leptin partially reversed the effects of smoke on NK cell function in both obese (p = 0.002) and lean controls (p = 0.01). CONCLUSIONS\\/SIGNIFICANCE: Obese subjects have impaired NK cell activity that is more susceptible to the detrimental effects of cigarette smoke compared to lean subjects. This may play a role in the increase of cancer and infection seen in this population. Adiponectin is capable of restoring NK cell activity and may have therapeutic potential for immunity in obese subjects and smokers.

  2. Natural killer cells in obesity: impaired function and increased susceptibility to the effects of cigarette smoke.

    LENUS (Irish Health Repository)

    O'Shea, Donal

    2010-01-01

    BACKGROUND: Obese individuals who smoke have a 14 year reduction in life expectancy. Both obesity and smoking are independently associated with increased risk of malignancy. Natural killer cells (NK) are critical mediators of anti-tumour immunity and are compromised in obese patients and smokers. We examined whether NK cell function was differentially affected by cigarette smoke in obese and lean subjects. METHODOLOGY AND PRINCIPAL FINDINGS: Clinical data and blood were collected from 40 severely obese subjects (BMI>40 kg\\/m(2)) and 20 lean healthy subjects. NK cell levels and function were assessed using flow cytometry and cytotoxicity assays. The effect of cigarette smoke on NK cell ability to kill K562 tumour cells was assessed in the presence or absence of the adipokines leptin and adiponectin. NK cell levels were significantly decreased in obese subjects compared to lean controls (7.6 vs 16.6%, p = 0.0008). NK function was also significantly compromised in obese patients (30% +\\/- 13% vs 42% +\\/-12%, p = 0.04). Cigarette smoke inhibited NK cell ability to kill tumour cell lines (p<0.0001). NK cells from obese subjects were even more susceptible to the inhibitory effects of smoke compared to lean subjects (33% vs 28%, p = 0.01). Cigarette smoke prevented NK cell activation, as well as perforin and interferon-gamma secretion upon tumour challenge. Adiponectin but not leptin partially reversed the effects of smoke on NK cell function in both obese (p = 0.002) and lean controls (p = 0.01). CONCLUSIONS\\/SIGNIFICANCE: Obese subjects have impaired NK cell activity that is more susceptible to the detrimental effects of cigarette smoke compared to lean subjects. This may play a role in the increase of cancer and infection seen in this population. Adiponectin is capable of restoring NK cell activity and may have therapeutic potential for immunity in obese subjects and smokers.

  3. Natural killer cells in obesity: impaired function and increased susceptibility to the effects of cigarette smoke.

    Directory of Open Access Journals (Sweden)

    Donal O'Shea

    Full Text Available BACKGROUND: Obese individuals who smoke have a 14 year reduction in life expectancy. Both obesity and smoking are independently associated with increased risk of malignancy. Natural killer cells (NK are critical mediators of anti-tumour immunity and are compromised in obese patients and smokers. We examined whether NK cell function was differentially affected by cigarette smoke in obese and lean subjects. METHODOLOGY AND PRINCIPAL FINDINGS: Clinical data and blood were collected from 40 severely obese subjects (BMI>40 kg/m(2 and 20 lean healthy subjects. NK cell levels and function were assessed using flow cytometry and cytotoxicity assays. The effect of cigarette smoke on NK cell ability to kill K562 tumour cells was assessed in the presence or absence of the adipokines leptin and adiponectin. NK cell levels were significantly decreased in obese subjects compared to lean controls (7.6 vs 16.6%, p = 0.0008. NK function was also significantly compromised in obese patients (30% +/- 13% vs 42% +/-12%, p = 0.04. Cigarette smoke inhibited NK cell ability to kill tumour cell lines (p<0.0001. NK cells from obese subjects were even more susceptible to the inhibitory effects of smoke compared to lean subjects (33% vs 28%, p = 0.01. Cigarette smoke prevented NK cell activation, as well as perforin and interferon-gamma secretion upon tumour challenge. Adiponectin but not leptin partially reversed the effects of smoke on NK cell function in both obese (p = 0.002 and lean controls (p = 0.01. CONCLUSIONS/SIGNIFICANCE: Obese subjects have impaired NK cell activity that is more susceptible to the detrimental effects of cigarette smoke compared to lean subjects. This may play a role in the increase of cancer and infection seen in this population. Adiponectin is capable of restoring NK cell activity and may have therapeutic potential for immunity in obese subjects and smokers.

  4. Increased cardiogenesis in P19-GFP teratocarcinoma cells expressing the propeptide IGF-1Ea

    International Nuclear Information System (INIS)

    Poudel, Bhawana; Bilbao, Daniel; Sarathchandra, Padmini; Germack, Renee; Rosenthal, Nadia; Santini, Maria Paola

    2011-01-01

    Highlights: ► In this study, we explored the function of IGF-1Ea propeptide in inducing cardiogenesis of stem cells. ► IGF-1Ea promoted cardiac mesodermal induction in uncommitted cells. ► Under differentiation condition, IGF-1Ea increased expression of cardiac differentiation markers. ► Furthermore, it promoted formation of finely organized sarcomeric structure. ► IGF-1Ea propeptide may be a good candidate to improve production of cardiomyocytes from pluripotent cells. -- Abstract: The mechanism implicated in differentiation of endogenous cardiac stem cells into cardiomyocytes to regenerate the heart tissue upon an insult remains elusive, limiting the therapeutical goals to exogenous cell injection and/or gene therapy. We have shown previously that cardiac specific overexpression of the insulin-like growth factor 1 propeptide IGF-1Ea induces beneficial myocardial repair after infarct. Although the mechanism is still under investigation, the possibility that this propeptide may be involved in promoting stem cell differentiation into the cardiac lineage has yet to be explored. To investigate whether IGF-1Ea promote cardiogenesis, we initially modified P19 embryonal carcinoma cells to express IGF-1Ea. Taking advantage of their cardiomyogenic nature, we analyzed whether overexpression of this propeptide affected cardiac differentiation program. The data herein presented showed for the first time that constitutively overexpressed IGF-1Ea increased cardiogenic differentiation program in both undifferentiated and DMSO-differentiated cells. In details, IGF-1Ea overexpression promoted localization of alpha-actinin in finely organized sarcomeric structure compared to control cells and upregulated the cardiac mesodermal marker NKX-2.5 and the ventricular structural protein MLC2v. Furthermore, activated IGF-1 signaling promoted cardiac mesodermal induction in undifferentiated cells independently of cell proliferation. This analysis suggests that IGF-1Ea may be a

  5. Increased phorbol 12,13-dibutyrate (PDBu) receptor function associated with sickle red cell membrane ghosts

    International Nuclear Information System (INIS)

    Ramachandran, M.; Nair, C.N.; Abraham, E.C.

    1987-01-01

    The biological receptor for tumor-promoting phorbol esters has been identified as the Ca 2+ /phospholipid dependent enzyme, protein kinase C. In the red cell, this enzyme is mainly cytosolic but becomes translocated to the membrane if the cellular Ca 2+ is allowed to rise. Since cellular Ca 2+ in sickle red cells is high, it was reasoned that this enzyme may become more membrane-bound. In fact, the authors noticed a four-fold increase in the binding of 3 H-PDBu by membrane ghosts isolated from sickle red cells compared to normal red cells (pmoles PDBu bound/mg protein; normal = 0.3 vs sickle cell = 1.4). Attempts to assay the enzyme directly as phospholipid-activated 32 P incorporation into the acid-precipitable membrane proteins also indicated a two-fold increase in the radiolabelling of sickle cell membrane ghosts. Autophosphorylation of membrane proteins and analysis of the phosphorylation profile by SDS-PAGE and autoradiography revealed phosphorylation predominantly of bands 3, 4.1 and 4.9 which are known protein kinase C substrates for the red cell enzyme. The increased membrane-associated protein kinase C in sickle red cells may have a bearing on the altered membrane properties reported in this condition

  6. N-acetylphytosphingosine enhances the radiosensitivity of tumor cells by increasing apoptosis

    International Nuclear Information System (INIS)

    Han, Y.; Kim, Y.; Yun, Y.; Jeon, S.; Kim, K.; Song, J.; Hong, S.H.; Park, C.

    2005-01-01

    Ceramides are well-known second messengers which mediate apoptosis, proliferation, differentiation in mammalian cells, but the physiological roles of phytosphingosines are poorly understood. We hypothesized that one of the phytosphingosine derivatives, N-acetylphytosphingosine (NAPS) can induce apoptosis in human leukemia Jurkat cell line and increase apoptosis in irradiated MDA-MB-231 cells. We first examined the effect of NAPS on apoptosis of Jurkat cells. NAPS had a more rapid and stronger apoptotic effect than C 2 -ceramide in Jurkat cells and significant increase of apoptosis was observed at 3 h after treatment. In contrast, the apoptosis induced by C2-ceramide was observed only after 16 h of treatment. NAPS induced apoptosis was mediated by caspase 3 and 8 activation and inhibited by z-VAD-fmk. Ceramide plays a pivotal role in radiation induced apoptosis. We postulated that exogenous treatment of NAPS sensitizes tumor cells to ionizing radiation, since NAPS might be used as a more effective alternative to C2-ceramide. As expected, NAPS decreased clonogenic survival of irradiated MDA-MB-231 cells dose dependently, and apoptosis of irradiated cells in the presence of NAPS was increased through the caspase activation. Taken together, NAPS is an effective apoptosis-inducing agent, which can be readily synthesized from yeast sources, and is a potent alternative to ceramide for the further study of ceramide associated signaling and the development of radiosensitizing agent. (orig.)

  7. Chronic exposure to trichloroethylene increases DNA methylation of the Ifng promoter in CD4+ T cells.

    Science.gov (United States)

    Gilbert, Kathleen M; Blossom, Sarah J; Erickson, Stephen W; Broadfoot, Brannon; West, Kirk; Bai, Shasha; Li, Jingyun; Cooney, Craig A

    2016-10-17

    CD4 + T cells in female MRL+/+ mice exposed to solvent and water pollutant trichloroethylene (TCE) skew toward effector/memory CD4 + T cells, and demonstrate seemingly non-monotonic alterations in IFN-γ production. In the current study we examined the mechanism for this immunotoxicity using effector/memory and naïve CD4 + T cells isolated every 6 weeks during a 40 week exposure to TCE (0.5mg/ml in drinking water). A time-dependent effect of TCE exposure on both Ifng gene expression and IFN-γ protein production was observed in effector/memory CD4 + T cells, with an increase after 22 weeks of exposure and a decrease after 40 weeks of exposure. No such effect of TCE was observed in naïve CD4 + T cells. A cumulative increase in DNA methylation in the CpG sites of the promoter of the Ifng gene was observed in effector/memory, but not naïve, CD4 + T cells over time. Also unique to the Ifng promoter was an increase in methylation variance in effector/memory compared to naïve CD4 + T cells. Taken together, the CpG sites of the Ifng promoter in effector/memory CD4 + T cells were especially sensitive to the effects of TCE exposure, which may help explain the regulatory effect of the chemical on this gene. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

  8. Active Prompting to Decrease Cell Phone Use and Increase Seat Belt Use While Driving

    OpenAIRE

    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 to 2,600 U.S. fatalities each year. An active prompting procedure was employed to increase seat belt use and decrease cell phone use among drivers ex...

  9. Increased Arf/p53 activity in stem cells, aging and cancer.

    Science.gov (United States)

    Carrasco-Garcia, Estefania; Moreno, Manuel; Moreno-Cugnon, Leire; Matheu, Ander

    2017-04-01

    Arf/p53 pathway protects the cells against DNA damage induced by acute stress. This characteristic is the responsible for its tumor suppressor activity. Moreover, it regulates the chronic type of stress associated with aging. This is the basis of its anti-aging activity. Indeed, increased gene dosage of Arf/p53 displays elongated longevity and delayed aging. At a cellular level, it has been recently shown that increased dosage of Arf/p53 delays age-associated stem cell exhaustion and the subsequent decline in tissue homeostasis and regeneration. However, p53 can also promote aging if constitutively activated. In this context, p53 reduces tissue regeneration, which correlates with premature exhaustion of stem cells. We discuss here the current evidence linking the Arf/p53 pathway to the processes of aging and cancer through stem cell regulation. © 2017 The Authors. Aging Cell published by the Anatomical Society and John Wiley & Sons Ltd.

  10. Chronic exposure to water pollutant trichloroethylene increased epigenetic drift in CD4(+) T cells.

    Science.gov (United States)

    Gilbert, Kathleen M; Blossom, Sarah J; Erickson, Stephen W; Reisfeld, Brad; Zurlinden, Todd J; Broadfoot, Brannon; West, Kirk; Bai, Shasha; Cooney, Craig A

    2016-05-01

    Autoimmune disease and CD4(+) T-cell alterations are induced in mice exposed to the water pollutant trichloroethylene (TCE). We examined here whether TCE altered gene-specific DNA methylation in CD4(+) T cells as a possible mechanism of immunotoxicity. Naive and effector/memory CD4(+) T cells from mice exposed to TCE (0.5 mg/ml in drinking water) for 40 weeks were examined by bisulfite next-generation DNA sequencing. A probabilistic model calculated from multiple genes showed that TCE decreased methylation control in CD4(+) T cells. Data from individual genes fitted to a quadratic regression model showed that TCE increased gene-specific methylation variance in both CD4 subsets. TCE increased epigenetic drift of specific CpG sites in CD4(+) T cells.

  11. Chronic exposure to water pollutant trichloroethylene increased epigenetic drift in CD4+ T cells

    Science.gov (United States)

    Gilbert, Kathleen M; Blossom, Sarah J; Erickson, Stephen W; Reisfeld, Brad; Zurlinden, Todd J; Broadfoot, Brannon; West, Kirk; Bai, Shasha; Cooney, Craig A

    2016-01-01

    Aim: Autoimmune disease and CD4+ T-cell alterations are induced in mice exposed to the water pollutant trichloroethylene (TCE). We examined here whether TCE altered gene-specific DNA methylation in CD4+ T cells as a possible mechanism of immunotoxicity. Materials & methods: Naive and effector/memory CD4+ T cells from mice exposed to TCE (0.5 mg/ml in drinking water) for 40 weeks were examined by bisulfite next-generation DNA sequencing. Results: A probabilistic model calculated from multiple genes showed that TCE decreased methylation control in CD4+ T cells. Data from individual genes fitted to a quadratic regression model showed that TCE increased gene-specific methylation variance in both CD4 subsets. Conclusion: TCE increased epigenetic drift of specific CpG sites in CD4+ T cells. PMID:27092578

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

    Science.gov (United States)

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

    2015-11-03

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

  13. Gambogic Acid Lysinate Induces Apoptosis in Breast Cancer MCF-7 Cells by Increasing Reactive Oxygen Species

    Directory of Open Access Journals (Sweden)

    Yong-Zhan Zhen

    2015-01-01

    Full Text Available Gambogic acid (GA inhibits the proliferation of various human cancer cells. However, because of its water insolubility, the antitumor efficacy of GA is limited. Objectives. To investigate the antitumor activity of gambogic acid lysinate (GAL and its mechanism. Methods. Inhibition of cell proliferation was determined by MTT assay; intracellular ROS level was detected by staining cells with DCFH-DA; cell apoptosis was determined by flow cytometer and the mechanism of GAL was investigated by Western blot. Results. GAL inhibited the proliferation of MCF-7 cells with IC50 values 1.46 μmol/L comparable with GA (IC50, 1.16 μmol/L. GAL promoted the production of ROS; however NAC could remove ROS and block the effect of GAL. GAL inhibited the expression of SIRT1 but increased the phosphorylation of FOXO3a and the expression of p27Kip1. At knockdown of FOXO3a, cell apoptosis induced by GAL can be partly blocked. In addition it also enhanced the cleavage of caspase-3. Conclusions. GAL inhibited MCF-7 cell proliferation and induced MCF-7 cell apoptosis by increasing ROS level which could induce cell apoptosis by both SIRT1/FOXO3a/p27Kip1 and caspase-3 signal pathway. These results suggested that GAL might be useful as a modulation agent in cancer chemotherapy.

  14. Increased Circulating Anti-inflammatory Cells in Marathon-trained Runners.

    Science.gov (United States)

    Rehm, K; Sunesara, I; Marshall, G D

    2015-10-01

    Exercise training can alter immune function. Marathon training has been associated with an increased susceptibility to infectious diseases and an increased activity of inflammatory-based diseases, but the precise mechanisms are unknown. The purpose of this study was to compare levels of circulating CD4+  T cell subsets in the periphery of marathon-trained runners and matched non-marathon controls. 19 recreational marathoners that were 4 weeks from running a marathon and 19 demographically-matched healthy control subjects had the percentage of CD4+ T cell subpopulations (T helper 1, T helper 2, T helper 1/T helper 2 ratio, regulatory T cells, CD4+ IL10+, and CD4+ TGFβ+ (Transforming Growth Factor-beta) measured by flow cytometry. Marathon-trained runners had significantly less T helper 1 and regulatory T cells and significantly more T helper 2, CD4+ IL10+, and TGFβ+ cells than the control subjects. The alterations in the percentage of T helper 1 and T helper 2 cells led to a significantly lower T helper 1/T helper 2 ratio in the marathon-trained runners. These data suggest that endurance-based training can increase the number of anti-inflammatory cells. This may be a potential mechanism for the increased incidence of both infectious and inflammatory diseases observed in endurance athletes. © Georg Thieme Verlag KG Stuttgart · New York.

  15. Y-27632 Increases Sensitivity of PANC-1 Cells to Epigallocatechin Gallate (EGCG) in Regulating Cell Proliferation and Migration

    Science.gov (United States)

    Liu, Xing; Bi, Yongyi

    2016-01-01

    Background The study aimed to investigate the inhibitory effect of (1R,4r)-4-((R)-1-aminoethyl)-N-(pyridin-4-yl) cyclohexanecarboxamide (Y-27632) and (−)-epigallocatechin-3-gallate (EGCG) on the proliferation and migration of PANC-1 cells. EGCG, found in green tea, has been previously shown to be one of the most abundant and powerful catechins in cancer prevention and treatment. Y-27632, a selective inhibitor of rho-associated protein kinase 1, is widely used in treating cardiovascular disease, inflammation, and cancer. Material/Methods PANC-1 cells, maintained in Dulbecco’s Modified Eagle’s Medium, were treated with dimethyl sulfoxide (control) as well as different concentrations (20, 40, 60, and 80 μg/mL) of EGCG for 48 h. In addition, PANC-1 cells were treated separately with 60 μg/mL EGCG, 20 μM Y-27632, and EGCG combined with Y-27632 (60 μg/mL EGCG + 20 μM Y-27632) for 48 h. The effect of EGCG and Y-27632 on the proliferation and migration of PANC-1 cells was evaluated using Cell Counting Kit-8 and transwell migration assays. The expression of peroxisome proliferator–activated receptor alpha (PPARα) and Caspase-3 mRNA was determined by Quantitative real-time polymerase chain reaction (RT-qPCR). Results EGCG (20–80 μg/mL) inhibited cell viability in a dose-dependent manner. Y-27632 enhanced the sensitivity of PANC-1 cells to EGCG (by increasing the expression of PPARα and Caspase-3 mRNA) and suppressed cell proliferation. PANC-1 cell migration was inhibited by treatment with a combination of EGCG and Y-27632. Conclusions Y-27632 increases the sensitivity of PANC-1 cells to EGCG in regulating cell proliferation and migration, which is likely to be related to the expression of PPARα mRNA and Caspase-3 mRNA. PMID:27694793

  16. Bifidobacterium breve attenuates murine dextran sodium sulfate-induced colitis and increases regulatory T cell responses.

    Science.gov (United States)

    Zheng, Bin; van Bergenhenegouwen, Jeroen; Overbeek, Saskia; van de Kant, Hendrik J G; Garssen, Johan; Folkerts, Gert; Vos, Paul; Morgan, Mary E; Kraneveld, Aletta D

    2014-01-01

    While some probiotics have shown beneficial effects on preventing or treating colitis development, others have shown no effects. In this study, we have assessed the immunomodulating effects of two probiotic strains, Lactobacillus rhamnosus (L. rhamnosus) and Bifidobacterium breve (B. breve) on T cell polarization in vitro, using human peripheral blood mononuclear cells (PBMC), and in vivo, using murine dextran sodium sulfate (DSS) colitis model. With respect to the latter, the mRNA expression of T cell subset-associated transcription factors and cytokines in the colon was measured and the T helper type (Th) 17 and regulatory T cell (Treg) subsets were determined in the Peyer's patches. Both L. rhamnosus and B. breve incubations in vitro reduced Th17 and increased Th2 cell subsets in human PBMCs. In addition, B. breve incubation was also able to reduce Th1 and increase Treg cell subsets in contrast to L. rhamnosus. In vivo intervention with B. breve, but not L. rhamnosus, significantly attenuated the severity of DSS-induced colitis. In DSS-treated C57BL/6 mice, intervention with B. breve increased the expression of mRNA encoding for Th2- and Treg-associated cytokines in the distal colon. In addition, intervention with B. breve led to increases of Treg and decreases of Th17 cell subsets in Peyer's patches of DSS-treated mice. B. breve modulates T cell polarization towards Th2 and Treg cell-associated responses in vitro and in vivo. In vivo B. breve intervention ameliorates DSS-induced colitis symptoms and this protective effect may mediated by its effects on the T-cell composition.

  17. Increased cell proliferation and mucocyte density in the sea anemone Aiptasia pallida recovering from bleaching.

    Directory of Open Access Journals (Sweden)

    David Fransolet

    Full Text Available Recovery of coral after bleaching episodes is a critical period for the health of the reef ecosystem. While events such as symbiont (genus Symbiodinium shifting/shuffling or tissue apoptosis have been demonstrated to occur following bleaching, little is known concerning tissue recovery or cell proliferation. Here, we studied the sea anemone Aiptasia pallida exposed to a transient elevation of water temperature combined with high illumination (33°C and 1900 µmol photons x m(-2 x s(-1 for 30 h. Following such treatment bleached anemones showed a significant reduction of their Symbiodinium density. Cell proliferation in the ectodermis and gastrodermis was determined by assessing the densities of cells labeled with a thymidine analogue (EdU. Cell proliferation significantly increased during the first day following stress in both tissue types. This increased cell proliferation returned to pre-stress values after one week. Although cell proliferation was higher in the ectodermis in absence of stress, it was relatively more pronounced in the gastrodermis of stressed anemones. In addition, the ratio of ectodermal mucocytes significantly increased three weeks after induced stress. These results suggest that thermal/photic stress coupled with the loss of the symbionts is able to enhance cell proliferation in both gastrodermis and ectodermis of cnidarians. While new cells formed in the gastrodermis are likely to host new Symbiodinium, the fate of new cells in the ectodermis was only partially revealed. Some new ectodermal cells may, in part, contribute to the increased number of mucocytes which could eventually help strengthen the heterotrophic state until restoration of the symbiosis.

  18. Bifidobacterium breve attenuates murine dextran sodium sulfate-induced colitis and increases regulatory T cell responses.

    Directory of Open Access Journals (Sweden)

    Bin Zheng

    Full Text Available While some probiotics have shown beneficial effects on preventing or treating colitis development, others have shown no effects. In this study, we have assessed the immunomodulating effects of two probiotic strains, Lactobacillus rhamnosus (L. rhamnosus and Bifidobacterium breve (B. breve on T cell polarization in vitro, using human peripheral blood mononuclear cells (PBMC, and in vivo, using murine dextran sodium sulfate (DSS colitis model. With respect to the latter, the mRNA expression of T cell subset-associated transcription factors and cytokines in the colon was measured and the T helper type (Th 17 and regulatory T cell (Treg subsets were determined in the Peyer's patches. Both L. rhamnosus and B. breve incubations in vitro reduced Th17 and increased Th2 cell subsets in human PBMCs. In addition, B. breve incubation was also able to reduce Th1 and increase Treg cell subsets in contrast to L. rhamnosus. In vivo intervention with B. breve, but not L. rhamnosus, significantly attenuated the severity of DSS-induced colitis. In DSS-treated C57BL/6 mice, intervention with B. breve increased the expression of mRNA encoding for Th2- and Treg-associated cytokines in the distal colon. In addition, intervention with B. breve led to increases of Treg and decreases of Th17 cell subsets in Peyer's patches of DSS-treated mice. B. breve modulates T cell polarization towards Th2 and Treg cell-associated responses in vitro and in vivo. In vivo B. breve intervention ameliorates DSS-induced colitis symptoms and this protective effect may mediated by its effects on the T-cell composition.

  19. MET signalling in primary colon epithelial cells leads to increased transformation irrespective of aberrant Wnt signalling

    Science.gov (United States)

    Boon, E M J; Kovarikova, M; Derksen, P W B; van der Neut, R

    2005-01-01

    It has been shown that in hereditary and most sporadic colon tumours, components of the Wnt pathway are mutated. The Wnt target MET has been implicated in the development of colon cancer. Here, we show that overexpression of wild-type or a constitutively activated form of MET in colon epithelial cells leads to increased transformation irrespective of Wnt signalling. Fetal human colon epithelial cells without aberrant Wnt signalling were transfected with wild-type or mutated MET constructs. Expression of these constructs leads to increased phosphorylation of MET and its downstream targets PKB and MAPK. Upon stimulation with HGF, the expression of E-cadherin is downregulated in wild-type MET-transfected cells, whereas cells expressing mutated MET show low E-cadherin levels independent of stimulation with ligand. This implies a higher migratory propensity of these cells. Furthermore, fetal human colon epithelial cells expressing the mutated form of MET have colony-forming capacity in soft agar, while cells expressing wild-type MET show an intermediate phenotype. Subcutaneous injection of mutated MET-transfected cells in nude mice leads to the formation of tumours within 12 days in all mice injected. At this time point, mock-transfected cells do not form tumours, while wild-type MET-transfected cells form subcutaneous tumours in one out of five mice. We thus show that MET signalling can lead to increased transformation of colon epithelial cells independent of Wnt signalling and in this way could play an essential role in the onset and progression of colorectal cancer. PMID:15785735

  20. Macrophage migration inhibitory factor stimulated by Helicobacter pylori increases proliferation of gastric epithelial cells

    Science.gov (United States)

    Xia, Harry Hua-Xiang; Lam, Shiu Kum; Chan, Annie O.O.; Lin, Marie Chia Mi; Kung, Hsiang Fu; Ogura, Keiji; Berg, Douglas E.; Wong, Benjamin C. Y.

    2005-01-01

    AIM: Helicobacter pylori (H pylori) is associated with increased gastric inflammatory and epithelial expression of macrophage migration inhibitory factor (MIF) and gastric epithelial cell proliferation. This study aimed at determining whether H pylori directly stimulates release of MIF in monocytes, whether the cag pathogenicity island (PAI) is involved for this function, and whether MIF stimulated by H pylori increases gastric epithelial cell proliferation in vitro. METHODS: A cytotoxic wild-type H pylori strain (TN2)and its three isogenic mutants (TN2△cag, TN2△cagA and TN2△cagE) were co-cultured with cells of a human monocyte cell line, THP-1, for 24 h at different organism/cell ratios. MIF in the supernatants was measured by an ELISA. Cells of a human gastric cancer cell line, MKN45, were then co-cultured with the supernatants, with and without monoclonal anti-MIF antibody for 24 h. The cells were further incubated for 12 h after addition of 3H-thymidine, and the levels of incorporation of 3H-thymidine were measured with a liquid scintillation counter. RESULTS: The wild-type strain and the isogenic mutants, TN2△cagA and TN2△cagE, increased MIF release at organism/cell ratios of 200/1 and 400/1, but not at the ratios of 50/1 and 100/1. However, the mutant TN2△cag did not increase the release of MIF at any of the four ratios. 3H-thymidine readings for MKN-45 cells were significantly increased with supernatants derived from the wild-type strain and the mutants TN2△cagA and TN2△cagE, but not from the mutant TN2△cag. Moreover, in the presence of monoclonal anti-MIF antibody, the stimulatory effects of the wild-type strain on cell proliferation disappeared. CONCLUSION: H pylori stimulates MIF release in monocytes, likely through its cag PAI, but not related to cagA or cagE. H pylori-stimulated monocyte culture supernatant increases gastric cell proliferation, which is blocked by anti-MIF antibody, suggesting that MIF plays an important role in H

  1. Human epithelial cells increase their rigidity with ageing in vitro: direct measurements

    International Nuclear Information System (INIS)

    Berdyyeva, Tamara K; Woodworth, Craig D; Sokolov, Igor

    2005-01-01

    The decrease in elasticity of epithelial tissues with ageing contributes to many human diseases. This change was previously attributed to increased crosslinking of extracellular matrix proteins. Here we show that individual human epithelial cells also become significantly more rigid during ageing in vitro. Using atomic force microscopy (AFM), we found that the Young's modulus of viable cells was consistently increased two- to four-fold in older versus younger cells. Direct visualization of the cytoskeleton using a novel method involving the AFM suggested that increased rigidity of ageing cells was due to a higher density of cytoskeletal fibres. Our results identify a unique mechanism that might contribute to the age-related loss of elasticity in epithelial tissues

  2. Overexpression of 15-lipoxygenase-1 in PC-3 human prostate cancer cells increases tumorigenesis.

    Science.gov (United States)

    Kelavkar, U P; Nixon, J B; Cohen, C; Dillehay, D; Eling, T E; Badr, K F

    2001-11-01

    The effect of overexpression of 15-lipoxygenase-1 (15-LO-1) was studied in the human prostate cancer cell line, PC-3. Stable PC-3 cell lines were generated by transfection with 15-LO-1-sense (15-LOS), 15-LO-1-antisense (15-LOAS) or vector (Zeo) and selection with Zeocin. After characterization by RT-PCR, western and HPLC, a PC3-15LOS clone was selected that possessed 10-fold 15-LO-1 enzyme activity compared with parental PC-3 cells. The PC3-15LOAS clone displayed little or no 15-LO-1 activity. These PC-3 cell lines were characterized for properties of tumorigenesis. The proliferation rates of the cell lines were as follows: PC3-15LOS > PC-3 = PC3-Zeo > PC3-15LOAS. Addition of a specific 15-LO-1 inhibitor, PD146176, caused a dose-dependent inhibition of proliferation in vitro. Overexpression of 15-LO-1 also caused [(3)H]thymidine incorporation to increase by 4.0-fold (P < 0.01). Compared with parental and PC-3-Zeo cells, PC3-15LOS enhanced whereas PC3-15LOAS reduced the ability of PC-3 cells to grow in an anchorage-independent manner, as assessed by colony formation in soft agar. These data suggested a pro-tumorigenic role for 15-LO-1 in PC-3 cells in vitro. Therefore, to clarify the role of 15-LO-1 in vivo, the effect of 15-LO-1 expression on the growth of tumors in nude mice was investigated. The PC-3 cell lines were inoculated subcutaneously into athymic nude mice. The frequency of tumor formation was increased and the sizes of the tumors formed were much larger in the PC3-15LOS compared with PC3-15LOAS, parental PC-3 and PC-3-Zeo cells. Immunohistochemistry for 15-LO-1 confirmed expression throughout the duration of the experiment. The expression of factor VIII, an angiogenesis marker, in tumor sections was increased in tumors derived from PC3-15LOS cells and decreased in those from PC3-15LOAS cells compared with tumors from parental or Zeo cells. These data further supported the evaluation by ELISA of vascular endothelial growth factor (VEGF) secretion by PC-3

  3. Endothelial progenitor cell mobilization and increased intravascular nitric oxide in patients undergoing cardiac rehabilitation.

    Science.gov (United States)

    Paul, Jonathan D; Powell, Tiffany M; Thompson, Michael; Benjamin, Moshe; Rodrigo, Maria; Carlow, Andrea; Annavajjhala, Vidhya; Shiva, Sruti; Dejam, Andre; Gladwin, Mark T; McCoy, J Philip; Zalos, Gloria; Press, Beverly; Murphy, Mandy; Hill, Jonathan M; Csako, Gyorgy; Waclawiw, Myron A; Cannon, Richard O

    2007-01-01

    We investigated whether cardiac rehabilitation participation increases circulating endothelial progenitor cells (EPCs) and benefits vasculature in patients already on stable therapy previously shown to augment EPCs and improve endothelial function. Forty-six of 50 patients with coronary artery disease completed a 36-session cardiac rehabilitation program: 45 were treated with HMG-CoA reductase inhibitor (statin) therapy > or = 1 month (average baseline low-density lipoprotein cholesterol = 81 mg/dL). Mononuclear cells isolated from blood were quantified for EPCs by flow cytometry (CD133/VEGFR-2 cells) and assayed in culture for EPC colony-forming units (CFUs). In 23 patients, EPCs were stained for annexin-V as a marker of apoptosis, and nitrite was measured in blood as an indicator of intravascular nitric oxide. Endothelial progenitor cells increased from 35 +/- 5 to 63 +/- 10 cells/mL, and EPC-CFUs increased from 0.9 +/- 0.2 to 3.1 +/- 0.6 per well (both P < .01), but 11 patients had no increase in either measure. Those patients whose EPCs increased from baseline showed significant increases in nitrite and reduction in annexin-V staining (both P < .01) versus no change in patients without increase in EPCs. Over the course of the program, EPCs increased prior to increase in nitrite in the blood. Cardiac rehabilitation in patients receiving stable statin therapy and with low-density lipoprotein cholesterol at goal increases EPC number, EPC survival, and endothelial differentiation potential, associated with increased nitric oxide in the blood. Although this response was observed in most patients, a significant minority showed neither EPC mobilization nor increased nitric oxide in the blood.

  4. A mechanically-induced colon cancer cell population shows increased metastatic potential

    KAUST Repository

    Tang, Xin; Kuhlenschmidt, Theresa B; Li, Qian; Ali, Shahjahan; Lezmi, Stephane; Chen, Hong; Pires-Alves, Melissa; Laegreid, William W; Saif, Taher A; Kuhlenschmidt, Mark S

    2014-01-01

    Background: Metastasis accounts for the majority of deaths from cancer. Although tumor microenvironment has been shown to have a significant impact on the initiation and/or promotion of metastasis, the mechanism remains elusive. We previously reported that HCT-8 colon cancer cells underwent a phenotypic transition from an adhesive epithelial type (E-cell) to a rounded dissociated type (R-cell) via soft substrate culture, which resembled the initiation of metastasis. The objective of current study was to investigate the molecular and metabolic mechanisms of the E-R transition.Methods: Global gene expressions of HCT-8 E and R cells were measured by RNA Sequencing (RNA-seq); and the results were further confirmed by real-time PCR. Reactive oxygen species (ROS), anoikis resistance, enzyme activity of aldehyde dehydrogenase 3 family, member A1 (ALDH3A1), and in vitro invasion assay were tested on both E and R cells. The deformability of HCT-8 E and R cells was measured by atomic force microscopy (AFM). To study the in vivo invasiveness of two cell types, athymic nude mice were intra-splenically injected with HCT-8 E or R cells and sacrificed after 9 weeks. Incidences of tumor development and metastasis were histologically evaluated and analyzed with Fisher's exact test.Results: Besides HCT-8, E-R transition on soft substrates was also seen in three other cancer cell lines (HCT116, SW480 colon and DU145 prostate cancer). The expression of some genes, such as ALDH3A1, TNS4, CLDN2, and AKR1B10, which are known to play important roles in cancer cell migration, invasion, proliferation and apoptosis, were increased in HCT-8 R cells. R cells also showed higher ALDH3A1 enzyme activity, higher ROS, higher anoikis resistance, and higher softness than E cells. More importantly, in vitro assay and in vivo animal models revealed that HCT-8 R cells were more invasive than E cells.Conclusions: Our comprehensive comparison of HCT-8 E and R cells revealed differences of molecular

  5. A mechanically-induced colon cancer cell population shows increased metastatic potential

    KAUST Repository

    Tang, Xin

    2014-05-29

    Background: Metastasis accounts for the majority of deaths from cancer. Although tumor microenvironment has been shown to have a significant impact on the initiation and/or promotion of metastasis, the mechanism remains elusive. We previously reported that HCT-8 colon cancer cells underwent a phenotypic transition from an adhesive epithelial type (E-cell) to a rounded dissociated type (R-cell) via soft substrate culture, which resembled the initiation of metastasis. The objective of current study was to investigate the molecular and metabolic mechanisms of the E-R transition.Methods: Global gene expressions of HCT-8 E and R cells were measured by RNA Sequencing (RNA-seq); and the results were further confirmed by real-time PCR. Reactive oxygen species (ROS), anoikis resistance, enzyme activity of aldehyde dehydrogenase 3 family, member A1 (ALDH3A1), and in vitro invasion assay were tested on both E and R cells. The deformability of HCT-8 E and R cells was measured by atomic force microscopy (AFM). To study the in vivo invasiveness of two cell types, athymic nude mice were intra-splenically injected with HCT-8 E or R cells and sacrificed after 9 weeks. Incidences of tumor development and metastasis were histologically evaluated and analyzed with Fisher\\'s exact test.Results: Besides HCT-8, E-R transition on soft substrates was also seen in three other cancer cell lines (HCT116, SW480 colon and DU145 prostate cancer). The expression of some genes, such as ALDH3A1, TNS4, CLDN2, and AKR1B10, which are known to play important roles in cancer cell migration, invasion, proliferation and apoptosis, were increased in HCT-8 R cells. R cells also showed higher ALDH3A1 enzyme activity, higher ROS, higher anoikis resistance, and higher softness than E cells. More importantly, in vitro assay and in vivo animal models revealed that HCT-8 R cells were more invasive than E cells.Conclusions: Our comprehensive comparison of HCT-8 E and R cells revealed differences of molecular

  6. Investigating the low-temperature impedance increase of lithium-ion cells

    International Nuclear Information System (INIS)

    Abraham, D. P.; Heaton, J. R.; Kang, S.-H.; Dees, D. W.; Jansen, A. N.; Chemical Engineering

    2008-01-01

    Low-temperature performance loss is a significant barrier to commercialization of lithium-ion cells in hybrid electric vehicles. Increased impedance, especially at temperatures below 0 C, reduces the cell pulse power performance required for cold engine starts, quick acceleration, or regenerative braking. Here we detail electrochemical impedance spectroscopy data on binder- and carbon-free layered-oxide and spinel-oxide electrodes, obtained over the +30 to ?30 C temperature range, in coin cells containing a lithium-preloaded Li 4/3 Ti 5/3 O 4 composite (LTOc) counter electrode and a LiPF 6 -bearing ethylene carbonate/ethyl methyl carbonate electrolyte. For all electrodes studied, the impedance increased with decreasing cell temperature; the increases observed in the midfrequency arc dwarfed the increases in ohmic resistance and diffusional impedance. Our data suggest that the movement of lithium ions across the electrochemical interface on the active material may have been increasingly hindered at lower temperatures, especially below 0 C. Low-temperature performance may be improved by modifying the electrolyte-active material interface (for example, through electrolyte composition changes). Increasing surface area of active particles (for example, through nanoparticle use) can lower the initial electrode impedance and lead to lower cell impedances at -30 C

  7. Baculovirus p35 increases pancreatic β-cell resistance to apoptosis

    International Nuclear Information System (INIS)

    Hollander, Kenneth; Bar-Chen, Michal; Efrat, Shimon

    2005-01-01

    β-cells die by apoptosis in type 1 diabetes as a result of autoimmune attack mediated by cytokines, and in type 2 diabetes by various perpetrators including human islet amyloid polypeptide (hIAPP). The cascade of apoptotic events induced by cytokines and hIAPP is mediated through caspases and reactive oxygen species. The baculovirus p35 protein is a potent anti-apoptotic agent shown to be effective in a variety of species and able to inhibit a number of apoptotic pathways. Here, we aimed at determining the protective potential of p35 in β-cells exposed to cytokines and hIAPP, as well as the effects of p35 on β-cell function. The p35 gene was introduced into βTC-tet cells, a differentiated murine β-cell line capable of undergoing inducible growth-arrest. Both proliferating and growth-arrested cells expressing p35 manifested increased resistance to cytokines and hIAPP, compared with control cells, as judged by cell viability, DNA fragmentation, and caspase-3 activity assays. p35 was significantly more protective in growth-arrested, compared with proliferating, cells. No significant differences were observed in proliferation and insulin content between cells expressing p35 and control cells. In contrast, p35 manifested a perturbing effect on glucose-induced insulin secretion. These findings suggest that p35 could be incorporated as part of a multi-pronged approach of immunoprotective strategies to provide protection from recurring autoimmunity for transplanted β-cells, as well as in preventive gene therapy in type 1 diabetes. p35 may also be protective from β-cell damage caused by hIAPP in type 2 diabetes

  8. Naive and effector B-cell subtypes are increased in chronic rhinosinusitis with polyps.

    Science.gov (United States)

    Miljkovic, Dijana; Psaltis, Alkis; Wormald, Peter-John; Vreugde, Sarah

    2018-01-01

    Recent studies demonstrated that B cells and their chemoattractants are elevated in the nasal mucosa of patients with chronic rhinosinusitis (CRS) with nasal polyposis (CRSwNP). However, the presence of naive B cells and of plasmablasts and memory B-cell subsets in the mucosa and periphery of the same patient with CRS is yet to be characterized. Here we sought to quantify naive, plasmablasts, and memory B cells in mucosal tissue and peripheral blood of patients with CRSwNP, patients with CRS without nasal polyps (CRSsNP), and control patients. Polyps, mucosa, and peripheral blood samples were prospectively collected from the patients with CRS and from the non-CRS controls. We used flow cytometry to distinguish among naive, plasmablast, and memory B cells in sinus tissue and peripheral blood. A total of 45 patients were recruited for the study. The patients with CRSwNP had significantly increased mucosal B-cell numbers versus the controls (3.39 ± 4.05% versus 0.39 ± 1.05% of live cells; p Kruskal-Wallis test), which included naive B cells (0.61 ± 0.94 versus 0.11 ± 0.24% of live cells; p Kruskal-Wallis test), plasmablasts (0.06 ± 0.26 versus 0.00 ± 0.00% of live cells; p Kruskal-Wallis test), and memory B cells (0.62 ± 1.26 versus 0.05 ± 0.15% of live cells; p Kruskal-Wallis test). Our study identified increased frequencies of different B-cell subtypes in the mucosa of patients with CRSwNP but not in the peripheral blood. We also found that patients with CRSwNP had significantly increased B-cell subtypes compared with the patients with CRSsNP and the controls. These results implied a potential role for mucosal B cells in the ongoing inflammation in patients with CRSwNP.

  9. Enhanced lysosomal acidification leads to increased chloroquine accumulation in CHO cells expressing the pfmdr1 gene

    NARCIS (Netherlands)

    van Es, H. H.; Renkema, H.; Aerts, H.; Schurr, E.

    1994-01-01

    Expression of the pfmdr1-encoded Pgh1 protein of Plasmodium falciparum in CHO cells confers a phenotype of increased sensitivity to chloroquine due to an increased Pgh1-mediated accumulation of this antimalarial. Pgh1 carrying amino acid substitutions associated with chloroquine resistance in P.

  10. Increasing the solar cell power output by coating with transition metal-oxide nanorods

    International Nuclear Information System (INIS)

    Kuznetsov, I.A.; Greenfield, M.J.; Mehta, Y.U.; Merchan-Merchan, W.; Salkar, G.; Saveliev, A.V.

    2011-01-01

    Highlights: → Nanoparticles enhance solar cell efficiency. → Solar cell power increase by nanorod coating. → Metal-oxide nanorods are prepared in flames. → Molybdenum oxide nanorods effectively scatter light on solar cell surface. → Scattering efficiency depends on coating density. -- Abstract: Photovoltaic cells produce electric current through interactions among photons from an ambient light source and electrons in the semiconductor layer of the cell. However, much of the light incident on the panel is reflected or absorbed without inducing the photovoltaic effect. Transition metal-oxide nanoparticles, an inexpensive product of a process called flame synthesis, can cause scattering of light. Scattering can redirect photon flux, increasing the fraction of light absorbed in the thin active layer of silicon solar cells. This research aims to demonstrate that the application of transition metal-oxide nanorods to the surface of silicon solar panels can enhance the power output of the panels. Several solar panels were coated with a nanoparticle-methanol suspension, and the power outputs of the panels before and after the treatment were compared. The results demonstrate an increase in power output of up to 5% after the treatment. The presence of metal-oxide nanorods on the surface of the coated solar cells is confirmed by electron microscopy.

  11. Universal method for protein bioconjugation with nanocellulose scaffolds for increased cell adhesion.

    Science.gov (United States)

    Kuzmenko, Volodymyr; Sämfors, Sanna; Hägg, Daniel; Gatenholm, Paul

    2013-12-01

    Bacterial nanocellulose (BNC) is an emerging biomaterial since it is biocompatible, integrates well with host tissue and can be biosynthesized in desired architecture. However, being a hydrogel, it exhibits low affinity for cell attachment, which is crucial for the cellular fate process. To increase cell attachment, the surface of BNC scaffolds was modified with two proteins, fibronectin and collagen type I, using an effective bioconjugation method applying 1-cyano-4-dimethylaminopyridinium (CDAP) tetrafluoroborate as the intermediate catalytic agent. The effect of CDAP treatment on cell adhesion to the BNC surface is shown for human umbilical vein endothelial cells and the mouse mesenchymal stem cell line C3H10T1/2. In both cases, the surface modification increased the number of cells attached to the surfaces. In addition, the morphology of the cells indicated more healthy and viable cells. CDAP activation of bacterial nanocellulose is shown to be a convenient method to conjugate extracellular proteins to the scaffold surfaces. CDAP treatment can be performed in a short period of time in an aqueous environment under heterogeneous and mild conditions preserving the nanofibrillar network of cellulose. © 2013.

  12. The oxysterol 27-hydroxycholesterol increases β-amyloid and oxidative stress in retinal pigment epithelial cells

    Directory of Open Access Journals (Sweden)

    Dasari Bhanu

    2010-09-01

    Full Text Available Abstract Background Alzheimer's disease (AD and age-related macular degeneration (AMD share several pathological features including β-amyloid (Aβ peptide accumulation, oxidative damage, and cell death. The causes of AD and AMD are not known but several studies suggest disturbances in cholesterol metabolism as a culprit of these diseases. We have recently shown that the cholesterol oxidation metabolite 27-hydroxycholesterol (27-OHC causes AD-like pathology in human neuroblastoma SH-SY5Y cells and in organotypic hippocampal slices. However, the extent to which and the mechanisms by which 27-OHC may also cause pathological hallmarks related to AMD are ill-defined. In this study, the effects of 27-OHC on AMD-related pathology were determined in ARPE-19 cells. These cells have structural and functional properties relevant to retinal pigmented epithelial cells, a target in the course of AMD. Methods ARPE-19 cells were treated with 0, 10 or 25 μM 27-OHC for 24 hours. Levels of Aβ peptide, mitochondrial and endoplasmic reticulum (ER stress markers, Ca2+ homeostasis, glutathione depletion, reactive oxygen species (ROS generation, inflammation and cell death were assessed using ELISA, Western blot, immunocytochemistry, and specific assays. Results 27-OHC dose-dependently increased Aβ peptide production, increased levels of ER stress specific markers caspase 12 and gadd153 (also called CHOP, reduced mitochondrial membrane potential, triggered Ca2+ dyshomeostasis, increased levels of the nuclear factor κB (NFκB and heme-oxygenase 1 (HO-1, two proteins activated by oxidative stress. Additionally, 27-OHC caused glutathione depletion, ROS generation, inflammation and apoptotic-mediated cell death. Conclusions The cholesterol metabolite 27-OHC is toxic to RPE cells. The deleterious effects of this oxysterol ranged from Aβ accumulation to oxidative cell damage. Our results suggest that high levels of 27-OHC may represent a common pathogenic factor for

  13. Suspension state increases reattachment of breast cancer cells by up-regulating lamin A/C.

    Science.gov (United States)

    Zhang, Xiaomei; Lv, Yonggang

    2017-12-01

    Extravasation is a rate-limiting step of tumor metastasis, for which adhesion to endothelium of circulating tumor cells (CTCs) is the prerequisite. The suspension state of CTCs undergoing detachment from primary tumor is a persistent biomechanical cue, which potentially regulates the biophysical characteristics and cellular behaviors of tumor cells. In this study, breast tumor cells MDA-MB-231 in suspension culture condition were used to investigate the effect of suspension state on reattachment of CTCs. Our study demonstrated that suspension state significantly increased the adhesion ability of breast tumor cells. In addition, suspension state markedly promoted the formation of stress fibers and focal adhesions and reduced the motility in reattached breast cancer cells. Moreover, lamin A/C was reversibly accumulated at posttranscriptional level under suspension state, improving the cell stiffness of reattached breast cancer cells. Disruption of actin cytoskeleton by cytochalasin D caused lamin A/C accumulation. Conversely, decreasing actomyosin contraction by ROCK inhibitor Y27632 reduced lamin A/C level. Knocking down lamin A/C weakened the suspension-induced increase of adhesion, and also abolished the suspension-induced decrease of motility and increase of stress fibers and focal adhesion in reattaching tumor cells, suggesting a crucial role of lamin A/C. In conclusion, it was demonstrated that suspension state promoted the reattachment of breast tumor cells by up-regulating lamin A/C via cytoskeleton disruption. These findings highlight the important role of suspension state for tumor cells in tumor metastasis. Copyright © 2017 Elsevier B.V. All rights reserved.

  14. Mechanical stretch increases CCN2/CTGF expression in anterior cruciate ligament-derived cells

    International Nuclear Information System (INIS)

    Miyake, Yoshiaki; Furumatsu, Takayuki; Kubota, Satoshi; Kawata, Kazumi; Ozaki, Toshifumi; Takigawa, Masaharu

    2011-01-01

    Highlights: → CCN2/CTGF localizes to the ligament-to-bone interface, but is not to the midsubstance region of human anterior cruciate ligament (ACL). → Mechanical stretch induces higher increase of CCN2/CTGF gene expression and protein secretion in ACL interface cells compared with ACL midsubstance cells. → CCN2/CTGF treatment stimulates the proliferation of ACL interface cells. -- Abstract: Anterior cruciate ligament (ACL)-to-bone interface serves to minimize the stress concentrations that would arise between two different tissues. Mechanical stretch plays an important role in maintaining cell-specific features by inducing CCN family 2/connective tissue growth factor (CCN2/CTGF). We previously reported that cyclic tensile strain (CTS) stimulates α1(I) collagen (COL1A1) expression in human ACL-derived cells. However, the biological function and stress-related response of CCN2/CTGF were still unclear in ACL fibroblasts. In the present study, CCN2/CTGF was observed in ACL-to-bone interface, but was not in the midsubstance region by immunohistochemical analyses. CTS treatments induced higher increase of CCN2/CTGF expression and secretion in interface cells compared with midsubstance cells. COL1A1 expression was not influenced by CCN2/CTGF treatment in interface cells despite CCN2/CTGF stimulated COL1A1 expression in midsubstance cells. However, CCN2/CTGF stimulated the proliferation of interface cells. Our results suggest that distinct biological function of stretch-induced CCN2/CTGF might regulate region-specific phenotypes of ACL-derived cells.

  15. Mechanical stretch increases CCN2/CTGF expression in anterior cruciate ligament-derived cells

    Energy Technology Data Exchange (ETDEWEB)

    Miyake, Yoshiaki [Department of Orthopaedic Surgery, Science of Functional Recovery and Reconstruction, Okayama University Graduate School of Medicine, Dentistry, and Pharmaceutical Sciences, 2-5-1 Shikata-cho, Kita-ku, Okayama (Japan); Department of Biochemistry and Molecular Dentistry, Okayama University Graduate School of Medicine, Dentistry, and Pharmaceutical Sciences, 2-5-1 Shikata-cho, Kita-ku, Okayama (Japan); Furumatsu, Takayuki, E-mail: matino@md.okayama-u.ac.jp [Department of Orthopaedic Surgery, Science of Functional Recovery and Reconstruction, Okayama University Graduate School of Medicine, Dentistry, and Pharmaceutical Sciences, 2-5-1 Shikata-cho, Kita-ku, Okayama (Japan); Kubota, Satoshi; Kawata, Kazumi [Department of Biochemistry and Molecular Dentistry, Okayama University Graduate School of Medicine, Dentistry, and Pharmaceutical Sciences, 2-5-1 Shikata-cho, Kita-ku, Okayama (Japan); Ozaki, Toshifumi [Department of Orthopaedic Surgery, Science of Functional Recovery and Reconstruction, Okayama University Graduate School of Medicine, Dentistry, and Pharmaceutical Sciences, 2-5-1 Shikata-cho, Kita-ku, Okayama (Japan); Takigawa, Masaharu [Department of Biochemistry and Molecular Dentistry, Okayama University Graduate School of Medicine, Dentistry, and Pharmaceutical Sciences, 2-5-1 Shikata-cho, Kita-ku, Okayama (Japan)

    2011-06-03

    Highlights: {yields} CCN2/CTGF localizes to the ligament-to-bone interface, but is not to the midsubstance region of human anterior cruciate ligament (ACL). {yields} Mechanical stretch induces higher increase of CCN2/CTGF gene expression and protein secretion in ACL interface cells compared with ACL midsubstance cells. {yields} CCN2/CTGF treatment stimulates the proliferation of ACL interface cells. -- Abstract: Anterior cruciate ligament (ACL)-to-bone interface serves to minimize the stress concentrations that would arise between two different tissues. Mechanical stretch plays an important role in maintaining cell-specific features by inducing CCN family 2/connective tissue growth factor (CCN2/CTGF). We previously reported that cyclic tensile strain (CTS) stimulates {alpha}1(I) collagen (COL1A1) expression in human ACL-derived cells. However, the biological function and stress-related response of CCN2/CTGF were still unclear in ACL fibroblasts. In the present study, CCN2/CTGF was observed in ACL-to-bone interface, but was not in the midsubstance region by immunohistochemical analyses. CTS treatments induced higher increase of CCN2/CTGF expression and secretion in interface cells compared with midsubstance cells. COL1A1 expression was not influenced by CCN2/CTGF treatment in interface cells despite CCN2/CTGF stimulated COL1A1 expression in midsubstance cells. However, CCN2/CTGF stimulated the proliferation of interface cells. Our results suggest that distinct biological function of stretch-induced CCN2/CTGF might regulate region-specific phenotypes of ACL-derived cells.

  16. A Lactobacillus rhamnosus strain induces a heme oxygenase dependent increase in Foxp3+ regulatory T cells.

    Directory of Open Access Journals (Sweden)

    Khalil Karimi

    Full Text Available We investigated the consequences of feeding with a Lactobacillus species on the immune environment in GALT, and the role of dendritic cells and heme oxygenase-1 in mediating these responses. Feeding with a specific strain of Lactobacillus rhamnosus induced a significant increase in CD4+CD25+Foxp3+ functional regulatory T cells in GALT. This increase was greatest in the mesenteric lymph nodes and associated with a marked decrease in TNF and IFNγ production. Dendritic cell regulatory function and HO-1 expression was also increased. The increase in Foxp3+ T cells could be prevented by treatment with a heme oxygenase inhibitor. However, neither inhibition of heme oxygenase nor blockade of IL-10 and TGFβ prevented the inhibition of inflammatory cytokine production. In conclusion Lactobacillus feeding induced a tolerogenic environment in GALT. HO-1 was critical to the enhancement of Foxp3+ regulatory T cells while additional, as yet unknown, pathways were involved in the down-regulation of inflammatory cytokine production by T cells.

  17. Epalrestat increases glutathione, thioredoxin, and heme oxygenase-1 by stimulating Nrf2 pathway in endothelial cells

    Directory of Open Access Journals (Sweden)

    Kaori Yama

    2015-04-01

    Full Text Available Epalrestat (EPS is the only aldose reductase inhibitor that is currently available for the treatment of diabetic neuropathy. Recently, we found that EPS at near-plasma concentration increases the intracellular levels of glutathione (GSH in rat Schwann cells. GSH plays a crucial role in protecting endothelial cells from oxidative stress, thereby preventing vascular diseases. Here we show that EPS increases GSH levels in not only Schwann cells but also endothelial cells. Treatment of bovine aortic endothelial cells (BAECs, an in vitro model of the vascular endothelium, with EPS caused a dramatic increase in intracellular GSH levels. This was concomitant with the up-regulation of glutamate cysteine ligase, an enzyme catalyzing the first and rate-limiting step in de novo GSH synthesis. Moreover, EPS stimulated the expression of thioredoxin and heme oxygenase-1, which have important redox regulatory functions in endothelial cells. Nuclear factor erythroid 2-related factor 2 (Nrf2 is a key transcription factor that regulates the expression of antioxidant genes. EPS increased nuclear Nrf2 levels in BAECs. Nrf2 knockdown by siRNA suppressed the EPS-induced glutamate cysteine ligase, thioredoxin-1, and heme oxygenase-1 expression. Interestingly, LY294002, an inhibitor of phosphatidylinositol 3-kinase, abolished the EPS-stimulated GSH synthesis, suggesting that the kinase is associated with Nrf2 activation induced by EPS. Furthermore, EPS reduced the cytotoxicity induced by H2O2 and tert-butylhydroperoxide, indicating that EPS plays a role in protecting cells from oxidative stress. Taken together, the results provide evidence that EPS exerts new beneficial effects on endothelial cells by increasing GSH, thioredoxin, and heme oxygenase-1 levels through the activation of Nrf2. We suggest that EPS has the potential to prevent several vascular diseases caused by oxidative stress.

  18. Pb exposure attenuates hypersensitivity in vivo by increasing regulatory T cells

    Energy Technology Data Exchange (ETDEWEB)

    Fang, Liang [Department of Immunology, Fourth Military Medical University, Xi' an 710032 (China); Zhao, Fang; Shen, Xuefeng [Department of Occupational and Environmental Health and the Ministry of Education Key Lab of Hazard Assessment and Control in Special Operational Environment, School of Public Health, Fourth Military Medical University, Xi' an 710032 (China); Ouyang, Weiming [Laboratory of Cell Biology, Division of Monoclonal Antibodies, Office of Biotechnology Products, Center for Drug Evaluation and Research, United States Food and Drug Administration, Bethesda, MD (United States); Liu, Xinqin; Xu, Yan; Yu, Tao [Department of Occupational and Environmental Health and the Ministry of Education Key Lab of Hazard Assessment and Control in Special Operational Environment, School of Public Health, Fourth Military Medical University, Xi' an 710032 (China); Jin, Boquan [Department of Immunology, Fourth Military Medical University, Xi' an 710032 (China); Chen, Jingyuan, E-mail: jy_chen@fmmu.edu.cn [Department of Occupational and Environmental Health and the Ministry of Education Key Lab of Hazard Assessment and Control in Special Operational Environment, School of Public Health, Fourth Military Medical University, Xi' an 710032 (China); Luo, Wenjing, E-mail: luowenj@fmmu.edu.cn [Department of Occupational and Environmental Health and the Ministry of Education Key Lab of Hazard Assessment and Control in Special Operational Environment, School of Public Health, Fourth Military Medical University, Xi' an 710032 (China)

    2012-12-01

    Pb is a common environmental pollutant affecting various organs. Exposure of the immune system to Pb leads to immunosuppression or immunodysregulation. Although previous studies showed that Pb exposure can modulate the function of helper T cells, Pb immunotoxicity remains incompletely understood. In this study, we investigated the effect of Pb exposure on T cell development, and the underlying mechanism of Pb-induced suppression of the delayed-type hypersensitivity (DTH) response in vivo. Sprague–Dawley rats were exposed to 300 ppm Pb-acetate solution via the drinking water for six weeks, and we found that Pb exposure significantly increased Pb concentrations in the blood by 4.2-fold (p < 0.05) as compared to those in the control rats. In Pb-exposed rats, the amount of thymic CD4{sup +}CD8{sup −} and peripheral CD4{sup +} T cells was significantly reduced, whereas, CD8{sup +} population was not affected. In contrast to conventional CD4{sup +} T cells, Foxp3{sup +} regulatory T cells (Tregs) were increased in both the thymus and peripheral lymphoid organs of Pb-exposed rats. In line with the increase of Tregs, the DTH response of Pb-exposed rats was markedly suppressed. Depletion of Tregs reversed the suppression of DTH response by Pb-exposed CD4{sup +} T cells in an adoptive transfer model, suggesting a critical role of the increased Tregs in suppressing the DTH response. Collectively, this study revealed that Pb-exposure may upregulate Tregs, thereby leading to immunosuppression. -- Highlights: ► Pb exposure impaired CD4{sup +} thymic T cell development. ► Peripheral T lymphocytes were reduced following Pb exposure. ► Pb exposure increases thymic and peripheral Treg cells in rats. ► Tregs played a critical role in Pb-exposure-induced immune suppression.

  19. Pb exposure attenuates hypersensitivity in vivo by increasing regulatory T cells

    International Nuclear Information System (INIS)

    Fang, Liang; Zhao, Fang; Shen, Xuefeng; Ouyang, Weiming; Liu, Xinqin; Xu, Yan; Yu, Tao; Jin, Boquan; Chen, Jingyuan; Luo, Wenjing

    2012-01-01

    Pb is a common environmental pollutant affecting various organs. Exposure of the immune system to Pb leads to immunosuppression or immunodysregulation. Although previous studies showed that Pb exposure can modulate the function of helper T cells, Pb immunotoxicity remains incompletely understood. In this study, we investigated the effect of Pb exposure on T cell development, and the underlying mechanism of Pb-induced suppression of the delayed-type hypersensitivity (DTH) response in vivo. Sprague–Dawley rats were exposed to 300 ppm Pb-acetate solution via the drinking water for six weeks, and we found that Pb exposure significantly increased Pb concentrations in the blood by 4.2-fold (p + CD8 − and peripheral CD4 + T cells was significantly reduced, whereas, CD8 + population was not affected. In contrast to conventional CD4 + T cells, Foxp3 + regulatory T cells (Tregs) were increased in both the thymus and peripheral lymphoid organs of Pb-exposed rats. In line with the increase of Tregs, the DTH response of Pb-exposed rats was markedly suppressed. Depletion of Tregs reversed the suppression of DTH response by Pb-exposed CD4 + T cells in an adoptive transfer model, suggesting a critical role of the increased Tregs in suppressing the DTH response. Collectively, this study revealed that Pb-exposure may upregulate Tregs, thereby leading to immunosuppression. -- Highlights: ► Pb exposure impaired CD4 + thymic T cell development. ► Peripheral T lymphocytes were reduced following Pb exposure. ► Pb exposure increases thymic and peripheral Treg cells in rats. ► Tregs played a critical role in Pb-exposure-induced immune suppression.

  20. Chemotherapy and Stem Cell Transplantation Increase p16INK4a Expression, a Biomarker of T-cell Aging

    Directory of Open Access Journals (Sweden)

    William A. Wood

    2016-09-01

    Full Text Available The expression of markers of cellular senescence increases exponentially in multiple tissues with aging. Age-related physiological changes may contribute to adverse outcomes in cancer survivors. To investigate the impact of high dose chemotherapy and stem cell transplantation on senescence markers in vivo, we collected blood and clinical data from a cohort of 63 patients undergoing hematopoietic cell transplantation. The expression of p16INK4a, a well-established senescence marker, was determined in T-cells before and 6 months after transplant. RNA sequencing was performed on paired samples from 8 patients pre- and post-cancer therapy. In patients undergoing allogeneic transplant, higher pre-transplant p16INK4a expression was associated with a greater number of prior cycles of chemotherapy received (p = 0.003, prior autologous transplantation (p = 0.01 and prior exposure to alkylating agents (p = 0.01. Transplantation was associated with a marked increase in p16INK4a expression 6 months following transplantation. Patients receiving autologous transplant experienced a larger increase in p16INK4a expression (3.1-fold increase, p = 0.002 than allogeneic transplant recipients (1.9-fold increase, p = 0.0004. RNA sequencing of T-cells pre- and post- autologous transplant or cytotoxic chemotherapy demonstrated increased expression of transcripts associated with cellular senescence and physiological aging. Cytotoxic chemotherapy, especially alkylating agents, and stem cell transplantation strongly accelerate expression of a biomarker of molecular aging in T-cells.

  1. BGLAP is expressed in pancreatic cancer cells and increases their growth and invasion

    Directory of Open Access Journals (Sweden)

    Michalski Christoph W

    2007-12-01

    Full Text Available Abstract Background Bone gamma-carboxyglutamate protein (BGLAP; osteocalcin is a small, highly conserved molecule first identified in the mineralized matrix of bone. It has been implicated in the pathophysiology of various malignancies. In this study, we analyzed the expression and role of BGLAP in the normal human pancreas, chronic pancreatitis (CP, and pancreatic ductal adenocarcinoma (PDAC using quantitative RT-PCR, immunohistochemistry, immunocytochemistry and enzyme immunoassays, as well as cell proliferation and invasion assays. Gene silencing was carried out using specific siRNA molecules. Results Compared to the normal pancreas, BGLAP mRNA and protein levels were not significantly different in CP and PDAC tissues. BGLAP was faintly present in the cytoplasm of normal acinar cells but was strongly expressed in the cytoplasm and nuclei of tubular complexes and PanIN lesions of CP and PDAC tissues. Furthermore, BGLAP expression was found in the cancer cells in PDAC tissues as well as in 4 cultured pancreatic cancer cell lines. TNFalpha reduced BGLAP mRNA and protein expression levels in pancreatic cancer cell lines. In addition, BGLAP silencing led to reduction of both cell growth and invasion in those cells. Conclusion BGLAP is expressed in pancreatic cancer cells, where it potentially increases pancreatic cancer cell growth and invasion through autocrine and/or paracrine mechanisms.

  2. Mesenchymal stem cells enhance autophagy and increase β-amyloid clearance in Alzheimer disease models

    Science.gov (United States)

    Shin, Jin Young; Park, Hyun Jung; Kim, Ha Na; Oh, Se Hee; Bae, Jae-Sung; Ha, Hee-Jin; Lee, Phil Hyu

    2014-01-01

    Current evidence suggests a central role for autophagy in Alzheimer disease (AD), and dysfunction in the autophagic system may lead to amyloid-β (Aβ) accumulation. Using in vitro and in vivo AD models, the present study investigated whether mesenchymal stem cells (MSCs) could enhance autophagy and thus exert a neuroprotective effect through modulation of Aβ clearance In Aβ-treated neuronal cells, MSCs increased cellular viability and enhanced LC3-II expression compared with cells treated with Aβ only. Immunofluorescence revealed that MSC coculture in Aβ-treated neuronal cells increased the number of LC3-II-positive autophagosomes that were colocalized with a lysosomal marker. Ultrastructural analysis revealed that most autophagic vacuoles (AVs) in Aβ-treated cells were not fused with lysosomes, whereas a large portion of autophagosomes were conjoined with lysosomes in MSCs cocultured with Aβ-treated neuronal cells. Furthermore, MSC coculture markedly increased Aβ immunoreactivity colocalized within lysosomes and decreased intracellular Aβ levels compared with Aβ-treated cells. In Aβ-treated animals, MSC administration significantly increased autophagosome induction, final maturation of late AVs, and fusion with lysosomes. Moreover, MSC administration significantly reduced the level of Aβ in the hippocampus, which was elevated in Aβ-treated mice, concomitant with increased survival of hippocampal neurons. Finally, MSC coculture upregulated BECN1/Beclin 1 expression in AD models. These results suggest that MSCs significantly enhance autolysosome formation and clearance of Aβ in AD models, which may lead to increased neuronal survival against Aβ toxicity. Modulation of the autophagy pathway to repair the damaged AD brain using MSCs would have a significant impact on future strategies for AD treatment. PMID:24149893

  3. Menstrual endometrial cells from women with endometriosis demonstrate increased adherence to peritoneal cells and increased expression of CD44 splice variants.

    Science.gov (United States)

    Griffith, Jason S; Liu, Ya-Guang; Tekmal, Rajeshwar R; Binkley, Peter A; Holden, Alan E C; Schenken, Robert S

    2010-04-01

    We previously demonstrated that adherence of endometrial epithelial (EECs) and stromal cells (ESCs) to peritoneal mesothelial cells (PMCs) is partly regulated by ESC/EEC CD44 interactions with PMC associated hyaluronan. CD44, a transmembrane glycoprotein and major ligand for hyaluronan, has numerous splice variants which may impact hyaluronan binding. Here, we assessed whether ESCs and EECs from women with endometriosis demonstrate increased adherence to PMCs and examined CD44 splice variants' potential role in this process. In vitro study. Academic medical center. Fertility patients with and without endometriosis. Menstrual endometrium was collected from women with and without endometriosis confirmed surgically. The adherence of ESC/EECs to PMCs was measured. The ESC/EEC CD44 splice variants were assessed using dot-blot analysis. The ESCs and EECs from women with endometriosis demonstrated increased adherence to PMCs. The predominant CD44 splice variants expressed by ESCs and EECs from women with and without endometriosis were v3, v6, v7, v8, v9, and v10. The ESCs and EECs from women with endometriosis were more likely to express v6, v7, v8, and v9. Increased eutopic endometrial-PMC adherence and CD44 splice variant expression may contribute to the histogenesis of endometriotic lesions. Elucidation of factors controlling this expression may lead to novel endometriosis therapies. Copyright 2010 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.

  4. 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.

  5. Electrical stimulation of schwann cells promotes sustained increases in neurite outgrowth.

    Science.gov (United States)

    Koppes, Abigail N; Nordberg, Andrea L; Paolillo, Gina M; Goodsell, Nicole M; Darwish, Haley A; Zhang, Linxia; Thompson, Deanna M

    2014-02-01

    Endogenous electric fields are instructive during embryogenesis by acting to direct cell migration, and postnatally, they can promote axonal growth after injury (McCaig 1991, Al-Majed 2000). However, the mechanisms for these changes are not well understood. Application of an appropriate electrical stimulus may increase the rate and success of nerve repair by directly promoting axonal growth. Previously, DC electrical stimulation at 50 mV/mm (1 mA, 8 h duration) was shown to promote neurite outgrowth and a more pronounced effect was observed if both peripheral glia (Schwann cells) and neurons were co-stimulated. If electrical stimulation is delivered to an injury site, both the neurons and all resident non-neuronal cells [e.g., Schwann cells, endothelial cells, fibroblasts] will be treated and this biophysical stimuli can influence axonal growth directly or indirectly via changes to the resident, non-neuronal cells. In this work, non-neuronal cells were electrically stimulated, and changes in morphology and neuro-supportive cells were evaluated. Schwann cell response (morphology and orientation) was examined after an 8 h stimulation over a range of DC fields (0-200 mV/mm, DC 1 mA), and changes in orientation were observed. Electrically prestimulating Schwann cells (50 mV/mm) promoted 30% more neurite outgrowth relative to co-stimulating both Schwann cells with neurons, suggesting that electrical stimulation modifies Schwann cell phenotype. Conditioned medium from the electrically prestimulated Schwann cells promoted a 20% increase in total neurite outgrowth and was sustained for 72 h poststimulation. An 11-fold increase in nerve growth factor but not brain-derived neurotrophic factor or glial-derived growth factor was found in the electrically prestimulated Schwann cell-conditioned medium. No significant changes in fibroblast or endothelial morphology and neuro-supportive behavior were observed poststimulation. Electrical stimulation is widely used in

  6. Sugars increase non-heme iron bioavailability in human epithelial intestinal and liver cells.

    Directory of Open Access Journals (Sweden)

    Tatiana Christides

    Full Text Available Previous studies have suggested that sugars enhance iron bioavailability, possibly through either chelation or altering the oxidation state of the metal, however, results have been inconclusive. Sugar intake in the last 20 years has increased dramatically, and iron status disorders are significant public health problems worldwide; therefore understanding the nutritional implications of iron-sugar interactions is particularly relevant. In this study we measured the effects of sugars on non-heme iron bioavailability in human intestinal Caco-2 cells and HepG2 hepatoma cells using ferritin formation as a surrogate marker for iron uptake. The effect of sugars on iron oxidation state was examined by measuring ferrous iron formation in different sugar-iron solutions with a ferrozine-based assay. Fructose significantly increased iron-induced ferritin formation in both Caco-2 and HepG2 cells. In addition, high-fructose corn syrup (HFCS-55 increased Caco-2 cell iron-induced ferritin; these effects were negated by the addition of either tannic acid or phytic acid. Fructose combined with FeCl3 increased ferrozine-chelatable ferrous iron levels by approximately 300%. In conclusion, fructose increases iron bioavailability in human intestinal Caco-2 and HepG2 cells. Given the large amount of simple and rapidly digestible sugars in the modern diet their effects on iron bioavailability may have important patho-physiological consequences. Further studies are warranted to characterize these interactions.

  7. IDH1R132H in Neural Stem Cells: Differentiation Impaired by Increased Apoptosis.

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    Kamila Rosiak

    Full Text Available The high frequency of mutations in the isocitrate dehydrogenase 1 (IDH1 gene in diffuse gliomas indicates its importance in the process of gliomagenesis. These mutations result in loss of the normal function and acquisition of the neomorphic activity converting α-ketoglutarate to 2-hydroxyglutarate. This potential oncometabolite may induce the epigenetic changes, resulting in the deregulated expression of numerous genes, including those related to the differentiation process or cell survivability.Neural stem cells were derived from human induced pluripotent stem cells following embryoid body formation. Neural stem cells transduced with mutant IDH1R132H, empty vector, non-transduced and overexpressing IDH1WT controls were differentiated into astrocytes and neurons in culture. The neuronal and astrocytic differentiation was determined by morphology and expression of lineage specific markers (MAP2, Synapsin I and GFAP as determined by real-time PCR and immunocytochemical staining. Apoptosis was evaluated by real-time observation of Caspase-3 activation and measurement of PARP cleavage by Western Blot.Compared with control groups, cells expressing IDH1R132H retained an undifferentiated state and lacked morphological changes following stimulated differentiation. The significant inhibitory effect of IDH1R132H on neuronal and astrocytic differentiation was confirmed by immunocytochemical staining for markers of neural stem cells. Additionally, real-time PCR indicated suppressed expression of lineage markers. High percentage of apoptotic cells was detected within IDH1R132H-positive neural stem cells population and their derivatives, if compared to normal neural stem cells and their derivatives. The analysis of PARP and Caspase-3 activity confirmed apoptosis sensitivity in mutant protein-expressing neural cells.Our study demonstrates that expression of IDH1R132H increases apoptosis susceptibility of neural stem cells and their derivatives. Robust

  8. Preferential retrotransposition in aging yeast mother cells is correlated with increased genome instability.

    Science.gov (United States)

    Patterson, Melissa N; Scannapieco, Alison E; Au, Pak Ho; Dorsey, Savanna; Royer, Catherine A; Maxwell, Patrick H

    2015-10-01

    Retrotransposon expression or mobility is increased with age in multiple species and could promote genome instability or altered gene expression during aging. However, it is unclear whether activation of retrotransposons during aging is an indirect result of global changes in chromatin and gene regulation or a result of retrotransposon-specific mechanisms. Retromobility of a marked chromosomal Ty1 retrotransposon in Saccharomyces cerevisiae was elevated in mother cells relative to their daughter cells, as determined by magnetic cell sorting of mothers and daughters. Retromobility frequencies in aging mother cells were significantly higher than those predicted by cell age and the rate of mobility in young populations, beginning when mother cells were only several generations old. New Ty1 insertions in aging mothers were more strongly correlated with gross chromosome rearrangements than in young cells and were more often at non-preferred target sites. Mother cells were more likely to have high concentrations and bright foci of Ty1 Gag-GFP than their daughter cells. Levels of extrachromosomal Ty1 cDNA were also significantly higher in aged mother cell populations than their daughter cell populations. These observations are consistent with a retrotransposon-specific mechanism that causes retrotransposition to occur preferentially in yeast mother cells as they begin to age, as opposed to activation by phenotypic changes associated with very old age. These findings will likely be relevant for understanding retrotransposons and aging in many organisms, based on similarities in regulation and consequences of retrotransposition in diverse species. Copyright © 2015 Elsevier B.V. All rights reserved.

  9. Down-regulation of lipoprotein lipase increases glucose uptake in L6 muscle cells

    Energy Technology Data Exchange (ETDEWEB)

    Lopez, Veronica; Saraff, Kumuda [Department of Chemistry and Biochemistry, California State University Northridge, Northridge, CA 91330-8262 (United States); Medh, Jheem D., E-mail: jheem.medh@csun.edu [Department of Chemistry and Biochemistry, California State University Northridge, Northridge, CA 91330-8262 (United States)

    2009-11-06

    Thiazolidinediones (TZDs) are synthetic hypoglycemic agents used to treat type 2 diabetes. TZDs target the peroxisome proliferator activated receptor-gamma (PPAR-{gamma}) and improve systemic insulin sensitivity. The contributions of specific tissues to TZD action, or the downstream effects of PPAR-{gamma} activation, are not very clear. We have used a rat skeletal muscle cell line (L6 cells) to demonstrate that TZDs directly target PPAR-{gamma} in muscle cells. TZD treatment resulted in a significant repression of lipoprotein lipase (LPL) expression in L6 cells. This repression correlated with an increase in glucose uptake. Down-regulation of LPL message and protein levels using siRNA resulted in a similar increase in insulin-dependent glucose uptake. Thus, LPL down-regulation improved insulin sensitivity independent of TZDs. This finding provides a novel method for the management of insulin resistance.

  10. Increased numbers of spleen colony forming units in B cell deficient CBA/N mice

    International Nuclear Information System (INIS)

    Wiktor-Jedrzejczak, W.; Krupienicz, A.; Scher, I.

    1986-01-01

    The formation of exogenous and endogenous spleen colonies was studied in immune-defective mice expressing the CBA/N X-linked xid gene. Bone marrow and spleen cells of immune deficient mice formed increased numbers of eight-day exogenous spleen colonies when transferred to either normal or B cell deficient lethally irradiated recipients. Moreover, defective mice showed increased formation of five-day endogenous spleen colonies (derived from transient endogenous colony forming units; T-CFU) and of ten-day endogenous spleen colonies (derived from CFU-S). Among the possible mechanisms responsible for the observed effects, the most probable appears the one in which decreased numbers of B cell precursors stimulate stem cell pools through a feedback mechanism. (orig.) [de

  11. TIMP-1 gene deficiency increases tumour cell sensitivity to chemotherapy-induced apoptosis

    DEFF Research Database (Denmark)

    Davidsen, Marie Louise; Würts, S.Ø.; Rømer, Maria Unni Koefoed

    2006-01-01

    deficiency increases the response to chemotherapy considerably, confirming that TIMP-1 protects the cells from apoptosis. This is to our knowledge the first study investigating TIMP-1 and chemotherapy-induced apoptosis employing a powerful model system comprising TIMP-1 gene-deficient cells...... this hypothesis, we have established TIMP-1 gene-deficient and TIMP-1 wild-type fibrosarcoma cells from mouse lung tissue. We have characterised these cells with regard to TIMP-1 genotype, TIMP-1 expression, malignant transformation and sensitivity to chemotherapy-induced apoptosis. We show that TIMP-1 gene...... and their genetically identical wild-type controls. For future studies, this cell system can be used to uncover the mechanisms and signalling pathways involved in the TIMP-1-mediated inhibition of apoptosis as well as to investigate the possibility of using TIMP-1 inhibitors to optimise the effect of conventional...

  12. Increased accumulation of dendritic cells in celiac disease associates with increased expression of autophagy protein LC3

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    Paramaguru Rajaguru

    2013-01-01

    Full Text Available Background: Celiac disease (CD an immune-mediated disorder associates with accumulation of dendritic cell (DC in duodenal mucosa. Autophagy has recently been implicated in autoantigen formation. However, its role in CD is still unknown. Aim: To examine role of autophagic protein LC3 expressed by activated DC in CD. Materials and Methods : Thirty CD patients were analyzed at initial presentation and after 6 months of gluten-free diet (GFD. Duodenal biopsies were studied for histological changes and CD11c, CD86, and MAP1LC3A expressions by double immunohistochemistry (IHC. Masson′s trichrome (MT staining was used to assess basement membrane (BM thickness and Oil Red O (ORO staining for mucosal lipid deposit. Polymerase chain reaction (PCR was performed for HLA-DQ system. Statistical analysis was done using paired and unpaired t test, chi-square test, Fisher′s exact test, and McNemar-Bowker test. A P-value <0.05 was considered statistically significant. Results: HLA-DQ2 and HLA-DQ8 alleles were present in all studied patients. Increased BM thickness was observed in 63% and 73% had ORO-positive lipid in surface lining epithelium. Pre-treatment biopsies showed increased DCs expressing LC3, which were significantly less in follow-up biopsies. The follow-up biopsies had shown significant reduction in BM thickness and ORO. Conclusion : Histological improvement in duodenal biopsies was associated with reduction in activated DCs expressing autophagic protein, which probably play important role in pathogenesis of an autoimmune disorder like CD.

  13. BRCA1 deficiency increases the sensitivity of ovarian cancer cells to auranofin

    International Nuclear Information System (INIS)

    Oommen, Deepu; Yiannakis, Dennis; Jha, Awadhesh N.

    2016-01-01

    Highlights: • BRCA1 deficient cancer cells exhibit increased DNA damage upon auranofin treatment. • Auranofin induces apoptosis in BRCA1 deficient cancer cells despite the activation of Nrf2. • Antioxidant protects BRCA1 deficient cancer cells from auranofin. - Abstract: Auranofin, a thioredoxin reductase inhibitor and an anti-rheumatic drug is currently undergoing phase 2 clinical studies for repurposing to treat recurrent epithelial ovarian cancer. Previous studies have established that auranofin exerts its cytotoxic activity by increasing the production of reactive oxygen species (ROS). Breast cancer 1, early onset (BRCA1) is a DNA repair protein whose functional status is critical in the prognosis of ovarian cancer. Apart from its key role in DNA repair, BRCA1 is also known to modulate cellular redox homeostasis by regulating the stability of anti-oxidant transcription factor, nuclear factor erythroid 2—related factor 2 (Nrf2) via direct protein–protein interaction. However, it is currently unknown whether BRCA1 modulates the sensitivity of ovarian cancer cells to auranofin. Here we report that BRCA1-depleted cells exhibited increased DNA double strand breaks (DSBs) and decreased clonogenic cell survival upon auranofin treatment. Interestingly, auranofin induced the expression of Nrf2 in BRCA1-depleted cells suggesting its regulation independent of BRCA1. Furthermore, anti-oxidant agent, N-acetyl cysteine (NAC) protected BRCA1-depleted cells from DNA damage and apoptosis induced by auranofin. Our study suggests that accumulated lethal DSBs resulting from the oxidative damage render BRCA1 deficient cells more sensitive to auranofin despite the activation of Nrf2.

  14. BRCA1 deficiency increases the sensitivity of ovarian cancer cells to auranofin

    Energy Technology Data Exchange (ETDEWEB)

    Oommen, Deepu [School of Biological Sciences, Plymouth University, Plymouth PL4 8AA (United Kingdom); Yiannakis, Dennis [Plymouth Oncology Centre, Derriford Hospital, Plymouth Hospitals NHS Trust, Plymouth PL6 8DH (United Kingdom); Jha, Awadhesh N., E-mail: a.jha@plymouth.ac.uk [School of Biological Sciences, Plymouth University, Plymouth PL4 8AA (United Kingdom)

    2016-02-15

    Highlights: • BRCA1 deficient cancer cells exhibit increased DNA damage upon auranofin treatment. • Auranofin induces apoptosis in BRCA1 deficient cancer cells despite the activation of Nrf2. • Antioxidant protects BRCA1 deficient cancer cells from auranofin. - Abstract: Auranofin, a thioredoxin reductase inhibitor and an anti-rheumatic drug is currently undergoing phase 2 clinical studies for repurposing to treat recurrent epithelial ovarian cancer. Previous studies have established that auranofin exerts its cytotoxic activity by increasing the production of reactive oxygen species (ROS). Breast cancer 1, early onset (BRCA1) is a DNA repair protein whose functional status is critical in the prognosis of ovarian cancer. Apart from its key role in DNA repair, BRCA1 is also known to modulate cellular redox homeostasis by regulating the stability of anti-oxidant transcription factor, nuclear factor erythroid 2—related factor 2 (Nrf2) via direct protein–protein interaction. However, it is currently unknown whether BRCA1 modulates the sensitivity of ovarian cancer cells to auranofin. Here we report that BRCA1-depleted cells exhibited increased DNA double strand breaks (DSBs) and decreased clonogenic cell survival upon auranofin treatment. Interestingly, auranofin induced the expression of Nrf2 in BRCA1-depleted cells suggesting its regulation independent of BRCA1. Furthermore, anti-oxidant agent, N-acetyl cysteine (NAC) protected BRCA1-depleted cells from DNA damage and apoptosis induced by auranofin. Our study suggests that accumulated lethal DSBs resulting from the oxidative damage render BRCA1 deficient cells more sensitive to auranofin despite the activation of Nrf2.

  15. Increased membrane cholesterol in lymphocytes diverts T-cells toward an inflammatory response.

    Directory of Open Access Journals (Sweden)

    Jacqueline Surls

    Full Text Available Cell signaling for T-cell growth, differentiation, and apoptosis is initiated in the cholesterol-rich microdomains of the plasma membrane known as lipid rafts. Herein, we investigated whether enrichment of membrane cholesterol in lipid rafts affects antigen-specific CD4 T-helper cell functions. Enrichment of membrane cholesterol by 40-50% following squalene administration in mice was paralleled by an increased number of resting CD4 T helper cells in periphery. We also observed sensitization of the Th1 differentiation machinery through co-localization of IL-2Rα, IL-4Rα, and IL-12Rβ2 subunits with GM1 positive lipid rafts, and increased STAT-4 and STAT-5 phosphorylation following membrane cholesterol enrichment. Antigen stimulation or CD3/CD28 polyclonal stimulation of membrane cholesterol-enriched, resting CD4 T-cells followed a path of Th1 differentiation, which was more vigorous in the presence of increased IL-12 secretion by APCs enriched in membrane cholesterol. Enrichment of membrane cholesterol in antigen-specific, autoimmune Th1 cells fostered their organ-specific reactivity, as confirmed in an autoimmune mouse model for diabetes. However, membrane cholesterol enrichment in CD4(+Foxp3(+ T-reg cells did not alter their suppressogenic function. These findings revealed a differential regulatory effect of membrane cholesterol on the function of CD4 T-cell subsets. This first suggests that membrane cholesterol could be a new therapeutic target to modulate the immune functions, and second that increased membrane cholesterol in various physiopathological conditions may bias the immune system toward an inflammatory Th1 type response.

  16. Novel nootropic dipeptide Noopept increases inhibitory synaptic transmission in CA1 pyramidal cells.

    Science.gov (United States)

    Kondratenko, Rodion V; Derevyagin, Vladimir I; Skrebitsky, Vladimir G

    2010-05-31

    Effects of newly synthesized nootropic and anxiolytic dipeptide Noopept on inhibitory synaptic transmission in hippocampal CA1 pyramidal cells were investigated using patch-clamp technique in whole-cell configuration. Bath application of Noopept (1 microM) significantly increased the frequency of spike-dependant spontaneous IPSCs whereas spike-independent mIPSCs remained unchanged. It was suggested that Noopept mediates its effect due to the activation of inhibitory interneurons terminating on CA1 pyramidal cells. Results of current clamp recording of inhibitory interneurons residing in stratum radiatum confirmed this suggestion. Copyright 2010 Elsevier Ireland Ltd. All rights reserved.

  17. Suppression of Cpn10 increases mitochondrial fission and dysfunction in neuroblastoma cells.

    Directory of Open Access Journals (Sweden)

    So Jung Park

    Full Text Available To date, several regulatory proteins involved in mitochondrial dynamics have been identified. However, the precise mechanism coordinating these complex processes remains unclear. Mitochondrial chaperones regulate mitochondrial function and structure. Chaperonin 10 (Cpn10 interacts with heat shock protein 60 (HSP60 and functions as a co-chaperone. In this study, we found that down-regulation of Cpn10 highly promoted mitochondrial fragmentation in SK-N-MC and SH-SY5Y neuroblastoma cells. Both genetic and chemical inhibition of Drp1 suppressed the mitochondrial fragmentation induced by Cpn10 reduction. Reactive oxygen species (ROS generation in 3-NP-treated cells was markedly enhanced by Cpn10 knock down. Depletion of Cpn10 synergistically increased cell death in response to 3-NP treatment. Furthermore, inhibition of Drp1 recovered Cpn10-mediated mitochondrial dysfunction in 3-NP-treated cells. Moreover, an ROS scavenger suppressed cell death mediated by Cpn10 knockdown in 3-NP-treated cells. Taken together, these results showed that down-regulation of Cpn10 increased mitochondrial fragmentation and potentiated 3-NP-mediated mitochondrial dysfunction in neuroblastoma cells.

  18. MutY-Homolog (MYH) inhibition reduces pancreatic cancer cell growth and increases chemosensitivity.

    Science.gov (United States)

    Sharbeen, George; Youkhana, Janet; Mawson, Amanda; McCarroll, Joshua; Nunez, Andrea; Biankin, Andrew; Johns, Amber; Goldstein, David; Phillips, Phoebe

    2017-02-07

    Patients with pancreatic ductal adenocarcinoma (PC) have a poor prognosis due to metastases and chemoresistance. PC is characterized by extensive fibrosis, which creates a hypoxic microenvironment, and leads to increased chemoresistance and intracellular oxidative stress. Thus, proteins that protect against oxidative stress are potential therapeutic targets for PC. A key protein that maintains genomic integrity against oxidative damage is MutY-Homolog (MYH). No prior studies have investigated the function of MYH in PC cells. Using siRNA, we showed that knockdown of MYH in PC cells 1) reduced PC cell proliferation and increased apoptosis; 2) further decreased PC cell growth in the presence of oxidative stress and chemotherapy agents (gemcitabine, paclitaxel and vincristine); 3) reduced PC cell metastatic potential; and 4) decreased PC tumor growth in a subcutaneous mouse model in vivo. The results from this study suggest MYH may be a novel therapeutic target for PC that could potentially improve patient outcome by reducing PC cell survival, increasing the efficacy of existing drugs and reducing metastatic spread.

  19. Arachidonate metabolism increases as rat alveolar type II cells differentiate in vitro

    International Nuclear Information System (INIS)

    Lipchik, R.J.; Chauncey, J.B.; Paine, R.; Simon, R.H.; Peters-Golden, M.

    1990-01-01

    Rat type II alveolar epithelial cells are known to undergo morphological and functional changes when maintained in culture for several days. Having previously demonstrated that these cells can deacylate free arachidonic acid (AA) and metabolize it to products of the cyclooxygenase pathway, the present study was undertaken to determine whether in vitro differentiation was accompanied by alterations in the availability and metabolism of AA. We assessed the constitutive and ionophore A23187-induced deacylation and metabolism of endogenous AA, as well as the metabolism of exogenously supplied AA, in primary cultures of rat type II cells at days 2, 4, and 7 after isolation. Levels of free endogenous AA were increased at day 4, whereas eicosanoid synthesis, predominantly prostaglandin E2 and prostacyclin, increased markedly only at day 7. A similar time course of augmentation of prostanoid release was seen in response to exogenous AA. Type II cells cultured on fibronectin, intended to hasten cell flattening and spreading, demonstrated accelerated increases in available free AA in response to A23187; cells cultured on basement membrane derived from Engelbreth-Holm-Swarm mouse sarcoma, known to maintain the type II phenotype, exhibited diminished levels of available free AA. From these findings, we conclude that alterations in arachidonate metabolism are linked to alterations in cellular phenotype. The potentiation of eicosanoid synthesis accompanying in vitro differentiation suggests a possible role for the alveolar epithelium in the modulation of inflammation and fibrosis in the distal lung

  20. Status epilepticus increases mature granule cells in the molecular layer of the dentate gyrus in rats★

    Science.gov (United States)

    Liang, Zhaoliang; Gao, Fei; Wang, Fajun; Wang, Xiaochen; Song, Xinyu; Liu, Kejing; Zhan, Ren-Zhi

    2013-01-01

    Enhanced neurogenesis in the dentate gyrus of the hippocampus following seizure activity, especially status epilepticus, is associated with ectopic residence and aberrant integration of newborn granule cells. Hilar ectopic granule cells may be detrimental to the stability of dentate circuitry by means of their electrophysiological properties and synaptic connectivity. We hypothesized that status epilepticus also increases ectopic granule cells in the molecular layer. Status epilepticus was induced in male Sprague-Dawley rats by intraperitoneal injection of pilocarpine. Immunostaining showed that many doublecortin-positive cells were present in the molecular layer and the hilus 7 days after the induction of status epilepticus. At least 10 weeks after status epilepticus, the estimated number of cells positive for both prospero homeobox protein 1 and neuron-specific nuclear protein in the hilus was significantly increased. A similar trend was also found in the molecular layer. These findings indicate that status epilepticus can increase the numbers of mature and ectopic newborn granule cells in the molecular layer. PMID:25206705

  1. Suppression of thymosin β10 increases cell migration and metastasis of cholangiocarcinoma

    International Nuclear Information System (INIS)

    Sribenja, Sirinapa; Sawanyawisuth, Kanlayanee; Kraiklang, Ratthaphol; Wongkham, Chaisiri; Vaeteewoottacharn, Kulthida; Obchoei, Sumalee; Yao, Qizhi; Wongkham, Sopit; Chen, Changyi

    2013-01-01

    Thymosin β10 (Tβ10) expression is associated with malignant phenotypes in many cancers. However, the role and mechanisms of Tβ10 in liver fluke-associated cholangiocarcinoma (CCA) are not fully understood. In this study, we investigated the expression of Tβ10 in CCA tumor tissues and cell lines as well as molecular mechanisms of Tβ10 in tumor metastasis of CCA cell lines. Tβ10 expression was determined by real time RT-PCR or immunocytochemistry. Tβ10 silence or overexpression in CCA cells was achieved using gene delivery techniques. Cell migration was assessed using modified Boyden chamber and wound healing assay. The effect of silencing Tβ10 on CCA tumor metastasis was determined in nude mice. Phosphorylation of ERK1/2 and the expression of EGR1, Snail and matrix metalloproteinases (MMPs) were studied. Ten pairs of CCA tissues (primary and metastatic tumors) and 5 CCA cell lines were studied. With real time RT-PCR and immunostaining analysis, Tβ10 was highly expressed in primary tumors of CCA; while it was relatively low in the metastatic tumors. Five CCA cell lines showed differential expression levels of Tβ10. Silence of Tβ10 significantly increased cell migration, invasion and wound healing of CCA cells in vitro; reversely, overexpression of Tβ10 reduced cell migration compared with control cells (P<0.05). In addition, silence of Tβ10 in CCA cells increased liver metastasis in a nude mouse model of CCA implantation into the spleen. Furthermore, silence of Tβ10 activated ERK1/2 and increased the expression of Snail and MMPs in CCA cell lines. Ras-GTPase inhibitor, FPT inhibitor III, effectively blocked Tβ10 silence-associated ERK1/2 activation, Snail expression and cell migration. Low expression of Tβ10 is associated with metastatic phenotype of CCA in vitro and in vivo, which may be mediated by the activation of Ras, ERK1/2 and upregulation of Snail and MMPs. This study suggests a new molecular pathway of CCA pathogenesis and a novel strategy to

  2. Overendocytosis of gold nanoparticles increases autophagy and apoptosis in hypoxic human renal proximal tubular cells

    Directory of Open Access Journals (Sweden)

    Ding F

    2014-09-01

    Full Text Available Fengan Ding,1 Yiping Li,1 Jing Liu,1 Lei Liu,1 Wenmin Yu,1 Zhi Wang,1 Haifeng Ni,2 Bicheng Liu,2 Pingsheng Chen1,2 1School of Medicine, Southeast University, Nanjing, People’s Republic of China; 2Institute of Nephrology, The Affiliated Zhongda Hospital, Southeast University, Nanjing, People’s Republic of China Background: Gold nanoparticles (GNPs can potentially be used in biomedical fields ranging from therapeutics to diagnostics, and their use will result in increased human exposure. Many studies have demonstrated that GNPs can be deposited in the kidneys, particularly in renal tubular epithelial cells. Chronic hypoxic is inevitable in chronic kidney diseases, and it results in renal tubular epithelial cells that are susceptible to different types of injuries. However, the understanding of the interactions between GNPs and hypoxic renal tubular epithelial cells is still rudimentary. In the present study, we characterized the cytotoxic effects of GNPs in hypoxic renal tubular epithelial cells.Results: Both 5 nm and 13 nm GNPs were synthesized and characterized using various biophysical methods, including transmission electron microscopy, dynamic light scattering, and ultraviolet–visible spectrophotometry. We detected the cytotoxicity of 5 and 13 nm GNPs (0, 1, 25, and 50 nM to human renal proximal tubular cells (HK-2 by Cell Counting Kit-8 assay and lactate dehydrogenase release assay, but we just found the toxic effect in the 5 nm GNP-treated cells at 50 nM dose under hypoxic condition. Furthermore, the transmission electron microscopy images revealed that GNPs were either localized in vesicles or free in the lysosomes in 5 nm GNPs-treated HK-2 cells, and the cellular uptake of the GNPs in the hypoxic cells was significantly higher than that in normoxic cells. In normoxic HK-2 cells, 5 nm GNPs (50 nM treatment could cause autophagy and cell survival. However, in hypoxic conditions, the GNP exposure at the same condition led to the

  3. Human embryonic stem cell derived islet progenitors mature inside an encapsulation device without evidence of increased biomass or cell escape.

    Science.gov (United States)

    Kirk, Kaitlyn; Hao, Ergeng; Lahmy, Reyhaneh; Itkin-Ansari, Pamela

    2014-05-01

    There are several challenges to successful implementation of a cell therapy for insulin dependent diabetes derived from human embryonic stem cells (hESC). Among these are development of functional insulin producing cells, a clinical delivery method that eliminates the need for chronic immunosuppression, and assurance that hESC derived tumors do not form in the patient. We and others have shown that encapsulation of cells in a bilaminar device (TheraCyte) provides immunoprotection in rodents and primates. Here we monitored human insulin secretion and employed bioluminescent imaging (BLI) to evaluate the maturation, growth, and containment of encapsulated islet progenitors derived from CyT49 hESC, transplanted into mice. Human insulin was detectable by 7 weeks post-transplant and increased 17-fold over the course of 8 weeks, yet during this period the biomass of encapsulated cells remained constant. Remarkably, by 20 weeks post-transplant encapsulated cells secreted sufficient levels of human insulin to ameliorate alloxan induced diabetes. Further, bioluminescent imaging revealed for the first time that hESCs remained fully contained in encapsulation devices for up to 150 days, the longest period tested. Collectively, the data suggest that encapsulated hESC derived islet progenitors hold great promise as an effective and safe cell replacement therapy for insulin dependent diabetes. Copyright © 2014. Published by Elsevier B.V.

  4. Sonic hedgehog signaling regulates mode of cell division of early cerebral cortex progenitors and increases astrogliogenesis

    Directory of Open Access Journals (Sweden)

    Geissy LL Araújo

    2014-03-01

    Full Text Available The morphogen Sonic Hedgehog (SHH plays a critical role in the development of different tissues. In the central nervous system, SHH is well known to contribute to the patterning of the spinal cord and separation of the brain hemispheres. In addition, it has recently been shown that SHH signaling also contributes to the patterning of the telencephalon and establishment of adult neurogenic niches. In this work, we investigated whether SHH signaling influences the behavior of neural progenitors isolated from the dorsal telencephalon, which generate excitatory neurons and macroglial cells in vitro. We observed that SHH increases proliferation of cortical progenitors and generation of astrocytes, whereas blocking SHH signaling with cyclopamine has opposite effects. In both cases, generation of neurons did not seem to be affected. However, cell survival was broadly affected by blockade of SHH signaling. SHH effects were related to three different cell phenomena: mode of cell division, cell cycle length and cell growth. Together, our data in vitro demonstrate that SHH signaling controls cell behaviors that are important for proliferation of cerebral cortex progenitors, as well as differentiation and survival of neurons and astroglial cells.

  5. MicroRNA-21 Increases Proliferation and Cisplatin Sensitivity of Osteosarcoma-Derived Cells.

    Directory of Open Access Journals (Sweden)

    Vanita Vanas

    Full Text Available Osteosarcoma is the most common primary bone tumor and poor prognosis for osteosarcoma patients is mainly due to chemotherapy resistance. MicroRNAs are important to maintain pathophysiological mechanisms of cancer and influence cell sensitivity to chemotherapy. In this study, we tested the functions of microRNA-21 for malignant features as well as for drug resistance of osteosarcoma. We used Northern blot to measure microRNA-21 levels in osteosarcoma-derived cell lines. MicroRNA-21 activity was modulated by either expressing a sponge to decrease its activity in an osteosarcoma-derived cell line expressing high levels of microRNA-21 or by introducing pri-microRNA-21 in a cell line with low endogenous levels. Cell migration was determined in a scratch assay and cell proliferation was measured by performing growth curve analysis. Sensitivity of the cells towards chemotherapeutics was investigated by performing cell viability assays and calculating the IC50 values. While cell migration was unaffected by modulated microRNA-21 levels, microRNA-21 inhibition slowed proliferation and exogenously expressed microRNA-21 promoted this process. Modulated microRNA-21 activity failed to effect sensitivity of osteosarcoma-derived cell lines to doxorubicin or methotrexate. Contrarily, reduction of microRNA-21 activity resulted in enhanced resistance towards cisplatin while ectopic expression of microRNA-21 showed the opposite effect. Increased microRNA-21 levels repressed the expression of Sprouty2 and ectopic expression of Sprouty2 was able to largely rescue the observed effects of microRNA-21 in osteosarcoma. In summary, our data indicate that in osteosarcoma microRNA-21 expression is an important component for regulation of cell proliferation and for determining sensitivity to cisplatin.

  6. Simultaneous Increases in Proliferation and Apoptosis of Vascular Smooth Muscle Cells Accelerate Diabetic Mouse Venous Atherosclerosis

    Science.gov (United States)

    Liu, Shuying; Zhang, Zhengyu; Wang, Jingjing; Zhou, Yuhuan; Liu, Kefeng; Huang, Jintao; Chen, Dadi; Wang, Junmei; Li, Chaohong

    2015-01-01

    Aims This study was designed to demonstrate simultaneous increases in proliferation and apoptosis of vascular smooth muscle cells (VSMCs) leading to accelerated vein graft remodeling and to explore the underlying mechanisms. Methods Vein grafts were performed in non-diabetic and diabetic mice. The cultured quiescent VSMCs were subjected to mechanical stretch stress (SS) and/or advanced glycosylation end products (AGEs). Harvested vein grafts and treated VSMCs were used to detect cell proliferation, apoptosis, mitogen-activated protein kinases (MAPKs) activation and SM-α-actin expression. Results Significantly thicker vessel walls and greater increases in proliferation and apoptosis were observed in diabetic vein grafts than those in non-diabetic. Both SS and AGEs were found to induce different activation of three members of MAPKs and simultaneous increases in proliferation and apoptosis of VSMCs, and combined treatment with both had a synergistic effect. VSMCs with strong SM-α-actin expression represented more activated JNKs or p38MAPK, and cell apoptosis, while the cells with weak SM-α-actin expression demonstrated preferential activation of ERKs and cell proliferation. In contrast, inhibition of MAPKs signals triggered significant decreases in VSMC proliferation, and apoptosis. Treatment of the cells with RNA interference of receptor of AGEs (RAGE) also resulted in significant decreases in both proliferation and apoptosis. Conclusions Increased pressure-induced SS triggers simultaneous increases in proliferation and apoptosis of VSMCs in the vein grafts leading to vein arterializations, which can be synergistically accelerated by high glucose-induced AGEs resulting in vein graft atherosclerosis. Either SS or AGEs and their combination induce simultaneous increases in proliferation and apoptosis of VSMCs via different activation of three members of MAPKs resulting from different VSMC subtypes classified by SM-α-actin expression levels. PMID:26488175

  7. Amiodarone increases the accumulation of DEA in a human alveolar epithelium-derived cell line.

    Science.gov (United States)

    Seki, Satoru; Itagaki, Shirou; Kobayashi, Masaki; Hirano, Takeshi; Iseki, Ken

    2008-07-01

    Amiodarone (AMD)-induced pulmonary toxicity (AIPT) is the most life-threatening side-effect of AMD treatment. N-Monodesethylamiodarone (DEA), an active metabolite of AMD, also exhibits cytotoxicity and tends to accumulate in the lung more intensively than AMD. In this study, we characterized the mechanism of DEA accumulation using A549 cells as a model of the alveolar epithelium. Typical ATP-depletion compounds caused an approximately 30% increase in the accumulation of DEA in A549 cells, although these effects were less than those in Caco-2 cells. Triiodothyronine (T(3)), which exhibited an inhibitory effect on DEA efflux in Caco-2 cells, did not affect the accumulation of DEA in A549 cells. On the other hand, 100 microM AMD caused an approximately 200% increase in DEA content in A549 cells, although AMD accumulation was not affected by 100 microM DEA. Since the reducing effect of AMD on cellular ATP levels and that of FCCP were similar, the mechanism by which DEA accumulation is increased by AMD might be different from the ATP-dependent DEA efflux mechanism. The decrease in cell viability by DEA in the presence of AMD (IC(50) value of DEA for A549 cell viability: 25.4+/-2.4 microM) was more pronounced than that by DEA alone (IC(50) value: 11.5+/-3.0 microM). This further DEA accumulation by AMD might be a factor responsible for the greater accumulation of DEA than that of AMD in the lung in long-term AMD-treated patients.

  8. Increased p38-MAPK is responsible for chemotherapy resistance in human gastric cancer cells

    International Nuclear Information System (INIS)

    Guo, Xianling; Zhang, Baihe; Wu, Mengchao; Wei, Lixin; Ma, Nannan; Wang, Jin; Song, Jianrui; Bu, Xinxin; Cheng, Yue; Sun, Kai; Xiong, Haiyan; Jiang, Guocheng

    2008-01-01

    Chemoresistance is one of the main obstacles to successful cancer therapy and is frequently associated with Multidrug resistance (MDR). Many different mechanisms have been suggested to explain the development of an MDR phenotype in cancer cells. One of the most studied mechanisms is the overexpression of P-glycoprotein (P-gp), which is a product of the MDR1 gene. Tumor cells often acquire the drug-resistance phenotype due to upregulation of the MDR1 gene. Overexpression of MDR1 gene has often been reported in primary gastric adenocarcinoma. This study investigated the role of p38-MAPK signal pathway in vincristine-resistant SGC7901/VCR cells. P-gp and MDR1 RNA were detected by Western blot analysis and RT-PCR amplification. Mitgen-activated protein kinases and function of P-gp were demonstrated by Western blot and FACS Aria cytometer analysis. Ap-1 activity and cell apoptosis were detected by Dual-Luciferase Reporter Assay and annexin V-PI dual staining. The vincristine-resistant SGC7901/VCR cells with increased expression of the multidrug-resistance 1 (MDR1) gene were resistant to P-gp-related drug and P-gp-unrelated drugs. Constitutive increases of phosphorylated p38-MAPK and AP-1 activities were also found in the drug-resistant cells. Inhibition of p38-MAPK by SB202190 reduced activator protein-1 (AP-1) activity and MDR1 expression levels and increased the sensitivity of SGC7901/VCR cells to chemotherapy. Activation of the p38-MAPK pathway might be responsible for the modulation of P-glycoprotein-mediated and P-glycoprotein-unmediated multidrug resistance in the SGC7901/VCR cell line

  9. Increased metastatic potential of tumor cells in von Willebrand factor-deficient mice.

    Science.gov (United States)

    Terraube, V; Pendu, R; Baruch, D; Gebbink, M F B G; Meyer, D; Lenting, P J; Denis, C V

    2006-03-01

    The key role played by von Willebrand factor (VWF) in platelet adhesion suggests a potential implication in various pathologies, where this process is involved. In cancer metastasis development, tumor cells interact with platelets and the vessel wall to extravasate from the circulation. As a potential mediator of platelet-tumor cell interactions, VWF could influence this early step of tumor spread and therefore play a role in cancer metastasis. To investigate whether VWF is involved in metastasis development. In a first step, we characterized the interaction between murine melanoma cells B16-BL6 and VWF in vitro. In a second step, an experimental metastasis model was used to compare the formation of pulmonary metastatic foci in C57BL/6 wild-type and VWF-null mice following the injection of B16-BL6 cells or Lewis lung carcinoma cells. In vitro adhesion assays revealed that VWF is able to promote a dose-dependent adhesion of B16-BL6 cells via its Arg-Gly-Asp (RGD) sequence. In the experimental metastasis model, we found a significant increase in the number of pulmonary metastatic foci in VWF-null mice compared with the wild-type mice, a phenotype that could be corrected by restoring VWF plasma levels. We also showed that increased survival of the tumor cells in the lungs during the first 24 h in the absence of VWF was the cause of this increased metastasis. These findings suggest that VWF plays a protective role against tumor cell dissemination in vivo. Underlying mechanisms remain to be investigated.

  10. Increased reactive oxygen species levels cause ER stress and cytotoxicity in andrographolide treated colon cancer cells.

    Science.gov (United States)

    Banerjee, Aditi; Banerjee, Vivekjyoti; Czinn, Steven; Blanchard, Thomas

    2017-04-18

    Chemotherapy continues to play an essential role in the management of many cancers including colon cancer, the third leading cause of death due to cancer in the United States. Many naturally occurring plant compounds have been demonstrated to possess anti-cancer cell activity and have the potential to supplement existing chemotherapy strategies. The plant metabolite andrographolide induces cell death in cancer cells and apoptosis is dependent upon the induction of endoplasmic reticulum stress (ER stress) leading to the unfolded protein response (UPR). The goal of the present study was to determine the mechanism by which andrographolide induces ER stress and to further evaluate its role in promoting cell death pathways. The T84 and COLO 205 cancer cell lines were used to demonstrate that andrographolide induces increased ROS levels, corresponding anti-oxidant response molecules, and reduced mitochondrial membrane potential. No increases in ROS levels were detected in control colon fibroblast cells. Andrographolide-induced cell death, UPR signaling, and CHOP, Bax, and caspase 3 apoptosis elements were all inhibited in the presence of the ROS scavenger NAC. Additionally, andrographolide-induced suppression of cyclins B1 and D1 were also reversed in the presence of NAC. Finally, Akt phosphorylation and phospho-mTOR levels that are normally suppressed by andrographolide were also expressed at normal levels in the absence of ROS. These data demonstrate that andrographolide induces ER stress leading to apoptosis through the induction of ROS and that elevated ROS also play an important role in down-regulating cell cycle progression and cell survival pathways as well.

  11. Haploidentical hematopoietic SCT increases graft-versus-tumor effect against renal cell carcinoma.

    Science.gov (United States)

    Budak-Alpdogan, T; Sauter, C T; Bailey, C P; Biswas, C S; Panis, M M; Civriz, S; Flomenberg, N; Alpdogan, O

    2013-08-01

    Allogeneic hematopoietic SCT (HSCT) has been shown to be an effective treatment option for advanced renal cell cancer (RCC). However, tumor resistance/relapse remains as the main post transplant issue. Therefore, enhancing graft-versus-tumor (GVT) activity without increasing GVHD is critical for improving the outcome of HSCT. We explored the GVT effect of haploidentical-SCT (haplo-SCT) against RCC in murine models. Lethally irradiated CB6F1 (H2K(b/d)) recipients were transplanted with T-cell-depleted BM cells from B6CBAF1 (H2K(b/k)) mice. Haplo-SCT combined with a low-dose haploidentical (HI) T-cell infusion (1 × 10(5)) successfully provided GVT activity without incurring GVHD. This effect elicited murine RCC growth control and consequently displayed a comparative survival advantage of haplo-SCT recipients when compared with MHC-matched (B6D2F1CB6F1) and parent-F1 (B6CB6F1) transplant recipients. Recipients of haplo-SCT had an increase in donor-derived splenic T-cell numbers, T-cell proliferation and IFN-γ-secreting donor-derived T-cells, a critical aspect for anti-tumor activity. The splenocytes from B6CBAF1 mice had a higher cytotoxicity against RENCA cells than the splenocytes from B6 and B6D2F1 donors after tumor challenge. These findings suggest that haplo-SCT might be an innovative immunotherapeutic platform for solid tumors, particularly for renal cell carcinoma.

  12. Minocycline causes widespread cell death and increases microglial labeling in the neonatal mouse brain.

    Science.gov (United States)

    Strahan, J Alex; Walker, William H; Montgomery, Taylor R; Forger, Nancy G

    2017-06-01

    Minocycline, an antibiotic of the tetracycline family, inhibits microglia in many paradigms and is among the most commonly used tools for examining the role of microglia in physiological processes. Microglia may play an active role in triggering developmental neuronal cell death, although findings have been contradictory. To determine whether microglia influence developmental cell death, we treated perinatal mice with minocycline (45 mg/kg) and quantified effects on dying cells and microglial labeling using immunohistochemistry for activated caspase-3 (AC3) and ionized calcium-binding adapter molecule 1 (Iba1), respectively. Contrary to our expectations, minocycline treatment from embryonic day 18 to postnatal day (P)1 caused a > tenfold increase in cell death 8 h after the last injection in all brain regions examined, including the primary sensory cortex, septum, hippocampus and hypothalamus. Iba1 labeling was also increased in most regions. Similar effects, although of smaller magnitude, were seen when treatment was delayed to P3-P5. Minocycline treatment from P3 to P5 also decreased overall cell number in the septum at weaning, suggesting lasting effects of the neonatal exposure. When administered at lower doses (4.5 or 22.5 mg/kg), or at the same dose 1 week later (P10-P12), minocycline no longer increased microglial markers or cell death. Taken together, the most commonly used microglial "inhibitor" increases cell death and Iba1 labeling in the neonatal mouse brain. Minocycline is used clinically in infant and pediatric populations; caution is warrented when using minocycline in developing animals, or extrapolating the effects of this drug across ages. © 2016 Wiley Periodicals, Inc. Develop Neurobiol 77: 753-766, 2017. © 2016 Wiley Periodicals, Inc.

  13. Increased risk of ALL among premature infants is not explained by increased prevalence of pre-leukemic cell clones

    DEFF Research Database (Denmark)

    Lausten-Thomsen, Ulrik; Madsen, Hans O.; Vestergaard, Therese Risom

    2010-01-01

    in the prevalence and magnitude of preleukaemic t(12;21)-positive cells compared to previously published data from mature children could be demonstrated. This indirectly supports the theory that prevalence and quantity of preleukaemic t(12;21)-positive cells peaks at term or early childhood and that exogenous...

  14. Curcumin induces apoptotic cell death of activated human CD4+ T cells via increasing endoplasmic reticulum stress and mitochondrial dysfunction.

    Science.gov (United States)

    Zheng, Min; Zhang, Qinggao; Joe, Yeonsoo; Lee, Bong Hee; Ryu, Do Gon; Kwon, Kang Beom; Ryter, Stefan W; Chung, Hun Taeg

    2013-03-01

    Curcumin, a natural polyphenolic antioxidant compound, exerts well-known anti-inflammatory and immunomodulatory effects, the latter which can influence the activation of immune cells including T cells. Furthermore, curcumin can inhibit the expression of pro-inflammatory cytokines and chemokines, through suppression of the NF-κB signaling pathway. The beneficial effects of curcumin in diseases such as arthritis, allergy, asthma, atherosclerosis, diabetes and cancer may be due to its immunomodulatory properties. We studied the potential of curcumin to modulate CD4+ T cells-mediated autoimmune disease, by examining the effects of this compound on human CD4+ lymphocyte activation. Stimulation of human T cells with PHA or CD3/CD28 induced IL-2 mRNA expression and activated the endoplasmic reticulum (ER) stress response. The treatment of T cells with curcumin induced the unfolded protein response (UPR) signaling pathway, initiated by the phosphorylation of PERK and IRE1. Furthermore, curcumin increased the expression of the ER stress associated transcriptional factors XBP-1, cleaved p50ATF6α and C/EBP homologous protein (CHOP) in human CD4+ and Jurkat T cells. In PHA-activated T cells, curcumin further enhanced PHA-induced CHOP expression and reduced the expression of the anti-apoptotic protein Bcl-2. Finally, curcumin treatment induced apoptotic cell death in activated T cells via eliciting an excessive ER stress response, which was reversed by the ER-stress inhibitor 4-phenylbutyric acid or transfection with CHOP-specific siRNA. These results suggest that curcumin can impact both ER stress and mitochondria functional pathways, and thereby could be used as a promising therapy in the context of Th1-mediated autoimmune diseases. Copyright © 2013 Elsevier B.V. All rights reserved.

  15. Increased cardiogenesis in P19-GFP teratocarcinoma cells expressing the propeptide IGF-1Ea

    Energy Technology Data Exchange (ETDEWEB)

    Poudel, Bhawana [Heart Science Centre, National Heart and Lung Institute, Imperial College, London (United Kingdom); Bilbao, Daniel [EMBL, Mouse Biology Unit, Monterotondo (Italy); Sarathchandra, Padmini; Germack, Renee [Heart Science Centre, National Heart and Lung Institute, Imperial College, London (United Kingdom); Rosenthal, Nadia [Heart Science Centre, National Heart and Lung Institute, Imperial College, London (United Kingdom); Australian Regenerative Medicine Institute, Monash University, Melbourne (Australia); Santini, Maria Paola, E-mail: m.santini@imperial.ac.uk [Heart Science Centre, National Heart and Lung Institute, Imperial College, London (United Kingdom)

    2011-12-16

    Highlights: Black-Right-Pointing-Pointer In this study, we explored the function of IGF-1Ea propeptide in inducing cardiogenesis of stem cells. Black-Right-Pointing-Pointer IGF-1Ea promoted cardiac mesodermal induction in uncommitted cells. Black-Right-Pointing-Pointer Under differentiation condition, IGF-1Ea increased expression of cardiac differentiation markers. Black-Right-Pointing-Pointer Furthermore, it promoted formation of finely organized sarcomeric structure. Black-Right-Pointing-Pointer IGF-1Ea propeptide may be a good candidate to improve production of cardiomyocytes from pluripotent cells. -- Abstract: The mechanism implicated in differentiation of endogenous cardiac stem cells into cardiomyocytes to regenerate the heart tissue upon an insult remains elusive, limiting the therapeutical goals to exogenous cell injection and/or gene therapy. We have shown previously that cardiac specific overexpression of the insulin-like growth factor 1 propeptide IGF-1Ea induces beneficial myocardial repair after infarct. Although the mechanism is still under investigation, the possibility that this propeptide may be involved in promoting stem cell differentiation into the cardiac lineage has yet to be explored. To investigate whether IGF-1Ea promote cardiogenesis, we initially modified P19 embryonal carcinoma cells to express IGF-1Ea. Taking advantage of their cardiomyogenic nature, we analyzed whether overexpression of this propeptide affected cardiac differentiation program. The data herein presented showed for the first time that constitutively overexpressed IGF-1Ea increased cardiogenic differentiation program in both undifferentiated and DMSO-differentiated cells. In details, IGF-1Ea overexpression promoted localization of alpha-actinin in finely organized sarcomeric structure compared to control cells and upregulated the cardiac mesodermal marker NKX-2.5 and the ventricular structural protein MLC2v. Furthermore, activated IGF-1 signaling promoted cardiac

  16. Micro-RNA 10a Is Increased in Feline T Regulatory Cells and Increases Foxp3 Protein Expression Following In Vitro Transfection

    Directory of Open Access Journals (Sweden)

    Yan Wang

    2017-02-01

    Full Text Available CD4+CD25+Foxp3+ T regulatory (Treg cells are activated during the course of lentiviral infection and exhibit heightened suppressor function when compared to Treg cells from uninfected controls. Foxp3 is essential to Treg cell function and multiple studies have documented that lentivirus-activated Treg cells exhibit heightened Foxp3 expression when compared to Treg cells from uninfected controls. Our hypothesis was that lentivirus-induced micro-RNAs (miRNAs contribute to heightened Treg cell suppressor function by stabilizing Foxp3 expression. We demonstrated that CD4+CD25+ T cells from both feline immunodeficiency virus infected (FIV+ cats and uninfected control cats exhibit increased miRNA 10a and 21 levels compared to autologous CD4+CD25− T cells but there was no difference in the levels of these miRNAs when Treg cells from FIV+ cats were compared to Treg cells from uninfected controls. Further, there was no increase in Foxp3 mRNA following transfection of miRNA 10a or 21 into a feline cell line. However, transfection with miRNA 10a resulted in increased Foxp3 protein expression.

  17. Bone marrow mesenchymal stem cells overexpressing human basic fibroblast growth factor increase vasculogenesis in ischemic rats

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, J.C. [Department of Vascular Surgery, The First Affiliated Hospital of Fujian Medical University, Fuzhou (China); Zheng, G.F. [Department of Vascular Surgery, The People' s Hospital of Ganzhou, Ganzhou (China); Wu, L.; Ou Yang, L.Y.; Li, W.X. [Department of Vascular Surgery, The First Affiliated Hospital of Fujian Medical University, Fuzhou (China)

    2014-08-08

    Administration or expression of growth factors, as well as implantation of autologous bone marrow cells, promote in vivo angiogenesis. This study investigated the angiogenic potential of combining both approaches through the allogenic transplantation of bone marrow-derived mesenchymal stem cells (MSCs) expressing human basic fibroblast growth factor (hbFGF). After establishing a hind limb ischemia model in Sprague Dawley rats, the animals were randomly divided into four treatment groups: MSCs expressing green fluorescent protein (GFP-MSC), MSCs expressing hbFGF (hbFGF-MSC), MSC controls, and phosphate-buffered saline (PBS) controls. After 2 weeks, MSC survival and differentiation, hbFGF and vascular endothelial growth factor (VEGF) expression, and microvessel density of ischemic muscles were determined. Stable hbFGF expression was observed in the hbFGF-MSC group after 2 weeks. More hbFGF-MSCs than GFP-MSCs survived and differentiated into vascular endothelial cells (P<0.001); however, their differentiation rates were similar. Moreover, allogenic transplantation of hbFGF-MSCs increased VEGF expression (P=0.008) and microvessel density (P<0.001). Transplantation of hbFGF-expressing MSCs promoted angiogenesis in an in vivo hind limb ischemia model by increasing the survival of transplanted cells that subsequently differentiated into vascular endothelial cells. This study showed the therapeutic potential of combining cell-based therapy with gene therapy to treat ischemic disease.

  18. Omega 3 fatty acids increase spontaneous release of cytosolic components from tumor cells

    International Nuclear Information System (INIS)

    Jenski, L.J.; Sturdevant, L.K.; Ehringer, W.D.; Stillwell, W.

    1991-01-01

    Mice fed menhaden (fish) oil or coconut oil-rich diets were inoculated intraperitoneally with a rapidly growing leukemia, T27A. After one week, the tumor cells were harvested, and 51Cr was used to label intracellular molecules. Spontaneous release of 51Cr was used as a measure of plasma membrane permeability. Compared to cells from mice fed coconut oil (rich in saturated fatty acids), tumor cells from mice fed menhaden oil (rich in long chain polyunsaturated omega 3 fatty acids) showed an increased level of spontaneous 51Cr release, which was exacerbated by increased temperature and reduced by extracellular protein. At physiological salt concentrations, the released 51Cr was detected in particles of approximately 2700 daltons. Enhanced permeability correlated with the incorporation of dietary (fish oil) omega 3 polyunsaturated fatty acids docosahexaenoic and eicosapentaenoic acid into the tumor cells. The results demonstrate that omega 3 fatty acids are incorporated into cellular constituents of tumor cells and change properties associated with the plasma membrane. This result suggests that dietary manipulation may be used to enhance tumor cell permeability and contribute to tumor eradication

  19. High Glucose Increases Metallothionein Expression in Renal Proximal Tubular Epithelial Cells

    Directory of Open Access Journals (Sweden)

    Daisuke Ogawa

    2011-01-01

    Full Text Available Metallothionein (MT is an intracellular metal-binding, cysteine-rich protein, and is a potent antioxidant that protects cells and tissues from oxidative stress. Although the major isoforms MT-1 and -2 (MT-1/-2 are highly inducible in many tissues, the distribution and role of MT-1/-2 in diabetic nephropathy are poorly understood. In this study, diabetes was induced in adult male rats by streptozotocin, and renal tissues were stained with antibodies for MT-1/-2. MT-1/-2 expression was also evaluated in mProx24 cells, a mouse renal proximal tubular epithelial cell line, stimulated with high glucose medium and pretreated with the antioxidant vitamin E. MT-1/-2 expression was gradually and dramatically increased, mainly in the proximal tubular epithelial cells and to a lesser extent in the podocytes in diabetic rats, but was hardly observed in control rats. MT-1/-2 expression was also increased by high glucose stimulation in mProx24 cells. Because the induction of MT was suppressed by pretreatment with vitamin E, the expression of MT-1/-2 is induced, at least in part, by high glucose-induced oxidative stress. These observations suggest that MT-1/-2 is induced in renal proximal tubular epithelial cells as an antioxidant to protect the kidney from oxidative stress, and may offer a novel therapeutic target against diabetic nephropathy.

  20. Bone marrow mesenchymal stem cells overexpressing human basic fibroblast growth factor increase vasculogenesis in ischemic rats

    Directory of Open Access Journals (Sweden)

    J.C. Zhang

    2014-10-01

    Full Text Available Administration or expression of growth factors, as well as implantation of autologous bone marrow cells, promote in vivo angiogenesis. This study investigated the angiogenic potential of combining both approaches through the allogenic transplantation of bone marrow-derived mesenchymal stem cells (MSCs expressing human basic fibroblast growth factor (hbFGF. After establishing a hind limb ischemia model in Sprague Dawley rats, the animals were randomly divided into four treatment groups: MSCs expressing green fluorescent protein (GFP-MSC, MSCs expressing hbFGF (hbFGF-MSC, MSC controls, and phosphate-buffered saline (PBS controls. After 2 weeks, MSC survival and differentiation, hbFGF and vascular endothelial growth factor (VEGF expression, and microvessel density of ischemic muscles were determined. Stable hbFGF expression was observed in the hbFGF-MSC group after 2 weeks. More hbFGF-MSCs than GFP-MSCs survived and differentiated into vascular endothelial cells (P<0.001; however, their differentiation rates were similar. Moreover, allogenic transplantation of hbFGF-MSCs increased VEGF expression (P=0.008 and microvessel density (P<0.001. Transplantation of hbFGF-expressing MSCs promoted angiogenesis in an in vivo hind limb ischemia model by increasing the survival of transplanted cells that subsequently differentiated into vascular endothelial cells. This study showed the therapeutic potential of combining cell-based therapy with gene therapy to treat ischemic disease.

  1. Irgm1-deficient mice exhibit Paneth cell abnormalities and increased susceptibility to acute intestinal inflammation.

    Science.gov (United States)

    Liu, Bo; Gulati, Ajay S; Cantillana, Viviana; Henry, Stanley C; Schmidt, Elyse A; Daniell, Xiaoju; Grossniklaus, Emily; Schoenborn, Alexi A; Sartor, R Balfour; Taylor, Gregory A

    2013-10-15

    Crohn's disease (CD) is a chronic, immune-mediated, inflammatory disorder of the intestine that has been linked to numerous susceptibility genes, including the immunity-related GTPase (IRG) M (IRGM). IRGs comprise a family of proteins known to confer resistance to intracellular infections through various mechanisms, including regulation of phagosome processing, cell motility, and autophagy. However, despite its association with CD, the role of IRGM and other IRGs in regulating intestinal inflammation is unclear. We investigated the involvement of Irgm1, an ortholog of IRGM, in the genesis of murine intestinal inflammation. After dextran sodium sulfate exposure, Irgm1-deficient [Irgm1 knockout (KO)] mice showed increased acute inflammation in the colon and ileum, with worsened clinical responses. Marked alterations of Paneth cell location and granule morphology were present in Irgm1 KO mice, even without dextran sodium sulfate exposure, and were associated with impaired mitophagy and autophagy in Irgm1 KO intestinal cells (including Paneth cells). This was manifested by frequent tubular and swollen mitochondria and increased LC3-positive autophagic structures. Interestingly, these LC3-positive structures often contained Paneth cell granules. These results suggest that Irgm1 modulates acute inflammatory responses in the mouse intestine, putatively through the regulation of gut autophagic processes, that may be pivotal for proper Paneth cell functioning.

  2. Glycocalyx Degradation Induces a Proinflammatory Phenotype and Increased Leukocyte Adhesion in Cultured Endothelial Cells under Flow.

    Directory of Open Access Journals (Sweden)

    Karli K McDonald

    Full Text Available Leukocyte adhesion to the endothelium is an early step in the pathogenesis of atherosclerosis. Effective adhesion requires the binding of leukocytes to their cognate receptors on the surface of endothelial cells. The glycocalyx covers the surface of endothelial cells and is important in the mechanotransduction of shear stress. This study aimed to identify the molecular mechanisms underlying the role of the glycocalyx in leukocyte adhesion under flow. We performed experiments using 3-D cell culture models, exposing human abdominal aortic endothelial cells to steady laminar shear stress (10 dynes/cm2 for 24 hours. We found that with the enzymatic degradation of the glycocalyx, endothelial cells developed a proinflammatory phenotype when exposed to uniform steady shear stress leading to an increase in leukocyte adhesion. Our results show an up-regulation of ICAM-1 with degradation compared to non-degraded controls (3-fold increase, p<0.05 and we attribute this effect to a de-regulation in NF-κB activity in response to flow. These results suggest that the glycocalyx is not solely a physical barrier to adhesion but rather plays an important role in governing the phenotype of endothelial cells, a key determinant in leukocyte adhesion. We provide evidence for how the destabilization of this structure may be an early and defining feature in the initiation of atherosclerosis.

  3. Addition of Wollastonite Fibers to Calcium Phosphate Cement Increases Cell Viability and Stimulates Differentiation of Osteoblast-Like Cells

    Directory of Open Access Journals (Sweden)

    Juliana Almeida Domingues

    2017-01-01

    Full Text Available Calcium phosphate cement (CPC that is based on α-tricalcium phosphate (α-TCP is considered desirable for bone tissue engineering because of its relatively rapid degradation properties. However, such cement is relatively weak, restricting its use to areas of low mechanical stress. Wollastonite fibers (WF have been used to improve the mechanical strength of biomaterials. However, the biological properties of WF remain poorly understood. Here, we tested the response of osteoblast-like cells to being cultured on CPC reinforced with 5% of WF (CPC-WF. We found that both types of cement studied achieved an ion balance for calcium and phosphate after 3 days of immersion in culture medium and this allowed subsequent long-term cell culture. CPC-WF increased cell viability and stimulated cell differentiation, compared to nonreinforced CPC. We hypothesize that late silicon release by CPC-WF induces increased cell proliferation and differentiation. Based on our findings, we propose that CPC-WF is a promising material for bone tissue engineering applications.

  4. Neurogenesis and Increase in Differentiated Neural Cell Survival via Phosphorylation of Akt1 after Fluoxetine Treatment of Stem Cells

    Directory of Open Access Journals (Sweden)

    Anahita Rahmani

    2013-01-01

    Full Text Available Fluoxetine (FLX is a selective serotonin reuptake inhibitor (SSRI. Its action is possibly through an increase in neural cell survival. The mechanism of improved survival rate of neurons by FLX may relate to the overexpression of some kinases such as Akt protein. Akt1 (a serine/threonine kinase plays a key role in the modulation of cell proliferation and survival. Our study evaluated the effects of FLX on mesenchymal stem cell (MSC fate and Akt1 phosphorylation levels in MSCs. Evaluation tests included reverse transcriptase polymerase chain reaction, western blot, and immunocytochemistry assays. Nestin, MAP-2, and β-tubulin were detected after neurogenesis as neural markers. Ten μM of FLX upregulated phosphorylation of Akt1 protein in induced hEnSC significantly. Also FLX did increase viability of these MSCs. Continuous FLX treatment after neurogenesis elevated the survival rate of differentiated neural cells probably by enhanced induction of Akt1 phosphorylation. This study addresses a novel role of FLX in neurogenesis and differentiated neural cell survival that may contribute to explaining the therapeutic action of fluoxetine in regenerative pharmacology.

  5. Increase in Dye:Dendrimer Ratio Decreases Cellular Uptake of Neutral Dendrimers in RAW Cells.

    Science.gov (United States)

    Vaidyanathan, Sriram; Kaushik, Milan; Dougherty, Casey; Rattan, Rahul; Goonewardena, Sascha N; Banaszak Holl, Mark M; Monano, Janet; DiMaggio, Stassi

    2016-09-12

    Neutral generation 3 poly(amidoamine) dendrimers were labeled with Oregon Green 488 (G3-OG n ) to obtain materials with controlled fluorophore:dendrimer ratios (n = 1-2), a mixture containing mostly 3 dyes per dendrimer, a mixture containing primarily 4 or more dyes per dendrimer ( n = 4+), and a stochastic mixture ( n = 4 avg ). The UV absorbance of the dye conjugates increased linearly as n increased and the fluorescence emission decreased linearly as n increased. Cellular uptake was studied in RAW cells and HEK 293A cells as a function of the fluorophore:dendrimer ratio (n). The cellular uptake of G3-OG n ( n = 3, 4+, 4 avg ) into RAW cells was significantly lower than G3-OG n ( n = 1, 2). The uptake of G3-OG n ( n = 3, 4+, 4 avg ) into HEK 293A cells was not significantly different from G3-OG 1 . Thus, the fluorophore:dendrimer ratio was observed to change the extent of uptake in the macrophage uptake mechanism but not in the HEK 293A cell. This difference in endocytosis indicates the presence of a pathway in the macrophage that is sensitive to hydrophobicity of the particle.

  6. Overexpression of amyloid precursor protein increases copper content in HEK293 cells

    International Nuclear Information System (INIS)

    Suazo, Miriam; Hodar, Christian; Morgan, Carlos; Cerpa, Waldo; Cambiazo, Veronica; Inestrosa, Nibaldo C.; Gonzalez, Mauricio

    2009-01-01

    Amyloid precursor protein (APP) is a transmembrane glycoprotein widely expressed in mammalian tissues and plays a central role in Alzheimer's disease. However, its physiological function remains elusive. Cu 2+ binding and reduction activities have been described in the extracellular APP135-156 region, which might be relevant for cellular copper uptake and homeostasis. Here, we assessed Cu 2+ reduction and 64 Cu uptake in two human HEK293 cell lines overexpressing APP. Our results indicate that Cu 2+ reduction increased and cells accumulated larger levels of copper, maintaining cell viability at supra-physiological levels of Cu 2+ ions. Moreover, wild-type cells exposed to both Cu 2+ ions and APP135-155 synthetic peptides increased copper reduction and uptake. Complementation of function studies in human APP751 transformed Fre1 defective Saccharomyces cerevisiae cells rescued low Cu 2+ reductase activity and increased 64 Cu uptake. We conclude that Cu 2+ reduction activity of APP facilitates copper uptake and may represent an early step in cellular copper homeostasis.

  7. Strategies for increasing the efficiency of heterojunction organic solar cells: material selection and device architecture.

    Science.gov (United States)

    Heremans, Paul; Cheyns, David; Rand, Barry P

    2009-11-17

    Thin-film blends or bilayers of donor- and acceptor-type organic semiconductors form the core of heterojunction organic photovoltaic cells. Researchers measure the quality of photovoltaic cells based on their power conversion efficiency, the ratio of the electrical power that can be generated versus the power of incident solar radiation. The efficiency of organic solar cells has increased steadily in the last decade, currently reaching up to 6%. Understanding and combating the various loss mechanisms that occur in processes from optical excitation to charge collection should lead to efficiencies on the order of 10% in the near future. In organic heterojunction solar cells, the generation of photocurrent is a cascade of four steps: generation of excitons (electrically neutral bound electron-hole pairs) by photon absorption, diffusion of excitons to the heterojunction, dissociation of the excitons into free charge carriers, and transport of these carriers to the contacts. In this Account, we review our recent contributions to the understanding of the mechanisms that govern these steps. Starting from archetype donor-acceptor systems of planar small-molecule heterojunctions and solution-processed bulk heterojunctions, we outline our search for alternative materials and device architectures. We show that non-planar phthalocynanines have appealing absorption characteristics but also have reduced charge carrier transport. As a result, the donor layer needs to be ultrathin, and all layers of the device have to be tuned to account for optical interference effects. Using these optimization techniques, we illustrate cells with 3.1% efficiency for the non-planar chloroboron subphthalocyanine donor. Molecules offering a better compromise between absorption and carrier mobility should allow for further improvements. We also propose a method for increasing the exciton diffusion length by converting singlet excitons into long-lived triplets. By doping a polymer with a

  8. Expression of p210 BCR/ABl increases hematopoietic progenitor cell radiosensitivity

    International Nuclear Information System (INIS)

    Santucci, M.A.; Anklesaria, P.; Das, I.J.; Sakakeeny, M.A.; FitzGerald, T.J.; Greenberger, J.S.; Laneuville, P.

    1993-01-01

    The cytogenetic finding of the Ph1+ chromosome and its molecular biologic marker bcr/abl gene rearrangement in cells from patients with chronic myeloid leukemia are associated with a proliferative advantage of the Ph1+ clone in vivo. Although the transition to the acute terminal phase or blastic crisis is often associated with additional cytogenetic abnormalities, the molecular events which correlate the initial cytogenetic lesion with the terminal phase are poorly understood. Defective cellular DNA repair capacity is often associated with chromosomal instability, increased mutation frequency, and biologic alterations. The authors tested whether the protein product of the bcr/abl translocation (p210) could alter DNA repair after gamma-irradiation of murine cell lines expressing the bcr/abl cDNA. The 32D cl 3 parent, 32D cl 3 pYN (containing the control vector plasmid) and each of two sources of 32D cl 3 cells expressing p210 cDNA (32D-PC1 cell line and 32D-LG7 subclone) showed a D 0 of 1.62, 1.57, 1.16, and 1.27 Gy, respectively. Thus, expression of the p210 product induced a significant increase in radiosensitivity at the clinically relevant radiation therapy dose-rate. The increased radiosensitivity of p210-expressing cells persisted if cells were held before plating in a density-inhibited state for 8 hr after gamma-irradiation, indicating little effect on the repair of potentially lethal gamma-irradiation damage. The IL-3 dependent parent 32D cl 3 cells demonstrated programmed cell death in the absence of growth factor or following gamma-irradiation to 200 cGy. Expression of p210 cDNA in the 32D-PC1 and 32D-LG7 subclones abrogated IL-3 requirement of these cell lines and inhibited gamma-irradiation induced programmed cell death. These data suggest a role for p210 in amplifying gamma-irradiation DNA damage or broadly inhibiting DNA repair, conditions that may stimulate further cytogenetic alterations in hematopoietic cells. 43 refs., 3 figs., 1 tab

  9. TIMP-1 increases expression and phosphorylation of proteins associated with drug resistance in breast cancer cells

    DEFF Research Database (Denmark)

    Hekmat, Omid; Munk, Stephanie; Fogh, Louise

    2013-01-01

    may explain the resistance phenotype to topoisomerase inhibitors that was observed in cells with high TIMP-1 levels. Pathway analysis showed an enrichment of proteins from functional categories such as apoptosis, cell cycle, DNA repair, transcription factors, drug targets and proteins associated......Tissue inhibitor of metalloproteinase 1 (TIMP-1) is a protein with a potential biological role in drug resistance. To elucidate the unknown molecular mechanisms underlying the association between high TIMP-1 levels and increased chemotherapy resistance, we employed SILAC-based quantitative mass...... spectrometry to analyze global proteome and phosphoproteome differences of MCF-7 breast cancer cells expressing high or low levels of TIMP-1. In TIMP-1 high expressing cells, 312 proteins and 452 phosphorylation sites were up-regulated. Among these were the cancer drug targets topoisomerase 1, 2A and 2B, which...

  10. Increasing Fuel Efficiency of Direct Methanol Fuel Cell Systems with Feedforward Control of the Operating Concentration

    Directory of Open Access Journals (Sweden)

    Youngseung Na

    2015-09-01

    Full Text Available Most of the R&D on fuel cells for portable applications concentrates on increasing efficiencies and energy densities to compete with other energy storage devices, especially batteries. To improve the efficiency of direct methanol fuel cell (DMFC systems, several modifications to system layouts and operating strategies are considered in this paper, rather than modifications to the fuel cell itself. Two modified DMFC systems are presented, one with an additional inline mixer and a further modification of it with a separate tank to recover condensed water. The set point for methanol concentration control in the solution is determined by fuel efficiency and varies with the current and other process variables. Feedforward concentration control enables variable concentration for dynamic loads. Simulation results were validated experimentally with fuel cell systems.

  11. Employing Si solar cell technology to increase efficiency of ultra-thin Cu(In,Ga)Se2 solar cells.

    Science.gov (United States)

    Vermang, Bart; Wätjen, Jörn Timo; Fjällström, Viktor; Rostvall, Fredrik; Edoff, Marika; Kotipalli, Ratan; Henry, Frederic; Flandre, Denis

    2014-10-01

    Reducing absorber layer thickness below 500 nm in regular Cu(In,Ga)Se 2 (CIGS) solar cells decreases cell efficiency considerably, as both short-circuit current and open-circuit voltage are reduced because of incomplete absorption and high Mo/CIGS rear interface recombination. In this work, an innovative rear cell design is developed to avoid both effects: a highly reflective rear surface passivation layer with nano-sized local point contact openings is employed to enhance rear internal reflection and decrease the rear surface recombination velocity significantly, as compared with a standard Mo/CIGS rear interface. The formation of nano-sphere shaped precipitates in chemical bath deposition of CdS is used to generate nano-sized point contact openings. Evaporation of MgF 2 coated with a thin atomic layer deposited Al 2 O 3 layer, or direct current magnetron sputtering of Al 2 O 3 are used as rear surface passivation layers. Rear internal reflection is enhanced substantially by the increased thickness of the passivation layer, and also the rear surface recombination velocity is reduced at the Al 2 O 3 /CIGS rear interface. (MgF 2 /)Al 2 O 3 rear surface passivated ultra-thin CIGS solar cells are fabricated, showing an increase in short circuit current and open circuit voltage compared to unpassivated reference cells with equivalent CIGS thickness. Accordingly, average solar cell efficiencies of 13.5% are realized for 385 nm thick CIGS absorber layers, compared with 9.1% efficiency for the corresponding unpassivated reference cells.

  12. Employing Si solar cell technology to increase efficiency of ultra-thin Cu(In,Ga)Se2 solar cells

    Science.gov (United States)

    Vermang, Bart; Wätjen, Jörn Timo; Fjällström, Viktor; Rostvall, Fredrik; Edoff, Marika; Kotipalli, Ratan; Henry, Frederic; Flandre, Denis

    2014-01-01

    Reducing absorber layer thickness below 500 nm in regular Cu(In,Ga)Se2 (CIGS) solar cells decreases cell efficiency considerably, as both short-circuit current and open-circuit voltage are reduced because of incomplete absorption and high Mo/CIGS rear interface recombination. In this work, an innovative rear cell design is developed to avoid both effects: a highly reflective rear surface passivation layer with nano-sized local point contact openings is employed to enhance rear internal reflection and decrease the rear surface recombination velocity significantly, as compared with a standard Mo/CIGS rear interface. The formation of nano-sphere shaped precipitates in chemical bath deposition of CdS is used to generate nano-sized point contact openings. Evaporation of MgF2 coated with a thin atomic layer deposited Al2O3 layer, or direct current magnetron sputtering of Al2O3 are used as rear surface passivation layers. Rear internal reflection is enhanced substantially by the increased thickness of the passivation layer, and also the rear surface recombination velocity is reduced at the Al2O3/CIGS rear interface. (MgF2/)Al2O3 rear surface passivated ultra-thin CIGS solar cells are fabricated, showing an increase in short circuit current and open circuit voltage compared to unpassivated reference cells with equivalent CIGS thickness. Accordingly, average solar cell efficiencies of 13.5% are realized for 385 nm thick CIGS absorber layers, compared with 9.1% efficiency for the corresponding unpassivated reference cells. PMID:26300619

  13. Increased percentage of Th17 cells in peritoneal fluid is associated with severity of endometriosis.

    Science.gov (United States)

    Gogacz, Marek; Winkler, Izabela; Bojarska-Junak, Agnieszka; Tabarkiewicz, Jacek; Semczuk, Andrzej; Rechberger, Tomasz; Adamiak, Aneta

    2016-09-01

    Th17 cells are a newly discovered T helper lymphocyte subpopulation, producing interleukin IL-17. Th17 cells are present in blood and peritoneal fluid (PF) at different stages of endometriosis. We aim to establish their potential importance in the pathogenesis and clinical features of the disease. The percentage of Th17 cells among T helper lymphocytes was determined in the PF and peripheral blood (PB) of patients with endometriosis and in the control group by flow cytometry using monoclonal antibodies: anti-CD-4-FITC, anti-CD-3-PE/Cy5, and anti-IL-17A-PE. Th17 percentage is increased in PF in comparison with PB in both endometriotic patients and in the control group. In severe endometriosis, the percentage of Th17 cells in PF was higher than with early (I/II stage) endometriosis. A positive correlation between the percentage of Th17 cells in PF and the white blood cell count in PB was found in patients with endometriosis. Targeting the activity of PF Th17 cells may have an influence on the proliferation of ectopic tissue and clinical manifestations of the disease. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

  14. Resveratrol sensitization of DU145 prostate cancer cells to ionizing radiation is associated to ceramide increase.

    Science.gov (United States)

    Scarlatti, Francesca; Sala, Giusy; Ricci, Clara; Maioli, Claudio; Milani, Franco; Minella, Marco; Botturi, Marco; Ghidoni, Riccardo

    2007-08-08

    Radiotherapy is an established therapeutic modality for prostate cancer. Since it is well known that radiotherapy is limited due to its severe toxicity towards normal cells at high dose and minimal effect at low dose, the search for biological compounds that increase the sensitivity of tumors cells to radiation may improve the efficacy of therapy. Resveratrol, a natural antioxidant, was shown to inhibit carcinogenesis in animal models, and to block the process of tumor initiation and progression. The purpose of this study was to examine whether or not resveratrol can sensitize DU145, an androgen-independent human prostate cancer cell line, to ionizing radiation. We report here that DU145 cells are resistant to ionizing radiation-induced cell death, but pretreatment with resveratrol significantly enhances cell death. Resveratrol acts synergistically with ionizing radiation to inhibit cell survival in vitro. Resveratrol also potentiates ionizing radiation-induced ceramide accumulation, by promoting its de novo biosynthesis. This confirms ceramide as an effective mediator of the anticancer potential induced by resveratrol.

  15. Oxidized-LDL induce morphological changes and increase stiffness of endothelial cells

    International Nuclear Information System (INIS)

    Chouinard, Julie A.; Grenier, Guillaume; Khalil, Abdelouahed; Vermette, Patrick

    2008-01-01

    There is increasing evidence suggesting that oxidized low-density lipoproteins (ox-LDL) play a critical role in endothelial injury contributing to the age-related physio-pathological process of atherosclerosis. In this study, the effects of native LDL and ox-LDL on the mechanical properties of living human umbilical vein endothelial cells (HUVEC) were investigated by atomic force microscopy (AFM) force measurements. The contribution of filamentous actin (F-actin) and vimentin on cytoskeletal network organization were also examined by fluorescence microscopy. Our results revealed that ox-LDL had an impact on the HUVEC shape by interfering with F-actin and vimentin while native LDL showed no effect. AFM colloidal force measurements on living individual HUVEC were successfully used to measure stiffness of cells exposed to native and ox-LDL. AFM results demonstrated that the cell body became significantly stiffer when cells were exposed for 24 h to ox-LDL while cells exposed for 24 h to native LDL displayed similar rigidity to that of the control cells. Young's moduli of LDL-exposed HUVEC were calculated using two models. This study thus provides quantitative evidence on biomechanical mechanisms related to endothelial cell dysfunction and may give new insight on strategies aiming to protect endothelial function in atherosclerosis

  16. Sulforaphane enhances irradiation effects in terms of perturbed cell cycle progression and increased DNA damage in pancreatic cancer cells.

    Directory of Open Access Journals (Sweden)

    Patrick Naumann

    Full Text Available Sulforaphane (SFN, an herbal isothiocyanate enriched in cruciferous vegetables like broccoli and cauliflower, has gained popularity for its antitumor effects in cell lines such as pancreatic cancer. Antiproliferative as well as radiosensitizing properties were reported for head and neck cancer but little is known about its effects in pancreatic cancer cells in combination with irradiation (RT.In four established pancreatic cancer cell lines we investigated clonogenic survival, analyzed cell cycle distribution and compared DNA damage via flow cytometry and western blot after treatment with SFN and RT.Both SFN and RT show a strong and dose dependent survival reduction in clonogenic assays, an induction of a G2/M cell cycle arrest and an increase in γH2AX protein level indicating DNA damage. Effects were more pronounced in combined treatment and both cell cycle perturbation and DNA damage persisted for a longer period than after SFN or RT alone. Moreover, SFN induced a loss of DNA repair proteins Ku 70, Ku 80 and XRCC4.Our results suggest that combination of SFN and RT exerts a more distinct DNA damage and growth inhibition than each treatment alone. SFN seems to be a viable option to improve treatment efficacy of chemoradiation with hopefully higher rates of secondary resectability after neoadjuvant treatment for pancreatic cancer.

  17. SUMO-modified insulin-like growth factor 1 receptor (IGF-1R) increases cell cycle progression and cell proliferation.

    Science.gov (United States)

    Lin, Yingbo; Liu, Hongyu; Waraky, Ahmed; Haglund, Felix; Agarwal, Prasoon; Jernberg-Wiklund, Helena; Warsito, Dudi; Larsson, Olle

    2017-10-01

    Increasing number of studies have shown nuclear localization of the insulin-like growth factor 1 receptor (nIGF-1R) in tumor cells and its links to adverse clinical outcome in various cancers. Any obvious cell physiological roles of nIGF-1R have, however, still not been disclosed. Previously, we reported that IGF-1R translocates to cell nucleus and modulates gene expression by binding to enhancers, provided that the receptor is SUMOylated. In this study, we constructed stable transfectants of wild type IGF1R (WT) and triple-SUMO-site-mutated IGF1R (TSM) using igf1r knockout mouse fibroblasts (R-). Cell clones (R-WT and R-TSM) expressing equal amounts of IGF-1R were selected for experiments. Phosphorylation of IGF-1R, Akt, and Erk upon IGF-1 stimulation was equal in R-WT and R-TSM. WT was confirmed to enter nuclei. TSM did also undergo nuclear translocation, although to a lesser extent. This may be explained by that TSM heterodimerizes with insulin receptor, which is known to translocate to cell nuclei. R-WT proliferated substantially faster than R-TSM, which did not differ significantly from the empty vector control. Upon IGF-1 stimulation G1-S-phase progression of R-WT increased from 12 to 38%, compared to 13 to 20% of R-TSM. The G1-S progression of R-WT correlated with increased expression of cyclin D1, A, and CDK2, as well as downregulation of p27. This suggests that SUMO-IGF-1R affects upstream mechanisms that control and coordinate expression of cell cycle regulators. Further studies to identify such SUMO-IGF-1R dependent mechanisms seem important. © 2017 The Authors. Journal of Cellular Physiology Published by Wiley Periodicals Inc.

  18. Hypoxia increases the metastatic ability of breast cancer cells via upregulation of CXCR4

    LENUS (Irish Health Repository)

    Cronin, Patricia A

    2010-05-21

    Abstract Background Chemokine SDF1α and its unique receptor CXCR4 have been implicated in organ-specific metastases of many cancers including breast cancer. Hypoxia is a common feature of solid tumors and is associated with their malignant phenotype. We hypothesized that hypoxia would upregulate CXCR4 expression and lead to increased chemotactic responsiveness to its specific ligand SDF1α. Methods Three breast cancer cell lines MDA-MB-231, MCF7 and 4T1 were subjected to 48 hrs of hypoxia or normoxia. Cell surface receptor expression was evaluated using flow cytometry. An extracellular matrix invasion assay and microporous migration assay was used to assess chemotactic response and metastatic ability. Results CXCR4 surface expression was significantly increased in the two human breast cancer cell lines, MDA-MB-231 and MCF7, following exposure to hypoxia. This upregulation of CXCR4 cell surface expression corresponded to a significant increase in migration and invasion in response to SDF1-α in vitro. The increase in metastatic potential of both the normoxic and the hypoxic treated breast cancer cell lines was attenuated by neutralization of CXCR4 with a CXCR4 neutralizing mAb, MAB172 or a CXCR4 antagonist, AMD3100, showing the relationship between CXCR4 overexpression and increased chemotactic responsiveness. Conclusions CXCR4 expression can be modulated by the tissue microenvironment such as hypoxia. Upregulation of CXCR4 is associated with increased migratory and invasive potential and this effect can be abrogated by CXCR4 inhibition. Chemokine receptor CXCR4 is a potential therapeutic target in the adjuvant treatment of breast cancer.

  19. Increased apoptosis and peripheral blood mononuclear cell suppression of bone marrow mesenchymal stem cells in severe aplastic anemia.

    Science.gov (United States)

    Chao, Yu-Hua; Lin, Chiao-Wen; Pan, Hui-Hsien; Yang, Shun-Fa; Weng, Te-Fu; Peng, Ching-Tien; Wu, Kang-Hsi

    2018-06-05

    Although immune-mediated pathogenesis is considered an important aspect of severe aplastic anemia (SAA), its underlying mechanisms remain unclear. Mesenchymal stem cells (MSCs) are essential to the formation of specialized microenvironments in the bone marrow (BM), and MSC insufficiency can trigger the development of SAA. To find MSC alterations in the SAA BM, we compared BM MSCs from five children with SAA and five controls. Peripheral blood mononuclear cells (PBMCs) were cocultured with MSCs to evaluate the supportive effects of MSCs on hematopoiesis. Cytometric bead array immunoassay was used to determine cytokine excretion by MSCs. The immune functions of MSCs and their conditioned medium (CM) were evaluated by PBMC proliferation assays. SAA MSCs were characterized by a high percentage of cells in the abnormal sub-G1 phase of the cell cycle, which suggests an increased rate of apoptosis in SAA MSCs. In comparison with control MSCs, PBMCs cocultured with SAA MSCs displayed significantly reduced PBMC proliferation (P = 0.009). Aberrant cytokine profiles were secreted by SAA MSCs, with increased concentrations of interleukin-6, interferon-γ, tumor necrosis factor-α, and interleukin-1β in the CM. PBMC proliferation assays demonstrated additional immunosuppressive effects of SAA MSCs (P = 0.016) and their CM (P = 0.013). Our data revealed increased apoptosis and PBMC suppression of SAA MSCs. The alterations of MSCs may contribute to the formation of functionally abnormal microenvironments in SAA BM. © 2018 Wiley Periodicals, Inc.

  20. Increased mast cell tryptase in sudden infant death - anaphylaxis, hypoxia or artefact?

    Science.gov (United States)

    Edston, E; Gidlund, E; Wickman, M; Ribbing, H; Van Hage-Hamsten, M

    1999-12-01

    Increased concentrations of mast cell tryptase in post mortem blood have frequently been observed in sudden infant deaths but the cause of this has not yet been clarified. The aim was to evaluate factors (immunological, morphological and anamnestic data) behind the observed increase in mast cell tryptase in sudden infant deaths with elevated tryptase. Mast cell tryptase and total immunoglobulin (Ig) E were measured in post mortem sera from 44 infants younger than 1.5 years. Radioallergosorbent tests were performed for possible allergens (mixture for relevant food allergens, Phadiatop and latex). IgG subclasses, IgM, and complement factors (C3, C4 and factor B) were measured with radial immunodiffusion. Mast cells, labelled with antibodies against mast cell tryptase, were counted in the lungs and heart. The circumstances of death and medical history of the deceased infant and family were obtained through police and hospital records. In 40% of the SIDS cases tryptase was elevated (>10 microg/L). Total IgE in serum was increased in 33% compared with clinical reference values but showed no association with mast cell tryptase. RAST tests were positive in three cases. In one of these cases both tryptase and total IgE were elevated. The only variable that was associated with high tryptase values was prone position at death (P Children with elevated total IgE also displayed high concentrations of IgG1 and IgG2. Infants who died in the spring had significantly higher IgE than the others (P < or = 0.05). The results do not support the hypothesis that the elevated tryptase concentrations in sudden infant death are caused by allergy. The association between prone position at death and elevated tryptase could hypothetically be explained by mast cell degranulation due to, for example, a hypoxic stimulus in these infants.

  1. Increased light harvesting in dye-sensitized solar cells with energy relay dyes

    KAUST Repository

    Hardin, Brian E.

    2009-06-21

    Conventional dye-sensitized solar cells have excellent charge collection efficiencies, high open-circuit voltages and good fill factors. However, dye-sensitized solar cells do not completely absorb all of the photons from the visible and near-infrared domain and consequently have lower short-circuit photocurrent densities than inorganic photovoltaic devices. Here, we present a new design where high-energy photons are absorbed by highly photoluminescent chromophores unattached to the titania and undergo Förster resonant energy transfer to the sensitizing dye. This novel architecture allows for broader spectral absorption, an increase in dye loading, and relaxes the design requirements for the sensitizing dye. We demonstrate a 26% increase in power conversion efficiency when using an energy relay dye (PTCDI) with an organic sensitizing dye (TT1). We estimate the average excitation transfer efficiency in this system to be at least 47%. This system offers a viable pathway to develop more efficient dye-sensitized solar cells.

  2. Azidothymidine and cisplatin increase p14ARF expression in OVCAR-3 ovarian cancer cell line

    International Nuclear Information System (INIS)

    Vaskivuo, Liisa; Rysae, Jaana; Koivuperae, Johanna; Myllynen, Paeivi; Vaskivuo, Tommi; Chvalova, Katerina; Serpi, Raisa; Savolainen, Eeva-Riitta; Puistola, Ulla; Vaehaekangas, Kirsi

    2006-01-01

    p14 ARF tumor suppressor protein regulates p53 by interfering with mdm2-p53 interaction. p14 ARF is activated in response to oncogenic stimuli but little is known of the responses of endogenous p14 ARF to different types of cellular stress or DNA damage. Azidothymidine (AZT) is being tested in several clinical trials as an enhancer of anticancer chemotherapy. However, the knowledge of the relationship between AZT and cellular pathways, e.g. p53 pathway, is very limited. In this study, we show that AZT, cisplatin (CDDP) and docetaxel (DTX) all induce unique molecular responses in OVCAR-3 ovarian carcinoma cells carrying a mutated p53, while in A2780, ovarian carcinoma and MCF-7 breast carcinoma cells with wild type p53, all of these drugs cause similar p53 responses. We found that endogenous p14 ARF protein in OVCAR-3 cells is down-regulated by DTX but induced by AZT and a short CDDP pulse treatment. In HT-29 colon carcinoma cells with a mutated p53, all treatments down-regulated p14 ARF protein. Both CDDP and AZT increased the expression of p14ARF mRNA in OVCAR-3 cells. Differences in cell death induced by these drugs did not explain the differences in protein and mRNA expressions. No increase in the level of either c-Myc or H-ras oncoproteins was seen in OVCAR-3 cells after AZT or CDDP-treatment. These results suggest that p14 ARF can respond to DNA damage without oncogene activation in cell lines without functional p53

  3. Reduction of radiation-induced cell cycle blocks by caffeine does not necessarily lead to increased cell killing

    Energy Technology Data Exchange (ETDEWEB)

    Musk, S.R. (Institute of Cancer Research, Sutton, Surrey (England))

    1991-03-01

    The effect of caffeine upon the radiosensitivities of three human tumor lines was examined and correlated with its action upon the radiation-induced S-phase and G2-phase blocks. Caffeine was found to reduce at least partially the S-phase and G2-phase blocks in all the cell lines examined but potentiated cytotoxicity in only one of the three tumor lines. That reductions have been demonstrated to occur in the absence of increased cell killing provides supporting evidence for the hypothesis that reductions may not be causal in those cases when potentiation of radiation-induced cytotoxicity is observed in the presence of caffeine.

  4. Perfusion of veins at arterial pressure increases the expression of KLF5 and cell cycle genes in smooth muscle cells

    International Nuclear Information System (INIS)

    Amirak, Emre; Zakkar, Mustafa; Evans, Paul C.; Kemp, Paul R.

    2010-01-01

    Vascular smooth muscle cell (VSMC) proliferation remains a major cause of veno-arterial graft failure. We hypothesised that exposure of venous SMCs to arterial pressure would increase KLF5 expression and that of cell cycle genes. Porcine jugular veins were perfused at arterial or venous pressure in the absence of growth factors. The KLF5, c-myc, cyclin-D and cyclin-E expression were elevated within 24 h of perfusion at arterial pressure but not at venous pressure. Arterial pressure also reduced the decline in SM-myosin heavy chain expression. These data suggest a role for KLF5 in initiating venous SMCs proliferation in response to arterial pressure.

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

    Science.gov (United States)

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

    2015-01-01

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

  6. 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.

  7. Enhanced nitrogen removal in single-chamber microbial fuel cells with increased gas diffusion areas

    KAUST Repository

    Yan, Hengjing; Regan, John M.

    2012-01-01

    significantly increased the ammonia and COD removal rates (by up to 115% and 39%), ammonia removal efficiency (by up to 134%), the cell voltage and cathode potentials, and the power densities (by a factor of approximately 2). When the COD/N ratio was lowered

  8. TH1 and TH2 cell polarization increases with aging and is modulated by zinc supplementation

    OpenAIRE

    2008-01-01

    TH1 and TH2 cell polarization increases with aging and is modulated by zinc supplementation correspondence: Corresponding author. Tel.: +49 241 8080208; fax: +49 241 8082613. (Rink, Lothar) (Rink, Lothar) Institute of Immunology, University Hospital, RWTH Aachen University - Aachen--> - GERMANY (Uciechowski, Peter) Institute of Immunology, University Hospital, RWTH Aachen University - Aachen--> - GERMAN...

  9. Exposure to TiO2 nanoparticles increases Staphylococcusaureusinfection of HeLa cells

    Science.gov (United States)

    Xu, Yan; Wei, Ming-Tzo; Walker, Stephen. G.; Wang, Hong Zhan; Gondon, Chris; Brink, Peter; Guterman, Shoshana; Zawacki, Emma; Applebaum, Eliana; Rafailovich, Miriam; Ou-Yang, H. Daniel; Mironava, Tatsiana

    TiO2 is one of the most common nanoparticles in industry from food additives to energy generation. Even though TiO2 is also used as an anti-bacterial agent in combination with UV, we found that, in the absence of UV, exposure of HeLa cells to TiO2 nanoparticles largely increased their risk of bacterial invasion. HeLa cells cultured with low dosage rutile and anatase TiO2 nanoparticles (0.1 mg/ml) for 24 hrs prior to exposure to bacteria had 350% and 250% respectively more bacteria infected per cell. The increase was attributed to increased LDH leakage, and changes in the mechanical response of the cell membrane. On the other hand, macrophages exposed to TiO2 particles ingested 40% fewer bacteria, further increasing the risk of infection. In combination, these two factors raise serious concerns regarding the impact of exposure to TiO2 nanoparticles on the ability of organisms to resist bacterial infection.

  10. Increase in natural killer cell activity during diethylcarbamazine treatment of patients with filariasis

    DEFF Research Database (Denmark)

    Pedersen, B K; Bygbjerg, Ib Christian; Svenson, M

    1987-01-01

    Two patients, one with Bancroftian filariasis and the other with onchocerciasis, and two healthy controls were treated with diethylcarbamazine (DEC). The natural killer (NK) cell activity of the two patients increased during DEC treatment to 2.5 and 2.8 times, respectively, while that of the cont...

  11. Increased memory phenotypes of CD4+ and CD8+ T cells in ...

    African Journals Online (AJOL)

    Conclusions: Children with SCA in Tanzania show an absolute increase in all leukocyte types, including lymphocytes, with skewing of both CD4+ and CD8+ T cells towards the memory phenotypes. These findings provide insights on the development of adaptive immunity which may have implications on vaccine ...

  12. Drug induced increases in CNS dopamine alter monocyte, macrophage and T cell functions: implications for HAND

    Science.gov (United States)

    Gaskill, Peter J.; Calderon, Tina M.; Coley, Jacqueline S.; Berman, Joan W.

    2013-01-01

    Central nervous system (CNS) complications resulting from HIV infection remain a major public health problem as individuals live longer due to the success of combined antiretroviral therapy (cART). As many as 70% of HIV infected people have HIV associated neurocognitive disorders (HAND). Many HIV infected individuals abuse drugs, such as cocaine, heroin or methamphetamine, that may be important cofactors in the development of HIV CNS disease. Despite different mechanisms of action, all drugs of abuse increase extracellular dopamine in the CNS. The effects of dopamine on HIV neuropathogenesis are not well understood, and drug induced increases in CNS dopamine may be a common mechanism by which different types of drugs of abuse impact the development of HAND. Monocytes and macrophages are central to HIV infection of the CNS and to HAND. While T cells have not been shown to be a major factor in HIV-associated neuropathogenesis, studies indicate that T cells may play a larger role in the development of HAND in HIV infected drug abusers. Drug induced increases in CNS dopamine may dysregulate functions of, or increase HIV infection in, monocytes, macrophages and T cells in the brain. Thus, characterizing the effects of dopamine on these cells is important for understanding the mechanisms that mediate the development of HAND in drug abusers. PMID:23456305

  13. Nitric Oxide Synthesis Is Increased in Cybrid Cells with m.3243A>G Mutation

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    Juliana Gamba

    2012-12-01

    Full Text Available Nitric oxide (NO is a free radical and a signaling molecule in several pathways, produced by nitric oxide synthase (NOS from the conversion of L-arginine to citrulline. Supplementation of L-arginine has been used to treat MELAS (mitochondrial encephalopathy with lactic acidosis and stroke like syndrome, a mitochondrial disease caused by the m.3243A>G mutation. Low levels of serum arginine and endothelium dysfunction have been reported in MELAS and this treatment may increase NO in endothelial cells and promote vasodilation, decreasing cerebral ischemia and strokes. Although clinical benefits have been reported, little is known about NO synthesis in MELAS. In this study we found that osteosarcoma derived cybrid cells with high levels of m.3243A>G had increased nitrite, an NO metabolite, and increased intracellular NO, demonstrated by an NO fluorescent probe (DAF-FM. Muscle vessels from patients with the same mutation had increased staining in NADPH diaphorase, suggestive of increased NOS. These results indicate increased production of NO in cells harboring the m.3243A>G, however no nitrated protein was detected by Western blotting. Further studies are necessary to clarify the exact mechanisms of L-arginine effect to determine the appropriate clinical use of this drug therapy.

  14. Pam2 lipopeptides systemically increase myeloid-derived suppressor cells through TLR2 signaling

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    Maruyama, Akira; Shime, Hiroaki, E-mail: shime@med.hokudai.ac.jp; Takeda, Yohei; Azuma, Masahiro; Matsumoto, Misako; Seya, Tsukasa, E-mail: seya-tu@pop.med.hokudai.ac.jp

    2015-02-13

    Myeloid-derived suppressor cells (MDSCs) are immature myeloid cells that exhibit potent immunosuppressive activity. They are increased in tumor-bearing hosts and contribute to tumor development. Toll-like receptors (TLRs) on MDSCs may modulate the tumor-supporting properties of MDSCs through pattern-recognition. Pam2 lipopeptides represented by Pam2CSK4 serve as a TLR2 agonist to exert anti-tumor function by dendritic cell (DC)-priming that leads to NK cell activation and cytotoxic T cell proliferation. On the other hand, TLR2 enhances tumor cell progression/invasion by activating tumor-infiltrating macrophages. How MDSCs respond to TLR2 agonists has not yet been determined. In this study, we found intravenous administration of Pam2CSK4 systemically up-regulated the frequency of MDSCs in EG7 tumor-bearing mice. The frequency of tumor-infiltrating MDSCs was accordingly increased in response to Pam2CSK4. MDSCs were not increased by Pam2CSK4 stimuli in TLR2 knockout (KO) mice. Adoptive transfer experiments using CFSE-labeled MDSCs revealed that the TLR2-positive MDSCs survived long in tumor-bearing mice in response to Pam2CSK4 treatment. Since the increased MDSC population sustained immune-suppressive properties, our study suggests that Pam2CSK4-triggered TLR2 activation enhances the MDSC potential and suppress antitumor immune response in tumor microenvironment. - Highlights: • Pam2CSK4 administration induces systemic accumulation of CD11b{sup +}Gr1{sup +} MDSCs. • TLR2 is essential for Pam2CSK4-induced accumulation of CD11b{sup +}Gr1{sup +} MDSCs. • Pam2CSK4 supports survival of CD11b{sup +}Gr1{sup +} MDSCs in vivo.

  15. Fractalkine is expressed in the human ovary and increases progesterone biosynthesis in human luteinised granulosa cells

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    Yan Jie

    2011-06-01

    Full Text Available Abstract Background Recent evidence from rodent ovaries has demonstrated expression of fractalkine and the existence of fractalkine receptor, and showed that there is a significant increase in steroidogenesis in response to fractalkine, yet the role of fractalkine and CX3CR1 in the human ovary is still unknown. This study aimed to determine the expression levels of fractalkine and CX3CR1 in the human ovary and to investigate their roles in sexual hormone biosynthesis by human luteinising granulosa cells. This is the first detailed report of fractalkine and CX3CR1 expression and function in the human ovary. Methods Fractalkine and CX3CR1 expression levels were measured by immunohistochemistry using ovarian tissue from pathological specimens from five individuals. Granulosa cells were obtained from patients during IVF treatment. They were cultured and treated with increasing doses of hCG with or without fractalkine. Media were collected to detect estradiol and progesterone by chemiluminescence. StAR, 3-βHSD and CYP11A expression were determined in granulosa cells treated with or without fractalkine by real-time RT-PCR. Results Fractalkine and CX3CR1 were expressed in the human ovary and in luteinising granulosa cells. However, fractalkine expression was stronger in luteinising granulosa cells. Treatment with fractalkine augmented hCG stimulation of progesterone production in a dose-dependent manner with concomitant increases in transcript levels for key steroidogenic enzymes (StAR, 3-βHSD and CYP11A but had no effect on estradiol biosynthesis(P Conclusions Fractalkine and CX3CR1 were found to express in human ovary and luteinising granulosa cells. Fractalkine can increase the biosynthesis of progesterone in a dose-dependent manner by enhancing transcript levels of key steroidogenic enzymes.

  16. EMT factor Zeb1 depletion in dendritic cells enhances Helminth clearance in mice by increasing Th2 cell differentiation

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    Shuchi Smita

    2017-10-01

    Full Text Available Dendritic cells are professional antigen presenting cells that act as bridging link between innate and adaptive immune system. They are equipped with pathogen recognition receptors (PRR to identify the pathogen associated molecular pattern (PAMPS on any antigen. DCs elicit an immune response through polarizing T cells towards various subtypes like Th1, Th2 & Tregs. Though DC-T cell interaction has been widely studied, but how this single DC molecule amalgamate various transcriptional signals for translating the message to the T cells and induce diverse immunological responses still needs to be unraveled. Therefore to identify the role of transcription factor in immune programming we have targeted the largest member of TFs family, Zinc Finger Transcription Factors (ZF-TFs. Among various ZF-TFs we have narrowed our study to three interesting candidates Zeb1, Zeb2 and Zbtb10 based on their expression in DCs from an unpublished microarray data. Here in this study we have tried to understand the role of Zeb1, master regulator of EMT program in orchestrating DC responses. Zeb1 links the epithelial – mesenchymal transition and has been widely studied molecule in cancer biology. Except for the fact that it act as transcriptional repressor and represses IL2 gene promoter no other reports are available in immune biology, thereby rendering it a perfect candidate to be used for detailed characterization in dendritic cells. In our study, we found that Zeb1 depleted CD8α+DCs shows an increase in co-stimulatory marker like CD80 & CD86 whereas there is a decrease in MHC class I & II molecule. Thereafter at transcript & protein level we found decrease in pro-inflammatory & anti-inflammatory cytokine like IL6 & IL10 respectively, the bioactive form of IL12 i.e. IL12p70 which polarizes T cells towards Th1 response showed a significant decrease in bio-plex when compared with control CD8α+DCs. The regulatory markers which develop regulatory T cells like Pdl1, IL

  17. Increased betulinic acid induced cytotoxicity and radiosensitivity in glioma cells under hypoxic conditions

    International Nuclear Information System (INIS)

    Bache, Matthias; Taubert, Helge; Vordermark, Dirk; Zschornak, Martin P; Passin, Sarina; Keßler, Jacqueline; Wichmann, Henri; Kappler, Matthias; Paschke, Reinhard; Kaluđerović, Goran N; Kommera, Harish

    2011-01-01

    Betulinic acid (BA) is a novel antineoplastic agent under evaluation for tumor therapy. Because of the selective cytotoxic effects of BA in tumor cells (including gliomas), the combination of this agent with conservative therapies (such as radiotherapy and chemotherapy) may be useful. Previously, the combination of BA with irradiation under hypoxic conditions had never been studied. In this study, the effects of 3 to 30 μM BA on cytotoxicity, migration, the protein expression of PARP, survivin and HIF-1α, as well as radiosensitivity under normoxic and hypoxic conditions were analyzed in the human malignant glioma cell lines U251MG and U343MG. Cytotoxicity and radiosensitivity were analyzed with clonogenic survival assays, migration was analyzed with Boyden chamber assays (or scratch assays) and protein expression was examined with Western blot analyses. Under normoxic conditions, a half maximal inhibitory concentration (IC 50 ) of 23 μM was observed in U251MG cells and 24 μM was observed in U343MG cells. Under hypoxic conditions, 10 μM or 15 μM of BA showed a significantly increased cytotoxicity in U251MG cells (p = 0.004 and p = 0.01, respectively) and U343MG cells (p < 0.05 and p = 0.01, respectively). The combination of BA with radiotherapy resulted in an additive effect in the U343MG cell line under normoxic and hypoxic conditions. Weak radiation enhancement was observed in U251MG cell line after treatment with BA under normoxic conditions. Furthermore, under hypoxic conditions, the incubation with BA resulted in increased radiation enhancement. The enhancement factor, at an irradiation dose of 15 Gy after treatment with 10 or 15 μM BA, was 2.20 (p = 0.02) and 4.50 (p = 0.03), respectively. Incubation with BA led to decreased cell migration, cleavage of PARP and decreased expression levels of survivin in both cell lines. Additionally, BA treatment resulted in a reduction of HIF-1α protein under hypoxic conditions. Our results suggest that BA is capable

  18. Trans fatty acids increase nitric oxide levels and pancreatic beta-cell necrosis in rats

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    Kusmiyati Tjahjono DK

    2013-04-01

    Full Text Available Background The prevalence of diabetes in Indonesia is increasing due to various factors, including life style changes such as trans fatty acid (TFA intake. High TFA intake is known to be related to blood lipid profile changes resulting in cardiovascular disorders. This study was to identify the effect of TFA on nitric oxide (NO production and on necrosis of pancreatic beta cells. Methods A study of randomized pre-test post–test design with control group. Thirty Sprague Dawley rats were divided into 3 groups, i.e. group K (control, group P1 receiving a diet with 5% TFA, and P2 receiving 10% TFA. The intervention was performed for 8 weeks. NO level and pancreatic beta-cell necrosis were analyzed using Pearson’s chi square test. Results After 4 weeks of treatment there was no change in NO levels in group K, but increased NO in P2 (2.6-3.8 ìM. At 8 weeks after treatment, NO levels in groups P1 and P2 increased to 2.6-3.4 ìM and 4.2-14.3 ìM, respectively, while in group K only 2 rats had increased NO levels of 2.8-2.9 ìM. With Pearson’s chi-square test, there was a signifant difference in the proportions of necrotic pancreatic beta cells after 4 weeks and 8 weeks (p=0.000. No necrosis of beta cells was found in group K, mild necrosis in group P1 (1-25% and moderate necrosis in group P2 (26-50%. Conclusion TFA consumption significantly increases NO levels in Sprague Dawley rats and also results in moderate grades of necrosis of pancreatic beta cells.

  19. Trans fatty acids increase nitric oxide levels and pancreatic beta-cell necrosis in rats

    Directory of Open Access Journals (Sweden)

    Kusmiyati Tjahjono DK

    2015-12-01

    Full Text Available BACKGROUND The prevalence of diabetes in Indonesia is increasing due to various factors, including life style changes such as trans fatty acid (TFA intake. High TFA intake is known to be related to blood lipid profile changes resulting in cardiovascular disorders. This study was to identify the effect of TFA on nitric oxide (NO production and on necrosis of pancreatic beta cells. METHODS A study of randomized pre-test post–test design with control group. Thirty Sprague Dawley rats were divided into 3 groups, i.e. group K (control, group P1 receiving a diet with 5% TFA, and P2 receiving 10% TFA. The intervention was performed for 8 weeks. NO level and pancreatic beta-cell necrosis were analyzed using Pearson’s chi square test. RESULTS After 4 weeks of treatment there was no change in NO levels in group K, but increased NO in P2 (2.6-3.8 ìM. At 8 weeks after treatment, NO levels in groups P1 and P2 increased to 2.6-3.4 ìM and 4.2-14.3 ìM, respectively, while in group K only 2 rats had increased NO levels of 2.8-2.9 ìM. With Pearson’s chi-square test, there was a signifant difference in the proportions of necrotic pancreatic beta cells after 4 weeks and 8 weeks (p= 0.000. No necrosis of beta cells was found in group K, mild necrosis in group P1 (1-25% and moderate necrosis in group P2 (26-50%. CONCLUSION TFA consumption significantly increases NO levels in Sprague Dawley rats and also results in moderate grades of necrosis of pancreatic beta cells

  20. Sorting cells of the microalga Chlorococcum littorale with increased triacylglycerol productivity.

    Science.gov (United States)

    Cabanelas, Iago Teles Dominguez; van der Zwart, Mathijs; Kleinegris, Dorinde M M; Wijffels, René H; Barbosa, Maria J

    2016-01-01

    Despite extensive research in the last decades, microalgae are still only economically feasible for high valued markets. Strain improvement is a strategy to increase productivities, hence reducing costs. In this work, we focus on microalgae selection: taking advantage of the natural biological variability of species to select variations based on desired characteristics. We focused on triacylglycerol (TAG), which have applications ranging from biodiesel to high-value omega-3 fatty-acids. Hence, we demonstrated a strategy to sort microalgae cells with increased TAG productivity. 1. We successfully identified sub-populations of cells with increased TAG productivity using Fluorescence assisted cell sorting (FACS). 2. We sequentially sorted cells after repeated cycles of N-starvation, resulting in five sorted populations (S1-S5). 3. The comparison between sorted and original populations showed that S5 had the highest TAG productivity [0.34 against 0.18 g l(-1) day(-1) (original), continuous light]. 4. Original and S5 were compared in lab-scale reactors under simulated summer conditions confirming the increased TAG productivity of S5 (0.4 against 0.2 g l(-1) day(-1)). Biomass composition analyses showed that S5 produced more biomass under N-starvation because of an increase only in TAG content and, flow cytometry showed that our selection removed cells with lower efficiency in producing TAGs. All combined, our results present a successful strategy to improve the TAG productivity of Chlorococcum littorale, without resourcing to genetic manipulation or random mutagenesis. Additionally, the improved TAG productivity of S5 was confirmed under simulated summer conditions, highlighting the industrial potential of S5 for microalgal TAG production.

  1. Estimating milk yield and value losses from increased somatic cell count on US dairy farms.

    Science.gov (United States)

    Hadrich, J C; Wolf, C A; Lombard, J; Dolak, T M

    2018-04-01

    Milk loss due to increased somatic cell counts (SCC) results in economic losses for dairy producers. This research uses 10 mo of consecutive dairy herd improvement data from 2013 and 2014 to estimate milk yield loss using SCC as a proxy for clinical and subclinical mastitis. A fixed effects regression was used to examine factors that affected milk yield while controlling for herd-level management. Breed, milking frequency, days in milk, seasonality, SCC, cumulative months with SCC greater than 100,000 cells/mL, lactation, and herd size were variables included in the regression analysis. The cumulative months with SCC above a threshold was included as a proxy for chronic mastitis. Milk yield loss increased as the number of test days with SCC ≥100,000 cells/mL increased. Results from the regression were used to estimate a monetary value of milk loss related to SCC as a function of cow and operation related explanatory variables for a representative dairy cow. The largest losses occurred from increased cumulative test days with a SCC ≥100,000 cells/mL, with daily losses of $1.20/cow per day in the first month to $2.06/cow per day in mo 10. Results demonstrate the importance of including the duration of months above a threshold SCC when estimating milk yield losses. Cows with chronic mastitis, measured by increased consecutive test days with SCC ≥100,000 cells/mL, resulted in higher milk losses than cows with a new infection. This provides farm managers with a method to evaluate the trade-off between treatment and culling decisions as it relates to mastitis control and early detection. Copyright © 2018 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

  2. Increased cell survival by inhibition of BRCA1 using an antisense approach in an estrogen responsive ovarian carcinoma cell line

    International Nuclear Information System (INIS)

    Annab, Lois A; Hawkins, Rebecca E; Solomon, Greg; Barrett, J Carl; Afshari, Cynthia A

    2000-01-01

    -day period (Fig. 2a). The BRCA1 antisense population also exhibited a threefold to sixfold increase in cell growth compared with control cells in the presence of estrogen treatment. BG-1 BRCA1 antisense clones demonstrated a similar response to pooled population studies, enhanced growth with estrogen, and failure to die upon estrogen depletion (Fig. 2b). The BRCA1 antisense clones were further examined for other associated tumorigenic properties. All of the antisense clones were able to form colonies in soft agar (2-23 colonies per 10 4 cells plated; data not shown), whereas control clones were deficient in their ability to form colonies (0-0.8 colonies per 10 4 cells plated). Table 1 shows, in the presence of estrogen, the clone with the lowest levels of BRCA1 (AS-4) produced significantly more colonies (133 ± 17.9 colonies per 10 4 cells plated) than the control clone (NEO; 6 ± 3.1 colonies per 10 4 cells plated). Clones AS-4 and NEO were also injected with matrigel subcutaneously into ovariectomized athymic mice. Almost twice as many sites were positive for the AS-4 clone (14 out of 14) as for the NEO clone (eight out of 14) 42 days after injection. In addition, BRCA1 antisense tumors averaged twice the size of control tumors. The BRCA1 reduced cells also formed tumors with half the latency of control cells in the presence of implanted estrogen (11 days versus 21 days until tumor formation). The present studies show that reduction in BRCA1 levels, using an antisense retroviral vector in the estrogen dependent BG-1 ovarian carcinoma cell line, contributes to confirmation of the hypothesis that BRCA1 plays a pivotal role in the balance between cell death and cell proliferation. BRCA1 RNA and protein levels were successfully reduced in populations and isolated clones of antisense infected BG-1 cells. Decreased BRCA1 levels rescued the BG-1 cells from growth arrest or cell death in adverse growth conditions in monolayer or soft agar conditions. Furthermore, a BRCA1

  3. The F309S mutation increases factor VIII secretion in human cell line

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    Daianne Maciely Carvalho Fantacini

    2016-06-01

    Full Text Available ABSTRACT OBJECTIVES: The capacity of a human cell line to secrete recombinant factor VIII with a F309S point mutation was investigated, as was the effect of the addition of chemical chaperones (betaine and sodium-4-phenylbutyrate on the secretion of factor VIII. METHODS: This work used a vector with a F309S mutation in the A1 domain to investigate FVIII production in the HEK 293 human cell line. Factor VIII activity was measured by chromogenic assay. Furthermore, the effects of chemical drugs on the culture were evaluated. RESULTS: The addition of the F309S mutation to a previously described FVIII variant increased FVIII secretion by 4.5 fold. Moreover, the addition of betaine or sodium-4-phenylbutyrate increased the secretion rate of FVIIIΔB proteins in HEK 293 cells, but the same effect was not seen for FVIIIΔB-F309S indicating that all the recombinant protein produced had been efficiently secreted. CONCLUSION: Bioengineering factor VIII expressed in human cells may lead to an efficient production of recombinant factor VIII and contribute toward low-cost coagulation factor replacement therapy for hemophilia A. FVIII-F309S produced in human cells can be effective in vivo.

  4. Functional Imaging of Proteolysis: Stromal and Inflammatory Cells Increase Tumor Proteolysis

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    Mansoureh Sameni

    2003-07-01

    Full Text Available The underlying basement membrane is degraded during progression of breast and colon carcinoma. Thus, we imaged degradation of a quenched fluorescent derivative of basement membrane type IV collagen (DQ-collagen IV by living human breast and colon tumor spheroids. Proteolysis of DQ-collagen IV by HCT 116 and HKh-2 human colon tumor spheroids was both intracellular and pericellular. In contrast, proteolysis of DQ-collagen IV by BT20 human breast tumor spheroids was pericellular. As stromal elements can contribute to proteolytic activities associated with tumors, we also examined degradation of DQ-collagen IV by human monocytes/macrophages and colon and breast fibroblasts. Fibroblasts themselves exhibited a modest amount of pericellular degradation. Degradation was increased 4–17-fold in cocultures of fibroblasts and tumor cells as compared to either cell type alone. Inhibitors of matrix metalloproteinases, plasmin, and the cysteine protease, cathepsin B, all reduced degradation in the cocultures. Monocytes did not degrade DQ-collagen IV; however, macrophages degraded DQ-collagen IV intracellularly. In coculture of tumor cells, fibroblasts, and macrophages, degradation of DQ-collagen IV was further increased. Imaging of living tumor and stromal cells has, thus, allowed us to establish that tumor proteolysis occurs pericellularly and intracellularly and that tumor, stromal, and inflammatory cells all contribute to degradative processes.

  5. Increased number of mast cells in the dermis in actinic keratosis lesions effectively treated with imiquimod.

    Science.gov (United States)

    Oyama, Satomi; Funasaka, Yoko; Tsuchiya, Shin-Ichi; Kawana, Seiji; Saeki, Hidehisa

    2017-08-01

    Actinic keratosis (AK) is a cutaneous cancer in situ which develops as a result of excessive exposure to ultraviolet (UV). Toll-like receptor (TLR)7 agonist imiquimod is a topical immune response modifier and is effective for the treatment of non-melanoma skin cancers. Recently, the diagnostic role of the dermatoscope has been reported in the course of treatment of AK. In addition, mast cells are now considered to contribute to both the innate and adaptive immune systems in topical imiquimod therapy. We assessed the effect of imiquimod treatment by dermatoscopic and immunohistochemical findings in 14 patients with a total of 21 AK lesions. With the dermatoscope, though the mean erythema score was not significantly different between the cured lesions and the unresponsive lesions, the erythema/red pseudo-network ("strawberry") pattern was decreased significantly in the cured lesions. By immunohistochemistry, the number of Ki-67-positive proliferative cells in the epidermis was decreased and that of CD117-positive mast cells in the dermis was increased in the responding lesions. To the best of our knowledge, this is the first study demonstrating that the number of mast cells in the dermis was increased in AK lesions effectively treated with imiquimod. Our present result suggests that mast cells may contribute an antitumor effect in human skin treated with topical imiquimod. © 2017 Japanese Dermatological Association.

  6. AM fungal exudates activate MAP kinases in plant cells in dependence from cytosolic Ca(2+) increase.

    Science.gov (United States)

    Francia, Doriana; Chiltz, Annick; Lo Schiavo, Fiorella; Pugin, Alain; Bonfante, Paola; Cardinale, Francesca

    2011-09-01

    The molecular dialogue occurring prior to direct contact between the fungal and plant partners of arbuscular-mycorrhizal (AM) symbioses begins with the release of fungal elicitors, so far only partially identified chemically, which can activate specific signaling pathways in the host plant. We show here that the activation of MAPK is also induced by exudates of germinating spores of Gigaspora margarita in cultured cells of the non-leguminous species tobacco (Nicotiana tabacum), as well as in those of the model legume Lotus japonicus. MAPK activity peaked about 15 min after the exposure of the host cells to the fungal exudates (FE). FE were also responsible for a rapid and transient increase in free cytosolic Ca(2+) in Nicotiana plumbaginifolia and tobacco cells, and pre-treatment with a Ca(2+)-channel blocker (La(3+)) showed that in these cells, MAPK activation was dependent on the cytosolic Ca(2+) increase. A partial dependence of MAPK activity on the common Sym pathway could be demonstrated for a cell line of L. japonicus defective for LjSym4 and hence unable to establish an AM symbiosis. Our results show that MAPK activation is triggered by an FE-induced cytosolic Ca(2+) transient, and that a Sym genetic determinant acts to modulate the intensity and duration of this activity. Copyright © 2011 Elsevier Masson SAS. All rights reserved.

  7. Enhanced caveolin-1 expression increases migration, anchorage-independent growth and invasion of endometrial adenocarcinoma cells

    International Nuclear Information System (INIS)

    Diaz-Valdivia, Natalia; Bravo, Denisse; Huerta, Hernán; Henriquez, Soledad; Gabler, Fernando; Vega, Margarita; Romero, Carmen; Calderon, Claudia; Owen, Gareth I.; Leyton, Lisette; Quest, Andrew F. G.

    2015-01-01

    Caveolin-1 (CAV1) has been implicated both in tumor suppression and progression, whereby the specific role appears to be context dependent. Endometrial cancer is one of the most common malignancies of the female genital tract; however, little is known about the role of CAV1 in this disease. Here, we first determined by immunohistochemistry CAV1 protein levels in normal proliferative human endometrium and endometrial tumor samples. Then using two endometrial cancer cell lines (ECC: Ishikawa and Hec-1A) we evaluated mRNA and protein levels of CAV1 by real time qPCR and Western blot analysis, respectively. The role of CAV1 expression in ECC malignancy was further studied by either inducing its expression in endometrial cancer cells with the tumor promotor 12-O-tetradecanoyl-phorbol-13-acetate (4β-TPA) or decreasing expression using short-hairpin RNA constructs, and then evaluating the effects of these changes on ECC proliferation, transmigration, matrigel invasion, and colony formation in soft agar. Immunohistochemical analysis of endometrial epithelia revealed that substantially higher levels of CAV1 were present in endometrial tumors than the normal proliferative epithelium. Also, in Ishikawa and Hec-1A endometrial cancer cells CAV1 expression was readily detectable. Upon treatment with 4β-TPA CAV1 levels increased and coincided with augmented cell transmigration, matrigel invasion, as well as colony formation in soft agar. Reduction of CAV1 expression using short-hairpin RNA constructs ablated these effects in both cell types whether treated or not with 4β-TPA. Alternatively, CAV1 expression appeared not to modulate significantly proliferation of these cells. Our study shows that elevated CAV1, observed in patients with endometrial cancer, is linked to enhanced malignancy of endometrial cancer cells, as evidenced by increased migration, invasion and anchorage-independent growth. The online version of this article (doi:10.1186/s12885-015-1477-5) contains

  8. Loss of P53 Function in Colon Cancer Cells Results in Increased Phosphocholine and Total Choline

    Directory of Open Access Journals (Sweden)

    Noriko Mori

    2004-10-01

    Full Text Available Mutations in the p53 gene are the most frequently observed genetic lesions in human cancers. Human cancers that contain a p53 mutation are more aggressive, more apt to metastasize, and more often fatal. p53 controls numerous downstream targets that can influence various outcomes such as apoptosis, growth arrest, and DNA repair. Based on previous observations using 1H magnetic resonance spectroscopy (MRS, we have identified choline phospholipid metabolite intensities typical of increased malignancy. Here we have used 1H MRS to characterize the choline phospholipid metabolite levels of p53+/+ and p53−/– cells, and demonstrated that loss of p53 function results in increased phosphocholine and total choline. These data suggest that the increased malignancy of cancer cells resulting from loss of p53 may be mediated, in part, through the choline phospholipid pathway.

  9. Net haemoglobin increase from reinfusion of refrigerated vs. frozen red blood cells after autologous blood transfusions

    DEFF Research Database (Denmark)

    Ashenden, M; Mørkeberg, Jakob Sehested

    2011-01-01

    BACKGROUND AND OBJECTIVES  Two main blood storage procedures can be used for storing red blood cells: refrigeration and freezing. Nevertheless, the efficiency of these procedures measured as the increase in haemoglobin after reinfusion compared with baseline has never been examined. The main...... objective was to examine which storage procedure yielded the largest increase in circulating haemoglobin after reinfusion compared to baseline. MATERIALS AND METHODS  Equal volumes of blood from 15 men were withdrawn and stored either frozen or refrigerated as packed red blood cells. Serial measures...... of circulating haemoglobin by carbon monoxide rebreathing provided an opportunity to monitor recovery from anaemia, as well as the net increase in circulating haemoglobin after transfusion. RESULTS  The post-thaw yield of haemoglobin in the bags was 72% after refrigerated storage compared with only 52% after...

  10. Sarcosine induces increase in HER2/neu expression in androgen-dependent prostate cancer cells

    DEFF Research Database (Denmark)

    Dahl, Malin; Bouchelouche, Pierre; Kramer-Marek, Gabriela

    2011-01-01

    Increasing evidence suggests that Human epidermal growth factor receptor 2 (HER2/neu) is involved in progression of prostate cancer. Recently, sarcosine was reported to be highly increased during prostate cancer progression, and exogenous sarcosine induces an invasive phenotype in benign prostate...... epithelial cells. The aim of this work was to investigate the effect of sarcosine on HER2/neu expression in prostate cancer cell lines LNCaP (androgen dependent), PC-3 and DU145 (both androgen independent). Relative amounts of HER2/neu and androgen receptor (AR) transcripts were determined using RT...... that sarcosine is involved in the regulation of the oncoprotein HER2/neu. Thus, sarcosine may induce prostate cancer progression by increased HER2/neu expression. However, detailed information on cellular mechanisms remains to be elucidated....

  11. Dragon enhances BMP signaling and increases transepithelial resistance in kidney epithelial cells.

    Science.gov (United States)

    Xia, Yin; Babitt, Jodie L; Bouley, Richard; Zhang, Ying; Da Silva, Nicolas; Chen, Shanzhuo; Zhuang, Zhenjie; Samad, Tarek A; Brenner, Gary J; Anderson, Jennifer L; Hong, Charles C; Schneyer, Alan L; Brown, Dennis; Lin, Herbert Y

    2010-04-01

    The neuronal adhesion protein Dragon acts as a bone morphogenetic protein (BMP) coreceptor that enhances BMP signaling. Given the importance of BMP signaling in nephrogenesis and its putative role in the response to injury in the adult kidney, we studied the localization and function of Dragon in the kidney. We observed that Dragon localized predominantly to the apical surfaces of tubular epithelial cells in the thick ascending limbs, distal convoluted tubules, and collecting ducts of mice. Dragon expression was weak in the proximal tubules and glomeruli. In mouse inner medullary collecting duct (mIMCD3) cells, Dragon generated BMP signals in a ligand-dependent manner, and BMP4 is the predominant endogenous ligand for the Dragon coreceptor. In mIMCD3 cells, BMP4 normally signaled through BMPRII, but Dragon enhanced its signaling through the BMP type II receptor ActRIIA. Dragon and BMP4 increased transepithelial resistance (TER) through the Smad1/5/8 pathway. In epithelial cells isolated from the proximal tubule and intercalated cells of collecting ducts, we observed coexpression of ActRIIA, Dragon, and BMP4 but not BMPRII. Taken together, these results suggest that Dragon may enhance BMP signaling in renal tubular epithelial cells and maintain normal renal physiology.

  12. Adipose progenitor cells increase fibronectin matrix strain and unfolding in breast tumors

    Science.gov (United States)

    Chandler, E. M.; Saunders, M. P.; Yoon, C. J.; Gourdon, D.; Fischbach, C.

    2011-02-01

    Increased stiffness represents a hallmark of breast cancer that has been attributed to the altered physicochemical properties of the extracellular matrix (ECM). However, the role of fibronectin (Fn) in modulating the composition and mechanical properties of the tumor-associated ECM remains unclear. We have utilized a combination of biochemical and physical science tools to evaluate whether paracrine signaling between breast cancer cells and adipose progenitor cells regulates Fn matrix assembly and stiffness enhancement in the tumor stroma. In particular, we utilized fluorescence resonance energy transfer imaging to map the molecular conformation and stiffness of Fn that has been assembled by 3T3-L1 preadipocytes in response to conditioned media from MDA-MB231 breast cancer cells. Our results reveal that soluble factors secreted by tumor cells promote Fn expression, unfolding, and stiffening by adipose progenitor cells and that transforming growth factor-β serves as a soluble cue underlying these changes. In vivo experiments using orthotopic co-transplantation of primary human adipose-derived stem cells and MDA-MB231 into SCID mice support the pathological relevance of our results. Insights gained by these studies advance our understanding of the role of Fn in mammary tumorigenesis and may ultimately lead to improved anti-cancer therapies.

  13. Adipose progenitor cells increase fibronectin matrix strain and unfolding in breast tumors

    International Nuclear Information System (INIS)

    Chandler, E M; Saunders, M P; Yoon, C J; Fischbach, C; Gourdon, D

    2011-01-01

    Increased stiffness represents a hallmark of breast cancer that has been attributed to the altered physicochemical properties of the extracellular matrix (ECM). However, the role of fibronectin (Fn) in modulating the composition and mechanical properties of the tumor-associated ECM remains unclear. We have utilized a combination of biochemical and physical science tools to evaluate whether paracrine signaling between breast cancer cells and adipose progenitor cells regulates Fn matrix assembly and stiffness enhancement in the tumor stroma. In particular, we utilized fluorescence resonance energy transfer imaging to map the molecular conformation and stiffness of Fn that has been assembled by 3T3-L1 preadipocytes in response to conditioned media from MDA-MB231 breast cancer cells. Our results reveal that soluble factors secreted by tumor cells promote Fn expression, unfolding, and stiffening by adipose progenitor cells and that transforming growth factor-β serves as a soluble cue underlying these changes. In vivo experiments using orthotopic co-transplantation of primary human adipose-derived stem cells and MDA-MB231 into SCID mice support the pathological relevance of our results. Insights gained by these studies advance our understanding of the role of Fn in mammary tumorigenesis and may ultimately lead to improved anti-cancer therapies

  14. Increased number of IgG4-positive plasma cells in chronic rhinosinusitis.

    Science.gov (United States)

    Ohno, Keiko; Kimura, Yurika; Matsuda, Yoko; Takahashi, Masatoki; Honjyou, Motomu; Arai, Tomio; Tsutsumi, Takeshi

    2017-02-01

    High levels of IgG4-positive plasma cells were observed in tissue samples from ∼30% of patients with chronic rhinosinusitis who satisfied the comprehensive diagnostic criteria for IgG4-related disease. Detection of increased numbers of IgG4-positive plasma cells in the nasal cavity or paranasal sinuses might not be sufficient to make a diagnosis of IgG4-related rhinosinusitis, and a comprehensive evaluation is required. This study aimed to clarify the clinicopathological characteristics of IgG4-positive plasma cells in patients with chronic rhinosinusitis. This study examined nasal mucosal specimens from 35 patients and assigned them to high-IgG4 and low-IgG4 groups based on infiltration of IgG4-positive plasma cells. It compared the pathological characteristics of the two groups, including the presence of fibrosis, phlebitis, hyperplasia of the nasal glands and infiltration of inflammatory cells. No cases of chronic rhinosinusitis showed storiform fibrosis or obliterative phlebitis. The mean number of IgG4-positive plasma cells in samples from all patients was 29.8 ± 40.3/high-power field. Eleven of the 35 cases (31.4%) were classified as high-IgG4. Hyperplasia of the nasal glands was observed significantly more frequently in the high-IgG4 group than in the low-IgG4 group (p = .03).

  15. Bone impairment in phenylketonuria is characterized by circulating osteoclast precursors and activated T cell increase.

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    Ilaria Roato

    Full Text Available BACKGROUND: Phenylketonuria (PKU is a rare inborn error of metabolism often complicated by a progressive bone impairment of uncertain etiology, as documented by both ionizing and non- ionizing techniques. METHODOLOGY: Peripheral blood mononuclear cell (PBMC cultures were performed to study osteoclastogenesis, in the presence or absence of recombinant human monocyte-colony stimulating factor (M-CSF and receptor activator of NFκB ligand (RANKL. Flow cytometry was utilized to analyze osteoclast precursors (OCPs and T cell phenotype. Tumour necrosis factor α (TNF-α, RANKL and osteoprotegerin (OPG were quantified in cell culture supernatants by ELISA. The effects of RANKFc and anti-TNF-α antibodies were also investigated to determine their ability to inhibit osteoclastogenesis. In addition, bone conditions and phenylalanine levels in PKU patients were clinically evaluated. PRINCIPAL FINDINGS: Several in vitro studies in PKU patients' cells identified a potential mechanism of bone formation inhibition commonly associated with this disorder. First, PKU patients disclosed an increased osteoclastogenesis compared to healthy controls, both in unstimulated and M-CSF/RANKL stimulated PBMC cultures. OCPs and the measured RANKL/OPG ratio were higher in PKU patients compared to healthy controls. The addition of specific antagonist RANKFc caused osteoclastogenesis inhibition, whereas anti-TNF-α failed to have this effect. Among PBMCs isolated from PKU patients, activated T cells, expressing CD69, CD25 and RANKL were identified. Confirmatory in vivo studies support this proposed model. These in vivo studies included the analysis of osteoclastogenesis in PKU patients, which demonstrated an inverse relation to bone condition assessed by phalangeal Quantitative Ultrasound (QUS. This was also directly related to non-compliance to therapeutic diet reflected by hyperphenylalaninemia. CONCLUSIONS: Our results indicate that PKU spontaneous osteoclastogenesis

  16. Tumor cell-macrophage interactions increase angiogenesis through secretion of EMMPRIN

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    Bat-Chen eAmit-Cohen

    2013-07-01

    Full Text Available Tumor macrophages are generally considered to be alternatively/M2 activated to induce secretion of pro-angiogenic factors such as VEGF and MMPs. EMMPRIN (CD147, basigin is overexpressed in many tumor types, and has been shown to induce fibroblasts and endothelial cell expression of MMPs and VEGF. We first show that tumor cell interactions with macrophages resulted in increased expression of EMMPRIN and induction of MMP-9 and VEGF. Human A498 renal carcinoma or MCF-7 breast carcinoma cell lines were co-cultured with the U937 monocytic-like cell line in the presence of TNFalpha (1 ng/ml. Membranal EMMPRIN expression was increased in the co-cultures (by 3-4 folds, p<0.01, as was the secretion of MMP-9 and VEGF (by 2-5 folds for both MMP-9 and VEGF, p<0.01, relative to the single cultures with TNFalpha. Investigating the regulatory mechanisms, we show that EMMPRIN was post-translationally regulated by miR-146a, as no change was observed in the tumoral expression of EMMPRIN mRNA during co-culture, expression of miR-146a was increased and its neutralization by its antagomir inhibited EMMPRIN expression. The secretion of EMMPRIN was also enhanced (by 2-3 folds, p<0.05, only in the A498 co-culture via shedding off of the membranal protein by a serine protease that is yet to be identified, as demonstrated by the use of wide range protease inhibitors. Finally, soluble EMMPRIN enhanced monocytic secretion of MMP-9 and VEGF, as inhibition of its expression levels by neutralizing anti-EMMPRIN or siRNA in the tumor cells lead to subsequent decreased induction of these two pro-angiogenic proteins. These results reveal a mechanism whereby tumor cell-macrophage interactions promote angiogenesis via an EMMPRIN-mediated pathway.

  17. Benfotiamine prevents increased β-amyloid production in HEK cells induced by high glucose.

    Science.gov (United States)

    Sun, Xiao-Jing; Zhao, Lei; Zhao, Na; Pan, Xiao-Li; Fei, Guo-Qiang; Jin, Li-Rong; Zhong, Chun-Jiu

    2012-10-01

    To determine whether high glucose enhances β-amyloid (Aβ) production in HEK293 Swedish mutant (APPsw) cells with Aβ precursor protein (APP) overexpression, and whether under this condition benfotiamine reduces the increased Aβ production. HEK293 APPsw cells were cultured with different concentrations of glucose for different times. The Aβ content in the supernatant was determined by ELISA. To investigate the mechanism by which benfotiamine reduced Aβ production, glycogen synthase kinase-3 (GSK-3) activity and expression were measured after the cells were cultured with 5.5 g/L glucose for 12 h. With 1.0, 3.0, 4.5, 5.5, 6.5, 7.5, 8.5, or 10.5 g/L glucose, Aβ production by HEK293 APPsw cells was highest in the presence of 5.5 g/L glucose for 6 and 12 h. The difference in Aβ content between 5.5 and 1.0 g/L was most marked after incubation for 12 h. Benfotiamine at 20 and 40 μg/mL significantly reduced Aβ production in cells incubated with 5.5 g/L glucose for 12 h. Moreover, 40 μg/mL benfotiamine significantly enhanced the ratio of phosphorylated GSK-3 to total GSK-3, together with consistent down-regulation of GSK-3 activity. High glucose increases Aβ production by HEK293 APPsw cells while benfotiamine prevents this increase. This is correlated with the modulation of GSK-3 activity.

  18. Oral Administration of Probiotics Increases Paneth Cells and Intestinal Antimicrobial Activity

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    Silvia I. Cazorla

    2018-04-01

    Full Text Available The huge amount of intestinal bacteria represents a continuing threat to the intestinal barrier. To meet this challenge, gut epithelial cells produce antimicrobial peptides (AMP that act at the forefront of innate immunity. We explore whether this antimicrobial activity and Paneth cells, the main intestinal cell responsible of AMP production, are influenced by probiotics administration, to avoid the imbalance of intestinal microbiota and preserve intestinal barrier. Administration of Lactobacillus casei CRL 431 (Lc 431 and L. paracasei CNCM I-1518 (Lp 1518 to 42 days old mice, increases the number of Paneth cells on small intestine, and the antimicrobial activity against the pathogens Staphylococcus aureus and Salmonella Typhimurium in the intestinal fluids. Specifically, strong damage of the bacterial cell with leakage of cytoplasmic content, and cellular fragmentation were observed in S. Typhimurium and S. aureus. Even more important, probiotics increase the antimicrobial activity of the intestinal fluids at the different ages, from weaning (21 days old to old age (180 days old. Intestinal antimicrobial activity stimulated by oral probiotics, do not influence significantly the composition of total anaerobic bacteria, lactobacilli and enterobacteria in the large intestine, at any age analyzed. This result, together with the antimicrobial activity observed against the same probiotic bacteria; endorse the regular consumption of probiotics without adverse effect on the intestinal homeostasis in healthy individuals. We demonstrate that oral probiotics increase intestinal antimicrobial activity and Paneth cells in order to strengthen epithelial barrier against pathogens. This effect would be another important mechanism by which probiotics protect the host mainly against infectious diseases.

  19. Acidosis increases the susceptibility of respiratory epithelial cells to Pseudomonas aeruginosa-induced cytotoxicity.

    Science.gov (United States)

    Torres, Iviana M; Demirdjian, Sally; Vargas, Jennifer; Goodale, Britton C; Berwin, Brent

    2017-07-01

    Bacterial infection can lead to acidosis of the local microenvironment, which is believed to exacerbate disease pathogenesis; however, the mechanisms by which changes in pH alter disease progression are poorly understood. We test the hypothesis that acidosis enhances respiratory epithelial cell death in response to infection with Pseudomonas aeruginosa Our findings support the idea that acidosis in the context of P. aeruginosa infection results in increased epithelial cell cytotoxicity due to ExoU intoxication. Importantly, enforced maintenance of neutral pH during P. aeruginosa infection demonstrates that cytotoxicity is dependent on the acidosis. Investigation of the underlying mechanisms revealed that host cell cytotoxicity correlated with increased bacterial survival during an acidic infection that was due to reduced bactericidal activity of host-derived antimicrobial peptides. These findings extend previous reports that the activities of antimicrobial peptides are pH-dependent and provide novel insights into the consequences of acidosis on infection-derived pathology. Therefore, this report provides the first evidence that physiological levels of acidosis increase the susceptibility of epithelial cells to acute Pseudomonas infection and demonstrates the benefit of maintaining pH homeostasis during a bacterial infection. Copyright © 2017 the American Physiological Society.

  20. Increased response to oxidative stress challenge of nano-copper-induced apoptosis in mesangial cells

    International Nuclear Information System (INIS)

    Xu, Pengjuan; Li, Zhigui; Zhang, Xiaochen; Yang, Zhuo

    2014-01-01

    Recently, many studies reported that nanosized copper particles (nano-Cu, the particle size was around 15–30 nm), one of the nanometer materials, could induce nephrotoxicity. To detect the effect of nano-Cu on mesangial cells (MCs), and investigate the underlying mechanism, MCs were treated with different concentrations of nano-Cu (1, 10, and 30 μg/mL) to determine the oxidative stress and apoptotic changes. It was revealed that nano-Cu could induce a decreased viability in MCs together with a significant increase in the number of apoptotic cells by using cell counting kit-8 assay and flow cytometry. The apoptotic morphological changes induced by nano-Cu in MCs were demonstrated by Hochest33342 staining. Results showed that nano-Cu induced the nuclear fragmentation in MCs. Meanwhile, nano-Cu significantly increased the levels of reactive oxygen species, especially increased the levels of H 2 O 2 . It also decreased the activity of total SOD enzyme. In addition, when pre-treated with N-(2-mercaptopropionyl)-glycine, the cell apoptosis induced by nano-Cu was significantly decreased. These results suggest that oxidative stress plays an important role in the nano-Cu toxicity in MCs, which may be the main mechanism of nano-Cu-induced nephrotoxicity

  1. Increase in intracellular PGE2 induces apoptosis in Bax-expressing colon cancer cell

    International Nuclear Information System (INIS)

    Lalier, Lisenn; Pedelaborde, François; Braud, Christophe; Menanteau, Jean; M Vallette, François; Olivier, Christophe

    2011-01-01

    NSAIDs exhibit protective properties towards some cancers, especially colon cancer. Yet, it is not clear how they play their protective role. PGE 2 is generally shown as the only target of the NSAIDs anticancerous activity. However, PGE 2 known targets become more and more manifold, considering both the molecular pathways involved and the target cells in the tumour. The role of PGE 2 in tumour progression thus appears complex and multipurpose. To gain understanding into the role of PGE 2 in colon cancer, we focused on the activity of PGE 2 in apoptosis in colon cancer cell lines. We observed that an increase in intracellular PGE 2 induced an apoptotic cell death, which was dependent on the expression of the proapoptotic protein Bax. This increase was induced by increasing PGE 2 intracellular concentration, either by PGE 2 microinjection or by the pharmacological inhibition of PGE 2 exportation and enzymatic degradation. We present here a new sight onto PGE 2 in colon cancer cells opening the way to a new prospective therapeutic strategy in cancer, alternative to NSAIDs

  2. Mesenchymal stem cells restore frataxin expression and increase hydrogen peroxide scavenging enzymes in Friedreich ataxia fibroblasts.

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    Kevin Kemp

    Full Text Available Dramatic advances in recent decades in understanding the genetics of Friedreich ataxia (FRDA--a GAA triplet expansion causing greatly reduced expression of the mitochondrial protein frataxin--have thus far yielded no therapeutic dividend, since there remain no effective treatments that prevent or even slow the inevitable progressive disability in affected individuals. Clinical interventions that restore frataxin expression are attractive therapeutic approaches, as, in theory, it may be possible to re-establish normal function in frataxin deficient cells if frataxin levels are increased above a specific threshold. With this in mind several drugs and cytokines have been tested for their ability to increase frataxin levels. Cell transplantation strategies may provide an alternative approach to this therapeutic aim, and may also offer more widespread cellular protective roles in FRDA. Here we show a direct link between frataxin expression in fibroblasts derived from FRDA patients with both decreased expression of hydrogen peroxide scavenging enzymes and increased sensitivity to hydrogen peroxide-mediated toxicity. We demonstrate that normal human mesenchymal stem cells (MSCs induce both an increase in frataxin gene and protein expression in FRDA fibroblasts via secretion of soluble factors. Finally, we show that exposure to factors produced by human MSCs increases resistance to hydrogen peroxide-mediated toxicity in FRDA fibroblasts through, at least in part, restoring the expression of the hydrogen peroxide scavenging enzymes catalase and glutathione peroxidase 1. These findings suggest, for the first time, that stem cells may increase frataxin levels in FRDA and transplantation of MSCs may offer an effective treatment for these patients.

  3. ATP Release from Chemotherapy-Treated Dying Leukemia Cells Elicits an Immune Suppressive Effect by Increasing Regulatory T Cells and Tolerogenic Dendritic Cells.

    Science.gov (United States)

    Lecciso, Mariangela; Ocadlikova, Darina; Sangaletti, Sabina; Trabanelli, Sara; De Marchi, Elena; Orioli, Elisa; Pegoraro, Anna; Portararo, Paola; Jandus, Camilla; Bontadini, Andrea; Redavid, Annarita; Salvestrini, Valentina; Romero, Pedro; Colombo, Mario P; Di Virgilio, Francesco; Cavo, Michele; Adinolfi, Elena; Curti, Antonio

    2017-01-01

    Chemotherapy-induced immunogenic cell death can favor dendritic cell (DC) cross-priming of tumor-associated antigens for T cell activation thanks to the release of damage-associated molecular patterns, including ATP. Here, we tested the hypothesis that in acute myeloid leukemia (AML), ATP release, along with its well-known immune stimulatory effect, may also contribute to the generation of an immune suppressive microenvironment. In a cohort of AML patients, undergoing combined daunorubicin and cytarabine chemotherapy, a population of T regulatory cells (Tregs) with suppressive phenotype, expressing the immune checkpoint programmed cell death protein 1 (PD-1), was significantly increased. Moving from these results, initial in vitro data showed that daunorubicin was more effective than cytarabine in modulating DC function toward Tregs induction and such difference was correlated with the higher capacity of daunorubicin to induce ATP release from treated AML cells. DCs cultured with daunorubicin-treated AML cells upregulated indoleamine 2,3-dioxygenase 1 (IDO1), which induced anti-leukemia Tregs. These data were confirmed in vivo as daunorubicin-treated mice show an increase in extracellular ATP levels with increased number of Tregs, expressing PD-1 and IDO1 + CD39 + DCs. Notably, daunorubicin failed to induce Tregs and tolerogenic DCs in mice lacking the ATP receptor P2X7. Our data indicate that ATP release from chemotherapy-treated dying cells contributes to create an immune suppressive microenvironment in AML.

  4. ATP Release from Chemotherapy-Treated Dying Leukemia Cells Elicits an Immune Suppressive Effect by Increasing Regulatory T Cells and Tolerogenic Dendritic Cells

    Directory of Open Access Journals (Sweden)

    Mariangela Lecciso

    2017-12-01

    Full Text Available Chemotherapy-induced immunogenic cell death can favor dendritic cell (DC cross-priming of tumor-associated antigens for T cell activation thanks to the release of damage-associated molecular patterns, including ATP. Here, we tested the hypothesis that in acute myeloid leukemia (AML, ATP release, along with its well-known immune stimulatory effect, may also contribute to the generation of an immune suppressive microenvironment. In a cohort of AML patients, undergoing combined daunorubicin and cytarabine chemotherapy, a population of T regulatory cells (Tregs with suppressive phenotype, expressing the immune checkpoint programmed cell death protein 1 (PD-1, was significantly increased. Moving from these results, initial in vitro data showed that daunorubicin was more effective than cytarabine in modulating DC function toward Tregs induction and such difference was correlated with the higher capacity of daunorubicin to induce ATP release from treated AML cells. DCs cultured with daunorubicin-treated AML cells upregulated indoleamine 2,3-dioxygenase 1 (IDO1, which induced anti-leukemia Tregs. These data were confirmed in vivo as daunorubicin-treated mice show an increase in extracellular ATP levels with increased number of Tregs, expressing PD-1 and IDO1+CD39+ DCs. Notably, daunorubicin failed to induce Tregs and tolerogenic DCs in mice lacking the ATP receptor P2X7. Our data indicate that ATP release from chemotherapy-treated dying cells contributes to create an immune suppressive microenvironment in AML.

  5. The cell wall stress response in Aspergillus niger involves increased expression of the glutamine: Fructose-6-phosphate amidotransferase-encoding gene (gfaA) and increased deposition of chitin in the cell wall

    NARCIS (Netherlands)

    Ram, A.F.J.; Arentshorst, M.; Damveld, R.A.; Kuyk, P.A. van; Klis, F.M.; Hondel, C.A.M.J.J. van den

    2004-01-01

    Perturbation of cell wall synthesis in Saccharomyces cerevisiae, either by mutations in cell wall synthesis-related genes or by adding compounds that interfere with normal cell wall assembly, triggers a compensatory response to ensure cell wall integrity. This response includes an increase in chitin

  6. Increased sensitivity of p53-deficient cells to anticancer agents due to loss of Pms2

    Science.gov (United States)

    Fedier, A; Ruefenacht, U B; Schwarz, V A; Haller, U; Fink, D

    2002-01-01

    A large fraction of human tumours carries mutations in the p53 gene. p53 plays a central role in controlling cell cycle checkpoint regulation, DNA repair, transcription, and apoptosis upon genotoxic stress. Lack of p53 function impairs these cellular processes, and this may be the basis of resistance to chemotherapeutic regimens. By virtue of the involvement of DNA mismatch repair in modulating cytotoxic pathways in response to DNA damaging agents, we investigated the effects of loss of Pms2 on the sensitivity to a panel of widely used anticancer agents in E1A/Ha-Ras-transformed p53-null mouse fibroblasts either proficient or deficient in Pms2. We report that lack of the Pms2 gene is associated with an increased sensitivity, ranging from 2–6-fold, to some types of anticancer agents including the topoisomerase II poisons doxorubicin, etoposide and mitoxantrone, the platinum compounds cisplatin and oxaliplatin, the taxanes docetaxel and paclitaxel, and the antimetabolite gemcitabine. In contrast, no change in sensitivity was found after treatment with 5-fluorouracil. Cell cycle analysis revealed that both, Pms2-deficient and -proficient cells, retain the ability to arrest at the G2/M upon cisplatin treatment. The data indicate that the concomitant loss of Pms2 function chemosensitises p53-deficient cells to some types of anticancer agents, that Pms2 positively modulates cell survival by mechanisms independent of p53, and that increased cytotoxicity is paralleled by increased apoptosis. Tumour-targeted functional inhibition of Pms2 may be a valuable strategy for increasing the efficacy of anticancer agents in the treatment of p53-mutant cancers. British Journal of Cancer (2002) 87, 1027–1033. doi:10.1038/sj.bjc.6600599 www.bjcancer.com © 2002 Cancer Research UK PMID:12434296

  7. Increased frequency of single base substitutions in a population of transcripts expressed in cancer cells

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    Bianchetti Laurent

    2012-11-01

    Full Text Available Abstract Background Single Base Substitutions (SBS that alter transcripts expressed in cancer originate from somatic mutations. However, recent studies report SBS in transcripts that are not supported by the genomic DNA of tumor cells. Methods We used sequence based whole genome expression profiling, namely Long-SAGE (L-SAGE and Tag-seq (a combination of L-SAGE and deep sequencing, and computational methods to identify transcripts with greater SBS frequencies in cancer. Millions of tags produced by 40 healthy and 47 cancer L-SAGE experiments were compared to 1,959 Reference Tags (RT, i.e. tags matching the human genome exactly once. Similarly, tens of millions of tags produced by 7 healthy and 8 cancer Tag-seq experiments were compared to 8,572 RT. For each transcript, SBS frequencies in healthy and cancer cells were statistically tested for equality. Results In the L-SAGE and Tag-seq experiments, 372 and 4,289 transcripts respectively, showed greater SBS frequencies in cancer. Increased SBS frequencies could not be attributed to known Single Nucleotide Polymorphisms (SNP, catalogued somatic mutations or RNA-editing enzymes. Hypothesizing that Single Tags (ST, i.e. tags sequenced only once, were indicators of SBS, we observed that ST proportions were heterogeneously distributed across Embryonic Stem Cells (ESC, healthy differentiated and cancer cells. ESC had the lowest ST proportions, whereas cancer cells had the greatest. Finally, in a series of experiments carried out on a single patient at 1 healthy and 3 consecutive tumor stages, we could show that SBS frequencies increased during cancer progression. Conclusion If the mechanisms generating the base substitutions could be known, increased SBS frequency in transcripts would be a new useful biomarker of cancer. With the reduction of sequencing cost, sequence based whole genome expression profiling could be used to characterize increased SBS frequency in patient’s tumor and aid diagnostic.

  8. Increased frequency of single base substitutions in a population of transcripts expressed in cancer cells

    International Nuclear Information System (INIS)

    Bianchetti, Laurent; Kieffer, David; Féderkeil, Rémi; Poch, Olivier

    2012-01-01

    Single Base Substitutions (SBS) that alter transcripts expressed in cancer originate from somatic mutations. However, recent studies report SBS in transcripts that are not supported by the genomic DNA of tumor cells. We used sequence based whole genome expression profiling, namely Long-SAGE (L-SAGE) and Tag-seq (a combination of L-SAGE and deep sequencing), and computational methods to identify transcripts with greater SBS frequencies in cancer. Millions of tags produced by 40 healthy and 47 cancer L-SAGE experiments were compared to 1,959 Reference Tags (RT), i.e. tags matching the human genome exactly once. Similarly, tens of millions of tags produced by 7 healthy and 8 cancer Tag-seq experiments were compared to 8,572 RT. For each transcript, SBS frequencies in healthy and cancer cells were statistically tested for equality. In the L-SAGE and Tag-seq experiments, 372 and 4,289 transcripts respectively, showed greater SBS frequencies in cancer. Increased SBS frequencies could not be attributed to known Single Nucleotide Polymorphisms (SNP), catalogued somatic mutations or RNA-editing enzymes. Hypothesizing that Single Tags (ST), i.e. tags sequenced only once, were indicators of SBS, we observed that ST proportions were heterogeneously distributed across Embryonic Stem Cells (ESC), healthy differentiated and cancer cells. ESC had the lowest ST proportions, whereas cancer cells had the greatest. Finally, in a series of experiments carried out on a single patient at 1 healthy and 3 consecutive tumor stages, we could show that SBS frequencies increased during cancer progression. If the mechanisms generating the base substitutions could be known, increased SBS frequency in transcripts would be a new useful biomarker of cancer. With the reduction of sequencing cost, sequence based whole genome expression profiling could be used to characterize increased SBS frequency in patient’s tumor and aid diagnostic

  9. A virulence-associated filamentous bacteriophage of Neisseria meningitidis increases host-cell colonisation.

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    Emmanuelle Bille

    2017-07-01

    Full Text Available Neisseria meningitidis is a commensal of human nasopharynx. In some circumstances, this bacteria can invade the bloodstream and, after crossing the blood brain barrier, the meninges. A filamentous phage, designated MDAΦ for Meningococcal Disease Associated, has been associated with invasive disease. In this work we show that the prophage is not associated with a higher virulence during the bloodstream phase of the disease. However, looking at the interaction of N. meningitidis with epithelial cells, a step essential for colonization of the nasopharynx, we demonstrate that the presence of the prophage, via the production of viruses, increases colonization of encapsulated meningococci onto monolayers of epithelial cells. The analysis of the biomass covering the epithelial cells revealed that meningococci are bound to the apical surface of host cells by few layers of heavily piliated bacteria, whereas, in the upper layers, bacteria are non-piliated but surrounded by phage particles which (i form bundles of filaments, and/or (ii are in some places associated with bacteria. The latter are likely to correspond to growing bacteriophages during their extrusion through the outer membrane. These data suggest that, as the biomass increases, the loss of piliation in the upper layers of the biomass does not allow type IV pilus bacterial aggregation, but is compensated by a large production of phage particles that promote bacterial aggregation via the formation of bundles of phage filaments linked to the bacterial cell walls. We propose that MDAΦ by increasing bacterial colonization in the mucosa at the site-of-entry, increase the occurrence of diseases.

  10. Oxidized low density lipoprotein increases RANKL level in human vascular cells. Involvement of oxidative stress

    Energy Technology Data Exchange (ETDEWEB)

    Mazière, Cécile, E-mail: maziere.cecile@chu-amiens.fr [Biochemistry Laboratory, South Hospital University, René Laennec Avenue, Amiens 80000 (France); Salle, Valéry [Internal Medicine, North Hospital University, Place Victor Pauchet, Amiens 80000 (France); INSERM U1088 (EA 4292), SFR CAP-Santé (FED 4231), University of Picardie – Jules Verne (France); Gomila, Cathy; Mazière, Jean-Claude [Biochemistry Laboratory, South Hospital University, René Laennec Avenue, Amiens 80000 (France)

    2013-10-18

    Highlights: •Oxidized LDL enhances RANKL level in human smooth muscle cells. •The effect of OxLDL is mediated by the transcription factor NFAT. •UVA, H{sub 2}O{sub 2} and buthionine sulfoximine also increase RANKL level. •All these effects are observed in human fibroblasts and endothelial cells. -- Abstract: Receptor Activator of NFκB Ligand (RANKL) and its decoy receptor osteoprotegerin (OPG) have been shown to play a role not only in bone remodeling but also in inflammation, arterial calcification and atherosclerotic plaque rupture. In human smooth muscle cells, Cu{sup 2+}-oxidized LDL (CuLDL) 10–50 μg/ml increased reactive oxygen species (ROS) and RANKL level in a dose-dependent manner, whereas OPG level was not affected. The lipid extract of CuLDL reproduced the effects of the whole particle. Vivit, an inhibitor of the transcription factor NFAT, reduced the CuLDL-induced increase in RANKL, whereas PKA and NFκB inhibitors were ineffective. LDL oxidized by myeloperoxidase (MPO-LDL), or other pro-oxidant conditions such as ultraviolet A (UVA) irradiation, incubation with H{sub 2}O{sub 2} or with buthionine sulfoximine (BSO), an inhibitor of glutathione synthesis{sub ,} also induced an oxidative stress and enhanced RANKL level. The increase in RANKL in pro-oxidant conditions was also observed in fibroblasts and endothelial cells. Since RANKL is involved in myocardial inflammation, vascular calcification and plaque rupture, this study highlights a new mechanism whereby OxLDL might, by generation of an oxidative stress, exert a deleterious effect on different cell types of the arterial wall.

  11. Attenuation of G2 cell cycle checkpoint control in human tumor cells is associated with increased frequencies of unrejoined chromosome breaks but not increased cytotoxicity following radiation exposure

    International Nuclear Information System (INIS)

    Schwartz, J.L.; Cowan, J.; Grdina, D.J.

    1997-01-01

    The contribution of G 2 cell cycle checkpoint control to ionizing radiation responses was examined in ten human tumor cell lines. Most of the delay in cell cycle progression seen in the first cell cycle following radiation exposure was due to blocks in G 2 and there were large cell line-to-cell line variations in the length of the G 2 block. Longer delays were seen in cell lines that had mutations in p53. There was a highly significant inverse correlation between the length of G 2 delay and the frequency of unrejoined chromosome breaks seen as chromosome terminal deletions in mitosis, and observation that supports the hypothesis that the signal for G 2 delay in mammalian cells is an unrejoined chromosome break. There were also an inverse correlation between the length of G 2 delay and the level of chromosome aneuploidy in each cell line, suggesting that the G 2 and mitotic spindel checkpoints may be linked to each other. Attenuation in G 2 checkpoint control was not associated with alterations in either the frequency of induced chromosome rearrangements or cell survival following radiation exposure suggesting that chromosome rearrangements, the major radiation-induced lethal lesion in tumor cells, form before cells enters G 2 . Thus, agents that act solely to override G 2 arrest should produce little radiosensitization in human tumor cells

  12. Nicotine-evoked cytosolic Ca2+ increase and cell depolarization in capillary endothelial cells of the bovine adrenal medulla

    Directory of Open Access Journals (Sweden)

    RAÚL VINET

    2009-01-01

    Full Text Available Endothelial cells are directly involved in many functions of the cardiovascular system by regulating blood flow and blood pressure through Ca2+ dependent exocitosis of vasoactive compounds. Using the Ca2+ indicator Fluo-3 and the patch-clamp technique, we show that bovine adrenal medulla capillary endothelial cells (B AMCECs respond to acetylcholine (ACh with a cytosolic Ca2+ increase and depolarization of the membrane potential (20.3±0.9 mV; n=23. The increase in cytosolic Ca2+ induced by 10µM ACh was mimicked by the same concentration of nicotine but not by muscarine and was blocked by 100 µM of hexamethonium. On the other hand, the increase in cytosolic Ca2+ could be depressed by nifedipine (0.01 -100 µM or withdrawal of extracellular Ca2+. Taken together, these results give evidence for functional nicotinic receptors (nAChRs in capillary endothelial cells of the adrenal medulla. It suggests that nAChRs in B AMCECs may be involved in the regulation of the adrenal gland's microcirculation by depolarizing the membrane potential, leading to the opening of voltage-activated Ca2+ channels, influx of external Ca2+ and liberation of vasoactive compounds.

  13. AKT increases VEGF expression in tumor cells by transactivating the proximal VEGF promoter

    International Nuclear Information System (INIS)

    Pore, N.; Bernhard, E.J.; Shu, H.-K.; Li, B.; O'Rourke, D.M.; Maity, A.; Haas-Kogan, D.

    2003-01-01

    Vascular endothelial growth factor (VEGF) is overexpressed in many cancers including glioblastomas and may contribute to their growth. Epidermal growth factor receptor (EGFR) amplification and loss of PTEN, commonly found in glioblastomas leading to increase phosphatidylinositol-3-kinase (PI3K) activity and VEGF expression. In the current study we show that AKT, which is downstream of PI3K, regulates VEGF expression. U87MG human glioblastoma cells lack wildtype PTEN and express high levels of phosphorylated AKT. Over expression of AKT either by stable expression in immortalized human astrocytes or by transduction with adenovirus containing activated myristoylated AKT in SF188 glioblastoma cells increases VEGF expression. Moreover the elevation of angiogenesis by constitutively expressed AKT is further confirmed by in vivo matrigel plug assay in nude mice. The upregulation of VEGF by AKT is mediated through a region in the proximal promoter located between -88 and -70 (+1 is transcription start site). In transient transfection activity of a luciferase reporter containing the -88/+54 region of the VEGF promoter is increased by cotransfection with myristoylated AKT and downregulated by a dominant negative AKT expression vector. Mutation of the putative Sp1 binding sites located in the -88/-70 region we show that AKT acts through Sp1 to transactivate the VEGF promoter. Cotransfection of the VEGF promoter reporter with both Sp1 and myristoylated AKT expression vectors increases promoter activity to a greater extent than either Sp1 or Akt by itself. In vivo phosphate labeling of proteins reveals that AKT leads to increased Sp1 phosphorylation. Gel shift assays using a radio labeled probe corresponding to nucleotides -88 through -66 in the promoter show increased binding with nuclear extracts from cells transduced with adenovirus expressing myristoylated AKT. In conclusion, our results suggest that loss of PTEN leads to increased VEGF expression by increasing AKT

  14. Angiotensin II type 1 receptor blockers increase tolerance of cells to copper and cisplatin

    Directory of Open Access Journals (Sweden)

    Pieter Spincemaille

    2014-10-01

    Full Text Available The human pathology Wilson disease (WD is characterized by toxic copper (Cu accumulation in brain and liver, resulting in, among other indications, mitochondrial dysfunction and apoptosis of hepatocytes. In an effort to identify novel compounds that can alleviate Cu-induced toxicity, we screened the Pharmakon 1600 repositioning library using a Cu-toxicity yeast screen. We identified 2 members of the drug class of Angiotensin II Type 1 receptor blockers (ARBs that could increase yeast tolerance to Cu, namely Candesartan and Losartan. Subsequently, we show that specific ARBs can increase yeast tolerance to Cu and/or the chemotherapeutic agent cisplatin (Cp. The latter also induces mitochondrial dysfunction and apoptosis in mammalian cells. We further demonstrate that specific ARBs can prevent the prevalence of Cu-induced apoptotic markers in yeast, with Candesartan Cilexetil being the ARB which demonstrated most pronounced reduction of apoptosis-related markers. Next, we tested the sensitivity of a selection of yeast knockout mutants affected in detoxification of reactive oxygen species (ROS and Cu for Candesartan Cilexetil rescue in presence of Cu. These data indicate that Candesartan Cilexetil increases yeast tolerance to Cu irrespectively of major ROS-detoxifying proteins. Finally, we show that specific ARBs can increase mammalian cell tolerance to Cu, as well as decrease the prevalence of Cu-induced apoptotic markers. All the above point to the potential of ARBs in preventing Cu-induced toxicity in yeast and mammalian cells.

  15. Increased pericarp cell length underlies a major quantitative trait locus for grain weight in hexaploid wheat.

    Science.gov (United States)

    Brinton, Jemima; Simmonds, James; Minter, Francesca; Leverington-Waite, Michelle; Snape, John; Uauy, Cristobal

    2017-08-01

    Crop yields must increase to address food insecurity. Grain weight, determined by grain length and width, is an important yield component, but our understanding of the underlying genes and mechanisms is limited. We used genetic mapping and near isogenic lines (NILs) to identify, validate and fine-map a major quantitative trait locus (QTL) on wheat chromosome 5A associated with grain weight. Detailed phenotypic characterisation of developing and mature grains from the NILs was performed. We identified a stable and robust QTL associated with a 6.9% increase in grain weight. The positive interval leads to 4.0% longer grains, with differences first visible 12 d after fertilization. This grain length effect was fine-mapped to a 4.3 cM interval. The locus also has a pleiotropic effect on grain width (1.5%) during late grain development that determines the relative magnitude of the grain weight increase. Positive NILs have increased maternal pericarp cell length, an effect which is independent of absolute grain length. These results provide direct genetic evidence that pericarp cell length affects final grain size and weight in polyploid wheat. We propose that combining genes that control distinct biological mechanisms, such as cell expansion and proliferation, will enhance crop yields. © 2017 The Authors. New Phytologist © 2017 New Phytologist Trust.

  16. Analysis and effects of cytosolic free calcium increases in response to elicitors in Nicotiana plumbaginifolia cells.

    Science.gov (United States)

    Lecourieux, David; Mazars, Christian; Pauly, Nicolas; Ranjeva, Raoul; Pugin, Alain

    2002-10-01

    Cell suspensions obtained from Nicotiana plumbaginifolia plants stably expressing the apoaequorin gene were used to analyze changes in cytosolic free calcium concentrations ([Ca(2+)](cyt)) in response to elicitors of plant defenses, particularly cryptogein and oligogalacturonides. The calcium signatures differ in lag time, peak time, intensity, and duration. The intensities of both signatures depend on elicitor concentration and extracellular calcium concentration. Cryptogein signature is characterized by a long-sustained [Ca(2+)](cyt) increase that should be responsible for sustained mitogen-activated protein kinase activation, microtubule depolymerization, defense gene activation, and cell death. The [Ca(2+)](cyt) increase in elicitor-treated cells first results from a calcium influx, which in turns leads to calcium release from internal stores and additional Ca(2+) influx. H(2)O(2) resulting from the calcium-dependent activation of the NADPH oxidase also participates in [Ca(2+)](cyt) increase and may activate calcium channels from the plasma membrane. Competition assays with different elicitins demonstrate that [Ca(2+)](cyt) increase is mediated by cryptogein-receptor interaction.

  17. Increasing the efficiency of silicon heterojunction solar cells and modules by light soaking

    KAUST Repository

    Kobayashi, Eiji

    2017-06-24

    Silicon heterojunction solar cells use crystalline silicon (c-Si) wafers as optical absorbers and employ bilayers of doped/intrinsic hydrogenated amorphous silicon (a-Si:H) to form passivating contacts. Recently, we demonstrated that such solar cells increase their operating voltages and thus their conversion efficiencies during light exposure. We found that this performance increase is due to improved passivation of the a-Si:H/c-Si interface and is induced by injected charge carriers (either by light soaking or forward-voltage biasing of the device). Here, we discuss this counterintuitive behavior and establish that: (i) the performance increase is observed in solar cells as well as modules; (ii) this phenomenon requires the presence of doped a-Si:H films, but is independent from whether light is incident from the a-Si:H(p) or the a-Si:H(n) side; (iii) UV and blue photons do not play a role in this effect; (iv) the performance increase can be observed under illumination intensities as low as 20Wm (0.02-sun) and appears to be almost identical in strength when under 1-sun (1000Wm); (v) the underlying physical mechanism likely differs from annealing-induced surface passivation.

  18. Radiosensitivity is increased by knockdown of FTS in uterine cervical cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Park, Wo Yoon; Anandharaj, Arunkumar; Cinghu, Senthikumar; Kim, Won Dong [Dept. of Radiation Oncology, Chungbuk National University College of Medicine, Cheongju (Korea, Republic of); Yu, Jae Ran [Dept. of Environmental and Tropical Medicine, Konkuk University College of Medicine, Seoul (Korea, Republic of)

    2012-04-15

    Uterine cervical cancer is still the second largest cancer in women worldwide, despite of effective screening methods. Radiotherapy is used to treat all the stages of cervical cancer and more than 60% of cervical cancer patients receive radiotherapy. New therapeutic targets or approaches are needed to further increase the results of radiotherapy. In the present study, we demonstrated the radiation induced overexpression and nuclear export of FTS in cervical cancer cells. Furthermore, we showed that silencing of FTS expression with FTS shRNA enhanced radiosensitivity of cervical cancer cells, induced cell cycle arrest and apoptosis FTS is involved in radioresistance of cervical cancer. Targeted inhibition of FTS can shutdown the key elemental characteristics of cervical cancer and could lead to an effective therapeutic strategy.

  19. Radiosensitivity is increased by knockdown of FTS in uterine cervical cancer cells

    International Nuclear Information System (INIS)

    Park, Wo Yoon; Anandharaj, Arunkumar; Cinghu, Senthikumar; Kim, Won Dong; Yu, Jae Ran

    2012-01-01

    Uterine cervical cancer is still the second largest cancer in women worldwide, despite of effective screening methods. Radiotherapy is used to treat all the stages of cervical cancer and more than 60% of cervical cancer patients receive radiotherapy. New therapeutic targets or approaches are needed to further increase the results of radiotherapy. In the present study, we demonstrated the radiation induced overexpression and nuclear export of FTS in cervical cancer cells. Furthermore, we showed that silencing of FTS expression with FTS shRNA enhanced radiosensitivity of cervical cancer cells, induced cell cycle arrest and apoptosis FTS is involved in radioresistance of cervical cancer. Targeted inhibition of FTS can shutdown the key elemental characteristics of cervical cancer and could lead to an effective therapeutic strategy

  20. Isoproterenol Increases RANKL Expression in a ATF4/NFATc1-Dependent Manner in Mouse Osteoblastic Cells

    Directory of Open Access Journals (Sweden)

    Kyunghwa Baek

    2017-10-01

    Full Text Available Sympathetic nervous system stimulation-induced β-adrenergic signal transduction is known to induce bone loss and increase of osteoclast activity. Although isoproterenol, a nonspecific β-adrenergic receptor agonist, has been shown to increase receptor activator of NF-κB ligand (RANKL, the details of the regulatory mechanisms remain unclear. In the present study, we investigated the role of the nuclear factor of activated T-cells (NFAT in isoproterenol-induced RANKL expression in C2C12 and in primary cultured mouse calvarial cells. Isoproterenol increased nuclear factor of activated T-cells cytoplasmic 1 (NFATc1 and RANKL expressions at both mRNA and protein levels and increased NFAT reporter activity. NFATc1 knockdown blocked isoproterenol-mediated RANKL expression. Isoproterenol also promoted cAMP response element-binding protein 1 (CREB1 and activating transcription factor 4 (ATF4 phosphorylation. Isoproterenol-mediated transcriptional activation of NFAT was blocked by protein kinase A (PKA inhibitor H89. Isoproterenol-induced CREB1, ATF4, NFATc1, and RANKL expressions were suppressed by H89. Mutations in cAMP response element-like or NFAT-binding element suppressed isoproterenol-induced RANKL promoter activity. Chromatin immunoprecipitation analysis demonstrated that isoproterenol increased NFAT-binding and ATF4-binding activities on the mouse RANKL promoter, but did not increase CREB1-binding activity. Association of NFATc1 and ATF4 was not observed in a co-immunoprecipitation study. ATF4 knockdown suppressed isoproterenol-induced NFAT binding to the RANKL promoter, whereas NFATc1 knockdown did not suppress isoproterenol-induced ATF4 binding to the RANKL promoter. ATF4 knockdown suppressed isoproterenol-induced expressions of NFATc1 and RANKL. These results suggest that isoproterenol increases RANKL expression in an ATF4/NFATc1-dependent manner.

  1. Mixed lactate and caffeine compound increases satellite cell activity and anabolic signals for muscle hypertrophy.

    Science.gov (United States)

    Oishi, Yoshimi; Tsukamoto, Hayato; Yokokawa, Takumi; Hirotsu, Keisuke; Shimazu, Mariko; Uchida, Kenji; Tomi, Hironori; Higashida, Kazuhiko; Iwanaka, Nobumasa; Hashimoto, Takeshi

    2015-03-15

    We examined whether a mixed lactate and caffeine compound (LC) could effectively elicit proliferation and differentiation of satellite cells or activate anabolic signals in skeletal muscles. We cultured C2C12 cells with either lactate or LC for 6 h. We found that lactate significantly increased myogenin and follistatin protein levels and phosphorylation of P70S6K while decreasing the levels of myostatin relative to the control. LC significantly increased protein levels of Pax7, MyoD, and Ki67 in addition to myogenin, relative to control. LC also significantly increased follistatin expression relative to control and stimulated phosphorylation of mTOR and P70S6K. In an in vivo study, male F344/DuCrlCrlj rats were assigned to control (Sed, n = 10), exercise (Ex, n = 12), and LC supplementation (LCEx, n = 13) groups. LC was orally administered daily. The LCEx and Ex groups were exercised on a treadmill, running for 30 min at low intensity every other day for 4 wk. The LCEx group experienced a significant increase in the mass of the gastrocnemius (GA) and tibialis anterior (TA) relative to both the Sed and Ex groups. Furthermore, the LCEx group showed a significant increase in the total DNA content of TA compared with the Sed group. The LCEx group experienced a significant increase in myogenin and follistatin expression of GA relative to the Ex group. These results suggest that administration of LC can effectively increase muscle mass concomitant with elevated numbers of myonuclei, even with low-intensity exercise training, via activated satellite cells and anabolic signals. Copyright © 2015 the American Physiological Society.

  2. Aging increases microglial proliferation, delays cell migration, and decreases cortical neurogenesis after focal cerebral ischemia.

    Science.gov (United States)

    Moraga, Ana; Pradillo, Jesús M; García-Culebras, Alicia; Palma-Tortosa, Sara; Ballesteros, Ivan; Hernández-Jiménez, Macarena; Moro, María A; Lizasoain, Ignacio

    2015-05-10

    Aging is not just a risk factor of stroke, but it has also been associated with poor recovery. It is known that stroke-induced neurogenesis is reduced but maintained in the aged brain. However, there is no consensus on how neurogenesis is affected after stroke in aged animals. Our objective is to determine the role of aging on the process of neurogenesis after stroke. We have studied neurogenesis by analyzing proliferation, migration, and formation of new neurons, as well as inflammatory parameters, in a model of cerebral ischemia induced by permanent occlusion of the middle cerebral artery in young- (2 to 3 months) and middle-aged mice (13 to 14 months). Aging increased both microglial proliferation, as shown by a higher number of BrdU(+) cells and BrdU/Iba1(+) cells in the ischemic boundary and neutrophil infiltration. Interestingly, aging increased the number of M1 monocytes and N1 neutrophils, consistent with pro-inflammatory phenotypes when compared with the alternative M2 and N2 phenotypes. Aging also inhibited (subventricular zone) SVZ cell proliferation by decreasing both the number of astrocyte-like type-B (prominin-1(+)/epidermal growth factor receptor (EGFR)(+)/nestin(+)/glial fibrillary acidic protein (GFAP)(+) cells) and type-C cells (prominin-1(+)/EGFR(+)/nestin(-)/Mash1(+) cells), and not affecting apoptosis, 1 day after stroke. Aging also inhibited migration of neuroblasts (DCX(+) cells), as indicated by an accumulation of neuroblasts at migratory zones 14 days after injury; consistently, aged mice presented a smaller number of differentiated interneurons (NeuN(+)/BrdU(+) and GAD67(+) cells) in the peri-infarct cortical area 14 days after stroke. Our data confirm that stroke-induced neurogenesis is maintained but reduced in aged animals. Importantly, we now demonstrate that aging not only inhibits proliferation of specific SVZ cell subtypes but also blocks migration of neuroblasts to the damaged area and decreases the number of new interneurons in

  3. Increased risk of gastric adenocarcinoma after treatment of primary gastric diffuse large B-cell lymphoma

    International Nuclear Information System (INIS)

    Inaba, Koji; Morota, Madoka; Mayahara, Hiroshi; Ito, Yoshinori; Sumi, Minako; Uno, Takashi; Itami, Jun; Kushima, Ryoji; Murakami, Naoya; Kuroda, Yuuki; Harada, Ken; Kitaguchi, Mayuka; Yoshio, Kotaro; Sekii, Shuhei; Takahashi, Kana

    2013-01-01

    There have been sporadic reports about synchronous as well as metachronous gastric adenocarcinoma and primary gastric lymphoma. Many reports have dealt with metachronous gastric adenocarcinoma in mucosa-associated lymphoid tissue lymphoma of stomach. But to our knowledge, there have been no reports that document the increased incidence of metachronous gastric adenocarcinoma in patients with gastric diffuse large B-cell lymphoma. This retrospective study was conducted to estimate the incidence of metachronous gastric adenocarcinoma after primary gastric lymphoma treatment, especially in diffuse large B-cell lymphoma. The retrospective cohort study of 139 primary gastric lymphoma patients treated with radiotherapy at our hospital. Mean observation period was 61.5 months (range: 3.7-124.6 months). Patients profile, characteristics of primary gastric lymphoma and metachronous gastric adenocarcinoma were retrieved from medical records. The risk of metachronous gastric adenocarcinoma was compared with the risk of gastric adenocarcinoma in Japanese population. There were 10 (7.2%) metachronous gastric adenocarcinoma patients after treatment of primary gastric lymphomas. It was quite high risk compared with the risk of gastric carcinoma in Japanese population of 54.7/100,000. Seven patients of 10 were diffuse large B-cell lymphoma and other 3 patients were mixed type of diffuse large B-cell lymphoma and mucosa associated lymphoid tissue lymphoma. Four patients of 10 metachronous gastric adenocarcinomas were signet-ring cell carcinoma and two patients died of gastric adenocarcinoma. Metachronous gastric adenocarcinoma may have a more malignant potential than sporadic gastric adenocarcinoma. Old age, Helicobacter pylori infection and gastric mucosal change of chronic gastritis and intestinal metaplasia were possible risk factors for metachronous gastric adenocarcinoma. There was an increased risk of gastric adenocarcinoma after treatment of primary gastric lymphoma

  4. Kaempferol increases apoptosis in human acute promyelocytic leukemia cells and inhibits multidrug resistance genes.

    Science.gov (United States)

    Moradzadeh, Maliheh; Tabarraei, Alijan; Sadeghnia, Hamid Reza; Ghorbani, Ahmad; Mohamadkhani, Ashraf; Erfanian, Saiedeh; Sahebkar, Amirhossein

    2018-02-01

    Acute promyelocytic leukemia (APL) is one of the most life-threatening hematological malignancies. Defects in the cell growth and apoptotic pathways are responsible for both disease pathogenesis and treatment resistance. Therefore, pro-apoptotic agents are potential candidates for APL treatment. Kaempferol is a flavonoid with antioxidant and anti-tumor properties. This study was designed to investigate the cytotoxic, pro-apoptotic, and differentiation-inducing effects of kaempferol on HL-60 and NB4 leukemia cells. Resazurin assay was used to determine cell viability following treatment with kaempferol (12.5-100 μM) and all-trans retinoic acid (ATRA; 10 μM; used as a positive control). Apoptosis and differentiation were also detected using propidium iodide and NBT staining techniques, respectively. Furthermore, the expression levels of genes involved in apoptosis (PI3 K, AKT, BCL2, BAX, p53, p21, PTEN, CASP3, CASP8, and CASP9), differentiation (PML-RAR and HDAC1), and multi-drug resistance (ABCB1 and ABCC1) were determined using quantitative real-time PCR. The protein expressions of Bax/Bcl2 and casp3 were confirmed using Western blot. The results showed that kaempferol decreased cell viability and increased subG1 population in the tested leukemic cells. This effect was associated with decreased expression of Akt, BCL2, ABCB1, and ABCC1 genes, while the expression of CASP3 and BAX/BCL-2 ratio were significantly increased at both gene and protein levels. Kaempferol promoted apoptosis and inhibited multidrug resistance in a concentration-dependent manner, without any differential effect on leukemic cells. In conclusion, this study suggested that kaempferol may be utilized as an appropriate alternative for ATRA in APL patients. © 2017 Wiley Periodicals, Inc.

  5. Short-term stress enhances cellular immunity and increases early resistance to squamous cell carcinoma.

    Science.gov (United States)

    Dhabhar, Firdaus S; Saul, Alison N; Daugherty, Christine; Holmes, Tyson H; Bouley, Donna M; Oberyszyn, Tatiana M

    2010-01-01

    In contrast to chronic/long-term stress that suppresses/dysregulates immune function, an acute/short-term fight-or-flight stress response experienced during immune activation can enhance innate and adaptive immunity. Moderate ultraviolet-B (UV) exposure provides a non-invasive system for studying the naturalistic emergence, progression and regression of squamous cell carcinoma (SCC). Because SCC is an immunoresponsive cancer, we hypothesized that short-term stress experienced before UV exposure would enhance protective immunity and increase resistance to SCC. Control and short-term stress groups were treated identically except that the short-term stress group was restrained (2.5h) before each of nine UV-exposure sessions (minimum erythemal dose, 3-times/week) during weeks 4-6 of the 10-week UV exposure protocol. Tumors were measured weekly, and tissue collected at weeks 7, 20, and 32. Chemokine and cytokine gene expression was quantified by real-time PCR, and CD4+ and CD8+ T cells by flow cytometry and immunohistochemistry. Compared to controls, the short-term stress group showed greater cutaneous T-cell attracting chemokine (CTACK)/CCL27, RANTES, IL-12, and IFN-gamma gene expression at weeks 7, 20, and 32, higher skin infiltrating T cell numbers (weeks 7 and 20), lower tumor incidence (weeks 11-20) and fewer tumors (weeks 11-26). These results suggest that activation of short-term stress physiology increased chemokine expression and T cell trafficking and/or function during/following UV exposure, and enhanced Type 1 cytokine-driven cell-mediated immunity that is crucial for resistance to SCC. Therefore, the physiological fight-or-flight stress response and its adjuvant-like immuno-enhancing effects, may provide a novel and important mechanism for enhancing immune system mediated tumor-detection/elimination that merits further investigation.

  6. Direct alcohol fuel cells: Increasing platinum performance by modification with sp-group metals

    Science.gov (United States)

    Figueiredo, Marta C.; Sorsa, Olli; Doan, Nguyet; Pohjalainen, Elina; Hildebrand, Helga; Schmuki, Patrik; Wilson, Benjamin P.; Kallio, Tanja

    2015-02-01

    By using sp group metals as modifiers, the catalytic properties of Pt can be improved toward alcohols oxidation. In this work we report the performance increase of direct alcohol fuel cells (DAFC) fuelled with ethanol or 2-propanol with platinum based anode electrodes modified with Bi and Sb adatoms. For example, by simply adding Sb to the Pt/C based anode ink during membrane electrode assembly fabrication of a direct ethanol fuel cell (DEFC) its performance is improved three-fold, with more than 100 mV increase in the open circuit potential. For the fuel cell fuelled with 2-propanol high power densities are obtained at very high potentials with these catalyst materials suggesting a great improvement for practical applications. Particularly in the case of Pt/C-Bi, the improvement is such that within 0.6 V (from 0.7 to 0.1 V) the power densities are between 7 and 9 mW/cm2. The results obtained with these catalysts are in the same range as those obtained with other bimetallic catalysts comprising of PtRu and PtSn, which are currently considered to be the best for these type of fuel cells and that are obtained by more complicated (and consequently more expensive) methods.

  7. Ferritin-iron increases killing of Chinese hamster ovary cells by X-irradiation

    International Nuclear Information System (INIS)

    Nelson, J.M.; Stevens, R.G.

    1992-01-01

    Stationary-phase Chinese hamster ovary cells were cultured in medium containing ferritin (∼19% iron by weight) added at concentrations ranging from 0 to 128 μg/ml. One set of cultures was unirradiated, another set exposed to 4.0 Gy of X-ray. Clonogenic cell survival was assessed in each set of cultures. In the absence of added ferritin, 4.0 Gy killed approximately 50% of the cells. In the absence of radiation, ferritin was not toxic at less than 48 μg/ml; above 48 μg/ml, toxicity increased with concentration. Apoferritin was not toxic at any concentration tested (up to 1000 μg/ml). Although 32 μg/ml ferritin, reflecting only a 3-6 fold increase in iron concentration over normal serum, was not toxic, it reduced survival of X-irradiated cells by an additional 75%. These results indicate that a sublethal concentration of ferritin can be a potent radiosensitizer. (Author)

  8. Cold stress increases reactive oxygen species formation via TRPA1 activation in A549 cells.

    Science.gov (United States)

    Sun, Wenwu; Wang, Zhonghua; Cao, Jianping; Cui, Haiyang; Ma, Zhuang

    2016-03-01

    Reactive oxygen species (ROS) are responsible for lung damage during inhalation of cold air. However, the mechanism of the ROS production induced by cold stress in the lung is still unclear. In this work, we measured the changes of ROS and the cytosolic Ca(2+) concentration ([Ca(2+)]c) in A549 cell. We observed that cold stress (from 20 to 5 °C) exposure of A549 cell resulted in an increase of ROS and [Ca(2+)]c, which was completely attenuated by removing Ca(2+) from medium. Further experiments showed that cold-sensing transient receptor potential subfamily member 1 (TRPA1) agonist (allyl isothiocyanate, AITC) increased the production of ROS and the level of [Ca(2+)]c in A549 cell. Moreover, HC-030031, a TRPA1 selective antagonist, significantly inhibited the enhanced ROS and [Ca(2+)]c induced by AITC or cold stimulation, respectively. Taken together, these data demonstrated that TRPA1 activation played an important role in the enhanced production of ROS induced by cold stress in A549 cell.

  9. Increased endothelial apoptotic cell density in human diabetic erectile tissue--comparison with clinical data.

    Science.gov (United States)

    Costa, Carla; Soares, Raquel; Castela, Angela; Adães, Sara; Hastert, Véronique; Vendeira, Pedro; Virag, Ronald

    2009-03-01

    Erectile dysfunction (ED) is a common complication of diabetes. Endothelial cell (EC) dysfunction is one of the main mechanisms of diabetic ED. However, loss of EC integrity has never been assessed in human diabetic corpus cavernosum. To identify and quantify apoptotic cells in human diabetic and normal erectile tissue and to compare these results with each patient's clinical data and erection status. Eighteen cavernosal samples were collected, 13 from diabetics with ED and 5 from nondiabetic individuals. Cavernosal structure and cell proliferation status were evaluated by immunohistochemistry. Tissue integrity was assessed by terminal transferase dUTP nick end labeling assay, an index of apoptotic cell density (ACD) established and compared with each patient age, type of diabetes, arterial risk factors number, arterial/veno-occlusive disease, response to intracavernous vasoactive injections (ICI), and penile nitric oxide release test (PNORT). Establish an index of ACD and correlate those results with patient clinical data. Nondiabetic samples presented few scattered cells in apoptosis and an ACD of 7.15 +/- 0.44 (mean apoptotic cells/tissue area mm(2) +/- standard error). The diabetic group showed an increased ACD of 23.82 +/- 1.53, and apoptotic cells were located specifically at vascular sites. Rehabilitation of these endothelial lesions seemed impaired, as no evidence of EC proliferation was observed. Furthermore, higher ACD in diabetic individuals correlated to poor response to PNORT and to ICI. We provided evidence for the first time that loss of cavernosal EC integrity is a crucial event involved in diabetic ED. Furthermore, we were able to establish a threshold between ACD values and cavernosal tissue functionality, as assessed by PNORT and vasoactive ICI.

  10. Increased Dickkopf-1 expression accelerates bone cell apoptosis in femoral head osteonecrosis.

    Science.gov (United States)

    Ko, Jih-Yang; Wang, Feng-Sheng; Wang, Ching-Jen; Wong, To; Chou, Wen-Yi; Tseng, Shin-Ling

    2010-03-01

    Intensive bone cell apoptosis contributes to osteonecrosis of femoral head (ONFH). Dickkopf-1 (DKK1) reportedly mediates various types of skeletal disorders. This study investigated whether DKK1 was linked to the occurrence of ONFH. Thirty-nine patients with various stages of ONFH were recruited. Bone specimens were harvested from 34 ONFH patients underwent hip arthroplasty, and from 10 femoral neck fracture patients. Bad, Bcl2 TNFalpha, DKK1, Wnt3a, LRP5, and Axin1 expressions were analyzed by quantitative RT-PCR and ELISA. Apoptotic cells were assayed using terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate-biotin nick end-labelling (TUNEL). Primary bone-marrow mesenchymal cells were treated with DKK1 RNA interference and recombinant DKK1 protein. ONFH patients with the histories of being administrated corticosteroids and excessive alcohol consumption had significantly higher Bad and DKK1 mRNA expressions in bone tissue and DKK1 abundances in serum than femoral neck fracture patients. Bone cells adjacent to osteonecrotic bone displayed strong DKK1 immunoreactivity and TUNEL staining. Increased DKK1 expression in bone tissue and serum correlated with Bad expression and TUNEL staining. Serum DKK1 abundance correlated with the severity of ONFH. The DKK1 RNA interference and recombinant DKK1 protein regulated Bad expression and apoptosis of primary bone-marrow mesenchymal cells. Knock down of DKK1 reduced dexamethasone-induced apoptosis of mesenchymal cells. Taken together, promoted DKK1 expression was associated with bone cell apoptosis in the occurrence of ONFH patients with the histories of corticosteroid and alcohol intake and progression of ONFH. DKK1 expression in injured tissue provides new insight into ONFH pathogenesis.

  11. Blood Outgrowth Endothelial Cells Increase Tumor Growth Rates and Modify Tumor Physiology: Relevance for Therapeutic Targeting

    Energy Technology Data Exchange (ETDEWEB)

    Pagan, Jonathan, E-mail: jdpagan@uams.edu; Przybyla, Beata; Jamshidi-Parsian, Azemat [Department of Radiation Oncology, University of Arkansas for Medical Sciences, 4301 West Markham Street, Little Rock, AR 72205 (United States); Gupta, Kalpna [Vascular Biology Center and Division of Hematology-Oncology Transplantation, Department of Medicine, University of Minnesota Medical School, MN 72223 (United States); Griffin, Robert J. [Department of Radiation Oncology, University of Arkansas for Medical Sciences, 4301 West Markham Street, Little Rock, AR 72205 (United States)

    2013-02-18

    Endothelial cell precursors from human peripheral blood have been shown to home to areas of neovascularization and may assist tumor growth by increasing or fortifying blood vessel growth. In the present study, the influence of these cells on tumor growth and physiology was investigated and the role of these cells as a therapeutic target or in determining treatment sensitivity was tested. After isolation from human blood and expansion in vitro, actively growing cells with verified endothelial phenotype (Blood Outgrowth Endothelial Cell, BOEC) were injected i.v. into tumor bearing mice for three consecutive days. The growth rate was significantly enhanced in relatively small RERF human lung tumors (i.e., less than 150 mm{sup 3}) grown in immunocompromised mice by an average of 1.5-fold while it had no effect when injections were given to animals bearing larger tumors. There were no signs of toxicity or unwanted systemic effects. We also observed evidence of increased perfusion, vessel number, response to 15 Gy radiation and oxygenation in RERF tumors of animals injected with BOECs compared to control tumors. In addition, FSaII murine fibrosarcoma tumors were found to grow faster upon injection of BOECs. When FSaII tumors were subjected to a partial thermal ablation treatment using high intensity focused ultrasound (HIFU) there was consistently elevated detection of fluorescently labeled and i.v. injected endothelial precursors in the tumor when analyzed with optical imaging and/or histological preparations. Importantly, we also observed that BOECs treated with the novel anti-angiogenic peptide anginex in-vitro, show decreased proliferation and increased sensitivity to radiation. In vivo, the normal increase in FSaII tumor growth induced by injected BOECs was blunted by the addition of anginex treatment. It appears that endothelial precursors may significantly contribute to tumor vessel growth, tumor progression and/or repair of tumor damage and may improve the

  12. Blood Outgrowth Endothelial Cells Increase Tumor Growth Rates and Modify Tumor Physiology: Relevance for Therapeutic Targeting

    International Nuclear Information System (INIS)

    Pagan, Jonathan; Przybyla, Beata; Jamshidi-Parsian, Azemat; Gupta, Kalpna; Griffin, Robert J.

    2013-01-01

    Endothelial cell precursors from human peripheral blood have been shown to home to areas of neovascularization and may assist tumor growth by increasing or fortifying blood vessel growth. In the present study, the influence of these cells on tumor growth and physiology was investigated and the role of these cells as a therapeutic target or in determining treatment sensitivity was tested. After isolation from human blood and expansion in vitro, actively growing cells with verified endothelial phenotype (Blood Outgrowth Endothelial Cell, BOEC) were injected i.v. into tumor bearing mice for three consecutive days. The growth rate was significantly enhanced in relatively small RERF human lung tumors (i.e., less than 150 mm 3 ) grown in immunocompromised mice by an average of 1.5-fold while it had no effect when injections were given to animals bearing larger tumors. There were no signs of toxicity or unwanted systemic effects. We also observed evidence of increased perfusion, vessel number, response to 15 Gy radiation and oxygenation in RERF tumors of animals injected with BOECs compared to control tumors. In addition, FSaII murine fibrosarcoma tumors were found to grow faster upon injection of BOECs. When FSaII tumors were subjected to a partial thermal ablation treatment using high intensity focused ultrasound (HIFU) there was consistently elevated detection of fluorescently labeled and i.v. injected endothelial precursors in the tumor when analyzed with optical imaging and/or histological preparations. Importantly, we also observed that BOECs treated with the novel anti-angiogenic peptide anginex in-vitro, show decreased proliferation and increased sensitivity to radiation. In vivo, the normal increase in FSaII tumor growth induced by injected BOECs was blunted by the addition of anginex treatment. It appears that endothelial precursors may significantly contribute to tumor vessel growth, tumor progression and/or repair of tumor damage and may improve the

  13. Hypomutability in Fanconi anemia cells is associated with increased deletion frequency at the HPRT locus

    International Nuclear Information System (INIS)

    Papadopoulo, D.; Guillouf, C.; Moustacchi, E.; Mohrenweiser, H.

    1990-01-01

    Fanconi anemia (FA) is an inherited human disorder associated with a predisposition to cancer and characterized by anomalies in the processing of DNA cross-links and certain monoadducts. The authors reported previously that the frequency of psoralen-photoinduced mutations at the HPRT locus is lower in FA cells than in normal cells. This hypomutability is shown here to be associated with an increased frequency of deletions in the HPRT gene when either a mixture of cross-links and monoadducts or monoadducts alone are induced. Molecular analysis of mutants in the HPRT gene was carried out. In normal cells the majority of spontaneous and induced mutants are point mutations whereas in FA deletion mutations predominate. In that case a majority of mutants were found to lack individual exons or small clusters of exons whereas in normal cells large (complete or major gene loss) and small deletions are almost equally represented. Thus they propose that the FA defect lies in a mutagenic pathway that, in normal cells, involves by passing lesions and subsequent gap filling by a recombinational process during replication

  14. Radioresistant DNA synthesis in cells of patients showing increased chromosomal sensitivity to ionizing radiation

    International Nuclear Information System (INIS)

    Barenfeld, L.S.; Pleskach, N.M.; Bildin, V.N.; Prokofjeva, V.V.; Mikhelson, V.M.

    1986-01-01

    The rate of DNA synthesis after γ-irradiation was studied either by analysis of the steady-state distribution of daughter [ 3 H]DNA in alkaline sucrose gradients or by direct assay of the amount of [ 3 H]thymidine incorporated into DNA of fibroblasts derived from a normal donor (LCH882) and from Down's syndrome (LCH944), Werner's syndrome (WS1LE) and xeroderma pigmentosum (XP2LE) patients with chromosomal sensitivity to ionizing radiation. Doses of γ-irradiation that markedly inhibited the rate of DNA synthesis in normal human cells caused almost no inhibition of DNA synthesis in the cells from the affected individuals. The radioresistant DNA synthesis in Down's syndrome cells was mainly due to a much lower inhibition of replicon initiation than that in normal cells; these cells were also more resistant to damage that inhibited replicon elongation. Our data suggest that radioresistant DNA synthesis may be an intrinsic feature of all genetic disorders showing increased radiosensitivity in terms of chromosome aberrations. (orig.)

  15. Natural Killer Cells Improve Hematopoietic Stem Cell Engraftment by Increasing Stem Cell Clonogenicity In Vitro and in a Humanized Mouse Model.

    Science.gov (United States)

    Escobedo-Cousin, Michelle; Jackson, Nicola; Laza-Briviesca, Raquel; Ariza-McNaughton, Linda; Luevano, Martha; Derniame, Sophie; Querol, Sergio; Blundell, Michael; Thrasher, Adrian; Soria, Bernat; Cooper, Nichola; Bonnet, Dominique; Madrigal, Alejandro; Saudemont, Aurore

    2015-01-01

    Cord blood (CB) is increasingly used as a source of hematopoietic stem cells (HSC) for transplantation. Low incidence and severity of graft-versus-host disease (GvHD) and a robust graft-versus-leukemia (GvL) effect are observed following CB transplantation (CBT). However, its main disadvantages are a limited number of HSC per unit, delayed immune reconstitution and a higher incidence of infection. Unmanipulated grafts contain accessory cells that may facilitate HSC engraftment. Therefore, the effects of accessory cells, particularly natural killer (NK) cells, on human CB HSC (CBSC) functions were assessed in vitro and in vivo. CBSC cultured with autologous CB NK cells showed higher levels of CXCR4 expression, a higher migration index and a higher number of colony forming units (CFU) after short-term and long-term cultures. We found that CBSC secreted CXCL9 following interaction with CB NK cells. In addition, recombinant CXCL9 increased CBSC clonogenicity, recapitulating the effect observed of CB NK cells on CBSC. Moreover, the co-infusion of CBSC with CB NK cells led to a higher level of CBSC engraftment in NSG mouse model. The results presented in this work offer the basis for an alternative approach to enhance HSC engraftment that could improve the outcome of CBT.

  16. Release of intracellular Calcium increase production of mitochondrial reactive oxygen species in renal distal epithelial cells

    DEFF Research Database (Denmark)

    Bjerregaard, Henning F.

    peroxide (H2O2) has traditionally been regarded as toxic by-products of aerobic metabolism. However, recent findings indicate that H2O2 act as a signalling molecule. The aim of the present study was to monitor, in real time, the rates of ROS generation in order to directly determine their production......Release of intracellular Calcium increase production of mitochondrial reactive oxygen species in renal distal epithelial cells. Henning F. Bjerregaard, Roskilde University, Department of Science, Systems and Models , 4000 Roskilde, Denmark. HFB@ RUC.DK Reactive oxygen species (ROS) like, hydrogen...... to G-protein stimulation of phospholipase C and release of inositol -3 phosphate. Cd (0.4 mM) treatment of A6 cells enhanced the ROS production after one minutes incubation. The production rate was constant for at least 10 to 20 min. Experiments showed that the Cd induced increase in ROS production...

  17. Increased efficiency of exogenous messenger RNA translation in a Krebs ascites cell lysate.

    Science.gov (United States)

    Metafora, S; Terada, M; Dow, L W; Marks, P A; Bank, A

    1972-05-01

    Addition of a 0.5 M KCl wash fraction from rabbit reticulocyte ribosomes causes a 3- to 10-fold increase in the extent of translation of natural mRNAs by Krebs-cell lysates. In the presence of the wash fraction, 1 pmol of rabbit or mouse 10S RNA directs the incorporation of 80 pmol of leucine into rabbit globin. The addition of human 10S RNA results in the synthesis of equal amounts of human alpha and beta chains, identified by column chromatography. The stimulation by the wash fraction is almost completely dependent on added mammalian tRNA. In contrast to the wash fraction from rabbit reticulocytes, the wash fraction isolated from Krebs-cell ribosomes is inhibitory to both endogenous and exogenous mRNA translation. The stimulation by the wash fraction from rabbit ribosomes is not specific for globin mRNAs, but also increases endogenous, phage Qbeta, and viral RNA-directed protein synthesis.

  18. Cisplatin-resistant cells express increased levels of a factor that recognizes damaged DNA

    International Nuclear Information System (INIS)

    Chu, G.; Chang, E.

    1990-01-01

    Cancer treatment with the drug cisplatin is often thwarted by the emergence of drug-resistant cells. To study this phenomenon, the authors identified two independent cellular factors that recognize cisplatin-damaged DNA. One of the two factors, designated XPE binding factor, is deficient in complementation group E of xeroderma pigmentosum, an inherited disease characterized by defective repair of DNA damaged by ultraviolet radiation, cisplatin, and other agents. Human tumor cell lines selected for resistance to cisplatin showed more efficient DNA repair and increased expression of XPE binding factor. These results suggest that XPE binding factor may be responsible, at least in part, for the development of cisplatin resistance in human tumors and that the mechanism may be increased DNA repair

  19. Increased proliferation of late-born retinal progenitor cells by gestational lead exposure delays rod and bipolar cell differentiation.

    Science.gov (United States)

    Chaney, Shawnta Y; Mukherjee, Shradha; Giddabasappa, Anand; Rueda, Elda M; Hamilton, W Ryan; Johnson, Jerry E; Fox, Donald A

    2016-01-01

    Studies of neuronal development in the retina often examine the stages of proliferation, differentiation, and synaptic development, albeit independently. Our goal was to determine if a known neurotoxicant insult to a population of retinal progenitor cells (RPCs) would affect their eventual differentiation and synaptic development. To that end, we used our previously published human equivalent murine model of low-level gestational lead exposure (GLE). Children and animals with GLE exhibit increased scotopic electroretinogram a- and b-waves. Adult mice with GLE exhibit an increased number of late-born RPCs, a prolonged period of RPC proliferation, and an increased number of late-born rod photoreceptors and rod and cone bipolar cells (BCs), with no change in the number of late-born Müller glial cells or early-born neurons. The specific aims of this study were to determine whether increased and prolonged RPC proliferation alters the spatiotemporal differentiation and synaptic development of rods and BCs in early postnatal GLE retinas compared to control retinas. C57BL/6N mouse pups were exposed to lead acetate via drinking water throughout gestation and until postnatal day 10, which is equivalent to the human gestation period for retinal neurogenesis. RT-qPCR, immunohistochemical analysis, and western blots of well-characterized, cell-specific genes and proteins were performed at embryonic and early postnatal ages to assess rod and cone photoreceptor differentiation, rod and BC differentiation and synaptic development, and Müller glial cell differentiation. Real-time quantitative PCR (RT-qPCR) with the rod-specific transcription factors Nrl , Nr2e3 , and Crx and the rod-specific functional gene Rho , along with central retinal confocal studies with anti-recoverin and anti-rhodopsin antibodies, revealed a two-day delay in the differentiation of rod photoreceptors in GLE retinas. Rhodopsin immunoblots supported this conclusion. No changes in glutamine synthetase gene

  20. Increased cell motility and invasion upon knockdown of lipolysis stimulated lipoprotein receptor (LSR in SW780 bladder cancer cells

    Directory of Open Access Journals (Sweden)

    Ørntoft Torben F

    2008-07-01

    Full Text Available Abstract Background Mechanisms underlying the malignant development in bladder cancer are still not well understood. Lipolysis stimulated lipoprotein receptor (LSR has previously been found to be upregulated by P53. Furthermore, we have previously found LSR to be differentially expressed in bladder cancer. Here we investigated the role of LSR in bladder cancer. Methods A time course siRNA knock down experiment was performed to investigate the functional role of LSR in SW780 bladder cancer cells. Since LSR was previously shown to be regulated by P53, siRNA against TP53 was included in the experimental setup. We used Affymetrix GeneChips for measuring gene expression changes and we used Ingenuity Pathway Analysis to investigate the relationship among differentially expressed genes upon siRNA knockdown. Results By Ingenuity Pathway analysis of the microarray data from the different timepoints we identified six gene networks containing genes mainly related to the functional categories "cancer", "cell death", and "cellular movement". We determined that genes annotated to the functional category "cellular movement" including "invasion" and "cell motility" were highly significantly overrepresented. A matrigel assay showed that 24 h after transfection the invasion capacity was significantly increased 3-fold (p Conclusion We conclude that LSR may impair bladder cancer cells from gaining invasive properties.

  1. Intracellular kinases mediate increased translation and secretion of netrin-1 from renal tubular epithelial cells.

    Directory of Open Access Journals (Sweden)

    Calpurnia Jayakumar

    Full Text Available BACKGROUND: Netrin-1 is a laminin-related secreted protein, is highly induced after tissue injury, and may serve as a marker of injury. However, the regulation of netrin-1 production is not unknown. Current study was carried out in mouse and mouse kidney cell line (TKPTS to determine the signaling pathways that regulate netrin-1 production in response to injury. METHODS AND PRINCIPAL FINDINGS: Ischemia reperfusion injury of the kidney was induced in mice by clamping renal pedicle for 30 minutes. Cellular stress was induced in mouse proximal tubular epithelial cell line by treating with pervanadate, cisplatin, lipopolysaccharide, glucose or hypoxia followed by reoxygenation. Netrin-1 expression was quantified by real time RT-PCR and protein production was quantified using an ELISA kit. Cellular stress induced a large increase in netrin-1 production without increase in transcription of netrin-1 gene. Mitogen activated protein kinase, ERK mediates the drug induced netrin-1 mRNA translation increase without altering mRNA stability. CONCLUSION: Our results suggest that netrin-1 expression is suppressed at the translational level and MAPK activation leads to rapid translation of netrin-1 mRNA in the kidney tubular epithelial cells.

  2. Intracellular Kinases Mediate Increased Translation and Secretion of Netrin-1 from Renal Tubular Epithelial Cells

    Science.gov (United States)

    Jayakumar, Calpurnia; Mohamed, Riyaz; Ranganathan, Punithavathi Vilapakkam; Ramesh, Ganesan

    2011-01-01

    Background Netrin-1 is a laminin-related secreted protein, is highly induced after tissue injury, and may serve as a marker of injury. However, the regulation of netrin-1 production is not unknown. Current study was carried out in mouse and mouse kidney cell line (TKPTS) to determine the signaling pathways that regulate netrin-1 production in response to injury. Methods and Principal Findings Ischemia reperfusion injury of the kidney was induced in mice by clamping renal pedicle for 30 minutes. Cellular stress was induced in mouse proximal tubular epithelial cell line by treating with pervanadate, cisplatin, lipopolysaccharide, glucose or hypoxia followed by reoxygenation. Netrin-1 expression was quantified by real time RT-PCR and protein production was quantified using an ELISA kit. Cellular stress induced a large increase in netrin-1 production without increase in transcription of netrin-1 gene. Mitogen activated protein kinase, ERK mediates the drug induced netrin-1 mRNA translation increase without altering mRNA stability. Conclusion Our results suggest that netrin-1 expression is suppressed at the translational level and MAPK activation leads to rapid translation of netrin-1 mRNA in the kidney tubular epithelial cells. PMID:22046354

  3. Increased Efficiency of Solar Cells Protected by Hydrophobic and Hydrophilic Anti-Reflecting Nanostructured Glasses.

    Science.gov (United States)

    Baquedano, Estela; Torné, Lorena; Caño, Pablo; Postigo, Pablo A

    2017-12-14

    We investigated the fabrication of large-area (cm²) nanostructured glasses for solar cell modules with hydrophobic and hydrophilic properties using soft lithography and colloidal lithography. Both of these techniques entail low-cost and ease of nanofabrication. We explored the use of simple 1D and 2D nanopatterns (nanowires and nanocones) and the effect of introducing disorder in the nanostructures. We observed an increase in the transmitted light for ordered nanostructures with a maximum value of 99% for wavelengths >600 nm when ordered nanocones are fabricated on the two sides of the solar glass. They produced an increment in the efficiency of the packaged solar cell with respect to the glass without nanostructures. On the one hand, the wettability properties showed that the ordering of the nanostructures improved the hydrophobicity of the solar glasses and increased their self-cleaning capacity. On the other hand, the disordered nanostructures improved the hydrophilic properties of solar glasses, increasing their anti-fogging capacity. The results show that by selecting the appropriate nanopattern, the wettability properties (hydrophobic or hydrophilic) can be easily improved without decreasing the efficiency of the solar cell underneath.

  4. Human Traumatic Brain Injury Results in Oligodendrocyte Death and Increases the Number of Oligodendrocyte Progenitor Cells.

    Science.gov (United States)

    Flygt, Johanna; Gumucio, Astrid; Ingelsson, Martin; Skoglund, Karin; Holm, Jonatan; Alafuzoff, Irina; Marklund, Niklas

    2016-06-01

    Oligodendrocyte (OL) death may contribute to white matter pathology, a common cause of network dysfunction and persistent cognitive problems in patients with traumatic brain injury (TBI). Oligodendrocyte progenitor cells (OPCs) persist throughout the adult CNS and may replace dead OLs. OL death and OPCs were analyzed by immunohistochemistry of human brain tissue samples, surgically removed due to life-threatening contusions and/or focal brain swelling at 60.6 ± 75 hours (range 4-192 hours) postinjury in 10 severe TBI patients (age 51.7 ± 18.5 years). Control brain tissue was obtained postmortem from 5 age-matched patients without CNS disorders. TUNEL and CC1 co-labeling was used to analyze apoptotic OLs, which were increased in injured brain tissue (p The OPC markers Olig2, A2B5, NG2, and PDGFR-α were used. In contrast to the number of single-labeled Olig2, A2B5, NG2, and PDGFR-α-positive cells, numbers of Olig2 and A2B5 co-labeled cells were increased in TBI samples (p human TBI results in OL death and increases in OPCs postinjury, which may influence white matter function following TBI. © 2016 American Association of Neuropathologists, Inc. All rights reserved.

  5. Kaempferol increases levels of coenzyme Q in kidney cells and serves as a biosynthetic ring precursor.

    Science.gov (United States)

    Fernández-Del-Río, Lucía; Nag, Anish; Gutiérrez Casado, Elena; Ariza, Julia; Awad, Agape M; Joseph, Akil I; Kwon, Ohyun; Verdin, Eric; de Cabo, Rafael; Schneider, Claus; Torres, Jorge Z; Burón, María I; Clarke, Catherine F; Villalba, José M

    2017-09-01

    Coenzyme Q (Q) is a lipid-soluble antioxidant essential in cellular physiology. Patients with Q deficiencies, with few exceptions, seldom respond to treatment. Current therapies rely on dietary supplementation with Q 10 , but due to its highly lipophilic nature, Q 10 is difficult to absorb by tissues and cells. Plant polyphenols, present in the human diet, are redox active and modulate numerous cellular pathways. In the present study, we tested whether treatment with polyphenols affected the content or biosynthesis of Q. Mouse kidney proximal tubule epithelial (Tkpts) cells and human embryonic kidney cells 293 (HEK 293) were treated with several types of polyphenols, and kaempferol produced the largest increase in Q levels. Experiments with stable isotope 13 C-labeled kaempferol demonstrated a previously unrecognized role of kaempferol as an aromatic ring precursor in Q biosynthesis. Investigations of the structure-function relationship of related flavonols showed the importance of two hydroxyl groups, located at C3 of the C ring and C4' of the B ring, both present in kaempferol, as important determinants of kaempferol as a Q biosynthetic precursor. Concurrently, through a mechanism not related to the enhancement of Q biosynthesis, kaempferol also augmented mitochondrial localization of Sirt3. The role of kaempferol as a precursor that increases Q levels, combined with its ability to upregulate Sirt3, identify kaempferol as a potential candidate in the design of interventions aimed on increasing endogenous Q biosynthesis, particularly in kidney. Copyright © 2017 Elsevier Inc. All rights reserved.

  6. Overexpression of PGC-1α Increases Fatty Acid Oxidative Capacity of Human Skeletal Muscle Cells

    Directory of Open Access Journals (Sweden)

    Nataša Nikolić

    2012-01-01

    Full Text Available We investigated the effects of PGC-1α (peroxisome proliferator-activated receptor γ coactivator-1α overexpression on the oxidative capacity of human skeletal muscle cells ex vivo. PGC-1α overexpression increased the oxidation rate of palmitic acid and mRNA expression of genes regulating lipid metabolism, mitochondrial biogenesis, and function in human myotubes. Basal and insulin-stimulated deoxyglucose uptake were decreased, possibly due to upregulation of PDK4 mRNA. Expression of fast fiber-type gene marker (MHCIIa was decreased. Compared to skeletal muscle in vivo, PGC-1α overexpression increased expression of several genes, which were downregulated during the process of cell isolation and culturing. In conclusion, PGC-1α overexpression increased oxidative capacity of cultured myotubes by improving lipid metabolism, increasing expression of genes involved in regulation of mitochondrial function and biogenesis, and decreasing expression of MHCIIa. These results suggest that therapies aimed at increasing PGC-1α expression may have utility in treatment of obesity and obesity-related diseases.

  7. Increasing Melanoma—Too Many Skin Cell Damages or Too Few Repairs?

    Directory of Open Access Journals (Sweden)

    Örjan Hallberg

    2013-02-01

    Full Text Available Skin melanoma rates have been increasing for a long time in many Western countries. The object of this study was to apply modern problem-solving theory normally used to clear industrial problems to search for roots and causes of this medical question. Increasing cancer rates can be due to too many cell damage incidents or to too few repairs. So far, it has been assumed that the melanoma epidemic mainly is caused by increasing sun tanning habits. In order to explore this problem in more detail, we used cancer statistics from several countries over time and space. Detailed analysis of data obtained and a model study to evaluate the effects from increased damages or decreased repairs clearly indicate that the main reason behind the melanoma problem is a disturbed immune system. The possibility to introduce efficient corrective actions is apparent.

  8. antiEGFR conjugated gold nanoparticles for increasing radiosensitivity in lung cancer cells

    International Nuclear Information System (INIS)

    Pujari, Geetanjali; Sarma, Asitikantha; Avasthi, Devesh K.

    2014-01-01

    One of the set back that lies in lung cancer treatment is the over expression of Epidermal Growth Factor Receptor (EGFR). EGFR is a transmembrane receptor that is highly expressed in lung cancer that leads to cell survival, proliferation and spread of the disease. Over the years, EGFR inhibitors, monoclonal antibodies, are being used in combination with radiotherapy in lung cancer patients so as to achieve better results. In the recent time, application of Au nanoparticles (AuNPs) in diagnosis and treatment of cancer has been extensively used in biomedical research. Among various applications, there is considerable use of AuNPs seen on the dose enhancement effect (radiosensitization) in radiation therapy of cancer. The conjugation of AuNP with monoclonal antibody antiEGFR (antiEGFR-AuNP) may provide excellent agent to sensitize the cells to heavy ion radiation. We synthesized AuNPs by citrate reduction method. Most of AuNPs were in the size range of 6-8 nm as studies by Transmission Electron Microscope (TEM). These AuNPs were found to be non toxic in A549 cells and thus biocompatible. Further, we conjugated AuNPs with antiEGFR (antiEGFR-AuNP). The conjugation was confirmed by UV-Vis spectroscopy. A549 cells were treated with antiEGFR-AuNP. TEM was carried out of ultrathin cross sections of antiEGFR-AuNP treated A549 cells to check the attachment internalization of AuNPs. We observed that the AuNPs are attached on the cell membrane as well as internalized in cytoplasm. Upon exposure of antiEGFR-AuNP treated cells to heavy ion 12 C beam, showed increase in radiosensitization as studied by survival assay and MTT assay. We will also explain the EGFR expression and cell cycle proliferation in A549 cells upon heavy ion beam irradiation of these. The study aims to overcome the current limitations of cancer-targeted therapies and improve the treatment modality of lung cancer. (author)

  9. Mitigation of radiation induced hematopoietic injury via regulation of Nrf-2 and increasing hematopoietic stem cells

    International Nuclear Information System (INIS)

    Patwardhan, R.S.; Sharma, Deepak; Checker, Rahul; Santosh Kumar, S.

    2014-01-01

    Therapeutic doses of ionizing radiation (IR) that can be delivered to tumors are restricted due to radiation induced damage to surrounding normal tissues thereby limiting the effectiveness of radiotherapy. Strategies to develop agents that selectively protect normal cells yielded limited success in the past. There is pressing need to develop safe, syndrome specific and effective radiation countermeasures to prevent or mitigate the harmful consequences of radiation exposure. Survival of bone marrow stem cells (HSCs) play a key role in protecting against IR induced hematopoietic injury. Many studies have shown manipulation of HSC frequency and/or survival as principal mechanism of radioprotection. It is known that, Nrf-2 plays crucial role in HSC survival and maintenance under oxidative stress conditions. In the present study, we have investigated the radioprotective ability of a flavonoid baicalein (5,6,7-trihydroxyflavone), extracted from the root of Scutellaria baicalensis Georgi, a medicinal plant traditionally used in Oriental medicine. There are numerous reports showing anti-inflammatory, anti-apoptotic, anti-oxidant, anti-cancer, anti-microbial, anti-mutagenic and neuroprotective properties of baicalein. Based on these reports, we have investigated the ability of baicalein to protect against radiation induced hematopoietic injury. Baicalein administration to mice protected against WBI induced mortality. Interestingly, the stem cell frequency increased in bone marrow cells obtained from baicalein administered mice as compared to vehicle treated mice. Baicalein treatment led to increased phospho-Nrf-2 levels in lineage negative BM-MNC. Administration of mice with Nrf-2 inhibitor prior to baicalein treatment led to significant abrogation of radioprotective ability of baicalein. This result suggests that, Nrf-2 may be playing a key role in baicalein mediated radioprotection. Here, we have shown that baicalein administration augments stem cell frequency, induces

  10. Protection against UVA-induced photooxidative damage in mammalian cell lines expressing increased levels of metallothionein

    International Nuclear Information System (INIS)

    Dudek, E.J.; Roth, R.M.

    1990-01-01

    Metallothionein (MT) is an endogenous low molecular weight protein that is inducible in a variety of eukaryotic cells and has the ability to selectivity bind heavy metal ions such as zinc and the cadmium. Although the exact physiological role of MT is still not understood, there is strong evidence that MT is involved in providing cellular resistance against the damaging effects of heavy metals and in the regulation of intracellular zinc and copper. Recently, it has been demonstrated that MT can scavenge radiation-induced reactive oxygen intermediates in vitro, specifically hydroxyl and superoxide radicals, and because of these observations it has been suggested that MT may provide protection against radiation-induced oxidative stress in vivo. Cell lines expressing increased levels of MT have demonstrated resistance to ionizing radiation, to ultraviolet radiation, and also to various DNA damaging agents including melphalan and cis-diaminedichloroplatinum. It is therefore important to gain some insight into the relationship between cellular MT content and cellular resistance to radiation and other DNA damaging agents. In this study we investigated the role of MT in providing protection against monochromatic 365-nm UVA radiation, which is known to generate intracellular reactive oxygen species that are involved in both DNA damage and cell killing. For this purpose, we used zinc acetate, a potent inducer of MT, to elevate MT levels in V79 Chinese hamster fibroblasts prior to UVA exposure and determined cell survival for uninduced and induced cultures. In order to eliminate any zinc effects other than MT induction, we also isolated and characterized cadmium chloride-resistant clones of V79 cells that have increased steady-state levels of both MT mRNA and protein, and we examined their survival characteristics against 365-nm radiation in the absence of zinc acetate. 14 refs., 3 figs

  11. Increased Intraepithelial Vα24 Invariant NKT Cells in the Celiac Duodenum

    Science.gov (United States)

    Montalvillo, Enrique; Bernardo, David; Martínez-Abad, Beatriz; Allegretti, Yessica; Fernández-Salazar, Luis; Calvo, Carmen; Chirdo, Fernando G.; Garrote, José A.; Arranz, Eduardo

    2015-01-01

    Celiac Disease (CD) is an interferon (IFN)γ-mediated duodenal hypersensitivity to wheat gluten occurring in genetically predisposed individuals. Gluten-free diet (GFD) leads to a complete remission of the disease. Vα24-restricted invariant NKT (iNKT) cells are important to maintain immune homeostasis in the gut mucosa because of their unique capacity to rapidly produce large quantities of both T-helper (Th)1 and Th2 cytokines upon stimulation. We studied the presence of these cells in the CD duodenum. Duodenal biopsies were obtained from 45 untreated-CD patients (uCD), 15 Gluten Free Diet-CD patients (GFD-CD), 44 non-inflamed non-CD controls (C-controls) and 15 inflamed non-CD controls (I-controls). Two populations from Spain and Argentina were recruited. Messenger RNA (mRNA) expression of Vα24-Jα18 (invariant TCRα chain of human iNKT cells), IFNγ and intracellular transcription factor Forkhead Box P3 (Foxp3), and flow cytometry intraepithelial lymphocyte (IEL) profile were determined. Both uCD and GFD-CD patients had higher Vα24-Jα18 mRNA levels than non-CD controls (I and C-controls). The expression of Vα24-Jα18 correlated with Marsh score for the severity of mucosal lesion and also with increased mRNA IFNγ levels. uCD and GFD-CD patients had decreased mRNA expression of FoxP3 but increased expression of Vα24-Jα18, which revealed a CD-like molecular profile. Increased numbers of iNKT cells were confirmed by flow cytometry within the intraepithelial lymphocyte compartment of uCD and GFD-CD patients and correlated with Vα24-Jα18 mRNA expression. In conclusion, we have found an increased number of iNKT cells in the duodenum from both uCD and GFD-CD patients, irrespective of the mucosal status. A CD-like molecular profile, defined by an increased mRNA expression of Vα24-Jα18 together with a decreased expression of FoxP3, may represent a pro-inflammatory signature of the CD duodenum. PMID:26529008

  12. Increased frequency of micronucleated exfoliated cells among humans exposed in vivo to mobile telephone radiations

    International Nuclear Information System (INIS)

    Manoj Kumar Sharma; Abhay Singh Yadav

    2007-01-01

    Complete text of publication follows. The health concerns have been raised following the enormous increase in the use of wireless mobile telephones through out the world. This investigation had been taken, with the motive to find out whether mobile phone radiations cause any in vivo effects on the frequency of micronucleated exfoliated cells in the exposed subjects. A total of 109 subjects including 85 regular mobile phone users (exposed) and 24 non-users (controls) had participated in this study. Exfoliated cells were obtained by swabbing the buccal-mucosa from exposed as well as sex-age-matched controls. One thousand exfoliated cells were screened from each individual for nuclear anomalies including micronuclei (MN), karyolysis (KL), karyorrhexis (KH), broken egg (BE) and bi-nucleated (BN) cells. The average daily duration of exposure to mobile phone radiations is 61.26 minutes with an overall average duration of exposure in term of years is 2.35 years in exposed subjects along with the 9.84±0.745 MNC (micronucleated cells) and 10.72±0.889 TMN (total micronuclei) as compared to zero duration of exposure along with average 3.75±0.774 MNC and 4.00±0.808 TMN in controls. The means are significantly different in case MNC and TMN at 0.01% level of significance. For all other nuclear anomalies (KL, KH, BE and BN cells) the means are found statistically nonsignificant. A positive correlation was found in the frequency of MNC and TMN with respect to duration of exposure time.

  13. The integrin alphav beta3 increases cellular stiffness and cytoskeletal remodeling dynamics to facilitate cancer cell invasion

    Science.gov (United States)

    Mierke, Claudia Tanja

    2013-01-01

    The process of cancer cell invasion through the extracellular matrix (ECM) of connective tissue plays a prominent role in tumor progression and is based fundamentally on biomechanics. Cancer cell invasion usually requires cell adhesion to the ECM through the cell-matrix adhesion receptors integrins. The expression of the αvβ3 integrin is increased in several tumor types and is consistently associated with increased metastasis formation in patients. The hypothesis was that the αvβ3 integrin expression increases the invasiveness of cancer cells through increased cellular stiffness, and increased cytoskeletal remodeling dynamics. Here, the invasion of cancer cells with different αvβ3 integrin expression levels into dense three-dimensional (3D) ECMs has been studied. Using a cell sorter, two subcell lines expressing either high or low amounts of αvβ3 integrins (αvβ3high or αvβ3low cells, respectively) have been isolated from parental MDA-MB-231 breast cancer cells. αvβ3high cells showed a threefold increased cell invasion compared to αvβ3low cells. Similar results were obtained for A375 melanoma, 786-O kidney and T24 bladder carcinoma cells, and cells in which the β3 integrin subunit was knocked down using specific siRNA. To investigate whether contractile forces are essential for αvβ3 integrin-mediated increased cellular stiffness and subsequently enhanced cancer cell invasion, invasion assays were performed in the presence of myosin light chain kinase inhibitor ML-7 and Rho kinase inhibitor Y27632. Indeed, cancer cell invasiveness was reduced after addition of ML-7 and Y27632 in αvβ3high cells but not in αvβ3low cells. Moreover, after addition of the contractility enhancer calyculin A, an increase in pre-stress in αvβ3low cells was observed, which enhanced cellular invasiveness. In addition, inhibition of the Src kinase, STAT3 or Rac1 strongly reduced the invasiveness of αvβ3high cells, whereas the invasiveness of β3 specific knock

  14. The integrin alphav beta3 increases cellular stiffness and cytoskeletal remodeling dynamics to facilitate cancer cell invasion

    International Nuclear Information System (INIS)

    Mierke, Claudia Tanja

    2013-01-01

    The process of cancer cell invasion through the extracellular matrix (ECM) of connective tissue plays a prominent role in tumor progression and is based fundamentally on biomechanics. Cancer cell invasion usually requires cell adhesion to the ECM through the cell-matrix adhesion receptors integrins. The expression of the αvβ3 integrin is increased in several tumor types and is consistently associated with increased metastasis formation in patients. The hypothesis was that the αvβ3 integrin expression increases the invasiveness of cancer cells through increased cellular stiffness, and increased cytoskeletal remodeling dynamics. Here, the invasion of cancer cells with different αvβ3 integrin expression levels into dense three-dimensional (3D) ECMs has been studied. Using a cell sorter, two subcell lines expressing either high or low amounts of αvβ3 integrins (αvβ3 high or αvβ3 low cells, respectively) have been isolated from parental MDA-MB-231 breast cancer cells. αvβ3 high cells showed a threefold increased cell invasion compared to αvβ3 low cells. Similar results were obtained for A375 melanoma, 786-O kidney and T24 bladder carcinoma cells, and cells in which the β3 integrin subunit was knocked down using specific siRNA. To investigate whether contractile forces are essential for αvβ3 integrin-mediated increased cellular stiffness and subsequently enhanced cancer cell invasion, invasion assays were performed in the presence of myosin light chain kinase inhibitor ML-7 and Rho kinase inhibitor Y27632. Indeed, cancer cell invasiveness was reduced after addition of ML-7 and Y27632 in αvβ3 high cells but not in αvβ3 low cells. Moreover, after addition of the contractility enhancer calyculin A, an increase in pre-stress in αvβ3 low cells was observed, which enhanced cellular invasiveness. In addition, inhibition of the Src kinase, STAT3 or Rac1 strongly reduced the invasiveness of αvβ3 high cells, whereas the invasiveness of β3 specific knock

  15. Increased sensitivity to ionizing radiation by targeting the homologous recombination pathway in glioma initiating cells.

    Science.gov (United States)

    Lim, Yi Chieh; Roberts, Tara L; Day, Bryan W; Stringer, Brett W; Kozlov, Sergei; Fazry, Shazrul; Bruce, Zara C; Ensbey, Kathleen S; Walker, David G; Boyd, Andrew W; Lavin, Martin F

    2014-12-01

    Glioblastoma is deemed the most malignant form of brain tumour, particularly due to its resistance to conventional treatments. A small surviving group of aberrant stem cells termed glioma initiation cells (GICs) that escape surgical debulking are suggested to be the cause of this resistance. Relatively quiescent in nature, GICs are capable of driving tumour recurrence and undergo lineage differentiation. Most importantly, these GICs are resistant to radiotherapy, suggesting that radioresistance contribute to their survival. In a previous study, we demonstrated that GICs had a restricted double strand break (DSB) repair pathway involving predominantly homologous recombination (HR) associated with a lack of functional G1/S checkpoint arrest. This unusual behaviour led to less efficient non-homologous end joining (NHEJ) repair and overall slower DNA DSB repair kinetics. To determine whether specific targeting of the HR pathway with small molecule inhibitors could increase GIC radiosensitivity, we used the Ataxia-telangiectasia mutated inhibitor (ATMi) to ablate HR and the DNA-dependent protein kinase inhibitor (DNA-PKi) to inhibit NHEJ. Pre-treatment with ATMi prior to ionizing radiation (IR) exposure prevented HR-mediated DNA DSB repair as measured by Rad51 foci accumulation. Increased cell death in vitro and improved in vivo animal survival could be observed with combined ATMi and IR treatment. Conversely, DNA-PKi treatment had minimal impact on GICs ability to resolve DNA DSB after IR with only partial reduction in cell survival, confirming the major role of HR. These results provide a mechanistic insight into the predominant form of DNA DSB repair in GICs, which when targeted may be a potential translational approach to increase patient survival. Copyright © 2014. Published by Elsevier B.V.

  16. Increasing the fidelity of noncanonical amino acid incorporation in cell-free protein synthesis.

    Science.gov (United States)

    Gan, Qinglei; Fan, Chenguang

    2017-11-01

    Cell-free protein synthesis provides a robust platform for co-translational incorporation of noncanonical amino acid (ncAA) into proteins to facilitate biological studies and biotechnological applications. Recently, eliminating the activity of release factor 1 has been shown to increase ncAA incorporation in response to amber codons. However, this approach could promote mis-incorporation of canonical amino acids by near cognate suppression. We performed a facile protocol to remove near cognate tRNA isoacceptors of the amber codon from total tRNAs, and used the phosphoserine (Sep) incorporation system as validation. By manipulating codon usage of target genes and tRNA species introduced into the cell-free protein synthesis system, we increased the fidelity of Sep incorporation at a specific position. By removing three near cognate tRNA isoacceptors of the amber stop codon [tRNA Lys , tRNA Tyr , and tRNA Gln (CUG)] from the total tRNA, the near cognate suppression decreased by 5-fold without impairing normal protein synthesis in the cell-free protein synthesis system. Mass spectrometry analyses indicated that the fidelity of ncAA incorporation was improved. Removal of near cognate tRNA isoacceptors of the amber codon could increase ncAA incorporation fidelity towards the amber stop codon in release factor deficiency systems. We provide a general strategy to improve fidelity of ncAA incorporation towards stop, quadruplet and sense codons in cell-free protein synthesis systems. This article is part of a Special Issue entitled "Biochemistry of Synthetic Biology - Recent Developments" Guest Editor: Dr. Ilka Heinemann and Dr. Patrick O'Donoghue. Copyright © 2016 Elsevier B.V. All rights reserved.

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

    Science.gov (United States)

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

    2013-11-01

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

  18. The increased concentration of 2,3-diphosphoglycerate in red blood cells of spontaneously hypertensive rats.

    Science.gov (United States)

    Przybylski, J; Skotnicka-Fedorowicz, B; Lisiecka, A; Siński, M; Abramczyk, P

    1997-12-01

    It has been recognised that high haemoglobin oxygen capacity is essential for the development of high blood pressure in spontaneously hypertensive rats. In the present study we have found increased concentration of 2,3 diphosphoglycerate (2,3-DPG) in red blood cells of spontaneously hypertensive rats (SHR) of Okamoto-Aoki strain. As 2,3-DPG is the major factor decreasing haemoglobin affinity to oxygen, our finding suggests that at given value of pO2 oxygen delivery to the tissue of SHR would be increased. Therefore increased concentration of 2,3-DPG in red blood cells of SHR would be of the pathophysiological meaning by promoting autoregulatory increase in total vascular resistance in this strain of rats. The mechanism responsible for enhanced synthesis of 2,3-DPG in SHR remains unclear. Intracellular alkalosis due to either hypocapnia and/or an enhanced activity of Na+/H+ antiporter occurring in SHR are the most plausible explanations for the above finding.

  19. Increased Cell Fusion in Cerebral Cortex May Contribute to Poststroke Regeneration

    Directory of Open Access Journals (Sweden)

    Alexander Paltsyn

    2013-01-01

    Full Text Available In this study, we used a model of a hemorrhagic stroke in a motor zone of the cortex in rats at the age of 3 months The report shows that cortical neurons can fuse with oligodendrocytes. In formed binuclear cells, the nucleus of an oligodendrocyte undergoes neuron specific reprogramming. It can be confirmed by changes in chromatin structure and in size of the second nucleus, by expression of specific neuronal markers and increasing total transcription rate. The nucleus of an oligodendrocyte likely transforms into a second neuronal nucleus. The number of binuclear neurons was validated with quantitative analysis. Fusion of neurons with oligodendrocytes might be a regenerative process in general and specifically following a stroke. The appearance of additional neuronal nuclei increases the functional outcome of the population of neurons. Participation of a certain number of binuclear cells in neuronal function might compensate for a functional deficit that arises from the death of a subset of neurons. After a stroke, the number of binuclear neurons increased in cortex around the lesion zone. In this case, the rate of recovery of stroke-damaged locomotor behavior also increased, which indicates the regenerative role of fusion.

  20. 'Cold shock' increases the frequency of homology directed repair gene editing in induced pluripotent stem cells.

    Science.gov (United States)

    Guo, Q; Mintier, G; Ma-Edmonds, M; Storton, D; Wang, X; Xiao, X; Kienzle, B; Zhao, D; Feder, John N

    2018-02-01

    Using CRISPR/Cas9 delivered as a RNA modality in conjunction with a lipid specifically formulated for large RNA molecules, we demonstrate that homology directed repair (HDR) rates between 20-40% can be achieved in induced pluripotent stem cells (iPSC). Furthermore, low HDR rates (between 1-20%) can be enhanced two- to ten-fold in both iPSCs and HEK293 cells by 'cold shocking' cells at 32 °C for 24-48 hours following transfection. This method can also increases the proportion of loci that have undergone complete sequence conversion across the donor sequence, or 'perfect HDR', as opposed to partial sequence conversion where nucleotides more distal to the CRISPR cut site are less efficiently incorporated ('partial HDR'). We demonstrate that the structure of the single-stranded DNA oligo donor can influence the fidelity of HDR, with oligos symmetric with respect to the CRISPR cleavage site and complementary to the target strand being more efficient at directing 'perfect HDR' compared to asymmetric non-target strand complementary oligos. Our protocol represents an efficient method for making CRISPR-mediated, specific DNA sequence changes within the genome that will facilitate the rapid generation of genetic models of human disease in iPSCs as well as other genome engineered cell lines.

  1. MicroRNA-101 inhibits cell proliferation, promotes cell apoptosis and increases sensitivity of breast cancer MDA-MB-231 cells to paclitaxel

    Directory of Open Access Journals (Sweden)

    Qiu-Lin Ke

    2016-02-01

    Full Text Available Objective: To explore the effect that miR-101 inhibits breast cancer MDA-MB-231 cell proliferation and increases the chemosensitivity of paclitaxel to breast cancer MDA-MB-231 cells and its influence on protein expression level of target gene Bcl2. Methods: miR-101 was artificially synthesized, it used liposome 3000 to transfect MDA-MB-231 cells, and experiment was divided into three groups: blank control group, negative control group and miR-101 group. MTT assay was used to detect the effect of miR-101 on MDA-MB-231 cell proliferation and chemosensitivity of paclitaxel-mediated MDA-MB-231 cells; flow cytometer was used to detect cell apoptosis. Real-time PCR and Western bloting were used to detect the changes of mRNA and protein expression levels of Bcl2. Results: After miR-101 transfected MDA-MB- 231 cells, cell proliferation ability significantly decreased compared with negative control group, and differences had statistical significance (P<0.01; after paclitaxel was used to process cells, IC50 of miR-101-processing group decreased by 2.45 times compared with blank control group, differences had statistical significance (P<0.05 and differences between blank control group and negative control group had no statistical significance; detection results by flow cytometer showed that both early-stage and late-stage apoptosis rates of MDA-MB-231 cells of miR-101 group were significantly higher than those of negative control group (P<0.05, and early-stage apoptosis rate was more significant (P<0.01; after transfection of miR-101, mRNA and protein levels of Bcl2 of MDA-MB-231 cells significantly decreased, and differences had statistical significance (P<0.05. Conclusion: miR-101 can inhibit breast cancer MDAMB- 231 cell proliferation through targeting and downregulating Bcl2, thereby increasing the chemosensitivity of breast cancer cells to paclitaxel and promoting cell apoptosis.

  2. Increased frequency of CD8+ and CD4+ regulatory T cells in chronic lymphocytic leukemia: association with disease progression.

    Science.gov (United States)

    Jadidi-Niaragh, Farhad; Yousefi, Mehdi; Memarian, Ali; Hojjat-Farsangi, Mohammad; Khoshnoodi, Jalal; Razavi, Seyed Mohsen; Jeddi-Tehrani, Mahmood; Shokri, Fazel

    2013-02-01

    Little is known regarding the immunobiology of regulatory T (Treg) cells in hematopoietic malignancies, particularly in chronic lymphocytic leukemia (CLL). In the present study, we showed that the frequencies of CD8(+) and CD4(+) Treg cells were significantly increased in progressive as compared with indolent CLL patients and normal subjects. Enriched CD4(+) Treg cells induced a similar level of inhibition in polyclonally activated B cells and effector T cells from CLL patients and normal subjects. Our results suggest that the increase in circulating Treg cells may result in downregulation of tumor-specific immune response, leading to tumor expansion and disease progression.

  3. Loss of CDKC;2 increases both cell division and drought tolerance in Arabidopsis thaliana.

    Science.gov (United States)

    Zhao, Lina; Li, Yaqiong; Xie, Qi; Wu, Yaorong

    2017-09-01

    Drought stress is one of the abiotic stresses that limit plant growth and agricultural productivity. To further understand the mechanism of drought tolerance and identify the genes involved in this process, a genetic screen for altered drought response was conducted in Arabidopsis. One mutant with enhanced drought tolerance was isolated and named Arabidopsis drought tolerance mutant 1 (atdtm1), which has larger lateral organs, prolonged growth duration, increased relative water content and a reduced leaf stomatal density compared with the wild type. The loss of AtDTM1 increases cell division during leaf development. The phenotype is caused by the loss of a T-DNA tagged gene encoding CYCLIN-DEPENDENT KINASE C;2 (CDKC;2), which functions in the regulation of transcription by influencing the phosphorylation status of RNA polymerase II (Pol II). Here, we show that CDKC;2 affects the transcription of downstream genes such as cell cycle genes and genes involved in stomatal development, resulting in altered plant organ size as well as drought tolerance of the plant. These results reveal the crucial role of CDKC;2 in modulating both cell division and the drought response in Arabidopsis. © 2017 The Authors The Plant Journal © 2017 John Wiley & Sons Ltd.

  4. Increased Fos expression among midbrain dopaminergic cell groups during birdsong tutoring.

    Science.gov (United States)

    Nordeen, E J; Holtzman, D A; Nordeen, K W

    2009-08-01

    During avian vocal learning, birds memorize conspecific song patterns and then use auditory feedback to match their vocal output to this acquired template. Some models of song learning posit that during tutoring, conspecific visual, social and/or auditory cues activate neuromodulatory systems that encourage acquisition of the tutor's song and attach incentive value to that specific acoustic pattern. This hypothesis predicts that stimuli experienced during social tutoring activate cell populations capable of signaling reward. Using immunocytochemistry for the protein product of the immediate early gene c-Fos, we found that brief exposure of juvenile male zebra finches to a live familiar male tutor increased the density of Fos+ cells within two brain regions implicated in reward processing: the ventral tegmental area (VTA) and substantia nigra pars compacta (SNc). This activation of Fos appears to involve both dopaminergic and non-dopaminergic VTA/SNc neurons. Intriguingly, a familiar tutor was more effective than a novel tutor in stimulating Fos expression within these regions. In the periaqueductal gray, a dopamine-enriched cell population that has been implicated in emotional processing, Fos labeling also was increased after tutoring, with a familiar tutor again being more effective than a novel conspecific. As several neural regions implicated in song acquisition receive strong dopaminergic projections from these midbrain nuclei, their activation in conjunction with hearing the tutor's song could help to establish sensory representations that later guide motor sequence learning.

  5. Circulating endothelial cells are increased in chronic myeloid leukemia blast crisis

    Directory of Open Access Journals (Sweden)

    C.R.T. Godoy

    2015-06-01

    Full Text Available We measured circulating endothelial precursor cells (EPCs, activated circulating endothelial cells (aCECs, and mature circulating endothelial cells (mCECs using four-color multiparametric flow cytometry in the peripheral blood of 84 chronic myeloid leukemia (CML patients and 65 healthy controls; and vascular endothelial growth factor (VEGF by quantitative real-time PCR in 50 CML patients and 32 healthy controls. Because of an increase in mCECs, the median percentage of CECs in CML blast crisis (0.0146% was significantly higher than in healthy subjects (0.0059%, P0.05. In addition, VEGF gene expression was significantly higher in all phases of CML: 0.245 in blast crisis, 0.320 in the active phase, and 0.330 in chronic phase patients than it was in healthy subjects (0.145. In conclusion, CML in blast crisis had increased levels of CECs and VEGF gene expression, which may serve as markers of disease progression and may become targets for the management of CML.

  6. Epithelial Cell Damage Activates Bactericidal/Permeability Increasing-Protein (BPI Expression in Intestinal Epithelium

    Directory of Open Access Journals (Sweden)

    Arjun Balakrishnan

    2017-08-01

    Full Text Available As the first line of defense against invading pathogen, intestinal epithelium produces various antimicrobial proteins (AMP that help in clearance of pathogen. Bactericidal/permeability-increasing protein (BPI is a 55 kDa AMP that is expressed in intestinal epithelium. Dysregulation of BPI in intestinal epithelium is associated with various inflammatory diseases like Crohn’s Disease, Ulcerative colitis, and Infectious enteritis’s. In this paper, we report a direct correlation between intestinal damage and BPI expression. In Caco-2 cells, we see a significant increase in BPI levels upon membrane damage mediated by S. aureus infection and pore-forming toxins (Streptolysin and Listeriolysin. Cells detect changes in potassium level as a Danger-associated molecular pattern associated with cell damage and induce BPI expression in a p38 dependent manner. These results are further supported by in vivo findings that the BPI expression in murine intestinal epithelium is induced upon infection with bacteria which cause intestinal damage (Salmonella Typhimurium and Shigella flexneri whereas mutants that do not cause intestinal damage (STM ΔfliC and STM ΔinvC did not induce BPI expression. Our results suggest that epithelial damage associated with infection act as a signal to induce BPI expression.

  7. Increased ATP generation in the host cell is required for efficient vaccinia virus production

    Directory of Open Access Journals (Sweden)

    Hsu Che-Fang

    2009-09-01

    Full Text Available Abstract To search for cellular genes up-regulated by vaccinia virus (VV infection, differential display-reverse transcription-polymerase chain reaction (ddRT-PCR assays were used to examine the expression of mRNAs from mock-infected and VV-infected HeLa cells. Two mitochondrial genes for proteins that are part of the electron transport chain that generates ATP, ND4 and CO II, were up-regulated after VV infection. Up-regulation of ND4 level by VV infection was confirmed by Western blotting analysis. Up-regulation of ND4 was reduced by the MAPK inhibitor, apigenin, which has been demonstrated elsewhere to inhibit VV replication. The induction of ND4 expression occurred after viral DNA replication since ara C, an inhibitor of poxviral DNA replication, could block this induction. ATP production was increased in the host cells after VV infection. Moreover, 4.5 μM oligomycin, an inhibitor of ATP production, reduced the ATP level 13 hr after virus infection to that of mock-infected cells and inhibited viral protein expression and virus production, suggesting that increased ATP production is required for efficient VV production. Our results further suggest that induction of ND4 expression is through a Bcl-2 independent pathway.

  8. Melatonin enhances neural stem cell differentiation and engraftment by increasing mitochondrial function.

    Science.gov (United States)

    Mendivil-Perez, Miguel; Soto-Mercado, Viviana; Guerra-Librero, Ana; Fernandez-Gil, Beatriz I; Florido, Javier; Shen, Ying-Qiang; Tejada, Miguel A; Capilla-Gonzalez, Vivian; Rusanova, Iryna; Garcia-Verdugo, José M; Acuña-Castroviejo, Darío; López, Luis Carlos; Velez-Pardo, Carlos; Jimenez-Del-Rio, Marlene; Ferrer, José M; Escames, Germaine

    2017-09-01

    Neural stem cells (NSCs) are regarded as a promising therapeutic approach to protecting and restoring damaged neurons in neurodegenerative diseases (NDs) such as Parkinson's disease and Alzheimer's disease (PD and AD, respectively). However, new research suggests that NSC differentiation is required to make this strategy effective. Several studies have demonstrated that melatonin increases mature neuronal markers, which reflects NSC differentiation into neurons. Nevertheless, the possible involvement of mitochondria in the effects of melatonin during NSC differentiation has not yet been fully established. We therefore tested the impact of melatonin on NSC proliferation and differentiation in an attempt to determine whether these actions depend on modulating mitochondrial activity. We measured proliferation and differentiation markers, mitochondrial structural and functional parameters as well as oxidative stress indicators and also evaluated cell transplant engraftment. This enabled us to show that melatonin (25 μM) induces NSC differentiation into oligodendrocytes and neurons. These effects depend on increased mitochondrial mass/DNA/complexes, mitochondrial respiration, and membrane potential as well as ATP synthesis in NSCs. It is also interesting to note that melatonin prevented oxidative stress caused by high levels of mitochondrial activity. Finally, we found that melatonin enriches NSC engraftment in the ND mouse model following transplantation. We concluded that a combined therapy involving transplantation of NSCs pretreated with pharmacological doses of melatonin could efficiently restore neuronal cell populations in PD and AD mouse models depending on mitochondrial activity promotion. © 2017 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  9. Exercise increases the frequency of circulating hematopoietic progenitor cells, but reduces hematopoietic colony-forming capacity.

    Science.gov (United States)

    Kroepfl, Julia Maria; Pekovits, Karin; Stelzer, Ingeborg; Fuchs, Robert; Zelzer, Sieglinde; Hofmann, Peter; Sedlmayr, Peter; Dohr, Gottfried; Wallner-Liebmann, Sandra; Domej, Wolfgang; Mueller, Wolfram

    2012-11-01

    Circulating hematopoietic progenitor cells (CPCs) may be triggered by physical exercise and/or normobaric hypoxia from the bone marrow. The aim of the study was to investigate the influence of physical exercise and normobaric hypoxia on CPC number and functionality in the peripheral blood as well as the involvement of oxidative stress parameters as possibly active agents. Ten healthy male subjects (25.3±4.4 years) underwent a standardized cycle incremental exercise test protocol (40 W+20 W/min) under either normoxic (FiO2 ∼0.21) or hypoxic conditions (FiO2exercise. The number of CPCs in the peripheral blood was analyzed by flow cytometry (CD34/CD45-positive cells). The functionality of cells present was addressed by secondary colony-forming unit-granulocyte macrophage (CFU-GM) assays. To determine a possible correlation between the mobilization of CPCs and reactive oxygen species, parameters for oxidative stress such as malondialdehyde (MDA) and myeloperoxidase (MPO) were obtained. Data showed a significant increase of CPC release under normoxic as well as hypoxic conditions after 10 min of recovery (Pexercise (Pexercise, possibly due to the influence of increased oxidative stress levels.

  10. Tumor necrosis factor alpha increases epithelial barrier permeability by disrupting tight junctions in Caco-2 cells

    Directory of Open Access Journals (Sweden)

    W. Cui

    2010-04-01

    Full Text Available The objectives of this study were to determine the effect of tumor necrosis factor alpha (TNF-α on intestinal epithelial cell permeability and the expression of tight junction proteins. Caco-2 cells were plated onto Transwell® microporous filters and treated with TNF-α (10 or 100 ng/mL for 0, 4, 8, 16, or 24 h. The transepithelial electrical resistance and the mucosal-to-serosal flux rates of the established paracellular marker Lucifer yellow were measured in filter-grown monolayers of Caco-2 intestinal cells. The localization and expression of the tight junction protein occludin were detected by immunofluorescence and Western blot analysis, respectively. SYBR-Green-based real-time PCR was used to measure the expression of occludin mRNA. TNF-α treatment produced concentration- and time-dependent decreases in Caco-2 transepithelial resistance and increases in transepithelial permeability to the paracellular marker Lucifer yellow. Western blot results indicated that TNF-α decreased the expression of phosphorylated occludin in detergent-insoluble fractions but did not affect the expression of non-phosphorylated occludin protein. Real-time RT-PCR data showed that TNF-α did not affect the expression of occludin mRNA. Taken together, our data demonstrate that TNF-α increases Caco-2 monolayer permeability, decreases occludin protein expression and disturbs intercellular junctions.

  11. Cells Lacking mtDNA Display Increased dNTP Pools upon DNA Damage

    DEFF Research Database (Denmark)

    Skovgaard, Tine; Rasmussen, Lene Juel; Munch-Petersen, Birgitte

    Imbalanced dNTP pools are highly mutagenic due to a deleterious effect on DNA polymerase fidelity. Mitochondrial DNA defects, including mutations and deletions, are commonly found in a wide variety of different cancer types. In order to further study the interconnection between dNTP pools...... and mitochondrial function we have examined the effect of DNA damage on dNTP pools in cells deficient of mtDNA. We show that DNA damage induced by UV irradiation, in a dose corresponding to LD50, induces an S phase delay in different human osteosarcoma cell lines. The UV pulse also has a destabilizing effect...... shows that normal mitochondrial function is prerequisite for retaining stable dNTP pools upon DNA damage. Therefore it is likely that mitochondrial deficiency defects may cause an increase in DNA mutations by disrupting dNTP pool balance....

  12. Increased DNA damage in blood cells of rat treated with lead as assessed by comet assay

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    Mohammad Arif

    2008-06-01

    Full Text Available A growing body of evidence suggests that oxidative stress is the key player in the pathogenesis of lead-induced toxicity. The present study investigated lead induced oxidative DNA damage, if any in rat blood cells by alkaline comet assay. Lead was administered intraperitoneally to rats at doses of 25, 50 and 100 mg/kg body weight for 5 days consecutively. Blood collected on day six from sacrificed lead-treated rats was used to assess the extent of DNA damage by comet assay which entailed measurement of comet length, olive tail moment, tail DNA (% and tail length. The results showed that treatment with lead significantly increased DNA damage in a dose-dependent manner. Therefore, our data suggests that lead treatment is associated with oxidative stress-induced DNA damage in rat blood cells which could be used as an early bio-marker of lead-toxicity.

  13. Increased numbers of Foxp3-positive regulatory T cells in gastritis, peptic ulcer and gastric adenocarcinoma

    Science.gov (United States)

    Cheng, Hsin-Hung; Tseng, Guan-Ying; Yang, Hsiao-Bai; Wang, Hung-Jung; Lin, Hwai-Jeng; Wang, Wen-Ching

    2012-01-01

    AIM: To determine the number of regulatory T cells (Tregs) in gastric mucosa of patients with gastritis, peptic ulcers and gastric cancer. METHODS: This study was a retrospective analysis of gastric antrum biopsy specimens from healthy controls (n = 22) and patients with gastritis (n = 30), peptic ulcer (n = 83), or gastric cancer (n = 32). Expression of CD4, CD25 and Foxp3 was determined by immunohistochemistry in three consecutive sections per sample. RESULTS: Compared with healthy controls, there was an increased number of CD25+ and Foxp3+ cells in patients with gastritis (P = 0.004 and P = 0.008), peptic ulcer (P gastritis (P gastritis and peptic ulcer groups. PMID:22228968

  14. EMMPRIN overexpression in SVZ neural progenitor cells increases their migration towards ischemic cortex.

    Science.gov (United States)

    Kanemitsu, Michiko; Tsupykov, Oleg; Potter, Gaël; Boitard, Michael; Salmon, Patrick; Zgraggen, Eloisa; Gascon, Eduardo; Skibo, Galina; Dayer, Alexandre G; Kiss, Jozsef Z

    2017-11-01

    Stimulation of endogenous neurogenesis and recruitment of neural progenitors from the subventricular zone (SVZ) neurogenic site may represent a useful strategy to improve regeneration in the ischemic cortex. Here, we tested whether transgenic overexpression of extracellular matrix metalloproteinase inducer (EMMPRIN), the regulator of matrix metalloproteinases (MMPs) expression, in endogenous neural progenitor cells (NPCs) in the subventricular zone (SVZ) could increase migration towards ischemic injury. For this purpose, we applied a lentivector-mediated gene transfer system. We found that EMMPRIN-transduced progenitors exhibited enhanced MMP-2 activity in vitro and showed improved motility in 3D collagen gel as well as in cortical slices. Using a rat model of neonatal ischemia, we showed that EMMPRIN overexpressing SVZ cells invade the injured cortical tissue more efficiently than controls. Our results suggest that EMMPRIN overexpression could be suitable approach to improve capacities of endogenous or transplanted progenitors to invade the injured cortex. Copyright © 2017 Elsevier Inc. All rights reserved.

  15. Oncogenic Nras has bimodal effects on stem cells that sustainably increase competitiveness.

    Science.gov (United States)

    Li, Qing; Bohin, Natacha; Wen, Tiffany; Ng, Victor; Magee, Jeffrey; Chen, Shann-Ching; Shannon, Kevin; Morrison, Sean J

    2013-12-05

    'Pre-leukaemic' mutations are thought to promote clonal expansion of haematopoietic stem cells (HSCs) by increasing self-renewal and competitiveness; however, mutations that increase HSC proliferation tend to reduce competitiveness and self-renewal potential, raising the question of how a mutant HSC can sustainably outcompete wild-type HSCs. Activating mutations in NRAS are prevalent in human myeloproliferative neoplasms and leukaemia. Here we show that a single allele of oncogenic Nras(G12D) increases HSC proliferation but also increases reconstituting and self-renewal potential upon serial transplantation in irradiated mice, all prior to leukaemia initiation. Nras(G12D) also confers long-term self-renewal potential to multipotent progenitors. To explore the mechanism by which Nras(G12D) promotes HSC proliferation and self-renewal, we assessed cell-cycle kinetics using H2B-GFP label retention and 5-bromodeoxyuridine (BrdU) incorporation. Nras(G12D) had a bimodal effect on HSCs, increasing the frequency with which some HSCs divide and reducing the frequency with which others divide. This mirrored bimodal effects on reconstituting potential, as rarely dividing Nras(G12D) HSCs outcompeted wild-type HSCs, whereas frequently dividing Nras(G12D) HSCs did not. Nras(G12D) caused these effects by promoting STAT5 signalling, inducing different transcriptional responses in different subsets of HSCs. One signal can therefore increase HSC proliferation, competitiveness and self-renewal through bimodal effects on HSC gene expression, cycling and reconstituting potential.

  16. Erythropoetin treatment can increase 2,3-diphosphoglycerate levels in red blood cells.

    Science.gov (United States)

    Birgegård, G; Sandhagen, B

    2001-01-01

    Some patients experience an improved well-being during treatment with recombinant human erythropoietin even with an unchanged Hb level. We have hypothesized that this may not be only a placebo effect. 2,3-diphosphoglycerate (2,3-DPG) in red blood cells increases in response to anaemia/hypoxia and causes a shift of the oxygen dissociation curve, allowing a more effective oxygen delivery. We have investigated red cell 2,3-DPG concentrations during erythropoietin treatment in healthy volunteers as a mediator of a possible physiological explanation. Thirteen healthy subjects with no iron deficiency were recruited and randomly assigned to a treatment group comprising five males and three females and a control group including three males and two females. The treatment group was treated with erythropoietin (Recormon), 20 IE/kg subcutaneously three times/week for 4 weeks. Blood samples were collected at each injection day and 10 days after the last injection and at corresponding times in the control group. B-Hb, red cell 2,3-DPG and P50 were measured by standard techniques and oxygen-releasing capacity was calculated. due to the sampling (26 ml each time, three times/week) the mean Hb level was lowered from 140.5 +/- 5.9 to 128.6 +/- 10.4 g/L in the control group whereas the erythropoietin treatment group maintained a mean Hb level of about 142 g/L (plevel curve as well as that for oxygen releasing capacity also differed significantly between the two groups (p levels. treatment with erythropoietin causes an increase in red cell 2,3-DPG levels.

  17. Ciprofloxacin Derivatives Affect Parasite Cell Division and Increase the Survival of Mice Infected with Toxoplasma gondii.

    Directory of Open Access Journals (Sweden)

    Erica S Martins-Duarte

    Full Text Available Toxoplasmosis, caused by the protozoan Toxoplasma gondii, is a worldwide disease whose clinical manifestations include encephalitis and congenital malformations in newborns. Previously, we described the synthesis of new ethyl-ester derivatives of the antibiotic ciprofloxacin with ~40-fold increased activity against T. gondii in vitro, compared with the original compound. Cipro derivatives are expected to target the parasite's DNA gyrase complex in the apicoplast. The activity of these compounds in vivo, as well as their mode of action, remained thus far uncharacterized. Here, we examined the activity of the Cipro derivatives in vivo, in a model of acute murine toxoplasmosis. In addition, we investigated the cellular effects T. gondii tachyzoites in vitro, by immunofluorescence and transmission electron microscopy (TEM. When compared with Cipro treatment, 7-day treatments with Cipro derivatives increased mouse survival significantly, with 13-25% of mice surviving for up to 60 days post-infection (vs. complete lethality 10 days post-infection, with Cipro treatment. Light microscopy examination early (6 and 24h post-infection revealed that 6-h treatments with Cipro derivatives inhibited the initial event of parasite cell division inside host cells, in an irreversible manner. By TEM and immunofluorescence, the main cellular effects observed after treatment with Cipro derivatives and Cipro were cell scission inhibition--with the appearance of 'tethered' parasites--malformation of the inner membrane complex, and apicoplast enlargement and missegregation. Interestingly, tethered daughter cells resulting from Cipro derivatives, and also Cipro, treatment did not show MORN1 cap or centrocone localization. The biological activity of Cipro derivatives against C. parvum, an apicomplexan species that lacks the apicoplast, is, approximately, 50 fold lower than that in T. gondii tachyzoites, supporting that these compounds targets the apicoplast. Our results

  18. Increased storage and secretion of phosphatidylcholines by senescent human peritoneal mesothelial cells.

    Science.gov (United States)

    Bartosova, Maria; Rudolf, Andras; Pichl, Sebastian; Schmidt, Kathrin; Okun, Jürgen G; Straub, Beate K; Rutkowski, Rafael; Witowski, Janusz; Schmitt, Claus P

    2016-08-01

    Human peritoneal mesothelial cells (HPMC) secrete phosphatidylcholines (PC) which form a lipid bilayer lining the peritoneum. They prevent frictions and adhesions and act as a barrier to the transport of water-soluble solutes while permitting water flux. PC may play an essential role in peritoneal integrity and function, the role of PD induced HPMC senescence on PC homeostasis, however, is unknown. HPMC cell lines were isolated from four non-uremic patients. Expression of the three PC synthesis genes (rt-PCR), and cellular storage and secretion of PC (ESI-mass-spectrometry) were analyzed in young and senescent HPMC (>Hayflick-limit). Senescent cells displayed significantly altered morphology; flow cytometry demonstrated extensive staining for senescence-associated beta galactosidase. Nine different PC were detected in HPMC with palmitoyl-myristoyl phosphatidylcholine (PMPC) being most abundant. In senescent HPMC mRNA expression of the three key PC synthesis genes was 1.5-, 2.4- and 6-fold increased as compared to young HPMC, with the latter, phosphatidylcholine cytidylyltransferase, being rate limiting. Intracellular storage of the nine PC was 75-450 % higher in senescent vs. young HPMC, PC secretion rates were 100-300 % higher. Intracellular PC concentrations were not correlated with the PC secretion rates. Electron microscopy demonstrated lamellar bodies, the primary storage site of PC, in senescent but not in young cells. Senescent HPMC store and secrete substantially more PC than young cells. Our findings indicate a novel protective mechanism, which should counteract peritoneal damage induced by chronic exposure to PD fluids.

  19. Increasing the operation temperature of polymer electrolyte membranes for fuel cells: From nanocomposites to hybrids

    Science.gov (United States)

    Licoccia, Silvia; Traversa, Enrico

    Among the possible systems investigated for energy production with low environmental impact, polymeric electrolyte membrane fuel cells (PEMFCs) are very promising as electrochemical power sources for application in portable technology and electric vehicles. For practical applications, operating FCs at temperatures above 100 °C is desired, both for hydrogen and methanol fuelled cells. When hydrogen is used as fuel, an increase of the cell temperature produces enhanced CO tolerance, faster reaction kinetics, easier water management and reduced heat exchanger requirement. The use of methanol instead of hydrogen as a fuel for vehicles has several practical benefits such as easy transport and storage, but the slow oxidation kinetics of methanol needs operating direct methanol fuel cells (DMFCs) at intermediate temperatures. For this reason, new membranes are required. Our strategy to achieve the goal of operating at temperatures above 120 °C is to develop organic/inorganic hybrid membranes. The first approach was the use of nanocomposite class I hybrids where nanocrystalline ceramic oxides were added to Nafion. Nanocomposite membranes showed enhanced characteristics, hence allowing their operation up to 130 °C when the cell was fuelled with hydrogen and up to 145 °C in DMFCs, reaching power densities of 350 mW cm -2. The second approach was to prepare Class II hybrids via the formation of covalent bonds between totally aromatic polymers and inorganic clusters. The properties of such covalent hybrids can be modulated by modifying the ratio between organic and inorganic groups and the nature of the chemical components allowing to reach high and stable conductivity values up to 6.4 × 10 -2 S cm -1 at 120 °C.

  20. Increased genome instability is not accompanied by sensitivity to DNA damaging agents in aged yeast cells

    NARCIS (Netherlands)

    Novarina, Daniele; Mavrova, Sara N.; Janssens, Georges E.; Rempel, Irina L.; Veenhoff, Liesbeth M.; Chang, Michael

    The budding yeast Saccharomyces cerevisiae divides asymmetrically, producing a new daughter cell from the original mother cell. While daughter cells are born with a full lifespan, a mother cell ages with each cell division and can only generate on average 25 daughter cells before dying. Aged yeast

  1. Lipofection indirectly increases expression of endogenous major histocompatibility complex class I molecules on tumor cells.

    Science.gov (United States)

    Fox, B A; Drury, M; Hu, H M; Cao, Z; Huntzicker, E G; Qie, W; Urba, W J

    1998-01-01

    Direct intratumoral injection of a lipid/DNA complex encoding an allogeneic major histocompatibility complex (MHC) class I molecule leads to regression of both an immunogenic murine tumor and also melanoma lesions in some patients. We have sought to understand the mechanism(s) for this augmentation of antitumor activity. While optimizing parameters for in vitro gene transfer into the D5 subclone of B16BL6, it was noted that lipofected tumors not only expressed the new alloantigen but also exhibited increased expression of endogenous MHC class I, both H-2 Kb and H-2 Db. This increase in expression was not restricted to the small percentage of cells that expressed the transfected gene, but appeared to affect the majority of cells in culture. Class I expression was not increased by lipopolysaccharide, DNA alone, lipid, or lipid/lipopolysaccharide mixtures. Enhanced class I expression required a DNA/lipid complex and was greatest when parameters optimized for gene transfer of the alloantigen were used. All DNA plasmids tested had this effect, including one plasmid whose DNA was not transcribed because it lacked an expression cassette. Because of the critical role that MHC class I antigens play in immune recognition, we propose that lipid complex-mediated gene transfer may provide immunological advantages beyond those that are attributable to expression of the specific gene transferred.

  2. Physiological red cell, 2,3-diphosphoglycerate increase by the sixth hour after birth.

    Science.gov (United States)

    Barretto, O C; Nonoyama, K; Deutsch, A D; Ramos, J L

    1995-01-01

    As the available hemoglobin A1 at birth ranges from 20 to 30% a possible mechanism to favor oxygen release to the tissues could be a decrease of hemoglobin A1 affinity to oxygen. This may be accomplished by an increase in blood pH soon after birth and by an elevation in red cell 2,3-diphosphoglycerate (2,3-DPG). This hypothesis is supported by Valleri and Hirsch, who described a rapid 2,3-DPG recovery of transfused depleted 2,3-DPG red cells. That being so, we carried out this current study by assaying the 2,3-DPG of cord blood from 22 newborns and at 6, 24 and 72 hours after birth, as well as those enzymes assumed to be envolved in the 2,3-DPG levels regulation. 2,3-DPG (nmoles g-1 Hb) demonstrated the following values: cord blood: 9,770 +/- 1,026; 6h: 12,773 +/- 1,726; 72 h: 11,990 +/- 728, unveiling a distinct behavior of a sharp increase of 30% by the sixth hour. This confirmed our hypothesis. Regarding the metabolic mechanisms which can account for the 2,3-DPG increase, besides the rise of blood pH, we detected a significant decrease of the 2,3-DPG phosphatase activity, which might diminish the 2,3-DPG breakdown.

  3. SadA-Expressing Staphylococci in the Human Gut Show Increased Cell Adherence and Internalization

    Directory of Open Access Journals (Sweden)

    Arif Luqman

    2018-01-01

    Full Text Available Summary: A subgroup of biogenic amines, the so-called trace amines (TAs, are produced by mammals and bacteria and can act as neuromodulators. In the genus Staphylococcus, certain species are capable of producing TAs through the activity of staphylococcal aromatic amino acid decarboxylase (SadA. SadA decarboxylates aromatic amino acids to produce TAs, as well as dihydroxy phenylalanine and 5-hydroxytryptophan to thus produce the neurotransmitters dopamine and serotonin. SadA-expressing staphylococci were prevalent in the gut of most probands, where they are part of the human intestinal microflora. Furthermore, sadA-expressing staphylococci showed increased adherence to HT-29 cells and 2- to 3-fold increased internalization. Internalization and adherence was also increased in a sadA mutant in the presence of tryptamine. The α2-adrenergic receptor is required for enhanced adherence and internalization. Thus, staphylococci in the gut might contribute to gut activity and intestinal colonization. : Luqman et al. examine the sadA gene and argue that it contributes to TAs. They found that neuromodulator-producing staphylococci were present in the gut of most probands. The produced neuromodulators enhanced the adherence and internalization of staphylococci to cells in culture. Keywords: adherence, aromatic amino acid decarboxylase, gut microbiota, internalization, neuromodulator, neurotransmitter, staphylococcus

  4. Fatty Acid Biosynthesis Inhibition Increases Reduction Potential in Neuronal Cells under Hypoxia

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    Stephen A Brose

    2016-11-01

    Full Text Available Recently, we have reported a novel neuronal specific pathway for adaptation to hypoxia through increased fatty acid (FA biosynthesis (FAS followed by esterification into lipids. However, the biological role of this pathway under hypoxia remains to be elucidated. In the presented study, we have tested our hypothesis that activation of FAS maintains reduction potential and reduces lactoacidosis in neuronal cells under hypoxia. To address this hypothesis, we measured the effect of FAS inhibition on NADH2+/NAD+ and NADPH2+/NADP+ ratios, and lactic acid levels in neuronal SH-SY5Y cells exposed to normoxic and hypoxic conditions. FAS inhibitors, TOFA (inhibits Acetyl-CoA carboxylase and cerulenin (inhibits FA synthase, increased NADH2+/NAD+ and NADPH2+/NADP+ ratios under hypoxia. Further, FAS inhibition increased lactic acid under both normoxic and hypoxic conditions, and caused cytotoxicity under hypoxia but not normoxia. These results indicate that FA may serve as hydrogen acceptors under hypoxia, thus supporting oxidation reactions including anaerobic glycolysis. These findings may help to identify a radically different approach to attenuate hypoxia related pathophysiology in the nervous system including stroke.

  5. Fatty Acid Biosynthesis Inhibition Increases Reduction Potential in Neuronal Cells under Hypoxia.

    Science.gov (United States)

    Brose, Stephen A; Golovko, Svetlana A; Golovko, Mikhail Y

    2016-01-01

    Recently, we have reported a novel neuronal specific pathway for adaptation to hypoxia through increased fatty acid (FA) biosynthesis followed by esterification into lipids. However, the biological role of this pathway under hypoxia remains to be elucidated. In the presented study, we have tested our hypothesis that activation of FA synthesis maintains reduction potential and reduces lactoacidosis in neuronal cells under hypoxia. To address this hypothesis, we measured the effect of FA synthesis inhibition on [Formula: see text]/NAD + and [Formula: see text]/NADP + ratios, and lactic acid levels in neuronal SH-SY5Y cells exposed to normoxic and hypoxic conditions. FA synthesis inhibitors, TOFA (inhibits Acetyl-CoA carboxylase) and cerulenin (inhibits FA synthase), increased [Formula: see text]/NAD + and [Formula: see text]/NADP + ratios under hypoxia. Further, FA synthesis inhibition increased lactic acid under both normoxic and hypoxic conditions, and caused cytotoxicity under hypoxia but not normoxia. These results indicate that FA may serve as hydrogen acceptors under hypoxia, thus supporting oxidation reactions including anaerobic glycolysis. These findings may help to identify a radically different approach to attenuate hypoxia related pathophysiology in the nervous system including stroke.

  6. Pseudogene PHBP1 promotes esophageal squamous cell carcinoma proliferation by increasing its cognate gene PHB expression.

    Science.gov (United States)

    Feng, Feiyue; Qiu, Bin; Zang, Ruochuan; Song, Peng; Gao, Shugeng

    2017-04-25

    Natural antisense transcripts (NATs) as one of the most diverse classes of long noncoding RNAs (lncRNAs), have been demonstrated involved in fundamental biological processes in human. Here, we reported that human prohibitin gene pseudogene 1 (PHBP1) was upregulated in ESCC, and increased PHBP1 expression in ESCC was associated with clinical advanced stage. Functional experiments showed that PHBP1 knockdown inhibited ESCC cells proliferation, colony formation and xenograft tumor growth in vitro and in vivo by causing cell-cycle arrest at the G1-G0 phase. Mechanisms analysis revealed that PHBP1 transcript as an antisense transcript of PHB is partially complementary to PHB mRNA and formed an RNA-RNA hybrid with PHB, consequently inducing an increase of PHB expression at both the mRNA and protein levels. Furthermore, PHBP1 expression is strongly correlated with PHB expression in ESCC tissues. Collectively, this study elucidates an important role of PHBP1 in promoting ESCC partly via increasing PHB expression.

  7. Protein Carbamylation: A Marker Reflecting Increased Age-Related Cell Oxidation

    Directory of Open Access Journals (Sweden)

    Julia Carracedo

    2018-05-01

    Full Text Available Carbamylation is a post-translational modification of proteins that may partake in the oxidative stress-associated cell damage, and its increment has been recently proposed as a “hallmark of aging”. The molecular mechanisms associated with aging are related to an increased release of free radicals. We have studied whether carbamylated proteins from the peripheral blood of healthy subjects are related to oxidative damage and aging, taking into account the gender and the immune profile of the subjects. The study was performed in healthy human volunteers. The detection of protein carbamylation and malondialdehyde (MDA levels was evaluated using commercial kits. The immune profile was calculated using parameters of immune cell function. The results show that the individuals from the elderly group (60–79 years old have increased carbamylated protein and MDA levels. When considered by gender, only men between 60 and 79 years old showed significantly increased carbamylated proteins and MDA levels. When those subjects were classified by their immune profile, the carbamylated protein levels were higher in those with an older immune profile. In conclusion, the carbamylation of proteins in peripheral blood is related to age-associated oxidative damage and to an aging functional immunological signature. Our results suggest that carbamylated proteins may play an important role at the cellular level in the aging process.

  8. Increased ICAM-1 Expression in Transformed Human Oral Epithelial Cells: Molecular Mechanism and Functional Role in Peripheral Blood Mononuclear Cell Adhesion and Lymphokine-Activated-Killer Cell Cytotoxicity

    Science.gov (United States)

    Huang, George T.-J.; Zhang, Xinli; Park, No-Hee

    2012-01-01

    The intercellular adhesion molecule-1 (ICAM-1, CD54) serves as a counter-receptor for the β2-integrins, LFA-1 and Mac-1, which are expressed on leukocytes. Although expression of ICAM-1 on tumor cells has a role in tumor progression and development, information on ICAM-1 expression and its role in oral cancer has not been established. Normal human oral keratinocytes (NHOK), human papilloma virus (HPV)-immortalized human oral keratinocyte lines (HOK-16B, HOK-18A, and HOK-18C), and six human oral neoplastic cell lines (HOK-16B-BaP-T1, SCC-4, SCC-9, HEp-2, Tu-177 and 1483) were used to study ICAM-1 expression and its functional role in vitro. Our results demonstrated that NHOK express negligible levels of ICAM-1, whereas immortalized human oral keratinocytes and cancer cells express significantly higher levels of ICAM-1, except for HOK-16B-BaP-T1 and HEp-2. Altered mRNA half-lives did not fully account for the increased accumulation of ICAM-1 mRNA. Adhesion of peripheral blood mononuclear cells (PBMC) to epithelial cells correlated with cell surface ICAM-1 expression levels. This adhesion was inhibited by antibodies specific for either ICAM-1 or LFA-1/Mac-1, suggesting a role for these molecules in adhesion. In contrast, lymphokine-activated-killer (LAK) cell cytotoxic killing of epithelial cells did not correlate with ICAM-1 levels or with adhesion. Nonetheless, within each cell line, blocking of ICAM-1 or LFA-1/Mac-1 reduced LAK cells killing, suggesting that ICAM-1 is involved in mediating this killing. PMID:10938387

  9. The hydroxypyridinone iron chelator CP94 increases methyl-aminolevulinate-based photodynamic cell killing by increasing the generation of reactive oxygen species

    Directory of Open Access Journals (Sweden)

    Yuktee Dogra

    2016-10-01

    Full Text Available Methyl-aminolevulinate-based photodynamic therapy (MAL-PDT is utilised clinically for the treatment of non-melanoma skin cancers and pre-cancers and the hydroxypyridinone iron chelator, CP94, has successfully been demonstrated to increase MAL-PDT efficacy in an initial clinical pilot study. However, the biochemical and photochemical processes leading to CP94-enhanced photodynamic cell death, beyond the well-documented increases in accumulation of the photosensitiser protoporphyrin IX (PpIX, have not yet been fully elucidated. This investigation demonstrated that MAL-based photodynamic cell killing of cultured human squamous carcinoma cells (A431 occurred in a predominantly necrotic manner following the generation of singlet oxygen and ROS. Augmenting MAL-based photodynamic cell killing with CP94 co-treatment resulted in increased PpIX accumulation, MitoSOX-detectable ROS generation (probably of mitochondrial origin and necrotic cell death, but did not affect singlet oxygen generation. We also report (to our knowledge, for the first time the detection of intracellular PpIX-generated singlet oxygen in whole cells via electron paramagnetic resonance spectroscopy in conjunction with a spin trap.

  10. Downregulated regulatory T cell function is associated with increased peptic ulcer in Helicobacter pylori-infection.

    Science.gov (United States)

    Bagheri, Nader; Shirzad, Hedayatollah; Elahi, Shokrollah; Azadegan-Dehkordi, Fatemeh; Rahimian, Ghorbanali; Shafigh, Mohammedhadi; Rashidii, Reza; Sarafnejad, Abdulfatah; Rafieian-Kopaei, Mahmoud; Faridani, Rana; Tahmasbi, Kamran; Kheiri, Soleiman; Razavi, Alireza

    2017-09-01

    Helicobacter pylori (H. pylori) chronically colonizes gastric/duodenal mucosa and induces gastroduodenal disease such as gastritis and peptic ulcer and induces vigorous innate and specific immune responses; however, the infection is not removed, a state of chronic active gastritis persists for life if untreated. The objective of this study was to determine the number of regulatory T cells (Tregs) in gastric mucosa of patients with gastritis and peptic ulcer and determined the relationship between main virulence factor of H. pylori and Tregs. A total of 89 patients with gastritis, 63 patients with peptic ulcer and 40 healthy, H. pylori-negative subjects were enrolled in this study. Expression of CD4 and Foxp3 was determined by immunohistochemistry. Antrum biopsy was obtained for detection of H. pylori, bacterial virulence factors and histopathological assessments. TGF-β1, IL-10 and FOXP3 expressions were determined by real-time polymerase chain reaction (qPCR). The numbers of CD4 + and Foxp3 + T cells as well as the expression of IL-10, TGF-β1, FOXP3, INF-γ and IL-17A in infected patients were significantly higher than the ones in uninfected patients. Also, the number of CD4 + T cells was independent on the vacuolating cytotoxin A (vacA) and outer inflammatory protein A (oipA), but it was positively correlated with cytotoxin-associated gene A (cagA). Instead, the number of Foxp3 + T cells was dependent on the vacA and oipA, but it was independent on cagA. The number of Foxp3 + T cells and the expression of IL-10, TGF-β1 and FOXP3 in infected patients with gastritis were significantly higher than the ones in infected patients with peptic ulcer. Moreover, the number of CD4 + T cells and the expression of IL-17A and INF-γ was the lowest in the gastritis patients, however, increased progressively in the peptic ulcer patients. Additionally, the numbers of CD4 + and Foxp3 + T cells as well as the expression of IL-10, TGF-β1, FOXP3 and INF-γ were positively

  11. Inhibition of the proliferation and acceleration of migration of vascular endothelial cells by increased cysteine-rich motor neuron 1

    Energy Technology Data Exchange (ETDEWEB)

    Nakashima, Yukiko; Morimoto, Mayuka [Department of Immunobiology, School of Pharmacy and Pharmaceutical Sciences, Mukogawa Women' s University, 11-68 Koshien Kyuban-cho, Nishinomiya, Hyogo 663-8179 (Japan); Toda, Ken-ichi [Department of Dermatology, Kitano Hospital, The Tazuke Kofukai Nedical Institute, 2-4-20 Ohgimachi, Kita-ku, Osaka 530-8480 (Japan); Shinya, Tomohiro; Sato, Keizo [Department of Clinical Biochemistry, School of Pharmaceutical Sciences, Kyushu University of Health and Welfare, Nobeoka, Miyazaki 882-8508 (Japan); Takahashi, Satoru, E-mail: imwalrus@mukogawa-u.ac.jp [Department of Immunobiology, School of Pharmacy and Pharmaceutical Sciences, Mukogawa Women' s University, 11-68 Koshien Kyuban-cho, Nishinomiya, Hyogo 663-8179 (Japan); Institute for Biosciences, Mukogawa Women' s University, 11-68 Koshien Kyuban-cho, Nishinomiya, Hyogo 663-8179 (Japan)

    2015-07-03

    Cysteine-rich motor neuron 1 (CRIM1) is upregulated only in extracellular matrix gels by angiogenic factors such as vascular endothelial growth factor (VEGF). It then plays a critical role in the tube formation of endothelial cells. In the present study, we investigated the effects of increased CRIM1 on other endothelial functions such as proliferation and migration. Knock down of CRIM1 had no effect on VEGF-induced proliferation or migration of human umbilical vein endothelial cells (HUVECs), indicating that basal CRIM1 is not involved in the proliferation or migration of endothelial cells. Stable CRIM1-overexpressing endothelial F-2 cells, termed CR1 and CR2, were constructed, because it was difficult to prepare monolayer HUVECs that expressed high levels of CRIM1. Proliferation was reduced and migration was accelerated in both CR1 and CR2 cells, compared with normal F-2 cells. Furthermore, the transient overexpression of CRIM1 resulted in decreased proliferation and increased migration of bovine aortic endothelial cells. In contrast, neither proliferation nor migration of COS-7 cells were changed by the overexpression of CRIM1. These results demonstrate that increased CRIM1 reduces the proliferation and accelerates the migration of endothelial cells. These CRIM1 effects might contribute to tube formation of endothelial cells. CRIM1 induced by angiogenic factors may serve as a regulator in endothelial cells to switch from proliferating cells to morphological differentiation. - Highlights: • CRIM1 was upregulated only in tubular endothelial cells, but not in monolayers. • Increased CRIM1 reduced the proliferation of endothelial cells. • Increased CRIM1 accelerated the migration of endothelial cells. • Increased CRIM1 had no effect on the proliferation or migration of COS-7 cells.

  12. Inhibition of the proliferation and acceleration of migration of vascular endothelial cells by increased cysteine-rich motor neuron 1

    International Nuclear Information System (INIS)

    Nakashima, Yukiko; Morimoto, Mayuka; Toda, Ken-ichi; Shinya, Tomohiro; Sato, Keizo; Takahashi, Satoru

    2015-01-01

    Cysteine-rich motor neuron 1 (CRIM1) is upregulated only in extracellular matrix gels by angiogenic factors such as vascular endothelial growth factor (VEGF). It then plays a critical role in the tube formation of endothelial cells. In the present study, we investigated the effects of increased CRIM1 on other endothelial functions such as proliferation and migration. Knock down of CRIM1 had no effect on VEGF-induced proliferation or migration of human umbilical vein endothelial cells (HUVECs), indicating that basal CRIM1 is not involved in the proliferation or migration of endothelial cells. Stable CRIM1-overexpressing endothelial F-2 cells, termed CR1 and CR2, were constructed, because it was difficult to prepare monolayer HUVECs that expressed high levels of CRIM1. Proliferation was reduced and migration was accelerated in both CR1 and CR2 cells, compared with normal F-2 cells. Furthermore, the transient overexpression of CRIM1 resulted in decreased proliferation and increased migration of bovine aortic endothelial cells. In contrast, neither proliferation nor migration of COS-7 cells were changed by the overexpression of CRIM1. These results demonstrate that increased CRIM1 reduces the proliferation and accelerates the migration of endothelial cells. These CRIM1 effects might contribute to tube formation of endothelial cells. CRIM1 induced by angiogenic factors may serve as a regulator in endothelial cells to switch from proliferating cells to morphological differentiation. - Highlights: • CRIM1 was upregulated only in tubular endothelial cells, but not in monolayers. • Increased CRIM1 reduced the proliferation of endothelial cells. • Increased CRIM1 accelerated the migration of endothelial cells. • Increased CRIM1 had no effect on the proliferation or migration of COS-7 cells

  13. Increased circulating fibrocytes are associated with higher reticulocyte percent in children with sickle cell anemia.

    Science.gov (United States)

    Karafin, Matthew S; Dogra, Shibani; Rodeghier, Mark; Burdick, Marie; Mehrad, Borna; Rose, C Edward; Strieter, Robert M; DeBaun, Michael R; Strunk, Robert C; Field, Joshua J

    2016-03-01

    Interstitial lung disease is common in patients with sickle cell anemia (SCA). Fibrocytes are circulating cells implicated in the pathogenesis of pulmonary fibrosis and airway remodeling in asthma. In this study, we tested the hypotheses that fibrocyte levels are: (1) increased in children with SCA compared to healthy controls, and (2) associated with pulmonary disease. Cross-sectional cohort study of children with SCA who participated in the Sleep Asthma Cohort Study. Fibrocyte levels were obtained from 45 children with SCA and 24 controls. Mean age of SCA cases was 14 years and 53% were female. In children with SCA, levels of circulating fibrocytes were greater than controls (P < 0.01). The fibrocytes expressed a hierarchy of chemokine receptors, with CXCR4 expressed on the majority of cells and CCR2 and CCR7 expressed on a smaller subset. Almost half of fibrocytes demonstrated α-smooth muscle actin activation. Increased fibrocyte levels were associated with a higher reticulocyte count (P = 0.03) and older age (P = 0.048) in children with SCA. However, children with increased levels of fibrocytes were not more likely to have asthma or lower percent predicted forced expiratory volume in 1 sec/forced vital capacity (FEV1 /FVC) or FEV1 than those with lower fibrocyte levels. Higher levels of fibrocytes in children with SCA compared to controls may be due to hemolysis. Longitudinal studies may be able to better assess the relationship between fibrocyte level and pulmonary dysfunction. © 2015 Wiley Periodicals, Inc.

  14. High-risk human papillomavirus E7 expression reduces cell-surface MHC class I molecules and increases susceptibility to natural killer cells

    DEFF Research Database (Denmark)

    Bottley, G; Watherston, O G; Hiew, Y-L

    2007-01-01

    a role for E7 in tumour immune evasion. We show that knockdown of E7 expression in HPV16- and HPV18-transformed cervical carcinoma cells by RNA interference increased expression of major histocompatibility complex (MHC) class I at the cell surface and reduced susceptibility of these cells to natural...... killer (NK) cells. Tetracycline-regulated induction of HPV16 E7 resulted in reduced expression of cell surface MHC class I molecules and increased NK cell killing. Our results suggest that, for HPV-associated malignancies, reduced MHC class I expression is the result of an active immune evasion strategy...

  15. Reduced tissue osmolarity increases TRPV4 expression and pro-inflammatory cytokines in intervertebral disc cells

    Directory of Open Access Journals (Sweden)

    BA Walter

    2016-07-01

    Full Text Available The mechanical behaviour and cellular metabolism of intervertebral discs (IVDs and articular cartilage are strongly influenced by their proteoglycan content and associated osmotic properties. This osmotic environment is a biophysical signal that changes with disease and may contribute to the elevated matrix breakdown and altered biologic response to loading observed in IVD degeneration and osteoarthritis. This study tested the hypothesis that changes in osmo-sensation by the transient receptor potential vallinoid-4 (TRPV4 ion channel occur with disease and contribute to the inflammatory environment found during degeneration. Immunohistochemistry on bovine IVDs from an inflammatory organ culture model were used to investigate if TRPV4 is expressed in the IVD and how expression changes with degeneration. Western blot, live-cell calcium imaging, and qRT-PCR were used to investigate whether osmolarity changes or tumour necrosis factor α (TNFα regulate TRPV4 expression, and how altered TRPV4 expression influences calcium signalling and pro-inflammatory cytokine expression. TRPV4 expression correlated with TNFα expression, and was increased when cultured in reduced medium osmolarity and unaltered with TNFα-stimulation. Increased TRPV4 expression increased the calcium flux following TRPV4 activation and increased interleukin-1β (IL-1β and IL-6 gene expression in IVD cells. TRPV4 expression was qualitatively elevated in regions of aggrecan depletion in degenerated human IVDs. Collectively, results suggest that reduced tissue osmolarity, likely following proteoglycan degradation, can increase TRPV4 signalling and enhance pro-inflammatory cytokine production, suggesting changes in TRPV4 mediated osmo-sensation may contribute to the progressive matrix breakdown in disease.

  16. Phospholipase D1 increases Bcl-2 expression during neuronal differentiation of rat neural stem cells.

    Science.gov (United States)

    Park, Shin-Young; Ma, Weina; Yoon, Sung Nyo; Kang, Min Jeong; Han, Joong-Soo

    2015-01-01

    We studied the possible role of phospholipase D1 (PLD1) in the neuronal differentiation, including neurite formation of neural stem cells. PLD1 protein and PLD activity increased during neuronal differentiation. Bcl-2 also increased. Downregulation of PLD1 by transfection with PLD1 siRNA or a dominant-negative form of PLD1 (DN-PLD1) inhibited both neurite outgrowth and Bcl-2 expression. PLD activity was dramatically reduced by a PLCγ (phospholipase Cγ) inhibitor (U73122), a Ca(2+)chelator (BAPTA-AM), and a PKCα (protein kinase Cα) inhibitor (RO320432). Furthermore, treatment with arachidonic acid (AA) which is generated by the action of PLA2 (phospholipase A2) on phosphatidic acid (a PLD1 product), increased the phosphorylation of p38 MAPK and CREB, as well as Bcl-2 expression, indicating that PLA2 is involved in the differentiation process resulting from PLD1 activation. PGE2 (prostaglandin E2), a cyclooxygenase product of AA, also increased during neuronal differentiation. Moreover, treatment with PGE2 increased the phosphorylation of p38 MAPK and CREB, as well as Bcl-2 expression, and this effect was inhibited by a PKA inhibitor (Rp-cAMP). As expected, inhibition of p38 MAPK resulted in loss of CREB activity, and when CREB activity was blocked with CREB siRNA, Bcl-2 production also decreased. We also showed that the EP4 receptor was required for the PKA/p38MAPK/CREB/Bcl-2 pathway. Taken together, these observations indicate that PLD1 is activated by PLCγ/PKCα signaling and stimulate Bcl-2 expression through PLA2/Cox2/EP4/PKA/p38MAPK/CREB during neuronal differentiation of rat neural stem cells.

  17. Increased Stiffness in Aged Skeletal Muscle Impairs Muscle Progenitor Cell Proliferative Activity.

    Directory of Open Access Journals (Sweden)

    Grégory Lacraz

    Full Text Available Skeletal muscle aging is associated with a decreased regenerative potential due to the loss of function of endogenous stem cells or myogenic progenitor cells (MPCs. Aged skeletal muscle is characterized by the deposition of extracellular matrix (ECM, which in turn influences the biomechanical properties of myofibers by increasing their stiffness. Since the stiffness of the MPC microenvironment directly impacts MPC function, we hypothesized that the increase in muscle stiffness that occurs with aging impairs the behavior of MPCs, ultimately leading to a decrease in regenerative potential.We showed that freshly isolated individual myofibers from aged mouse muscles contain fewer MPCs overall than myofibers from adult muscles, with fewer quiescent MPCs and more proliferative and differentiating MPCs. We observed alterations in cultured MPC behavior in aged animals, where the proliferation and differentiation of MPCs were lower and higher, respectively. These alterations were not linked to the intrinsic properties of aged myofibers, as shown by the similar values for the cumulative population-doubling values and fusion indexes. However, atomic force microscopy (AFM indentation experiments revealed a nearly 4-fold increase in the stiffness of the MPC microenvironment. We further showed that the increase in stiffness is associated with alterations to muscle ECM, including the accumulation of collagen, which was correlated with higher hydroxyproline and advanced glycation end-product content. Lastly, we recapitulated the impaired MPC behavior observed in aging using a hydrogel substrate that mimics the stiffness of myofibers.These findings provide novel evidence that the low regenerative potential of aged skeletal muscle is independent of intrinsic MPC properties but is related to the increase in the stiffness of the MPC microenvironment.

  18. Electrical Stimulation of Schwann Cells Promotes Sustained Increases in Neurite Outgrowth

    OpenAIRE

    Koppes, Abigail N.; Nordberg, Andrea L.; Paolillo, Gina M.; Goodsell, Nicole M.; Darwish, Haley A.; Zhang, Linxia; Thompson, Deanna M.

    2013-01-01

    Endogenous electric fields are instructive during embryogenesis by acting to direct cell migration, and postnatally, they can promote axonal growth after injury (McCaig 1991, Al-Majed 2000). However, the mechanisms for these changes are not well understood. Application of an appropriate electrical stimulus may increase the rate and success of nerve repair by directly promoting axonal growth. Previously, DC electrical stimulation at 50 mV/mm (1 mA, 8 h duration) was shown to promote neurite ou...

  19. Increased power generation from primary sludge by a submersible microbial fuel cell and optimum operational conditions

    DEFF Research Database (Denmark)

    Vologni, Valentina; Kakarla, Ramesh; Angelidaki, Irini

    2013-01-01

    Microbial fuel cells (MFCs) have received attention as a promising renewable energy technology for waste treatment and energy recovery. We tested a submersible MFC with an innovative design capable of generating a stable voltage of 0.250 ± 0.008 V (with a fixed 470 Ω resistor) directly from prima...... prolonged the current generation and increased the power density by 7 and 1.5 times, respectively, in comparison with raw primary sludge. These findings suggest that energy recovery from primary sludge can be maximized using an advanced MFC system with optimum conditions....

  20. Shape Memory Polymers Containing Higher Acrylate Content Display Increased Endothelial Cell Attachment

    Science.gov (United States)

    Govindarajan, Tina; Shandas, Robin

    2018-01-01

    Shape Memory Polymers (SMPs) are smart materials that can recall their shape upon the application of a stimulus, which makes them appealing materials for a variety of applications, especially in biomedical devices. Most prior SMP research has focused on tuning bulk properties; studying surface effects of SMPs may extend the use of these materials to blood-contacting applications, such as cardiovascular stents, where surfaces that support rapid endothelialization have been correlated to stent success. Here, we evaluate endothelial attachment onto the surfaces of a family of SMPs previously developed in our group that have shown promise for biomedical devices. Nine SMP formulations containing varying amounts of tert-Butyl acrylate (tBA) and Poly(ethylene glycol) dimethacrylate (PEGDMA) were analyzed for endothelial cell attachment. Dynamic mechanical analysis (DMA), contact angle studies, and atomic force microscopy (AFM) were used to verify bulk and surface properties of the SMPs. Human umbilical vein endothelial cell (HUVEC) attachment and viability was verified using fluorescent methods. Endothelial cells preferentially attached to SMPs with higher tBA content, which have rougher, more hydrophobic surfaces. HUVECs also displayed an increased metabolic activity on these high tBA SMPs over the course of the study. This class of SMPs may be promising candidates for next generation blood-contacting devices. PMID:29707382

  1. α-Synuclein overexpression increases dopamine toxicity in BE(2-M17 cells

    Directory of Open Access Journals (Sweden)

    Miller David W

    2010-03-01

    Full Text Available Abstract Background Oxidative stress has been proposed to be involved in the pathogenesis of Parkinson's disease (PD. A plausible source of oxidative stress in nigral dopaminergic neurons is the redox reactions that specifically involve dopamine and produce various toxic molecules, i.e., free radicals and quinone species. α-Synuclein, a protein found in Lewy bodies characteristic of PD, is also thought to be involved in the pathogenesis of PD and point mutations and multiplications in the gene coding for α-synuclein have been found in familial forms of PD. Results We used dopaminergic human neuroblastoma BE(2-M17 cell lines stably transfected with WT or A30P mutant α-synuclein to characterize the effect of α-synuclein on dopamine toxicity. Cellular toxicity was analyzed by lactate dehydrogenase assay and by fluorescence-activated cell sorter analysis. Increased expression of either wild-type or mutant α-synuclein enhances the cellular toxicity induced by the accumulation of intracellular dopamine or DOPA. Conclusions Our results suggest that an interplay between dopamine and α-synuclein can cause cell death in a neuron-like background. The data presented here are compatible with several models of cytotoxicity, including the formation of α-synuclein oligomers and impairment of the lysosomal degradation.

  2. Ox-LDL increases OX40L in endothelial cells through a LOX-1-dependent mechanism

    Energy Technology Data Exchange (ETDEWEB)

    Dong, Q.; Xiang, R.; Zhang, D.Y.; Qin, S. [Department of Cardiology, The First Affiliated Hospital, Chongqing Medical University, Chongqing (China)

    2013-09-19

    Oxidative low-density lipoprotein (Ox-LDL) is a key risk factor for the development of atherosclerosis, and it can stimulate the expression of a variety of inflammatory signals. As a new and highly sensitive inflammation index, OX40L may be a key to understanding the mechanisms that regulate interactions between cells within the vessel wall and inflammatory mediators during the development of atherosclerosis. To investigate whether Ox-LDL regulates OX40L expression through an oxidized LDL-1 receptor (LOX-1)-mediated mechanism, we investigated the effect of different concentrations of Ox-LDL (50, 100, 150 µg/mL) on endothelial cell proliferation and apoptosis. Stimulation with Ox-LDL increased OX40L protein 1.44-fold and mRNA 4.0-fold in endothelial cells, and these effects were inhibited by blocking LOX-1. These results indicate that LOX-1 plays an important role in the chronic inflammatory process in blood vessel walls. Inhibiting LOX-1 may reduce blood vessel inflammation and provide a therapeutic option to limit atherosclerosis progression.

  3. Shape Memory Polymers Containing Higher Acrylate Content Display Increased Endothelial Cell Attachment

    Directory of Open Access Journals (Sweden)

    Tina Govindarajan

    2017-11-01

    Full Text Available Shape Memory Polymers (SMPs are smart materials that can recall their shape upon the application of a stimulus, which makes them appealing materials for a variety of applications, especially in biomedical devices. Most prior SMP research has focused on tuning bulk properties; studying surface effects of SMPs may extend the use of these materials to blood-contacting applications, such as cardiovascular stents, where surfaces that support rapid endothelialization have been correlated to stent success. Here, we evaluate endothelial attachment onto the surfaces of a family of SMPs previously developed in our group that have shown promise for biomedical devices. Nine SMP formulations containing varying amounts of tert-Butyl acrylate (tBA and Poly(ethylene glycol dimethacrylate (PEGDMA were analyzed for endothelial cell attachment. Dynamic mechanical analysis (DMA, contact angle studies, and atomic force microscopy (AFM were used to verify bulk and surface properties of the SMPs. Human umbilical vein endothelial cell (HUVEC attachment and viability was verified using fluorescent methods. Endothelial cells preferentially attached to SMPs with higher tBA content, which have rougher, more hydrophobic surfaces. HUVECs also displayed an increased metabolic activity on these high tBA SMPs over the course of the study. This class of SMPs may be promising candidates for next generation blood-contacting devices.

  4. Increasing the electric efficiency of a fuel cell system by recirculating the anodic offgas

    Science.gov (United States)

    Heinzel, A.; Roes, J.; Brandt, H.

    The University of Duisburg-Essen and the Center for Fuel Cell Technology (ZBT Duisburg GmbH) have developed a compact multi-fuel steam reformer suitable for natural gas, propane and butane. Fuel processor prototypes based on this concept were built up in the power range from 2.5 to 12.5 kW thermal hydrogen power for different applications and different industrial partners. The fuel processor concept contains all the necessary elements, a prereformer step, a primary reformer, water gas shift reactors, a steam generator, internal heat exchangers, in order to achieve an optimised heat integration and an external burner for heat supply as well as a preferential oxidation step (PrOx) as CO purification. One of the built fuel processors is designed to deliver a thermal hydrogen power output of 2.5 kW according to a PEM fuel cell stack providing about 1 kW electrical power and achieves a thermal efficiency of about 75% (LHV basis after PrOx), while the CO content of the product gas is below 20 ppm. This steam reformer has been combined with a 1 kW PEM fuel cell. Recirculating the anodic offgas results in a significant efficiency increase for the fuel processor. The gross efficiency of the combined system was already clearly above 30% during the first tests. Further improvements are currently investigated and developed at the ZBT.

  5. Ox-LDL increases OX40L in endothelial cells through a LOX-1-dependent mechanism

    International Nuclear Information System (INIS)

    Dong, Q.; Xiang, R.; Zhang, D.Y.; Qin, S.

    2013-01-01

    Oxidative low-density lipoprotein (Ox-LDL) is a key risk factor for the development of atherosclerosis, and it can stimulate the expression of a variety of inflammatory signals. As a new and highly sensitive inflammation index, OX40L may be a key to understanding the mechanisms that regulate interactions between cells within the vessel wall and inflammatory mediators during the development of atherosclerosis. To investigate whether Ox-LDL regulates OX40L expression through an oxidized LDL-1 receptor (LOX-1)-mediated mechanism, we investigated the effect of different concentrations of Ox-LDL (50, 100, 150 µg/mL) on endothelial cell proliferation and apoptosis. Stimulation with Ox-LDL increased OX40L protein 1.44-fold and mRNA 4.0-fold in endothelial cells, and these effects were inhibited by blocking LOX-1. These results indicate that LOX-1 plays an important role in the chronic inflammatory process in blood vessel walls. Inhibiting LOX-1 may reduce blood vessel inflammation and provide a therapeutic option to limit atherosclerosis progression

  6. Patients with sepsis exhibit increased mitochondrial respiratory capacity in peripheral blood immune cells

    DEFF Research Database (Denmark)

    Sjövall, Fredrik; Morota, Saori; Persson, Johan Mikael

    2013-01-01

    to 7). Mitochondrial DNA (mtDNA), cytochrome c (Cyt c), and citrate synthase (CS) were measured as indicators of cellular mitochondrial content. RESULTS: In intact PBICs with endogenous substrates, a gradual increase in cellular respiration reached 173% of controls after 1 week (P = 0......INTRODUCTION: In sepsis, mitochondria have been associated with both initial dysfunction and subsequent upregulation (biogenesis). However, the evolvement of mitochondrial function in sepsis over time is largely unknown, and we therefore investigated mitochondrial respiration in peripheral blood.......001). In permeabilized cells, respiration using substrates of complex I, II, and IV were significantly increased days 1 to 2, reaching 137%, 130%, and 173% of controls, respectively. In parallel, higher levels of CS activity, mtDNA, and Cyt c content in PBICs (211%, 243%, and 331% of controls for the respective...

  7. Damage to the microbial cell membrane during pyrolytic sugar utilization and strategies for increasing resistance.

    Science.gov (United States)

    Jin, Tao; Rover, Marjorie R; Petersen, Elspeth M; Chi, Zhanyou; Smith, Ryan G; Brown, Robert C; Wen, Zhiyou; Jarboe, Laura R

    2017-09-01

    Lignocellulosic biomass is an appealing feedstock for the production of biorenewable fuels and chemicals, and thermochemical processing is a promising method for depolymerizing it into sugars. However, trace compounds in this pyrolytic sugar syrup are inhibitory to microbial biocatalysts. This study demonstrates that hydrophobic inhibitors damage the cell membrane of ethanologenic Escherichia coli KO11+lgk. Adaptive evolution was employed to identify design strategies for improving pyrolytic sugar tolerance and utilization. Characterization of the resulting evolved strain indicates that increased resistance to the membrane-damaging effects of the pyrolytic sugars can be attributed to a glutamine to leucine mutation at position 29 of carbon storage regulator CsrA. This single amino acid change is sufficient for decreasing EPS protein production and increasing membrane integrity when exposed to pyrolytic sugars.

  8. Suppression of Peroxiredoxin 4 in Glioblastoma Cells Increases Apoptosis and Reduces Tumor Growth

    Science.gov (United States)

    Kim, Tae Hyong; Song, Jieun; Alcantara Llaguno, Sheila R.; Murnan, Eric; Liyanarachchi, Sandya; Palanichamy, Kamalakannan; Yi, Ji-Yeun; Viapiano, Mariano Sebastian; Nakano, Ichiro; Yoon, Sung Ok; Wu, Hong; Parada, Luis F.; Kwon, Chang-Hyuk

    2012-01-01

    Glioblastoma multiforme (GBM), the most common and aggressive primary brain malignancy, is incurable despite the best combination of current cancer therapies. For the development of more effective therapies, discovery of novel candidate tumor drivers is urgently needed. Here, we report that peroxiredoxin 4 (PRDX4) is a putative tumor driver. PRDX4 levels were highly increased in a majority of human GBMs as well as in a mouse model of GBM. Reducing PRDX4 expression significantly decreased GBM cell growth and radiation resistance in vitro with increased levels of ROS, DNA damage, and apoptosis. In a syngenic orthotopic transplantation model, Prdx4 knockdown limited GBM infiltration and significantly prolonged mouse survival. These data suggest that PRDX4 can be a novel target for GBM therapies in the future. PMID:22916164

  9. Suppression of peroxiredoxin 4 in glioblastoma cells increases apoptosis and reduces tumor growth.

    Directory of Open Access Journals (Sweden)

    Tae Hyong Kim

    Full Text Available Glioblastoma multiforme (GBM, the most common and aggressive primary brain malignancy, is incurable despite the best combination of current cancer therapies. For the development of more effective therapies, discovery of novel candidate tumor drivers is urgently needed. Here, we report that peroxiredoxin 4 (PRDX4 is a putative tumor driver. PRDX4 levels were highly increased in a majority of human GBMs as well as in a mouse model of GBM. Reducing PRDX4 expression significantly decreased GBM cell growth and radiation resistance in vitro with increased levels of ROS, DNA damage, and apoptosis. In a syngenic orthotopic transplantation model, Prdx4 knockdown limited GBM infiltration and significantly prolonged mouse survival. These data suggest that PRDX4 can be a novel target for GBM therapies in the future.

  10. Removal of glycosaminoglycans from bovine granulosa cells contributes to increased binding of hydrogen-3 heparin

    Energy Technology Data Exchange (ETDEWEB)

    Ax, R.L.; Stodd, C.M.; Boehm, S.K.; Bellin, M.E.

    1986-02-01

    Granulosa cells from small or large bovine follicles were pretreated with enzymes that hydrolyze various glycosaminoglycans, and binding of (/sup 3/H)-heparin to the granulosa was measured. Binding of (/sup 3/H) heparin increased significantly after enzymatic pretreatments with chondroitinase ABC and fungal hyaluronidase, and similar results were obtained with granulosa from small and large follicles. No changes in binding of (/sup 3/H) heparin were detected after hydrolyses with chondroitinase AC and heparinase in either follicle size. Heparitinase, which hydrolyzes heparan sulfate, led to a significant 50% increase in binding of (/sup 3/H) heparin to granulosa from large follicles but was without effect in small follicles. These results suggest that the lower binding of (/sup 3/H) heparin, which has been reported with follicular enlargement, may be due to heparan sulfate occupying or obstructing binding sites for heparin on granulosa from large follicles.

  11. Early Increases in Superantigen-Specific Foxp3+ Regulatory T Cells during Mouse Mammary Tumor Virus Infection▿ †

    Science.gov (United States)

    Cabrera, Gabriel; Burzyn, Dalia; Mundiñano, Juliana; Courreges, M. Cecilia; Camicia, Gabriela; Lorenzo, Daniela; Costa, Héctor; Ross, Susan R.; Nepomnaschy, Irene; Piazzon, Isabel

    2008-01-01

    Mouse mammary tumor virus (MMTV) is a milk-borne betaretrovirus that has developed strategies to exploit and subvert the host immune system. Here, we show in a natural model of MMTV infection that the virus causes early and progressive increases in superantigen (SAg)-specific Foxp3+ regulatory T cells (Treg) in Peyer's patches (PP). These increases were shown to be dependent on the presence of dendritic cells. CD4+ CD25+ T cells from the PP of infected mice preferentially suppress the proliferative response of T cells to SAg-expressing antigen-presenting cells ex vivo. We investigated the influence of the depletion of CD25+ cells at different stages of the infection. When CD25+ cells were depleted before MMTV infection, an increase in the number of PP SAg-cognate Foxp3− T cells was found at day 6 of infection. Since the SAg response is associated with viral amplification, the possibility exists that Treg cells attenuate the increase in viral load at the beginning of the infection. In contrast, depletion of CD25+ cells once the initial SAg response has developed caused a lower viral load, suggesting that at later stages Treg cells may favor viral persistence. Thus, our results indicated that Treg cells play an important and complex role during MMTV infection. PMID:18495774

  12. Hydrogen Sulfide Increases Nitric Oxide Production and Subsequent S-Nitrosylation in Endothelial Cells

    Directory of Open Access Journals (Sweden)

    Ping-Ho Chen

    2014-01-01

    Full Text Available Hydrogen sulfide (H2S and nitric oxide (NO, two endogenous gaseous molecules in endothelial cells, got increased attention with respect to their protective roles in the cardiovascular system. However, the details of the signaling pathways between H2S and NO in endothelia cells remain unclear. In this study, a treatment with NaHS profoundly increased the expression and the activity of endothelial nitric oxide synthase. Elevated gaseous NO levels were observed by a novel and specific fluorescent probe, 5-amino-2-(6-hydroxy-3-oxo-3H-xanthen-9-ylbenzoic acid methyl ester (FA-OMe, and quantified by flow cytometry. Further study indicated an increase of upstream regulator for eNOS activation, AMP-activated protein kinase (AMPK, and protein kinase B (Akt. By using a biotin switch, the level of NO-mediated protein S-nitrosylation was also enhanced. However, with the addition of the NO donor, NOC-18, the expressions of cystathionine-γ-lyase, cystathionine-β-synthase, and 3-mercaptopyruvate sulfurtransferase were not changed. The level of H2S was also monitored by a new designed fluorescent probe, 4-nitro-7-thiocyanatobenz-2-oxa-1,3-diazole (NBD-SCN with high specificity. Therefore, NO did not reciprocally increase the expression of H2S-generating enzymes and the H2S level. The present study provides an integrated insight of cellular responses to H2S and NO from protein expression to gaseous molecule generation, which indicates the upstream role of H2S in modulating NO production and protein S-nitrosylation.

  13. Electrospun nanofibrous scaffolds increase the efficacy of stem cell-mediated therapy of surgically resected glioblastoma

    Science.gov (United States)

    Bagó, Juli R.; Pegna, Guillaume J.; Okolie, Onyi; Mohiti-Asli, Mahsa; Loboa, Elizabeth G.; Hingtgen, Shawn D.

    2017-01-01

    Engineered stem cell (SC)-based therapy holds enormous promise for treating the incurable brain cancer glioblastoma (GBM). Retaining the cytotoxic SCs in the surgical cavity after GBM resection is one of the greatest challenges to this approach. Here, we describe a biocompatible electrospun nanofibrous scaffold (bENS) implant capable of delivering and retaining tumor-homing cytotoxic stem cells that suppress recurrence of post-surgical GBM. As a new approach to GBM therapy, we created poly(l-lactic acid) (PLA) bENS bearing drug-releasing human mesenchymal stem cells (hMSCs). We discovered that bENS-based implant increased hMSC retention in the surgical cavity 5-fold and prolonged persistence 3-fold compared to standard direct injection using our mouse model of GBM surgical resection/recurrence. Time-lapse imaging showed cytotoxic hMSC/bENS treatment killed co-cultured human GBM cells, and allowed hMSCs to rapidly migrate off the scaffolds as they homed to GBMs. In vivo, bENS loaded with hMSCs releasing the anti-tumor protein TRAIL (bENSsTR) reduced the volume of established GBM xenografts 3-fold. Mimicking clinical GBM patient therapy, lining the post-operative GBM surgical cavity with bENSsTR implants inhibited the re-growth of residual GBM foci 2.3-fold and prolonged post-surgical median survival from 13.5 to 31 days in mice. These results suggest that nanofibrous-based SC therapies could be an innovative new approach to improve the outcomes of patients suffering from terminal brain cancer. PMID:27016620

  14. Extended passaging increases the efficiency of neural differentiation from induced pluripotent stem cells

    Directory of Open Access Journals (Sweden)

    Koehler Karl R

    2011-08-01

    Full Text Available Abstract Background The use of induced pluripotent stem cells (iPSCs for the functional replacement of damaged neurons and in vitro disease modeling is of great clinical relevance. Unfortunately, the capacity of iPSC lines to differentiate into neurons is highly variable, prompting the need for a reliable means of assessing the differentiation capacity of newly derived iPSC cell lines. Extended passaging is emerging as a method of ensuring faithful reprogramming. We adapted an established and efficient embryonic stem cell (ESC neural induction protocol to test whether iPSCs (1 have the competence to give rise to functional neurons with similar efficiency as ESCs and (2 whether the extent of neural differentiation could be altered or enhanced by increased passaging. Results Our gene expression and morphological analyses revealed that neural conversion was temporally delayed in iPSC lines and some iPSC lines did not properly form embryoid bodies during the first stage of differentiation. Notably, these deficits were corrected by continual passaging in an iPSC clone. iPSCs with greater than 20 passages (late-passage iPSCs expressed higher expression levels of pluripotency markers and formed larger embryoid bodies than iPSCs with fewer than 10 passages (early-passage iPSCs. Moreover, late-passage iPSCs started to express neural marker genes sooner than early-passage iPSCs after the initiation of neural induction. Furthermore, late-passage iPSC-derived neurons exhibited notably greater excitability and larger voltage-gated currents than early-passage iPSC-derived neurons, although these cells were morphologically indistinguishable. Conclusions These findings strongly suggest that the efficiency neuronal conversion depends on the complete reprogramming of iPSCs via extensive passaging.

  15. Hyperglycosylated human chorionic gonadotropin does not increase progesterone production by luteinized granulosa cells.

    Science.gov (United States)

    Crochet, John R; Shah, Anish A; Schomberg, David W; Price, Thomas M

    2012-09-01

    Trophoblast-derived human chorionic gonadotropin (hCG) promotes corpus luteum progesterone (P4) production, and wide ranges of serum P4 levels are noted in various pregnancy outcomes, despite similar hCG concentrations. There are five unique biologically active hCG variants in human pregnancy urine, and previous studies of P4 production in response to hCG have used only preparations containing all isoforms. Understanding exactly which hCG variant is primarily responsible for stimulating corpus luteum steroidogenesis may have great clinical and diagnostic implications, including in the setting of ectopic pregnancy. Our objective was to delineate the role of the standard and hyperglycosylated (H)-hCG isoforms in stimulating P4 production by luteinized granulosa cells. Cell culture, ELISA, and fluorometric-based protein assays were done at Duke University Medical Center. Patients were anonymous oocyte donors. Cultured luteinized granulosa cells were treated with 0.25, 0.5, and 1.0 ng/ml total hCG, which contains all isoforms, purified standard hCG (37.1 kDa), and purified H-hCG (42.8 kDa). P4 produced per total cellular protein (nanograms per microgram) was measured via ELISA and fluorometric protein determination kits. Both total hCG (P = 0.0003) and purified standard hCG (P production. Purified H-hCG did not change the P4 produced per total cellular protein response (P value not significant). Standard hCG stimulated P4 production by cultured granulosa cells and likely supports corpus luteum function via interactions with the LH/hCG receptor. In contrast, H-hCG did not increase P4 production, which indicates a nonsteroidogenic role for this protein during early gestation.

  16. Increased cell proliferation in spleen and lymph nodes peripheral to contact allergen application site

    International Nuclear Information System (INIS)

    Chipinda, Itai; Anderson, Stacey E.; Butterworth, Leon F.; Beezhold, Donald; Siegel, Paul D.

    2009-01-01

    The local lymph node assay (LLNA) is widely used to identify chemicals that are contact sensitizers. The assay involves dosing mice with the chemical on both ears and pooling the superficial parotid lymph nodes for assessment of lymphocyte proliferation as a marker of sensitization. The present study explored potential reduction in animal usage by dosing one ear with the allergen and the other with vehicle-only. The respective draining lymph nodes were processed separately for tritiated thymidine ( 3 H-TdR) incorporation. Cell proliferation in proper axillary and renal nodes, as well as in the spleen was also assessed. Cross-contamination of the chemicals from the dosed ears to other parts of the body via preening was prevented by dosing restrained animals and washing off the residual chemical with saline after 4 h. Dosing the left ear with 0.02% oxazolone (OX) on unrestrained animals resulted in marked cell proliferation in its draining lymph node (stimulation index, SI = 12.8) and in the lymph node draining the contra-lateral vehicle-dosed ear (SI = 6), as well as the proper axillary lymph nodes (SI = 3.3). Increased 3 H-TdR incorporation was not observed in the renal lymph nodes (SI = 1.1). Similar stimulation of cells was observed in the lymph node draining the ear contra-lateral to the 30% hexylcinnamaldehyde (HCA)-dosed ear. Increased proliferative activity was observed in contra-lateral draining lymph nodes of restrained mice demonstrating that these results cannot be attributed to cross-contamination of adjacent skin. A significant increase in proliferation of splenocytes was also observed. It is concluded that dermal application of a contact allergen, as exemplified by OX and HCA, may induce cell proliferation in the neighboring lymph nodes and spleen indicative of hapten and/or haptenated proteins diffusing through the skin to peripheral nodes and the blood to produce systemic sensitization. It is also possible that lymphatic capillaries may communicate

  17. Inhibition of Hsp90 acts synergistically with topoisomerase II poisons to increase the apoptotic killing of cells due to an increase in topoisomerase II mediated DNA damage.

    Science.gov (United States)

    Barker, Catherine R; McNamara, Anne V; Rackstraw, Stephen A; Nelson, David E; White, Mike R; Watson, Alastair J M; Jenkins, John R

    2006-01-01

    Topoisomerase II plays a crucial role during chromosome condensation and segregation in mitosis and meiosis and is a highly attractive target for chemotherapeutic agents. We have identified previously topoisomerase II and heat shock protein 90 (Hsp90) as part of a complex. In this paper we demonstrate that drug combinations targeting these two enzymes cause a synergistic increase in apoptosis. The objective of our study was to identify the mode of cell killing and the mechanism behind the increase in topoisomerase II mediated DNA damage. Importantly we demonstrate that Hsp90 inhibition results in an increased topoiosmerase II activity but not degradation of topoisomerase II and it is this, in the presence of a topoisomerase II poison that causes the increase in cell death. Our results suggest a novel mechanism of action where the inhibition of Hsp90 disrupts the Hsp90-topoisomerase II interaction leading to an increase in and activation of unbound topoisomerase II, which, in the presence of a topoisomerase II poison leads to the formation of an increased number of cleavable complexes ultimately resulting in rise in DNA damage and a subsequent increase cell death.

  18. Enhanced nitrogen removal in single-chamber microbial fuel cells with increased gas diffusion areas

    KAUST Repository

    Yan, Hengjing

    2012-11-23

    Single-chamber microbial fuel cells (MFCs) with nitrifiers pre-enriched at the air cathodes have previously been demonstrated as a passive strategy for integrating nitrogen removal into current-generating bioelectrochemical systems. To further define system design parameters for this strategy, we investigated in this study the effects of oxygen diffusion area and COD/N ratio in continuous-flow reactors. Doubling the gas diffusion area by adding an additional air cathode or a diffusion cloth significantly increased the ammonia and COD removal rates (by up to 115% and 39%), ammonia removal efficiency (by up to 134%), the cell voltage and cathode potentials, and the power densities (by a factor of approximately 2). When the COD/N ratio was lowered from 13 to 3, we found up to 244% higher ammonia removal rate but at least 19% lower ammonia removal efficiency. An increase of COD removal rate by up to 27% was also found when the COD/N ratio was lowered from 11 to 3. The Coulombic efficiency was not affected by the additional air cathode, but decreased by an average of 11% with the addition of a diffusion cloth. Ammonia removal by assimilation was also estimated to understand the ammonia removal mechanism in these systems. These results showed that the doubling of gas diffusion area enhanced N and COD removal rates without compromising electrochemical performance. © 2012 Wiley Periodicals, Inc.

  19. Single-molecule photobleaching reveals increased MET receptor dimerization upon ligand binding in intact cells

    International Nuclear Information System (INIS)

    Dietz, Marina S; Haße, Daniel; Ferraris, Davide M; Göhler, Antonia; Niemann, Hartmut H; Heilemann, Mike

    2013-01-01

    The human receptor tyrosine kinase MET and its ligand hepatocyte growth factor/scatter factor are essential during embryonic development and play an important role during cancer metastasis and tissue regeneration. In addition, it was found that MET is also relevant for infectious diseases and is the target of different bacteria, amongst them Listeria monocytogenes that induces bacterial uptake through the surface protein internalin B. Binding of ligand to the MET receptor is proposed to lead to receptor dimerization. However, it is also discussed whether preformed MET dimers exist on the cell membrane. To address these issues we used single-molecule fluorescence microscopy techniques. Our photobleaching experiments show that MET exists in dimers on the membrane of cells in the absence of ligand and that the proportion of MET dimers increases significantly upon ligand binding. Our results indicate that partially preformed MET dimers may play a role in ligand binding or MET signaling. The addition of the bacterial ligand internalin B leads to an increase of MET dimers which is in agreement with the model of ligand-induced dimerization of receptor tyrosine kinases.

  20. Arsenic induces diabetic effects through beta-cell dysfunction and increased gluconeogenesis in mice

    Science.gov (United States)

    Liu, Su; Guo, Xuechao; Wu, Bing; Yu, Haiyan; Zhang, Xuxiang; Li, Mei

    2014-11-01

    Arsenic as a potential risk factor for type 2 diabetes has been received attention recently. However, the roles of arsenic on development of diabetes are unclear. In this study, we compared the influences of inorganic arsenic (iAs) on normal and diabetic mice by systems toxicology approaches. Although iAs exposure did not change glucose tolerance in normal mice, it caused the pancreatic β-cell dysfunction and increased gluconeogenesis and oxidative damages in liver. However, iAs exposure worsened the glucose tolerance in diabetic mice, which might be due to increased gluconeogenesis and impairment of pancreatic β-cell function. It is interesting that iAs exposure could improve the insulin sensitivity based on the insulin tolerance testing by the activation of glucose uptake-related genes and enzymes in normal and diabetic individuals. Our data suggested that iAs exposure could cause pre-diabetic effects by altering the lipid metabolism, gluconeogenesis and insulin secretion in normal individual, and worsen diabetic effects in diabetes individual by these processes. Insulin resistance might be not the reason of diabetic effects caused by iAs, indicating that mechanism of the diabetogenic effects of iAs exposure is different from the mechanism associated with traditional risk factors (such as obesity)-reduced type 2 diabetes.

  1. Increased NY-ESO-1 Expression and Reduced Infiltrating CD3+ T Cells in Cutaneous Melanoma

    Directory of Open Access Journals (Sweden)

    Mara Giavina-Bianchi

    2015-01-01

    Full Text Available NY-ESO-1 is a cancer-testis antigen aberrantly expressed in melanomas, which may serve as a robust and specific target in immunotherapy. NY-ESO-1 antigen expression, tumor features, and the immune profile of tumor infiltrating lymphocytes were assessed in primary cutaneous melanoma. NY-ESO-1 protein was detected in 20% of invasive melanomas (16/79, rarely in in situ melanoma (1/10 and not in benign nevi (0/20. Marked intratumoral heterogeneity of NY-ESO-1 protein expression was observed. NY-ESO-1 expression was associated with increased primary tumor thickness (P=0.007 and inversely correlated with superficial spreading melanoma (P<0.02. NY-ESO-1 expression was also associated with reduced numbers and density of CD3+ tumor infiltrating lymphocytes (P=0.017. When NY-ESO-1 protein was expressed, CD3+ T cells were less diffusely infiltrating the tumor and were more often arranged in small clusters (P=0.010 or as isolated cells (P=0.002 than in large clusters of more than five lymphocytes. No correlation of NY-ESO-1 expression with gender, age, tumor site, ulceration, lymph node sentinel status, or survival was observed. NY-ESO-1 expression in melanoma was associated with tumor progression, including increased tumor thickness, and with reduced tumor infiltrating lymphocytes.

  2. Arsenic induces diabetic effects through beta-cell dysfunction and increased gluconeogenesis in mice.

    Science.gov (United States)

    Liu, Su; Guo, Xuechao; Wu, Bing; Yu, Haiyan; Zhang, Xuxiang; Li, Mei

    2014-11-04

    Arsenic as a potential risk factor for type 2 diabetes has been received attention recently. However, the roles of arsenic on development of diabetes are unclear. In this study, we compared the influences of inorganic arsenic (iAs) on normal and diabetic mice by systems toxicology approaches. Although iAs exposure did not change glucose tolerance in normal mice, it caused the pancreatic β-cell dysfunction and increased gluconeogenesis and oxidative damages in liver. However, iAs exposure worsened the glucose tolerance in diabetic mice, which might be due to increased gluconeogenesis and impairment of pancreatic β-cell function. It is interesting that iAs exposure could improve the insulin sensitivity based on the insulin tolerance testing by the activation of glucose uptake-related genes and enzymes in normal and diabetic individuals. Our data suggested that iAs exposure could cause pre-diabetic effects by altering the lipid metabolism, gluconeogenesis and insulin secretion in normal individual, and worsen diabetic effects in diabetes individual by these processes. Insulin resistance might be not the reason of diabetic effects caused by iAs, indicating that mechanism of the diabetogenic effects of iAs exposure is different from the mechanism associated with traditional risk factors (such as obesity)-reduced type 2 diabetes.

  3. Nivalenol induces oxidative stress and increases deoxynivalenol pro-oxidant effect in intestinal epithelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Del Regno, Marisanta; Adesso, Simona; Popolo, Ada [Department of Pharmacy, School of Pharmacy, University of Salerno, Via Giovanni Paolo II, 132–84084 Fisciano, Salerno (Italy); Quaroni, Andrea [Department of Biomedical Sciences, Cornell University, Veterinary Research Tower, Cornell University, Ithaca, NY 14853–6401 (United States); Autore, Giuseppina [Department of Pharmacy, School of Pharmacy, University of Salerno, Via Giovanni Paolo II, 132–84084 Fisciano, Salerno (Italy); Severino, Lorella [Department of Pathology and Animal Health, Division of Toxicology, School of Veterinary Medicine, University of Naples “Federico II”, Via Delpino 1, 80137 Naples (Italy); Marzocco, Stefania, E-mail: smarzocco@unisa.it [Department of Pharmacy, School of Pharmacy, University of Salerno, Via Giovanni Paolo II, 132–84084 Fisciano, Salerno (Italy)

    2015-06-01

    Mycotoxins are secondary fungal metabolites often found as contaminants in almost all agricultural commodities worldwide, and the consumption of food or feed contaminated by mycotoxins represents a major risk for human and animal health. Reactive oxygen species are normal products of cellular metabolism. However, disproportionate generation of reactive oxygen species poses a serious problem to bodily homeostasis and causes oxidative tissue damage. In this study we analyzed the effect of two trichothecenes mycotoxins: nivalenol and deoxynivalenol, alone and in combination, on oxidative stress in the non-tumorigenic intestinal epithelial cell line IEC-6. Our results indicate the pro-oxidant nivalenol effect in IEC-6, the stronger pro-oxidant effect of nivalenol when compared to deoxynivalenol and, interestingly, that nivalenol increases deoxynivalenol pro-oxidative effects. Mechanistic studies indicate that the observed effects were mediated by NADPH oxidase, calcium homeostasis alteration, NF-kB and Nrf2 pathways activation and by iNOS and nitrotyrosine formation. The toxicological interaction by nivalenol and deoxynivalenol reported in this study in IEC-6, points out the importance of the toxic effect of these mycotoxins, mostly in combination, further highlighting the risk assessment process of these toxins that are of growing concern. - Highlights: • Nivalenol induces oxidative stress in intestinal epithelial cells (IECs). • Nivalenol increases deoxynivalenol pro-oxidant effects in IECs. • Nivalenol and deoxynivalenol trigger antioxidant response IECs. • These results indicate the importance of mycotoxins co-contamination.

  4. Three dimensional live-cell STED microscopy at increased depth using a water immersion objective

    Science.gov (United States)

    Heine, Jörn; Wurm, Christian A.; Keller-Findeisen, Jan; Schönle, Andreas; Harke, Benjamin; Reuss, Matthias; Winter, Franziska R.; Donnert, Gerald

    2018-05-01

    Modern fluorescence superresolution microscopes are capable of imaging living cells on the nanometer scale. One of those techniques is stimulated emission depletion (STED) which increases the microscope's resolution many times in the lateral and the axial directions. To achieve these high resolutions not only close to the coverslip but also at greater depths, the choice of objective becomes crucial. Oil immersion objectives have frequently been used for STED imaging since their high numerical aperture (NA) leads to high spatial resolutions. But during live-cell imaging, especially at great penetration depths, these objectives have a distinct disadvantage. The refractive index mismatch between the immersion oil and the usually aqueous embedding media of living specimens results in unwanted spherical aberrations. These aberrations distort the point spread functions (PSFs). Notably, during z- and 3D-STED imaging, the resolution increase along the optical axis is majorly hampered if at all possible. To overcome this limitation, we here use a water immersion objective in combination with a spatial light modulator for z-STED measurements of living samples at great depths. This compact design allows for switching between objectives without having to adapt the STED beam path and enables on the fly alterations of the STED PSF to correct for aberrations. Furthermore, we derive the influence of the NA on the axial STED resolution theoretically and experimentally. We show under live-cell imaging conditions that a water immersion objective leads to far superior results than an oil immersion objective at penetration depths of 5-180 μm.

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

    Science.gov (United States)

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

    2016-01-01

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

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

    Directory of Open Access Journals (Sweden)

    John D Lyons

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

  7. Increase of NK-T cells in aged depressed patients not treated with antidepressive drugs

    NARCIS (Netherlands)

    Flentge, F; van den Berg, MD; Bouhuys, AL; The, HT

    2000-01-01

    Background: A change in number and/or activity of natural killer cells has repeatedly been reported in depressive illness. Much less attention has yet been given to the subgroup of natural killer cells that are positive Sor the T-cell marker CD3 (NK-T cells). These cells possibly have important

  8. An increase in immature β-cells lacking Glut2 precedes the expansion of β-cell mass in the pregnant mouse.

    Directory of Open Access Journals (Sweden)

    Christine A Beamish

    Full Text Available A compensatory increase in β-cell mass occurs during pregnancy to counter the associated insulin resistance, and a failure in adaptation is thought to contribute to gestational diabetes. Insulin-expressing but glucose-transporter-2-low (Ins+Glut2LO progenitor cells are present in mouse and human pancreas, being predominantly located in extra-islet β-cell clusters, and contribute to the regeneration of the endocrine pancreas following induced ablation. We therefore sought to investigate the contribution of Ins+Glut2LO cells to β-cell mass expansion during pregnancy. Female C57Bl/6 mice were time mated and pancreata were collected at gestational days (GD 6, 9, 12, 15, and 18, and postpartum D7 (n = 4/time-point and compared to control (non-pregnant animals. Beta cell mass, location, proliferation (Ki67+, and proportion of Ins+Glut2LO cells were measured using immunohistochemistry and bright field or confocal microscopy. Beta cell mass tripled by GD18 and β-cell proliferation peaked at GD12 in islets (≥6 β-cells and small β-cell clusters (1-5 β-cells. The proportion and fraction of Ins+Glut2LO cells undergoing proliferation increased significantly at GD9 in both islets and clusters, preceding the increase in β-cell mass and proliferation, and their proliferation within clusters persisted until GD15. The overall number of clusters increased significantly at GD9. Quantitative PCR showed a significant increase in Pdx1 presence at GD9 vs. GD18 or control pancreas, and Pdx1 was visualized by immunohistochemistry within both Ins+Glut2LO and Ins+Glut2HI cells within clusters. These results indicate that Ins+Glut2LO cells are likely to contribute to β-cell mass expansion during pregnancy.

  9. The histone deacetylase inhibitor vorinostat (SAHA) increases the susceptibility of uninfected CD4+ T cells to HIV by increasing the kinetics and efficiency of postentry viral events.

    Science.gov (United States)

    Lucera, Mark B; Tilton, Carisa A; Mao, Hongxia; Dobrowolski, Curtis; Tabler, Caroline O; Haqqani, Aiman A; Karn, Jonathan; Tilton, John C

    2014-09-01

    Latently infected cells remain a primary barrier to eradication of HIV-1. Over the past decade, a better understanding of the molecular mechanisms by which latency is established and maintained has led to the discovery of a number of compounds that selectively reactivate latent proviruses without inducing polyclonal T cell activation. Recently, the histone deacetylase (HDAC) inhibitor vorinostat has been demonstrated to induce HIV transcription from latently infected cells when administered to patients. While vorinostat will be given in the context of antiretroviral therapy (ART), infection of new cells by induced virus remains a clinical concern. Here, we demonstrate that vorinostat significantly increases the susceptibility of CD4(+) T cells to infection by HIV in a dose- and time-dependent manner that is independent of receptor and coreceptor usage. Vorinostat does not enhance viral fusion with cells but rather enhances the kinetics and efficiency of postentry viral events, including reverse transcription, nuclear import, and integration, and enhances viral production in a spreading-infection assay. Selective inhibition of the cytoplasmic class IIb HDAC6 with tubacin recapitulated the effect of vorinostat. These findings reveal a previously unknown cytoplasmic effect of HDAC inhibitors promoting productive infection of CD4(+) T cells that is distinct from their well-characterized effects on nuclear histone acetylation and long-terminal-repeat (LTR) transcription. Our results indicate that careful monitoring of patients and ART intensification are warranted during vorinostat treatment and indicate that HDAC inhibitors that selectively target nuclear class I HDACs could reactivate latent HIV without increasing the susceptibility of uninfected cells to HIV. HDAC inhibitors, particularly vorinostat, are currently being investigated clinically as part of a "shock-and-kill" strategy to purge latent reservoirs of HIV. We demonstrate here that vorinostat increases the

  10. Direct in vivo evidence for increased proliferation of CLL cells in lymph nodes compared to bone marrow and peripheral blood

    DEFF Research Database (Denmark)

    Herndon, Thomas M; Chen, Shih-Ann; Saba, Nakhle S

    2017-01-01

    of active cell proliferation remains to be definitively determined. Based on findings that CLL cells in LNs have increased expression of B-cell activation genes, we tested the hypothesis that the fraction of 'newly born' cells would be highest in the LNs. Using a deuterium oxide ((2)H) in vivo labeling...... method in which patients consumed deuterated (heavy) water ((2)H2O), we determined CLL cell kinetics in concurrently obtained samples from LN, PB and BM. The LN was identified as the anatomical site harboring the largest fraction of newly born cells, compared to PB and BM. In fact, the calculated birth...

  11. Chewing suppresses the stress-induced increase in the number of pERK-immunoreactive cells in the periaqueductal grey.

    Science.gov (United States)

    Yamada, Kentaro; Narimatsu, Yuri; Ono, Yumie; Sasaguri, Ken-Ichi; Onozuka, Minoru; Kawata, Toshitsugu; Yamamoto, Toshiharu

    2015-07-10

    We investigated the effects of chewing under immobilization stress on the periaqueductal gray (PAG) matter using phosphorylated extracellular signal-regulated kinase (pERK) as a marker of responding cells. Immobilization stress increased pERK-immunoreactive cells in the PAG. Among four subdivisions of the PAG, the increase of immunoreactive cells was remarkable in the dorsolateral and ventrolateral subdivisions. However, increase of pERK-immunoreactive cells by the immobilization stress was not so evident in the dorsomedial and lateral subdivisions. The chewing under immobilization stress prevented the stress-induced increase of pERK-immunoreactive cells in the dorsolateral and ventrolateral subdivisions with statistical significances (p<0.05). Again, chewing effects on pERK-immunoreactive cells were not visible in the dorsomedial and lateral subdivisions. These results suggest that the chewing alleviates the PAG (dorsolateral and ventrolateral subdivisions) responses to stress. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

  12. Epalrestat increases intracellular glutathione levels in Schwann cells through transcription regulation

    Directory of Open Access Journals (Sweden)

    Keisuke Sato

    2014-01-01

    Full Text Available Epalrestat (EPS, approved in Japan, is the only aldose reductase inhibitor that is currently available for the treatment of diabetic neuropathy. Here we report that EPS at near-plasma concentration increases the intracellular levels of glutathione (GSH, which is important for protection against oxidative injury, through transcription regulation. Treatment of Schwann cells with EPS caused a dramatic increase in intracellular GSH levels. EPS increased the mRNA levels of γ-glutamylcysteine synthetase (γ-GCS, the enzyme catalyzing the first and rate-limiting step in de novo GSH synthesis. Nuclear factor erythroid 2-related factor 2 (Nrf2 is a key transcription factor that plays a central role in regulating the expression of γ-GCS. ELISA revealed that EPS increased nuclear Nrf2 levels. Knockdown of Nrf2 by siRNA suppressed the EPS-induced GSH biosynthesis. Furthermore, pretreatment with EPS reduced the cytotoxicity induced by H2O2, tert-butylhydroperoxide, 2,2'-azobis (2-amidinopropane dihydrochloride, and menadione, indicating that EPS plays a role in protecting against oxidative stress. This is the first study to show that EPS induces GSH biosynthesis via the activation of Nrf2. We suggest that EPS has new beneficial properties that may prevent the development and progression of disorders caused by oxidative stress.

  13. Peripheral Mononuclear Cell Resistin mRNA Expression Is Increased in Type 2 Diabetic Women

    Directory of Open Access Journals (Sweden)

    Panayoula C. Tsiotra

    2008-01-01

    Full Text Available Resistin has been shown to cause insulin resistance and to impair glucose tolerance in rodents, but in humans its physiological role still remains elusive. The aim of this study was to examine whether resistin mRNA expression in human peripheral mononuclear cells (PBMCs and its corresponding plasma levels are altered in type 2 diabetes. Resistin mRNA levels were easily detectable in human PBMC, and found to be higher in DM2 compared to healthy women (P=.05. Similarly, mononuclear mRNA levels of the proinflammatory cytokines IL-1β, TNF-α, and IL-6 were all significantly higher in DM2 compared to control women (P<.001. The corresponding plasma resistin levels were slightly, but not significantly, increased in DM2 women (P=.051, and overall, they correlated significantly with BMI (r=0.406, P=.010 and waist circumference (r=0.516, P=.003, but not with fasting insulin levels or HOMA-IR. Resistin mRNA expression is increased in PBMC from DM2 women, together with increased expression of the inflammatory cytokines IL-1β, TNF-α, and IL-6, independent of obesity. These results suggest that resistin and cytokines might contribute to the low-grade inflammation and the increased atherogenic risk observed in these patients.

  14. Mesenchymal stem cells increase antioxidant capacity in intestinal ischemia/reperfusion damage.

    Science.gov (United States)

    Inan, M; Bakar, E; Cerkezkayabekir, A; Sanal, F; Ulucam, E; Subaşı, C; Karaöz, E

    2017-07-01

    Mesenchymal stem cells (MSCs) may have beneficial effects in reversing intestinal damage resulting from circulatory disorders. The hypothesis of this study is that MSCs increase antioxidant capacity of small bowel tissue following intestinal ischemia reperfusion (I/R) damage. A total of 100 rats were used for the control group and three experimental groups, as follows: the sham control, local MSC, and systemic MSC groups. Each group consisted of 10 animals on days 1, 4, and 7 of the experiment. Ischemia was established by clamping the superior mesenteric artery (SMA) for 45min; following this, reperfusion was carried out for 1, 4, and 7days in all groups. In the local and systemic groups, MSCs were administered intravenously and locally just after the ischemia, and they were investigated after 1, 4, and 7days. The superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (Gpx) activities, as well as malondialdehyde (MDA) and total protein levels, were measured. Histopathological analysis was performed using light and electron microscopy. The indicators of proliferation from the effects of anti- and pro-inflammatory cytokines were evaluated using immunohistochemistry. MDA was increased (Pcytokines interleukin-1β (IL1β), transforming growth factor-β1 (TGFβ1), tumor necrosis factor-α (TNFα), IL6, MIP2, and MPO decreased (Pcytokines EP3 and IL1ra increased (poxygen radicals, suppression of pro-inflammatory cytokines, and increasing the expression of anti-inflammatory cytokines. Copyright © 2017 Elsevier Inc. All rights reserved.

  15. Overexpression of α-catenin increases osteoblastic differentiation in mouse mesenchymal C3H10T1/2 cells

    International Nuclear Information System (INIS)

    Kim, Dohee; Yang, Jae-Yeon; Shin, Chan Soo

    2009-01-01

    α- and β-Catenin link cadherins to the actin-based cytoskeleton at adherens junctions and regulate cell-cell adhesion. Although roles of cadherins and canonical Wnt-/β-catenin-signaling in osteoblastic differentiation have been extensively studied, the role of α-catenin is not known. Murine embryonic mesenchymal stem cells, C3H10T1/2 cells, were transduced with retrovirus encoding α-catenin (MSCV-α-catenin-HA-GFP). In the presence of Wnt-3A conditioned medium or osteogenic medium (β-glycerol phosphate and ascorbic acid), cells overexpressing α-catenin showed enhanced osteoblastic differentiation as measured by alkaline phosphatase (ALP) staining and ALP activity assay compared to cells transduced with empty virus (MSCV-GFP). In addition, mRNA expression of osteocalcin and Runx2 was significantly increased compared to control. Cell aggregation assay revealed that α-catenin overexpression has significantly increased cell-cell aggregation. However, cellular β-catenin levels (total, cytoplasmic-nuclear ratio) and β-catenin-TCF/LEF transcriptional activity did not change by overexpression of α-catenin. Knock-down of α-catenin using siRNA decreased osteoblastic differentiation as measured by ALP assay. These results suggest that α-catenin overexpression increases osteoblastic differentiation by increasing cell-cell adhesion rather than Wnt-/β-catenin-signaling.

  16. Macroautophagy-generated increase of lysosomal amyloid β-protein mediates oxidant-induced apoptosis of cultured neuroblastoma cells

    DEFF Research Database (Denmark)

    Zheng, Lin; Terman, Alexei; Hallbeck, Martin

    2011-01-01

    and accumulation of Aβ within lysosomes, induced apoptosis in differentiated SH-SY5Y neuroblastoma cells. Cells under hyperoxia showed: (1) increased numbers of autophagic vacuoles that contained amyloid precursor protein (APP) as well as Aβ monomers and oligomers, (2) increased reactive oxygen species production...... and resulting lysosomal Aβ accumulation are essential for oxidant-induced apoptosis in cultured neuroblastoma cells and provide additional support for the interactive role of oxidative stress and the lysosomal system in AD-related neurodegeneration....

  17. Marijuana use and increased risk of squamous cell carcinoma of the head and neck.

    Science.gov (United States)

    Zhang, Z F; Morgenstern, H; Spitz, M R; Tashkin, D P; Yu, G P; Marshall, J R; Hsu, T C; Schantz, S P

    1999-12-01

    Marijuana is the most commonly used illegal drug in the United States. In some subcultures, it is widely perceived to be harmless. Although the carcinogenic properties of marijuana smoke are similar to those of tobacco, no epidemiological studies of the relationship between marijuana use and head and neck cancer have been published. The relationship between marijuana use and head and neck cancer was investigated by a case-control study of 173 previously untreated cases with pathologically confirmed diagnoses of squamous cell carcinoma of the head and neck and 176 cancer-free controls at Memorial Sloan-Kettering Cancer Center between 1992 and 1994. Epidemiological data were collected by using a structured questionnaire, which included history of tobacco smoking, alcohol use, and marijuana use. The associations between marijuana use and head and neck cancer were analyzed by Mantel-Haenszel methods and logistic regression models. Controlling for age, sex, race, education, alcohol consumption, pack-years of cigarette smoking, and passive smoking, the risk of squamous cell carcinoma of the head and neck was increased with marijuana use [odds ratio (OR) comparing ever with never users, 2.6; 95% confidence interval (CI), 1.1-6.6]. Dose-response relationships were observed for frequency of marijuana use/day (P for trend marijuana use (P for trend marijuana use were observed with cigarette smoking, mutagen sensitivity, and to a lesser extent, alcohol use. Our results suggest that marijuana use may increase the risk of head and neck cancer with a strong dose-response pattern. Our analysis indicated that marijuana use may interact with mutagen sensitivity and other risk factors to increase the risk of head and neck cancer. The results need to be interpreted with some caution in drawing causal inferences because of certain methodological limitations, especially with regard to interactions.

  18. Cold hypersensitivity increases with age in mice with sickle cell disease

    Science.gov (United States)

    Zappia, Katherine J.; Garrison, Sheldon R.; Hillery, Cheryl A.; Stucky, Cheryl L.

    2014-01-01

    Sickle cell disease (SCD) is associated with acute vaso-occlusive crises that trigger painful episodes and frequently involves ongoing, chronic pain. Additionally, both humans and mice with SCD experience heighted cold sensitivity. However, studies have not addressed the mechanism(s) underlying the cold sensitization, nor its progression with age. Here we measured thermotaxis behavior in young and aged mice with severe SCD. Sickle mice had a marked increase in cold sensitivity measured by a cold preference test. Further, cold hypersensitivity worsened with advanced age. We assessed whether enhanced peripheral input contributes to the chronic cold pain behavior by recording from C fibers, many of which are cold-sensitive, in skin-nerve preparations. We observed that C fibers from sickle mice displayed a shift to warmer (more sensitive) cold-detection thresholds. To address mechanisms underlying the cold sensitization in primary afferent neurons, we quantified mRNA expression levels for ion channels thought to be involved in cold detection. These included the Transient Receptor Potential Melastatin 8 (Trpm8) and TRP Ankyrin 1 (Trpa1) channels, as well as the two-pore domain potassium channels, TREK-1 (Kcnk2), TREK-2 (Kcnk4), and TRAAK (Kcnk10). Surprisingly, transcript expression levels of all of these channels were comparable between sickle and control mice. We further examined transcript expression of 83 additional pain-related genes and found increased mRNA levels for endothelin 1 and tachykinin receptor 1. These factors may contribute to hypersensitivity in sickle mice at both the afferent and behavioral levels. Sensory neurons from sickle cell disease mice are sensitized to cold, mirroring behavioral observations, and have increased expression of endothelin 1 and tachykinin receptor 1. PMID:24953902

  19. Noise-induced nitrotyrosine increase and outer hair cell death in guinea pig cochlea.

    Science.gov (United States)

    Han, Wei-ju; Shi, Xiao-rui; Nuttall, Alfred

    2013-01-01

    Modern research has provided new insights into the biological mechanisms of noise-induced hearing loss, and a number of studies showed the appearance of increased reactive oxygen species (ROS) and reactive nitrogen species (RNS) during and after noise exposure. This study was designed to investigate the noise exposure induced nitrotyrosine change and the mechanism of outer hair cells death in guinea pig cochlea. Thirty guinea pigs were used in this study. The experimental animals were either exposed for 4 hours per day to broadband noise at 122 dB SPL (A-weighted) for 2 consecutive days or perfused cochleae with 5 mg/ml of the SIN1 solutions, an exogenous NO and superoxide donor, for 30 minutes. Then the cochleae of the animals were dissected. Propidium iodide (PI), a DNA intercalating fluorescent probe, was used to trace morphological changes in OHC nuclei. The distribution of nitrotyrosine (NT) in the organ of Corti and the cochlear lateral wall tissue from the guinea pigs were examined using fluorescence immunohistochemistry method. Whole mounts of organ of Corti were prepared. Morphological and fluorescent changes were examined under a confocal microscope. Either after noise exposure or after SIN1 perfusion, outer hair cells (OHCs) death with characteristics of both apoptotic and necrotic degradation appeared. Nitrotyrosine immunolabeling could be observed in the OHCs from the control animals. After noise exposure, NT immunostaining became much greater than the control animals in OHCs. The apoptotic OHC has significant increase of nitrotyrosine in and around the nucleus following noise exposure. In the normal later wall of cochleae, relatively weak nitrotyrosine immunolabeling could be observed. After noise exposure, nitrotyrosine immunoactivity became stronger in stria vascularis. Noise exposure induced increase of nitrotyrosine production is associated with OHCs death suggesting reactive nitrogen species participation in the cochlear pathophysiology of noise

  20. Increased mitochondrial mass in cells with functionally compromised mitochondria after exposure to both direct gamma radiation and bystander factors.

    LENUS (Irish Health Repository)

    Nugent, Sharon M E

    2007-07-01

    The bystander effect describes radiation-like damage in unirradiated cells either in the vicinity of irradiated cells or exposed to medium from irradiated cells. This study aimed to further characterize the poorly understood mitochondrial response to both direct irradiation and bystander factor(s) in human keratinocytes (HPV-G) and Chinese hamster ovarian cells (CHO-K1). Oxygen consumption rates were determined during periods of state 4, state 3 and uncoupled respiration. Mitochondrial mass was determined using MitoTracker FM. CHO-K1 cells showed significantly reduced oxygen consumption rates 4 h after exposure to 5 Gy direct radiation and irradiated cell conditioned medium (ICCM) and an apparent recovery 12-24 h later. The apparent recovery was likely due to the substantial increase in mitochondrial mass observed in these cells as soon as 4 h after exposure. HPV-G cells, on the other hand, showed a sustained increase in oxygen consumption rates after ICCM exposure and a transient increase 4 h after exposure to 5 Gy direct radiation. A significant increase in mitochondrial mass per HPV-G cell was observed after exposure to both direct radiation and ICCM. These findings are indicative of a stress response to mitochondrial dysfunction that increases the number of mitochondria per cell.

  1. Calcimimetic R-568 and its enantiomer S-568 increase nitric oxide release in human endothelial cells.

    Directory of Open Access Journals (Sweden)

    Mario Bonomini

    Full Text Available Calcimimetics, such as R-568, are thought to activate G protein-linked Ca(2+-sensing receptor (CaSR by allosterically increasing the affinity of the receptor for Ca(2+ allowing for efficient control of uremic hyperparathyroidism. Several recent studies suggest they possess additional vascular actions. Although it has been postulated that calcimimetics may have a direct effect on CaSR in the blood vessels, further studies are needed to elucidate their vascular CaSR-dependent versus CaSR-independent effects.Focusing on human umbilical vein endothelial cells (HUVECs, we studied the CaSR expression and distribution by Immunofluorescence and Western Blot analysis. CaSR function was evaluated by measuring the potential effect of calcimimetic R-568 and its enantiomer S-568 upon the modulation of intracellular Ca(2+ levels (using a single cell approach and FURA-2AM, in the presence or absence of Calhex-231, a negative modulator of CaSR. To address their potential vascular functions, we also evaluated R- and S-568-stimulated enzymatic release of Nitric Oxide (NO by DAF-2DA, by Nitric Oxide Synthase (NOS radiometric assay (both in HUVECs and in Human Aortic Endothelial Cells and by measuring eNOS-ser1177 phosphorylation levels (Immunoblotting. We show that, although the CaSR protein was expressed in HUVECs, it was mainly distributed in cytoplasm while the functional CaSR dimers, usually localized on the plasma membrane, were absent. In addition, regardless of the presence or absence of Calhex-231, both R- and S-568 significantly increased intracellular Ca(2+ levels by mobilization of Ca(2+ from intracellular stores, which in turn augmented NO release by a time- and Ca(2+-dependent increase in eNOS-ser1177 phosphorylation levels.Taken together, these data indicate that in human endothelium there is no stereoselectivity in the responses to calcimimetics and that CaSR is probably not involved in the action of R- and S-568. This suggests an additional

  2. Increased Numbers of NK Cells, NKT-Like Cells, and NK Inhibitory Receptors in Peripheral Blood of Patients with Chronic Obstructive Pulmonary Disease

    Directory of Open Access Journals (Sweden)

    Ying Tang

    2013-01-01

    Full Text Available T cells and B cells participate in the pathogenesis of COPD. Currently, NK cells and NKT cells have gained increasing attention. In the present study, 19 COPD patients and 12 healthy nonsmokers (HNS were recruited, and their pulmonary function was assessed. The frequencies of CD3+ T, CD4+ T, CD8+ T, B, NK, and NKT-like cells were determined using flow cytometry. The frequencies of spontaneous and inducible IFN-γ+ or CD107a+ NK and NKT-like cells as well as activating or inhibitory receptors were also detected. The potential association of lymphocyte subsets with disease severity was further analyzed. Significantly decreased numbers of CD3+ and CD4+ T cells, and the CD4+/CD8+ ratio, but increased numbers of CD3−CD56+ NK and CD3+CD56+ NKT-like cells were observed in COPD patients compared to HNS. The frequencies of inducible IFN-γ-secreting NK and NKT-like cells were less in COPD patients. The frequencies of CD158a and CD158b on NK cells and CD158b on NKT-like cells were greater. The frequency of CD158b+ NK cells was negatively correlated with FEV1% prediction and FEV1/FVC. Our data indicate that COPD patients have immune dysfunction, and higher frequencies of inhibitory NK cells and NKT-like cells may participate in the pathogenesis of COPD.

  3. MG132, a proteasome inhibitor, induces human pulmonary fibroblast cell death via increasing ROS levels and GSH depletion.

    Science.gov (United States)

    Park, Woo Hyun; Kim, Suhn Hee

    2012-04-01

    MG132 as a proteasome inhibitor can induce apoptotic cell death in lung cancer cells. However, little is known about the toxicological cellular effects of MG132 on normal primary lung cells. Here, we investigated the effects of N-acetyl cysteine (NAC) and vitamin C (well known antioxidants) or L-buthionine sulfoximine (BSO; an inhibitor of GSH synthesis) on MG132-treated human pulmonary fibroblast (HPF) cells in relation to cell death, reactive oxygen species (ROS) and glutathione (GSH). MG132 induced growth inhibition and death in HPF cells, accompanied by the loss of mitochondrial membrane potential (MMP; ∆ψm). MG132 increased ROS levels and GSH-depleted cell numbers in HPF cells. Both antioxidants, NAC and vitamin C, prevented growth inhibition, death and MMP (∆ψm) loss in MG132-treated HPF cells and also attenuated ROS levels in these cells. BSO showed a strong increase in ROS levels in MG132-treated HPF cells and slightly enhanced the growth inhibition, cell death, MMP (∆ψm) loss and GSH depletion. In addition, NAC decreased anonymous ubiquitinated protein levels in MG132-treated HPF cells. Furthermore, superoxide dismutase (SOD) 2, catalase (CTX) and GSH peroxidase (GPX) siRNAs enhanced HPF cell death by MG132, which was not correlated with ROS and GSH level changes. In conclusion, MG132 induced the growth inhibition and death of HPF cells, which were accompanied by increasing ROS levels and GSH depletion. Both NAC and vitamin C attenuated HPF cell death by MG132, whereas BSO slightly enhanced the death.

  4. BIX-01294 increases pig cloning efficiency by improving epigenetic reprogramming of somatic cell nuclei.

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    Huang, Jiaojiao; Zhang, Hongyong; Yao, Jing; Qin, Guosong; Wang, Feng; Wang, Xianlong; Luo, Ailing; Zheng, Qiantao; Cao, Chunwei; Zhao, Jianguo

    2016-01-01

    Accumulating evidence suggests that faulty epigenetic reprogramming leads to the abnormal development of cloned embryos and results in the low success rates observed in all mammals produced through somatic cell nuclear transfer (SCNT). The aberrant methylation status of H3K9me and H3K9me2 has been reported in cloned mouse embryos. To explore the role of H3K9me2 and H3K9me in the porcine somatic cell nuclear reprogramming, BIX-01294, known as a specific inhibitor of G9A (histone-lysine methyltransferase of H3K9), was used to treat the nuclear-transferred (NT) oocytes for 14-16 h after activation. The results showed that the developmental competence of porcine SCNT embryos was significantly enhanced both in vitro (blastocyst rate 16.4% vs 23.2%, Pcloning rate 1.59% vs 2.96%) after 50 nm BIX-01294 treatment. BIX-01294 treatment significantly decreased the levels of H3K9me2 and H3K9me at the 2- and 4-cell stages, which are associated with embryo genetic activation, and increased the transcriptional expression of the pluripotency genes SOX2, NANOG and OCT4 in cloned blastocysts. Furthermore, the histone acetylation levels of H3K9, H4K8 and H4K12 in cloned embryos were decreased after BIX-01294 treatment. However, co-treatment of activated NT oocytes with BIX-01294 and Scriptaid rescued donor nuclear chromatin from decreased histone acetylation of H4K8 that resulted from exposure to BIX-01294 only and consequently improved the preimplantation development of SCNT embryos (blastocyst formation rates of 23.7% vs 21.5%). These results indicated that treatment with BIX-01294 enhanced the developmental competence of porcine SCNT embryos through improvements in epigenetic reprogramming and gene expression. © 2016 Society for Reproduction and Fertility.

  5. Hypoxia/reoxygenation increases the permeability of endothelial cell monolayers: Role of oxygen radicals

    International Nuclear Information System (INIS)

    Inauen, W.; Payne, D.K.; Kvietys, P.R.; Granger, D.N.

    1990-01-01

    We assessed the effect of hypoxia/reoxygenation on 14C-albumin flux across endothelial monolayers. Cultured bovine pulmonary artery endothelial cells were grown to confluence on nitrocellulose filters (pore size 12 microns). The endothelialized filters were mounted in Ussing-type chambers which were filled with cell culture medium (M 199). Equimolar amounts (33 nM) of 14C-labeled and unlabeled albumin were added to the hot and cold chambers, respectively. The monolayers were then exposed to successive periods (90 min) of normoxia (pO2 145 mmHg), hypoxia (pO2 20 mmHg), and reoxygenation (pO2 145 mmHg). A gas bubbling system was used to control media pO2 and to ensure adequate mixing. Four aliquots of culture media were taken during each period in order to calculate the 14C-albumin permeability across the endothelialized filter. In some experiments, either the xanthine oxidase inhibitor, oxypurinol (10 microM), or superoxide dismutase (600 U/mL), was added to the media immediately prior to the experiments. As compared to the normoxic control period, albumin permeability was 1.5 times higher during hypoxia (p less than 0.01) and 2.3 times higher during reoxygenation (p less than 0.01). The reoxygenation-induced increase in albumin permeability was prevented by either oxypurinol or superoxide dismutase. These data indicate that xanthine oxidase-derived oxygen radicals contribute to the hypoxia/reoxygenation-induced endothelial cell dysfunction. The altered endothelial barrier function induced by hypoxia/reoxygenation is consistent with the microvascular dysfunction observed following reperfusion of ischemic tissues

  6. Centella asiatica increases B-cell lymphoma 2 expression in rat prefrontal cortex

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    Kuswati

    2015-04-01

    Full Text Available Background Stress is one of the factors that cause apoptosis in neuronal cells. Centella asiatica has a neuroprotective effect that can inhibit apoptosis. This study aimed to examine the effect of Centella asiatica ethanol extract on B-cell lymphoma 2 (Bcl-2 protein expression in the prefrontal cortex of rats. Methods An experimental study was conducted on 34 brain tissue samples from male Sprague Dawley rats exposed to chronic restraint stress for 21 days. The samples were taken from following groups: non-stress group K, negative control group P1 (stress + arabic gum powder, P2 (stress + C.asiatica at 150 mg/kgBW, P3 (stress + C.asiatica at 300 mg/kg BW, P4 (stress + C.asiatica at 600 mg/kg body weight and positive control group P5 (stress + fluoxetine at 10 mg/kgBW. The samples were made into sections that were stained immunohistochemically using Bcl-2 antibody to determine the percentage of cells expressing Bcl-2. Data were analyzed using one way ANOVA test followed by a post - hoc test. Results There were significant differences in mean Bcl-2 expression between the groups receiving Centella asiatica compared with the non-stress group and stress-only group (negative control group (p<0.05. The results were comparable to those of the fluoxetine treatment group. Conclusion The Centella asiatica ethanol extract was able to increase Bcl-2 expression in the prefrontal cortex of Sprague Dawley rats exposed to restraint stress. This study suggests that Centella asiatica may be useful in the treatment of cerebral stress.

  7. Increased Number of Langerhans Cells in the Epidermis of Diabetic Foot Ulcers Correlates with Healing Outcome

    Science.gov (United States)

    Stojadinovic, Olivera; Yin, Natalie; Lehmann, Janin; Pastar, Irena; Kirsner, Robert S.; Tomic-Canic, Marjana

    2015-01-01

    Langerhans cells (LCs) are a specialized subset of epidermal dendritic cells. They represent one of the first cells of immunological barrier and play an important role during the inflammatory phase of acute wound healing. Despite considerable progress in our understanding of the immunopathology of diabetes mellitus and its associated co-morbidities such as diabetic foot ulcers (DFUs), considerable gaps in our knowledge exist. In this study, we utilized the human ex vivo wound model and confirmed the increased epidermal LCs at wound edges during early phases of wound healing. Next, we aimed to determine differences in quantity of LCs between normal human and diabetic foot skin and to learn if the presence of LCs correlates with the healing outcome in DFUs. We utilized immunofluorescence to detect CD207+ LCs in specimens from normal and diabetic foot skin and DFU wound edges. Specimens from DFUs were collected at the initial visit and 4 weeks at the time when the healing outcome was determined. DFUs that decreased in size by >50% were considered to be healing, while DFUs with a size reduction of healing. Quantitative assessment of LCs showed a higher number of LCs in healing when compared to non–healing DFU’s. Our findings provide evidence that LCs are present in higher number in diabetic feet than normal foot skin. Healing DFUs show a higher number of LCs compared to non-healing DFUs. These findings indicate that the epidermal immune barrier plays an important role in the DFU healing outcome and may offer new therapeutic avenues targeting LC in non-healing DFUs. PMID:24277309

  8. Merkel cell carcinoma: Current US incidence and projected increases based on changing demographics.

    Science.gov (United States)

    Paulson, Kelly G; Park, Song Youn; Vandeven, Natalie A; Lachance, Kristina; Thomas, Hannah; Chapuis, Aude G; Harms, Kelly L; Thompson, John A; Bhatia, Shailender; Stang, Andreas; Nghiem, Paul

    2018-03-01

    Merkel cell carcinoma (MCC) incidence rates are rising and strongly age-associated, relevant for an aging population. Determine MCC incidence in the United States and project incident cases through the year 2025. Registry data were obtained from the SEER-18 Database, containing 6600 MCC cases. Age- and sex-adjusted projections were generated using US census data. During 2000-2013, the number of reported solid cancer cases increased 15%, melanoma cases increased 57%, and MCC cases increased 95%. In 2013, the MCC incidence rate was 0.7 cases/100,000 person-years in the United States, corresponding to 2488 cases/year. MCC incidence increased exponentially with age, from 0.1 to 1.0 to 9.8 (per 100,000 person-years) among age groups 40-44 years, 60-64 years, and ≥85 years, respectively. Due to aging of the Baby Boomer generation, US MCC incident cases are predicted to climb to 2835 cases/year in 2020 and 3284 cases/year in 2025. We assumed that the age-adjusted incidence rate would stabilize, and thus, the number of incident cases we projected might be an underestimate. An aging population is driving brisk increases in the number of new MCC cases in the United States. This growing impact combined with the rapidly evolving therapeutic landscape warrants expanded awareness of MCC diagnosis and management. Copyright © 2017 American Academy of Dermatology, Inc. Published by Elsevier Inc. All rights reserved.

  9. Peripheral mononuclear cell resistin mRNA expression is increased in type 2 diabetic women.

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    Tsiotra, Panayoula C; Tsigos, Constantine; Anastasiou, Eleni; Yfanti, Eleni; Boutati, Eleni; Souvatzoglou, Emmanouil; Kyrou, Ioannis; Raptis, Sotirios A

    2008-01-01

    Resistin has been shown to cause insulin resistance and to impair glucose tolerance in rodents, but in humans its physiological role still remains elusive. The aim of this study was to examine whether resistin mRNA expression in human peripheral mononuclear cells (PBMCs) and its corresponding plasma levels are altered in type 2 diabetes. Resistin mRNA levels were easily detectable in human PBMC, and found to be higher in DM2 compared to healthy women (P = .05). Similarly, mononuclear mRNA levels of the proinflammatory cytokines IL-1beta, TNF-alpha, and IL-6 were all significantly higher in DM2 compared to control women (P DM2 women (P = .051), and overall, they correlated significantly with BMI (r = 0.406, P = .010) and waist circumference (r = 0.516, P = .003), but not with fasting insulin levels or HOMA-IR. Resistin mRNA expression is increased in PBMC from DM2 women, together with increased expression of the inflammatory cytokines IL-1beta, TNF-alpha, and IL-6, independent of obesity. These results suggest that resistin and cytokines might contribute to the low-grade inflammation and the increased atherogenic risk observed in these patients.

  10. Increasing power generation for scaling up single-chamber air cathode microbial fuel cells

    KAUST Repository

    Cheng, Shaoan; Logan, Bruce E.

    2011-01-01

    Scaling up microbial fuel cells (MFCs) requires a better understanding the importance of the different factors such as electrode surface area and reactor geometry relative to solution conditions such as conductivity and substrate concentration. It is shown here that the substrate concentration has significant effect on anode but not cathode performance, while the solution conductivity has a significant effect on the cathode but not the anode. The cathode surface area is always important for increasing power. Doubling the cathode size can increase power by 62% with domestic wastewater, but doubling the anode size increases power by 12%. Volumetric power density was shown to be a linear function of