Sample records for a549 sk-ov-3 sk-mel-2

  1. Stably Fluorescent Cell Line of Human Ovarian Epithelial Cancer Cells SK-OV-3ip-red.

    PubMed

    Konovalova, E V; Shulga, A A; Chumakov, S P; Khodarovich, Yu M; Woo, Eui-Jeon; Deev, S M

    2017-11-01

    Stable red fluorescing line of human ovarian epithelial cancer cells SK-OV-3ip-red was generated expressing gene coding for protein TurboFP635 (Katushka) fluorescing in the far-red spectrum region with excitation and emission peaks at 588 and 635 nm, respectively. Fluorescence of SK-OV-3ip-red line remained high during long-term cell culturing and after cryogenic freezing. The obtained cell line SK-OV-3ip-red can serve a basis for a model of a scattered tumor with numerous/extended metastases and used both for testing anticancer drugs inhibiting metastasis growth and for non-invasive monitoring of the growth dynamics with high precision.

  2. Synthesis and evaluation of (Z)-2,3-diphenylacrylonitrile analogs as anti-cancer and anti-microbial agents.

    PubMed

    Alam, Mohammad Sayed; Nam, Young-Joo; Lee, Dong-Ung

    2013-11-01

    In the present study, a series of (Z)-2,3-diphenylacrylonitrile analogs were synthesized and then evaluated in terms of their cytotoxic activities against four human cancer cell lines, e.g. lung cancer (A549), ovarian cancer (SK-OV-3), skin cancer (SK-MEL-2), and colon cancer (HCT15), as well as anti-microbial activities against three microbes, e.g. Staphylococcus aureus, Salmonella typhi, and Aspergillus niger. The title compounds were synthesized by Knoevenagel condensation reaction of benzyl cyanide or p-nitrobenzyl cyanide with substituted benzaldehydes in good yields. Most of the compounds exhibited significant suppressive activities against the growth of all cancer cell lines. Compound 3c was most active in inhibiting the growth of A549, SK-OV-3, SK-MEL-2, and HCT15 cells lines with IC50 values of 0.57, 0.14, 0.65, and 0.34 mg/mL, respectively, followed by compounds 3f, 3i, and 3h. Compound 3c exhibited 2.4 times greater cytotoxic activity against HCT15 cells, whereas it showed similar potency against SK-OV-3 cells to that of the standard anti-cancer agent doxorubicin. Structure-activity relationship study revealed that electron-donating groups at the para-position of phenyl ring B were more favorable for improved cytotoxic activity, whereas the presence of electron-withdrawing groups was unfavorable compare to unsubstituted acrylonitrile. An optimal electron density on phenyl ring A of (Z)-2,3-diphenylacrylonitrile analogs was crucial for their cytotoxic activities against human cancer cell lines used in the present study. Qualitative structure-cytotoxic activity relationships were studied using physicochemical parameters; a good correlation between calculated polar surface area (PSA), a lipophobic parameter, and cytotoxic activity was found. Moreover, all compounds showed significant anti-bacterial activities against S. typhi, whereas compound 3k showed potent inhibition against both S. aureus and S. typhi bacterial strains. Copyright © 2013 Elsevier

  3. Pharbinilic acid, an allogibberic acid from morning glory (Pharbitis nil).

    PubMed

    Kim, Ki Hyun; Choi, Sang Un; Son, Mi Won; Choi, Sang Zin; Clardy, Jon; Lee, Kang Ro

    2013-07-26

    Pharbinilic acid (1), the first naturally occurring allogibberic acid, was isolated from ethanol extracts of morning glory (Pharbitis nil) seeds. Its absolute configuration was determined by NOESY NMR and ECD experiments. Compound 1 showed weak cytotoxicity against A549, SK-OV-3, SK-MEL-2, and HCT-15 cells and weakly inhibited nitric oxide production in lipopolysaccharide-activated BV-2 microglia cells.

  4. Specific pesticide-dependent increases in α-synuclein levels in human neuroblastoma (SH-SY5Y) and melanoma (SK-MEL-2) cell lines.

    PubMed

    Chorfa, Areski; Bétemps, Dominique; Morignat, Eric; Lazizzera, Corinne; Hogeveen, Kevin; Andrieu, Thibault; Baron, Thierry

    2013-06-01

    Epidemiological studies indicate a role of genetic and environmental factors in Parkinson's disease involving alterations of the neuronal α-synuclein (α-syn) protein. In particular, a relationship between Parkinson's disease and occupational exposure to pesticides has been repeatedly suggested. Our objective was to precisely assess changes in α-syn levels in human neuroblastoma (SH-SY5Y) and melanoma (SK-MEL-2) cell lines following acute exposure to pesticides (rotenone, paraquat, maneb, and glyphosate) using Western blot and flow cytometry. These human cell lines express α-syn endogenously, and overexpression of α-syn (wild type or mutated A53T) can be obtained following recombinant adenoviral transduction. We found that endogenous α-syn levels in the SH-SY5Y neuroblastoma cell line were markedly increased by paraquat, and to a lesser extent by rotenone and maneb, but not by glyphosate. Rotenone also clearly increased endogenous α-syn levels in the SK-MEL-2 melanoma cell line. In the SH-SY5Y cell line, similar differences were observed in the α-syn adenovirus-transduced cells, with a higher increase of the A53T mutated protein. Paraquat markedly increased α-syn in the SK-MEL-2 adenovirus-transduced cell line, similarly for the wild-type or A53T proteins. The observed differences in the propensities of pesticides to increase α-syn levels are in agreement with numerous reports that indicate a potential role of exposure to certain pesticides in the development of Parkinson's disease. Our data support the hypothesis that pesticides can trigger some molecular events involved in this disease and also in malignant melanoma that consistently shows a significant but still unexplained association with Parkinson's disease.

  5. Process optimization and photostability of silymarin nanostructured lipid carriers: effect on UV-irradiated rat skin and SK-MEL 2 cell line.

    PubMed

    Singh, Pooja; Singh, Mahendra; Kanoujia, Jovita; Arya, Malti; Saraf, Shailendra K; Saraf, Shubhini A

    2016-10-01

    The objective of the present work was to formulate a novel stable delivery system which would not only overcome the solubility issue of silymarin, but also help to increase the therapeutic value by better permeation, anticancer action and reduced toxicity. This was envisaged through the recent developments in nanotechnology, combined with the activity of the phytoconstituent silymarin. A 2(3) full factorial design based on three independent variables was used for process optimization of nanostructured lipid carriers (NLC). Developed formulations were evaluated on the basis of particle size, morphology, in vitro drug release, photostability and cell line studies. Optimized silymarin-NLC was incorporated into carbopol gel and further assessed for rheological parameters. Stable behaviour in presence of light was proven by photostability testing of formulation. Permeability parameters were significantly higher in NLC as compared to marketed phytosome formulation. The NLC based gel described in this study showed faster onset, and prolonged activity up to 24 h and better action against edema as compared to marketed formulation. In case of anticancer activity of silymarin-NLC against SK-MEL 2 cell lines, silymarin-NLC proved to possess anticancer activity in a dose-dependent manner (10-80 μM) and induced apoptosis at 80 μM in SK-MEL 2 cancer cells. This work documents for the first time that silymarin can be formulated into nanostructured lipoidal carrier system for enhanced permeation, greater stability as well as anticancer activity for skin.

  6. Estimates of (239+240)Pu inventories in Gdańsk Bay and Gdańsk basin.

    PubMed

    Skwarzec, Bogdan; Strumińska, Dagmara I; Prucnal, Małgorzata

    2003-01-01

    This paper presents and discusses the results of (239+240)Pu determinations in different components of Gdańsk bay and Gdańsk basin ecosystem, as well as estimated sources and inventories of plutonium in these basins. The total plutonium (239+240)Pu activities deposited in Gdańsk bay and Gdańsk basin sediments are 1.18 TBq and 3.77 TBq, respectively. Two rivers, the Vistula and Neman rivers, and atmospheric fallout were distinguished as the main sources of plutonium in these basins. In seawater (with suspended matter included) there is about 2.33 GBq (239+240)Pu (0.2% of total activity) in Gdańsk bay and 9.92 GBq (239+240)Pu (0.3% of total activity) in Gdańsk basin. In both cases, 56% of (239+240)Pu is associated with suspended matter. Organisms contain 3.81 MBq in Gdańsk bay and 7.45 MBq (239+240)Pu in Gdańsk basin. From this value in Gdańsk bay 82.1% of plutonium is associated with zoobenthos, 13.6% with phytobenthos, 1.6% with phytoplankton, 1.5% with zooplankton and 1.2% with fish. In Gdańsk basin, 83.2% is associated with zoobenthos, 7.5% with phytobenthos, 3.6% with phytoplankton, 3.2% with zooplankton and 2.5% with fish.

  7. MuSK induced experimental autoimmune myasthenia gravis does not require IgG1 antibody to MuSK.

    PubMed

    Küçükerden, Melike; Huda, Ruksana; Tüzün, Erdem; Yılmaz, Abdullah; Skriapa, Lamprini; Trakas, Nikos; Strait, Richard T; Finkelman, Fred D; Kabadayı, Sevil; Zisimopoulou, Paraskevi; Tzartos, Socrates; Christadoss, Premkumar

    2016-06-15

    Sera of myasthenia gravis (MG) patients with muscle-specific receptor kinase-antibody (MuSK-Ab) predominantly display the non-complement fixing IgG4 isotype. Similarly, mouse IgG1, which is the analog of human IgG4, is the predominant isotype in mice with experimental autoimmune myasthenia gravis (EAMG) induced by MuSK immunization. The present study was performed to determine whether IgG1 anti-MuSK antibody is required for immunized mice to develop EAMG. Results demonstrated a significant correlation between clinical severity of EAMG and levels of MuSK-binding IgG1+, IgG2+ and IgG3+ peripheral blood B cells in MuSK-immunized wild-type (WT) mice. Moreover, MuSK-immunized IgG1 knockout (KO) and WT mice showed similar EAMG severity, serum MuSK-Ab levels, muscle acetylcholine receptor concentrations, neuromuscular junction immunoglobulin and complement deposit ratios. IgG1 and IgG3 were the predominant anti-MuSK isotypes in WT and IgG1 KO mice, respectively. These observations demonstrate that non-IgG1 isotypes can mediate MuSK-EAMG pathogenesis. Copyright © 2016 Elsevier B.V. All rights reserved.

  8. SK2 channels regulate mitochondrial respiration and mitochondrial Ca2+ uptake.

    PubMed

    Honrath, Birgit; Matschke, Lina; Meyer, Tammo; Magerhans, Lena; Perocchi, Fabiana; Ganjam, Goutham K; Zischka, Hans; Krasel, Cornelius; Gerding, Albert; Bakker, Barbara M; Bünemann, Moritz; Strack, Stefan; Decher, Niels; Culmsee, Carsten; Dolga, Amalia M

    2017-05-01

    Mitochondrial calcium ([Ca 2+ ] m ) overload and changes in mitochondrial metabolism are key players in neuronal death. Small conductance calcium-activated potassium (SK) channels provide protection in different paradigms of neuronal cell death. Recently, SK channels were identified at the inner mitochondrial membrane, however, their particular role in the observed neuroprotection remains unclear. Here, we show a potential neuroprotective mechanism that involves attenuation of [Ca 2+ ] m uptake upon SK channel activation as detected by time lapse mitochondrial Ca 2+ measurements with the Ca 2+ -binding mitochondria-targeted aequorin and FRET-based [Ca 2+ ] m probes. High-resolution respirometry revealed a reduction in mitochondrial respiration and complex I activity upon pharmacological activation and overexpression of mitochondrial SK2 channels resulting in reduced mitochondrial ROS formation. Overexpression of mitochondria-targeted SK2 channels enhanced mitochondrial resilience against neuronal death, and this effect was inhibited by overexpression of a mitochondria-targeted dominant-negative SK2 channel. These findings suggest that SK channels provide neuroprotection by reducing [Ca 2+ ] m uptake and mitochondrial respiration in conditions, where sustained mitochondrial damage determines progressive neuronal death.

  9. SK2 channels regulate mitochondrial respiration and mitochondrial Ca2+ uptake

    PubMed Central

    Honrath, Birgit; Matschke, Lina; Meyer, Tammo; Magerhans, Lena; Perocchi, Fabiana; Ganjam, Goutham K; Zischka, Hans; Krasel, Cornelius; Gerding, Albert; Bakker, Barbara M; Bünemann, Moritz; Strack, Stefan; Decher, Niels; Culmsee, Carsten; Dolga, Amalia M

    2017-01-01

    Mitochondrial calcium ([Ca2+]m) overload and changes in mitochondrial metabolism are key players in neuronal death. Small conductance calcium-activated potassium (SK) channels provide protection in different paradigms of neuronal cell death. Recently, SK channels were identified at the inner mitochondrial membrane, however, their particular role in the observed neuroprotection remains unclear. Here, we show a potential neuroprotective mechanism that involves attenuation of [Ca2+]m uptake upon SK channel activation as detected by time lapse mitochondrial Ca2+ measurements with the Ca2+-binding mitochondria-targeted aequorin and FRET-based [Ca2+]m probes. High-resolution respirometry revealed a reduction in mitochondrial respiration and complex I activity upon pharmacological activation and overexpression of mitochondrial SK2 channels resulting in reduced mitochondrial ROS formation. Overexpression of mitochondria-targeted SK2 channels enhanced mitochondrial resilience against neuronal death, and this effect was inhibited by overexpression of a mitochondria-targeted dominant-negative SK2 channel. These findings suggest that SK channels provide neuroprotection by reducing [Ca2+]m uptake and mitochondrial respiration in conditions, where sustained mitochondrial damage determines progressive neuronal death. PMID:28282037

  10. SK3 is an important component of K(+) channels mediating the afterhyperpolarization in cultured rat SCG neurones.

    PubMed

    Hosseini, R; Benton, D C; Dunn, P M; Jenkinson, D H; Moss, G W

    2001-09-01

    1. Our aim was to identify the small-conductance Ca(2+)-activated K(+) channel(s) (SK) underlying the apamin-sensitive afterhyperpolarization (AHP) in rat superior cervical ganglion (SCG) neurones. 2. Degenerate oligonucleotide primers designed to the putative calmodulin-binding domain conserved in all mammalian SK channel sequences were employed to detect SK DNA in a cDNA library from rat SCG. Only a single band, corresponding to a fragment of the rSK3 gene, was amplified. 3. Northern blot analysis employing a PCR-generated rSK3 fragment showed the presence of mRNA coding for SK3 in SCG as well in other rat peripheral tissues including adrenal gland and liver. 4. The same rSK3 fragment enabled the isolation of a full-length rSK3 cDNA from the library. Its sequence was closely similar to, but not identical with, that of the previously reported rSK3 gene. 5. Expression of the rSK3 gene in mammalian cell lines (CHO, HEK cells) caused the appearance of a K(+) conductance with SK channel properties. 6. The application of selective SK blocking agents (including apamin, scyllatoxin and newer non-peptidic compounds) showed these homomeric SK3 channels to have essentially the same pharmacological characteristics as the SCG afterhyperpolarization, but to differ from those of homomeric SK1 and SK2 channels. 7. Immunohistochemistry using a rSK3 antipeptide antibody revealed the presence of SK3 protein in the cell bodies and processes of cultured SCG neurones. 8. Taken together, these results identify SK3 as a major component of the SK channels responsible for the afterhyperpolarization of cultured rat SCG neurones.

  11. Reconstitution of a functional 7SK snRNP

    PubMed Central

    Brogie, John E.

    2017-01-01

    Abstract The 7SK small nuclear ribonucleoprotein (snRNP) plays a central role in RNA polymerase II elongation control by regulating the availability of active P-TEFb. We optimized conditions for analyzing 7SK RNA by SHAPE and demonstrated a hysteretic effect of magnesium on 7SK folding dynamics including a 7SK GAUC motif switch. We also found evidence that the 5΄ end pairs alternatively with two different regions of 7SK giving rise to open and closed forms that dictate the state of the 7SK motif. We then used recombinant P-TEFb, HEXIM1, LARP7 and MEPCE to reconstruct a functional 7SK snRNP in vitro. Stably associated P-TEFb was highly inhibited, but could still be released and activated by HIV-1 Tat. Notably, P-TEFb association with both in vitro-reconstituted and cellular snRNPs led to similar changes in SHAPE reactivities, confirming that 7SK undergoes a P-TEFb-dependent structural change. We determined that the xRRM of LARP7 binds to the 3΄ stem loop of 7SK and inhibits the methyltransferase activity of MEPCE through a C-terminal MEPCE interaction domain (MID). Inhibition of MEPCE is dependent on the structure of the 3΄ stem loop and the closed form of 7SK RNA. This study provides important insights into intramolecular interactions within the 7SK snRNP. PMID:28431135

  12. The SK3 channel promotes placental vascularization by enhancing secretion of angiogenic factors.

    PubMed

    Rada, Cara C; Murray, Grace; England, Sarah K

    2014-11-15

    Proper placental perfusion is essential for fetal exchange of oxygen, nutrients, and waste with the maternal circulation. Impairment of uteroplacental vascular function can lead to pregnancy complications, including preeclampsia and intrauterine growth restriction (IUGR). Potassium channels have been recognized as regulators of vascular proliferation, angiogenesis, and secretion of vasoactive factors, and their dysfunction may underlie pregnancy-related vascular diseases. Overexpression of one channel in particular, the small-conductance calcium-activated potassium channel 3 (SK3), is known to increase vascularization in mice, and mice overexpressing the SK3 channel (SK3(T/T) mice) have a high rate of fetal demise and IUGR. Here, we show that overexpression of SK3 causes fetal loss through abnormal placental vascularization. We previously reported that, at pregnancy day 14, placentas isolated from SK3(T/T) mice are smaller than those obtained from wild-type mice. In this study, histological analysis reveals that SK3(T/-) placentas at this stage have abnormal placental morphology, and microcomputed tomography shows that these placentas have significantly larger and more blood vessels than those from wild-type mice. To identify the mechanism by which these vascularization defects occur, we measured levels of vascular endothelial growth factor (VEGF), placental growth factor, and the soluble form of VEGF receptor 1 (sFlt-1), which must be tightly regulated to ensure proper placental development. Our data reveal that overexpression of SK3 alters systemic and placental ratios of the angiogenic factor VEGF to antiangiogenic factor sFlt-1 throughout pregnancy. Additionally, we observe increased expression of hypoxia-inducing factor 2α in SK3(T/-) placentas. We conclude that the SK3 channel modulates placental vascular development and fetal health by altering VEGF signaling. Copyright © 2014 the American Physiological Society.

  13. Feruloylserotonin inhibits hydrogen peroxide-induced melanogenesis and apoptosis in B16F10 and SK-Mel-2 melanoma cells.

    PubMed

    Cho, Hyejoung; Kim, Okjoon; Lee, Younghee; Kang, Li-Jung; Nguyen, Cam Ngoc; Ishihara, Atsushi; Kim, Hye-Eun

    2017-09-30

    Feruloylserotonin (FS) is a major bioactive component of safflower seeds, with documented strong antibacterial, anti-inflammatory, and free radical scavenging activities. Reactive oxygen species (ROS) can strongly induce melanogenesis and cell apoptosis. The present study aimed to investigate the ability of FS in preventing hydrogen peroxide (H 2 O 2 )-induced melanogenesis and cell apoptosis. Melanogenesis and apoptotic cell death were induced by transient exposure to H 2 O 2 in B16F10 and SK-Mel-2 melanoma cells. FS significantly inhibited melanogenesis and cell death in both cell lines. FS inhibited H 2 O 2 -induced melanin production by down-regulating CREB/MITF/TYR signaling via inhibited intracellular cAMP accumulation. Additionally, FS induced extracellular regulated kinase activation, which led to the degradation of MITF and consequently decreased TYR expression and melanin production in H 2 O 2 -stimulated cells. Furthermore, FS inhibited H 2 O 2 -induced apoptotic cell death by maintaining mitochondrial membrane potential. Therefore, FS might have potential use for cosmetic whitening and as a therapeutic agent for hyperpigmentation disorder. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

  14. MuSK myasthenia gravis and pregnancy.

    PubMed

    Santos, Ernestina; Braga, Antonio; Gabriel, Denis; Duarte, Sara; Martins da Silva, Ana; Matos, Ilda; Freijo, Marta; Martins, Joao; Silveira, Fernando; Nadais, Goreti; Sousa, Filipa; Fraga, Carla; Santos Silva, Rosa; Lopes, Carlos; Gonçalves, Guilherme; Pinto, Clara; Sousa Braga, Jorge; Leite, Maria Isabel

    2018-02-01

    Muscle specific kinase (MuSK) myasthenia gravis (MG, MuSK-MG) is a rare subgroup of MG affecting mainly women during childbearing years. We investigated the influence of pregnancy in the course of MuSK-MG and pregnancy outcomes in females with MuSK-MG. A multicentre cohort of 17 women with MuSK-MG was studied retrospectively; 13 of them with ≥1 pregnancy. MuSK-MG onset age was 35,4 years; 23,0% had other autoimmune disorder; 46,2% were treatment refractory. Thirteen women experienced 27 pregnancies, either after MG onset (group I) (n = 4; maternal age at conception = 29.8 years) or before MG onset (group II) (n = 23; maternal age at conception = 26.2 years). In group I pregnancy occurred in average 9.8 years after the MG onset; it occurred in average 17.0 years before MG in group II. In group I, all were on steroids at time of conception, one on azathioprine and another receiving IVIG regularly. There were mild exacerbations that responded to treatment adjustments. There were no relapses in the 12 months following the delivery. There was no pre-eclampsia, birth defects or stillbirths in either group; 3 miscarriages in group II. One case of neonatal MG was recorded. In this small series, pregnancy did not seem to precipitate MuSK-MG or to have a major influence in the MuSK-MG course, and there was no apparent negative impact in pregnancy outcomes in those where pregnancy followed the MG onset. The weight was lower in the newborn of the group I mothers, although none had low birth weight. Copyright © 2017 Elsevier B.V. All rights reserved.

  15. Sequencing and molecular dissection of Sk-2 in Neurospora

    USDA-ARS?s Scientific Manuscript database

    Neurospora Spore killer-2 is a selfish meiotic drive element that kills its non-Sk-2 siblings during sexual sporulation. Although the location of Sk-2 has been mapped to a large recombination-suppressed region of chromosome III, the physical length and exact borders of the element have remained unde...

  16. Developmental profile of SK2 channel expression and function in CA1 neurons

    PubMed Central

    Ballesteros-Merino, Carmen; Lin, Mike; Wu, Wendy W.; Ferrandiz-Huertas, Clotilde; Cabañero, María J.; Watanabe, Masahiko; Fukazawa, Yugo; Shigemoto, Ryuichi; Maylie, James; Adelman, John P.; Luján, Rafael

    2012-01-01

    We investigated the temporal and spatial expression of SK2 in the developing mouse hippocampus using molecular and biochemical techniques, quantitative immunogold electron microscopy and electrophysiology. The mRNA encoding SK2 was expressed in the developing and adult hippocampus. Western blotting and immunohistochemistry showed that SK2 protein increased with age. This was accompanied by a shift in subcellular localization. Early in development (P5), SK2 was predominantly localized to the endoplasmic reticulum in the pyramidal cell layer. But by P30 SK2 was almost exclusively expressed in the dendrites and spines. The level of SK2 at the postsynaptic density (PSD) also increased during development. In the adult, SK2 expression on the spine plasma membrane showed a proximal-to-distal gradient. Consistent with this redistribution and gradient of SK2, the selective SK channel blocker apamin increased evoked excitatory postsynaptic potentials (EPSPs) only in CA1 pyramidal neurons from mice older than P15. However, the effect of apamin on EPSPs was not different between synapses in proximal or distal stratum radiatum or stratum lacunosum-moleculare in adult. These results show a developmental increase and gradient in SK2-containing channel surface expression that underlie their influence on neurotransmission, and that may contribute to increased memory acquisition during early development. PMID:22072564

  17. SK4 channels modulate Ca2+ signalling and cell cycle progression in murine breast cancer.

    PubMed

    Steudel, Friederike A; Mohr, Corinna J; Stegen, Benjamin; Nguyen, Hoang Y; Barnert, Andrea; Steinle, Marc; Beer-Hammer, Sandra; Koch, Pierre; Lo, Wing-Yee; Schroth, Werner; Hoppe, Reiner; Brauch, Hiltrud; Ruth, Peter; Huber, Stephan M; Lukowski, Robert

    2017-09-01

    Oncogenic signalling via Ca 2+ -activated K + channels of intermediate conductance (SK4, also known as K Ca 3.1 or IK) has been implicated in different cancer entities including breast cancer. Yet, the role of endogenous SK4 channels for tumorigenesis is unclear. Herein, we generated SK4-negative tumours by crossing SK4-deficient (SK4 KO) mice to the polyoma middle T-antigen (PyMT) and epidermal growth factor receptor 2 (cNeu) breast cancer models in which oncogene expression is driven by the retroviral promoter MMTV. Survival parameters and tumour progression were studied in cancer-prone SK4 KO in comparison with wild-type (WT) mice and in a syngeneic orthotopic mouse model following transplantation of SK4-negative or WT tumour cells. SK4 activity was modulated by genetic or pharmacological means using the SK4 inhibitor TRAM-34 in order to establish the role of breast tumour SK4 for cell growth, electrophysiological signalling, and [Ca 2+ ] i oscillations. Ablation of SK4 and TRAM-34 treatment reduced the SK4-generated current fraction, growth factor-dependent Ca 2+ entry, cell cycle progression and the proliferation rate of MMTV-PyMT tumour cells. In vivo, PyMT oncogene-driven tumorigenesis was only marginally affected by the global lack of SK4, whereas tumour progression was significantly delayed after orthotopic implantation of MMTV-PyMT SK4 KO breast tumour cells. However, overall survival and progression-free survival time in the MMTV-cNeu mouse model were significantly extended in the absence of SK4. Collectively, our data from murine breast cancer models indicate that SK4 activity is crucial for cell cycle control. Thus, the modulation of this channel should be further investigated towards a potential improvement of existing antitumour strategies in human breast cancer. © 2017 The Authors. Published by FEBS Press and John Wiley & Sons Ltd.

  18. BMP9 Inhibits Proliferation and Metastasis of HER2-Positive SK-BR-3 Breast Cancer Cells through ERK1/2 and PI3K/AKT Pathways

    PubMed Central

    Ren, Wei; Liu, Yuehong; Wan, Shaoheng; Fei, Chang; Wang, Wei; Chen, Yingying; Zhang, Zhihui; Wang, Ting; Wang, Jinshu; Zhou, Lan; Weng, Yaguang; He, Tongchuan; Zhang, Yan

    2014-01-01

    Bone morphogenetic protein 9 (BMP9), a member of TGF-β superfamily, is reported to inhibit the growth and migration of prostate cancer, osteosarcoma and triple-negative MDA-MB-231 breast cancer cells. However, little is known about the effect of on the biological behaviors of HER2-positive SK-BR-3 breast cancer cells and the underlying mechanisms. This study aimed to investigate the effects of BMP9 on the proliferation and metastasis of SK-BR-3 cells with BMP9 over-expression or BMP9 down-regulated expression. Results indicated that exogenously expressed BMP9 inhibited the proliferation and metastasis of SK-BR-3 cells while decreased endogenous BMP9 expression in SK-BR-3 cells promoted the proliferation and migration of breast cancer cells in vitro and in vivo. In SK-BR-3 cells with BMP9 over-expression, the phosphorylation of HER2, ERK1/2 and AKT was markedly suppressed and the HER2 expression decreased at both mRNA and protein levels, while opposite results were observed in SK-BR-3 cells with BMP9 knock down. When the phosphorylation of ERK1/2 and PI3K/AKT was inhibited by PD98059 and LY294002, respectively, the decreased proliferation and invasion induced by BMP9 knock down were eliminated. These findings suggest that BMP9 can inhibit the proliferation and metastasis of SK-BR-3 cells via inactivating ERK1/2 and PI3K/AKT signaling pathways. Thus, BMP9 may serve as a useful agent in the treatment of HER-2 positive breast cancer. PMID:24805814

  19. BMP9 inhibits proliferation and metastasis of HER2-positive SK-BR-3 breast cancer cells through ERK1/2 and PI3K/AKT pathways.

    PubMed

    Ren, Wei; Liu, Yuehong; Wan, Shaoheng; Fei, Chang; Wang, Wei; Chen, Yingying; Zhang, Zhihui; Wang, Ting; Wang, Jinshu; Zhou, Lan; Weng, Yaguang; He, Tongchuan; Zhang, Yan

    2014-01-01

    Bone morphogenetic protein 9 (BMP9), a member of TGF-β superfamily, is reported to inhibit the growth and migration of prostate cancer, osteosarcoma and triple-negative MDA-MB-231 breast cancer cells. However, little is known about the effect of on the biological behaviors of HER2-positive SK-BR-3 breast cancer cells and the underlying mechanisms. This study aimed to investigate the effects of BMP9 on the proliferation and metastasis of SK-BR-3 cells with BMP9 over-expression or BMP9 down-regulated expression. Results indicated that exogenously expressed BMP9 inhibited the proliferation and metastasis of SK-BR-3 cells while decreased endogenous BMP9 expression in SK-BR-3 cells promoted the proliferation and migration of breast cancer cells in vitro and in vivo. In SK-BR-3 cells with BMP9 over-expression, the phosphorylation of HER2, ERK1/2 and AKT was markedly suppressed and the HER2 expression decreased at both mRNA and protein levels, while opposite results were observed in SK-BR-3 cells with BMP9 knock down. When the phosphorylation of ERK1/2 and PI3K/AKT was inhibited by PD98059 and LY294002, respectively, the decreased proliferation and invasion induced by BMP9 knock down were eliminated. These findings suggest that BMP9 can inhibit the proliferation and metastasis of SK-BR-3 cells via inactivating ERK1/2 and PI3K/AKT signaling pathways. Thus, BMP9 may serve as a useful agent in the treatment of HER-2 positive breast cancer.

  20. NS309 decreases rat detrusor smooth muscle membrane potential and phasic contractions by activating SK3 channels

    PubMed Central

    Parajuli, Shankar P; Hristov, Kiril L; Soder, Rupal P; Kellett, Whitney F; Petkov, Georgi V

    2013-01-01

    Background and Purpose Overactive bladder (OAB) is often associated with abnormally increased detrusor smooth muscle (DSM) contractions. We used NS309, a selective and potent opener of the small or intermediate conductance Ca2+-activated K+ (SK or IK, respectively) channels, to evaluate how SK/IK channel activation modulates DSM function. Experimental Approach We employed single-cell RT-PCR, immunocytochemistry, whole cell patch-clamp in freshly isolated rat DSM cells and isometric tension recordings of isolated DSM strips to explore how the pharmacological activation of SK/IK channels with NS309 modulates DSM function. Key Results We detected SK3 but not SK1, SK2 or IK channels expression at both mRNA and protein levels by RT-PCR and immunocytochemistry in DSM single cells. NS309 (10 μM) significantly increased the whole cell SK currents and hyperpolarized DSM cell resting membrane potential. The NS309 hyperpolarizing effect was blocked by apamin, a selective SK channel inhibitor. NS309 inhibited the spontaneous phasic contraction amplitude, force, frequency, duration and tone of isolated DSM strips in a concentration-dependent manner. The inhibitory effect of NS309 on spontaneous phasic contractions was blocked by apamin but not by TRAM-34, indicating no functional role of the IK channels in rat DSM. NS309 also significantly inhibited the pharmacologically and electrical field stimulation-induced DSM contractions. Conclusions and Implications Our data reveal that SK3 channel is the main SK/IK subtype in rat DSM. Pharmacological activation of SK3 channels with NS309 decreases rat DSM cell excitability and contractility, suggesting that SK3 channels might be potential therapeutic targets to control OAB associated with detrusor overactivity. PMID:23145946

  1. Coupling of SK channels, L-type Ca2+ channels, and ryanodine receptors in cardiomyocytes.

    PubMed

    Zhang, Xiao-Dong; Coulibaly, Zana A; Chen, Wei Chun; Ledford, Hannah A; Lee, Jeong Han; Sirish, Padmini; Dai, Gu; Jian, Zhong; Chuang, Frank; Brust-Mascher, Ingrid; Yamoah, Ebenezer N; Chen-Izu, Ye; Izu, Leighton T; Chiamvimonvat, Nipavan

    2018-03-16

    Small-conductance Ca 2+ -activated K + (SK) channels regulate the excitability of cardiomyocytes by integrating intracellular Ca 2+ and membrane potentials on a beat-to-beat basis. The inextricable interplay between activation of SK channels and Ca 2+ dynamics suggests the pathology of one begets another. Yet, the exact mechanistic underpinning for the activation of cardiac SK channels remains unaddressed. Here, we investigated the intracellular Ca 2+ microdomains necessary for SK channel activation. SK currents coupled with Ca 2+ influx via L-type Ca 2+ channels (LTCCs) continued to be elicited after application of caffeine, ryanodine or thapsigargin to deplete SR Ca 2+ store, suggesting that LTCCs provide the immediate Ca 2+ microdomain for the activation of SK channels in cardiomyocytes. Super-resolution imaging of SK2, Ca v 1.2 Ca 2+ channel, and ryanodine receptor 2 (RyR2) was performed to quantify the nearest neighbor distances (NND) and localized the three molecules within hundreds of nanometers. The distribution of NND between SK2 and RyR2 as well as SK2 and Ca v 1.2 was bimodal, suggesting a spatial relationship between the channels. The activation mechanism revealed by our study paved the way for the understanding of the roles of SK channels on the feedback mechanism to regulate the activities of LTCCs and RyR2 to influence local and global Ca 2+ signaling.

  2. MuSK IgG4 autoantibodies cause myasthenia gravis by inhibiting binding between MuSK and Lrp4

    PubMed Central

    Huijbers, Maartje G.; Zhang, Wei; Klooster, Rinse; Niks, Erik H.; Friese, Matthew B.; Straasheijm, Kirsten R.; Thijssen, Peter E.; Vrolijk, Hans; Plomp, Jaap J.; Vogels, Pauline; Losen, Mario; Van der Maarel, Silvère M.; Burden, Steven J.; Verschuuren, Jan J.

    2013-01-01

    Myasthenia gravis (MG) is a severely debilitating autoimmune disease that is due to a decrease in the efficiency of synaptic transmission at neuromuscular synapses. MG is caused by antibodies against postsynaptic proteins, including (i) acetylcholine receptors, the neurotransmitter receptor, (ii) muscle-specific kinase (MuSK), a receptor tyrosine kinase essential for the formation and maintenance of neuromuscular synapses, and (iii) low-density lipoprotein receptor-related protein 4 (Lrp4), which responds to neural Agrin by binding and stimulating MuSK. Passive transfer studies in mice have shown that IgG4 antibodies from MuSK MG patients cause disease without requiring complement or other immune components, suggesting that these MuSK antibodies cause disease by directly interfering with MuSK function. Here we show that pathogenic IgG4 antibodies to MuSK bind to a structural epitope in the first Ig-like domain of MuSK, prevent binding between MuSK and Lrp4, and inhibit Agrin-stimulated MuSK phosphorylation. In contrast, these IgG4 antibodies have no direct effect on MuSK dimerization or MuSK internalization. These results provide insight into the unique pathogenesis of MuSK MG and provide clues toward development of specific treatment options. PMID:24297891

  3. Two cytotoxic sesquiterpene lactones from the leaves of Xanthium strumarium and their in vitro inhibitory activity on farnesyltransferase.

    PubMed

    Kim, Young Sup; Kim, Jeoung Seob; Park, Sung-Hee; Choi, Sang-Un; Lee, Chong Ock; Kim, Seong-Kie; Kim, Young-Kyoon; Kim, Sung Hoon; Ryu, Shi Yong

    2003-04-01

    Two xanthanolide sesquiterpene lactones, 8- epi-xanthatin (1) and 8- epi-xanthatin epoxide (2), isolated from the leaves of Xanthium strumarium (Compositae), demonstrated a significant inhibition on the proliferation of cultured human tumor cells, i. e., A549 (non-small cell lung), SK-OV-3 (ovary), SK-MEL-2 (melanoma), XF498 (central nervous system) and HCT-15 (colon) in vitro. They were also found to inhibit the farnesylation process of human lamin-B by farnesyltransferase (FTase), in a dose-dependent manner in vitro (IC 50 value was calculated as 64 and 58 microM, respectively). Due to the relatively high concentrations of 1 and 2 required to obtain an FTase inhibition as compared with those necessary for a cytotoxic effect on tumor cells, it remains unclear whether a relationship between these two activities exists.

  4. Xylitol induces cell death in lung cancer A549 cells by autophagy.

    PubMed

    Park, Eunjoo; Park, Mi Hee; Na, Hee Sam; Chung, Jin

    2015-05-01

    Xylitol is a widely used anti-caries agent that has anti-inflammatory effects. We have evaluated the potential of xylitol in cancer treatment. It's effects on cell proliferation and cytotoxicity were measured by MTT assay and LDH assay. Cell morphology and autophagy were examined by immunostaining and immunoblotting. Xylitol inhibited cell proliferation in a dose-dependent manner in these cancer cells: A549, Caki, NCI-H23, HCT-15, HL-60, K562, and SK MEL-2. The IC50 of xylitol in human gingival fibroblast cells was higher than in cancer cells, indicating that it is more specific for cancer cells. Moreover, xylitol induced autophagy in A549 cells that was inhibited by 3-methyladenine, an autophagy inhibitor. These results indicate that xylitol has potential in therapy against lung cancer by inhibiting cell proliferation and inducing autophagy of A549 cells.

  5. [Roles of KLF5 in inhibition TNFα-induced SK-BR-3 breast cancer cell apoptosis].

    PubMed

    Shi, Jianhong; Liu, Caiyun; Zhang, Anyi; Cui, Naipeng; Wang, Bing; Chen, Baoping; Ma, Zhenfeng

    2014-07-08

    To explore the expression levels and roles of Krüpple-like factor 5 (KLF5) in tumor necrosis factor α (TNFα)-induced SK-BR-3 breast cancer cells. SK-BR-3 breast cancer cells were stimulated by TNFα at different concentrations (0, 1, 5, 10, 20 µg/L) for specified durations (0, 6, 12, 24, 36 h). Western blot was performed to detect KLF5 protein levels. Then Western blot and quantitative real-time PCR (qRT-PCR) were used to detect the expression levels of apoptosis genes. Flow cytometry and qRT-PCR were used to observe the effects of exogenous KLF5 on TNFα-induced apoptosis of SK-BR-3 breast cancer cell. KLF5 expression levels significantly decreased in TNFα-stimulated SK-BR-3 breast cancer cells in a concentration- and time-dependent manner. Quantitative RT-PCR results showed that TNFα up-regulate apoptosis gene caspase 3, caspase 9 and bax expression levels and down-regulate bcl-1 level in SK-BR-3 cells. Adenovirus expression vectors of pAd-GFP and pAd-GFP-KLF5 were constructed and used to infect SK-BR-3 breast cancer cells. Over-expression of GFP-KLF5 inhibited apoptosis in TNFα-stimulated SK-BR-3 breast cancer cells. TNFα reduces KLF5 expression in SK-BR-3 breast cancer cells and KLF5 participates in TNFα-induced SK-BR-3 cell apoptosis.

  6. Overexpression of YB1 C-terminal domain inhibits proliferation, angiogenesis and tumorigenicity in a SK-BR-3 breast cancer xenograft mouse model.

    PubMed

    Shi, Jian-Hong; Cui, Nai-Peng; Wang, Shuo; Zhao, Ming-Zhi; Wang, Bing; Wang, Ya-Nan; Chen, Bao-Ping

    2016-01-01

    Y-box-binding protein 1 (YB1) is a multifunctional transcription factor with vital roles in proliferation, differentiation and apoptosis. In this study, we have examined the role of its C-terminal domain (YB1 CTD) in proliferation, angiogenesis and tumorigenicity in breast cancer. Breast cancer cell line SK-BR-3 was infected with GFP-tagged YB1 CTD adenovirus expression vector. An 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS) proliferation assay showed that YB1 CTD decreased SK-BR-3 cell proliferation, and down-regulated cyclin B1 and up-regulated p21 levels in SK-BR-3 cells. YB1 CTD overexpression changed the cytoskeletal organization and slightly inhibited the migration of SK-BR-3 cells. YB1 CTD also inhibited secreted VEGF expression in SK-BR-3 cells, which decreased SK-BR-3-induced EA.hy926 endothelial cell angiogenesis in vitro. YB1 CTD overexpression attenuated the ability of SK-BR-3 cells to form tumours in nude mice, and decreased in vivo VEGF levels and angiogenesis in the xenografts in SK-BR-3 tumour-bearing mice. Taken together, our findings demonstrate the vital role of YB1 CTD overexpression in inhibiting proliferation, angiogenesis and tumorigenicity of breast cancer cell line SK-BR-3.

  7. The SigmaR1 chaperone drives breast and colorectal cancer cell migration by tuning SK3-dependent Ca2+ homeostasis.

    PubMed

    Gueguinou, M; Crottès, D; Chantôme, A; Rapetti-Mauss, R; Potier-Cartereau, M; Clarysse, L; Girault, A; Fourbon, Y; Jézéquel, P; Guérin-Charbonnel, C; Fromont, G; Martin, P; Pellissier, B; Schiappa, R; Chamorey, E; Mignen, O; Uguen, A; Borgese, F; Vandier, C; Soriani, O

    2017-06-22

    The remodeling of calcium homeostasis contributes to the cancer hallmarks and the molecular mechanisms involved in calcium channel regulation in tumors remain to be characterized. Here, we report that SigmaR1, a stress-activated chaperone, is required to increase calcium influx by triggering the coupling between SK3, a Ca 2+ -activated K + channel (KCNN3) and the voltage-independent calcium channel Orai1. We show that SigmaR1 physically binds SK3 in BC cells. Inhibition of SigmaR1 activity, either by molecular silencing or by the use of sigma ligand (igmesine), decreased SK3 current and Ca 2+ entry in breast cancer (BC) and colorectal cancer (CRC) cells. Interestingly, SigmaR1 inhibition diminished SK3 and/or Orai1 levels in lipid nanodomains isolated from BC cells. Analyses of tissue microarray from CRC patients showed higher SigmaR1 expression levels in cancer samples and a correlation with tumor grade. Moreover, the exploration of a cohort of 4937 BC patients indicated that high expression of SigmaR1 and Orai1 channels was significantly correlated to a lower overall survival. As the SK3/Orai1 tandem drives invasive process in CRC and bone metastasis progression in BC, our results may inaugurate innovative therapeutic approaches targeting SigmaR1 to control the remodeling of Ca 2+ homeostasis in epithelial cancers.

  8. Alternative splice isoforms of small conductance calcium-activated SK2 channels differ in molecular interactions and surface levels

    PubMed Central

    Scholl, Elizabeth Storer; Pirone, Antonella; Cox, Daniel H; Duncan, R Keith; Jacob, Michele H

    2014-01-01

    Small conductance Ca2+-sensitive potassium (SK2) channels are voltage-independent, Ca2+-activated ion channels that conduct potassium cations and thereby modulate the intrinsic excitability and synaptic transmission of neurons and sensory hair cells. In the cochlea, SK2 channels are functionally coupled to the highly Ca2+ permeant α9/10-nicotinic acetylcholine receptors (nAChRs) at olivocochlear postsynaptic sites. SK2 activation leads to outer hair cell hyperpolarization and frequency-selective suppression of afferent sound transmission. These inhibitory responses are essential for normal regulation of sound sensitivity, frequency selectivity, and suppression of background noise. However, little is known about the molecular interactions of these key functional channels. Here we show that SK2 channels co-precipitate with α9/10-nAChRs and with the actin-binding protein α-actinin-1. SK2 alternative splicing, resulting in a 3 amino acid insertion in the intracellular 3′ terminus, modulates these interactions. Further, relative abundance of the SK2 splice variants changes during developmental stages of synapse maturation in both the avian cochlea and the mammalian forebrain. Using heterologous cell expression to separately study the 2 distinct isoforms, we show that the variants differ in protein interactions and surface expression levels, and that Ca2+ and Ca2+-bound calmodulin differentially regulate their protein interactions. Our findings suggest that the SK2 isoforms may be distinctly modulated by activity-induced Ca2+ influx. Alternative splicing of SK2 may serve as a novel mechanism to differentially regulate the maturation and function of olivocochlear and neuronal synapses. PMID:24394769

  9. [Roles of Y box-binding protein 1 in SK-BR-3 breast cancer proliferation].

    PubMed

    Shi, Jianhong; Lü, Xinrui; Wang, Bing; Daudan, Lin; Yanan, Wang; Yuhui, Bu; Zhenfeng, Ma

    2014-09-30

    To explore the roles of Y box-binding protein 1 (YB-1) in breast cancer cell proliferation. Twenty cases of surgical removal of breast cancer tissue (diagnosed with invasive ductal carcinoma, stage II, by postoperative paraffin pathology) and normal breast tissues adjacent to carcinoma were collected during June 2013 to August 2013.Quantitative real-time PCR (qRT-PCR) was performed to detect the YB1 mRNA levels. Cultured mammary epithelial cells (HBL-100) and breast cancer cells (MCF7, MDA-MB-231 & SK-BR-3 cells) were harvested and qRT-PCR was performed to detect the YB1 mRNA levels.SK-BR-3 cells were stimulated with various concentrations of PDGF-BB and YB1 expression levels were detected by qRT-PCR. Down-regulation or over-expression of YB1 by si-YB1 or Ad-GFP-YB1 was detected in SK-BR-3 cells. And MTS cell proliferation assay kit was used to detect cell proliferation. YB1 mRNA levels were significantly higher in breast cancer tissues and MDA-MB-231 and SK-BR-3 breast cancer cell lines than that in adjacent normal breast tissues and HBL-100 mammary epithelial cells respectively (P < 0.05).YB1 expression levels increased in PDGF-BB stimulated SK-BR-3 cells in a dose-dependent manner. A down-regulation of endogenous YB1 decreases and an over-expression of exogenous YB1 promotes the proliferation activity in SK-BR-3 cells.

  10. Selective activation of heteromeric SK channels contributes to action potential repolarization in mouse atrial myocytes.

    PubMed

    Hancock, Jane M; Weatherall, Kate L; Choisy, Stéphanie C; James, Andrew F; Hancox, Jules C; Marrion, Neil V

    2015-05-01

    Activation of small conductance calcium-activated potassium (SK) channels is proposed to contribute to repolarization of the action potential in atrial myocytes. This role is controversial, as these cardiac SK channels appear to exhibit an uncharacteristic pharmacology. The objectives of this study were to resolve whether activation of SK channels contributes to atrial action potential repolarization and to determine the likely subunit composition of the channel. The effect of 2 SK channel inhibitors was assessed on outward current evoked in voltage clamp and on action potential duration in perforated patch and whole-cell current clamp recording from acutely isolated mouse atrial myocytes. The presence of SK channel subunits was assessed using immunocytochemistry. A significant component of outward current was reduced by the SK channel blockers apamin and UCL1684. Block by apamin displayed a sensitivity indicating that this current was carried by homomeric SK2 channels. Action potential duration was significantly prolonged by UCL1684, but not by apamin. This effect was accompanied by an increase in beat-to-beat variability and action potential triangulation. This pharmacology was matched by that of expressed heteromeric SK2-SK3 channels in HEK293 cells. Immunocytochemistry showed that atrial myocytes express both SK2 and SK3 channels with an overlapping expression pattern. Only proposed heteromeric SK2-SK3 channels are physiologically activated to contribute to action potential repolarization, which is indicated by the difference in pharmacology of evoked outward current and prolongation of atrial action potential duration. The effect of blocking this channel on the action potential suggests that SK channel inhibition during cardiac function has the potential to be proarrhythmic. Copyright © 2015 Heart Rhythm Society. Published by Elsevier Inc. All rights reserved.

  11. Altered expression and function of small-conductance (SK) Ca2+-activated K+ channels in pilocarpine-treated epileptic rats

    PubMed Central

    Oliveira, Mauro S.; Skinner, Frank; Arshadmansab, Massoud F.; Garcia, Ileana; Mello, Carlos F.; Knaus, Hans-Günther; Ermolinsky, Boris S.; Pacheco Otalora, Luis F.; Garrido-Sanabria, Emilio R.

    2010-01-01

    Small conductance calcium (Ca2+) activated SK channels are critical regulators of neuronal excitability in hippocampus. Accordingly, these channels are thought to play a key role in controlling neuronal activity in acute models of epilepsy. In this study, we investigate the expression and function of SK channels in the pilocarpine model of mesial temporal lobe epilepsy. For this purpose, protein expression was assessed using western blotting assays and gene expression was analyzed using TaqMan-based probes and the quantitative real-time polymerase chain reaction (qPCR) comparative method delta-delta cycle threshold (ΔΔCT) in samples extracted from control and epileptic rats. In addition, the effect of SK channel antagonist UCL1684 and agonist NS309 on CA1 evoked population spikes was studied in hippocampal slices. Western blotting analysis showed a significant reduction in the expression of SK1 and SK2 channels at 10 days following status epilepticus (SE), but levels recovered at 1 month and at more than 2 months after SE. In contrast, a significant down-regulation of SK3 channels was detected after 10 days of SE. Analysis of gene expression by qPCR revealed a significant reduction of transcripts for SK2 (Kcnn1) and SK3 (Kcnn3) channels as early as 10 days following pilocarpine-induced SE and during the chronic phase of the pilocarpine model. Moreover, bath application of UCL1684 (100 nM for 15 min) induced a significant increase of the population spike amplitude and number of spikes in the hippocampal CA1 area of slices obtained control and chronic epileptic rats. This effect was obliterated by co-administration of UCL1684 with SK channel agonist NS309 (1 μM). Application of NS309 failed to modify population spikes in the CA1 area of slices taken from control and epileptic rats. These data indicate an abnormal expression of SK channels and a possible dysfunction of these channels in experimental MTLE. PMID:20553876

  12. Muscle-specific kinase (MuSK) autoantibodies suppress the MuSK pathway and ACh receptor retention at the mouse neuromuscular junction

    PubMed Central

    Ghazanfari, Nazanin; Morsch, Marco; Reddel, Stephen W; Liang, Simon X; Phillips, William D

    2014-01-01

    Muscle-specific kinase (MuSK) autoantibodies from myasthenia gravis patients can block the activation of MuSK in vitro and/or reduce the postsynaptic localization of MuSK. Here we use a mouse model to examine the effects of MuSK autoantibodies upon some key components of the postsynaptic MuSK pathway and upon the regulation of junctional ACh receptor (AChR) numbers. Mice became weak after 14 daily injections of anti-MuSK-positive patient IgG. The intensity and area of AChR staining at the motor endplate was markedly reduced. Pulse-labelling of AChRs revealed an accelerated loss of pre-existing AChRs from postsynaptic AChR clusters without a compensatory increase in incorporation of (newly synthesized) replacement AChRs. Large, postsynaptic AChR clusters were replaced by a constellation of tiny AChR microaggregates. Puncta of AChR staining also appeared in the cytoplasm beneath the endplate. Endplate staining for MuSK, activated Src, rapsyn and AChR were all reduced in intensity. In the tibialis anterior muscle there was also evidence that phosphorylation of the AChR β-subunit-Y390 was reduced at endplates. In contrast, endplate staining for β-dystroglycan (through which rapsyn couples AChR to the synaptic basement membrane) remained intense. The results suggest that anti-MuSK IgG suppresses the endplate density of MuSK, thereby down-regulating MuSK signalling activity and the retention of junctional AChRs locally within the postsynaptic membrane scaffold. PMID:24860174

  13. Muscle-specific kinase (MuSK) autoantibodies suppress the MuSK pathway and ACh receptor retention at the mouse neuromuscular junction.

    PubMed

    Ghazanfari, Nazanin; Morsch, Marco; Reddel, Stephen W; Liang, Simon X; Phillips, William D

    2014-07-01

    Muscle-specific kinase (MuSK) autoantibodies from myasthenia gravis patients can block the activation of MuSK in vitro and/or reduce the postsynaptic localization of MuSK. Here we use a mouse model to examine the effects of MuSK autoantibodies upon some key components of the postsynaptic MuSK pathway and upon the regulation of junctional ACh receptor (AChR) numbers. Mice became weak after 14 daily injections of anti-MuSK-positive patient IgG. The intensity and area of AChR staining at the motor endplate was markedly reduced. Pulse-labelling of AChRs revealed an accelerated loss of pre-existing AChRs from postsynaptic AChR clusters without a compensatory increase in incorporation of (newly synthesized) replacement AChRs. Large, postsynaptic AChR clusters were replaced by a constellation of tiny AChR microaggregates. Puncta of AChR staining also appeared in the cytoplasm beneath the endplate. Endplate staining for MuSK, activated Src, rapsyn and AChR were all reduced in intensity. In the tibialis anterior muscle there was also evidence that phosphorylation of the AChR β-subunit-Y390 was reduced at endplates. In contrast, endplate staining for β-dystroglycan (through which rapsyn couples AChR to the synaptic basement membrane) remained intense. The results suggest that anti-MuSK IgG suppresses the endplate density of MuSK, thereby down-regulating MuSK signalling activity and the retention of junctional AChRs locally within the postsynaptic membrane scaffold. © 2014 The Authors. The Journal of Physiology © 2014 The Physiological Society.

  14. Crucial importance of the endothelial K+ channel SK3 and connexin40 in arteriolar dilations during skeletal muscle contraction.

    PubMed

    Milkau, Malte; Köhler, Ralf; de Wit, Cor

    2010-09-01

    Skeletal muscle activity requires substantial increases in blood flow, and the underlying vasodilation involves endothelial activity, but the contribution of the endothelium-dependent hyperpolarizing factor (EDHF) is only poorly defined. In EDHF signaling, endothelial hyperpolarization mediated by the Ca(2+)-activated K(+) channels SK3 and IK1 is a key step and also initiates gap junction-dependent conducted dilations. We assessed the role of SK3, IK1, and connexin40 (Cx40) in muscular contraction-induced dilations in the microcirculation in vivo. Hitherto, arterioles were observed in the electrically stimulated cremaster skeletal muscle of anesthetized mice lacking SK3, IK1, or Cx40 using intravital microscopy. Genetic deficiency of SK3, but not of IK1, strongly attenuated dilations to muscular contraction. Similarly, pharmacologic blockade of SK3 by the specific blocker UCL1684 impaired such dilations in wild-type and IK1-deficient mice. In contrast, IK1 was required for acetylcholine-induced dilations. Genetic deficiency of Cx40 also attenuated dilations induced by muscular contraction but not by acetylcholine. These data support the concept that endothelial hyperpolarization through activation of SK3 contributes to exercise hyperemia and the hyperpolarization ascends the vascular tree through gap junctions formed by Cx40 to orchestrate dilation. The differential impact of SK3- and IK1-deficiency on dilations to distinct stimuli suggests stimulus-dependent activation of these endothelial channels.

  15. Computational simulations assessment of mutations impact on streptokinase (SK) from a group G streptococci with enhanced activity - insights into the functional roles of structural dynamics flexibility of SK and stabilization of SK-μplasmin catalytic complex.

    PubMed

    Kazemi, Faegheh; Arab, Seyed Shahriar; Mohajel, Nasir; Keramati, Malihe; Niknam, Niloofar; Aslani, Mohammad Mehdi; Roohvand, Farzin

    2018-05-28

    Streptokinase (SK), a plasminogen activator (PA) that converts inactive plasminogen (Pg) to plasmin (Pm), is a protein secreted by groups A, C, and G streptococci (GAS, GCS, and GGS, respectively), with high sequence divergence and functional heterogeneity. While roles of some residual changes in altered SK functionality are shown, the underlying structural mechanisms are less known. Herein, using computational approaches, we analyzed the conformational basis for the increased activity of SK from a GGS (SKG132) isolate with four natural residual substitutions (Ile33Phe, Arg45Gln, Asn228Lys, Phe287Ile) compared to the standard GCS (SKC). Using the crystal structure of SK.Pm catalytic complex as main template SKC.μPm catalytic complex was modeled through homology modeling process and validated by several online validation servers. Subsequently, SKG132.μPm structure was constructed by altering the corresponding residual substitutions. Results of three independent MD simulations showed increased RMSF values for SKG132.μPm, indicating the enhanced structural flexibility compared to SKC.μPm, specially in 170 and 250 loops and three regions: R1 (149-161), R2 (182-215) and R3 (224-229). In parallel, the average number of Hydrogen bonds in 170 loop, R2 and R3 (especially for Asn228Lys) of SKG132 compared to that of the SKC was decreased. Accordingly, residue interaction networks (RINs) analyses indicated that Asn228Lys might induce more level of structural flexibility by generation of free Lys256, while Phe287Ile and Ile33Phe enhanced the stabilization of the SKG132.μPm catalytic complex. These results denoted the potential role of the optimal dynamic state and stabilized catalytic complex for increased PA potencies of SK as a thrombolytic drug.

  16. [Effect of 3-bromopyruvate on mitochondrial membrane potential and apoptosis of human breast carcinoma SK-BR-3 cells].

    PubMed

    Zhang, Yuanyuan; Liu, Zhe; Zhang, Qianwen; Chao, Zhenhua; Zhang, Pei; Xia, Fei; Jiang, Chenchen; Liu, Hao; Jiang, Zhiwen

    2013-09-01

    To study the effect of glycolysis inhibitor 3-bromopyruvate (3-BrPA) in inducing apoptosis of human breast carcinoma cells SK-BR-3 and the possible mechanism. MTT assay was used to detect the growth inhibition induced by 3-BrPA in breast cancer cells SK-BR-3. The apoptotic cells were detected by flow cytometry with propidium iodide (PI). ATP levels in the cells were detected by ATP assay kit, and DHE fluorescent probe technique was used to determine superoxide anion levels; the mitochondrial membrane potential was assessed using JC-1 staining assay. MTT assay showed that the proliferation of SK-BR-3 cells was inhibited by 3-BrPA in a time- and concentration-dependent manner. Exposure to 80, 160, and 320 µmol·L(-1) 3-BrPA for 24 h resulted in cell apoptosis rates of 6.7%, 22.3%, and 79.6%, respectively, and the intracellular ATP levels of SK-BR-3 cells treated with 80, 160, 320 µmol·L(-1) 3-BrPA for 5 h were 87.7%, 60.6%, and 23.7% of the control levels. 3-BrPA at 160 µmol·L(-1) increased reactive oxygen levels and lowered mitochondrial membrane potential of SK-BR-3 cells. 3-BrPA can inhibit cell proliferation, reduce the mitochondrial membrane potential and induce apoptosis in SK-BR-3 cells, the mechanism of which may involve a reduced ATP level by inhibiting glycolysis and increasing the reactive oxygen level in the cells.

  17. 7SK-BAF axis controls pervasive transcription at enhancers

    PubMed Central

    Flynn, Ryan A.; Do, Brian T.; Rubin, Adam J.; Calo, Eliezer; Lee, Byron; Kuchelmeister, Hannes; Rale, Michael; Chu, Ci; Kool, Eric T.; Wysocka, Joanna; Khavari, Paul A.

    2016-01-01

    RNA functions at enhancers remain mysterious. Here we show that the 7SK small nuclear RNA (snRNA) inhibits enhancer transcription by modulating nucleosome position. 7SK occupies enhancers and super enhancers genome-wide in mouse and human cells, and 7SK is required to limit eRNA initiation and synthesis in a manner distinct from promoter pausing. Clustered elements at super enhancers uniquely require 7SK to prevent convergent transcription and DNA damage signaling. 7SK physically interacts with the BAF chromatin remodeling complex, recruit BAF to enhancers, and inhibits enhancer transcription by modulating chromatin structure. In turn, 7SK occupancy at enhancers coincides with Brd4 and is exquisitely sensitive to the bromodomain inhibitor JQ1. Thus, 7SK employs distinct mechanisms to counteract diverse consequences of pervasive transcription that distinguish super enhancers, enhancers, and promoters. PMID:26878240

  18. METCAM/MUC18 promoted tumorigenesis of human breast cancer SK-BR-3 cells in a dosage-specific manner.

    PubMed

    Huang, Chang-Yu; Wu, Guang-Jer

    2016-04-01

    Overexpression of METCAM/MUC18, an immunoglobulin-like cell-adhesion molecule, promotes tumorigenesis and progression of human breast cancer cells. We also observed an intriguing phenomenon that a high-expressing SK-BR-3 clone manifested a transient tumor suppression effect in vivo. The purpose of this study was to understand if this was caused by clonal variation, METCAM/MUC18-dosage effect, or the number of cells injected. Several G418-resistant clones of SK-BR-3, expressing different levels of METCAM/MUC18, were obtained for testing effects of human METCAM/MUC18 on in vitro motility, invasiveness, and anchorage-independent colony formation (in vitro tumorigenicity) and in vivo tumorigenesis in female Balb/C athymic nude mice. Tumor sections were made for histology and immunohistochemistry analyses, and tumor lysates for Western blot analysis to determine the effects of human METCAM/MUC18 expression on levels of various downstream effectors. METCAM/MUC18 promoted in vitro motility, invasiveness, and in vitro tumorigenicity of SK-BR-3 cells in a dosage-specific manner. Overexpression of METCAM/MUC18 could promote in vivo tumorigenesis of SK-BR-3 cells even when one tenth of the previously used cell number (5 × 10(5)) was injected and in vivo tumorigenesis of SK-BR-3 cells was directly proportional to the dosage of the protein. The previously observed transient tumor suppression effect from the same clone was no longer observed. The downstream effector, such as phospho-AKT/AKT ratio, was elevated in the tumors. Transient suppression observed previously in the clone was caused by injection of a high cell number (2 × 10(6)-5 × 10(6)). METCAM/MUC18 positively promotes tumorigenesis of SK-BR-3 cells by increasing the survival and proliferation pathway. Copyright © 2016. Published by Elsevier B.V.

  19. The hSK4 (KCNN4) isoform is the Ca2+-activated K+ channel (Gardos channel) in human red blood cells.

    PubMed

    Hoffman, Joseph F; Joiner, William; Nehrke, Keith; Potapova, Olga; Foye, Kristen; Wickrema, Amittha

    2003-06-10

    The question is, does the isoform hSK4, also designated KCNN4, represent the small conductance, Ca2+-activated K+ channel (Gardos channel) in human red blood cells? We have analyzed human reticulocyte RNA by RT-PCR, and, of the four isoforms of SK channels known, only SK4 was found. Northern blot analysis of purified and synchronously growing human erythroid progenitor cells, differentiating from erythroblasts to reticulocytes, again showed only the presence of SK4. Western blot analysis, with an anti-SK4 antibody, showed that human erythroid progenitor cells and, importantly, mature human red blood cell ghost membranes, both expressed the SK4 protein. The Gardos channel is known to turn on, given inside Ca2+, in the presence but not the absence of external Ko+ and remains refractory to Ko+ added after exposure to inside Ca2+. Heterologously expressed SK4, but not SK3, also shows this behavior. In inside-out patches of red cell membranes, the open probability (Po) of the Gardos channel is markedly reduced when the temperature is raised from 27 to 37 degrees C. Net K+ efflux of intact red cells is also reduced by increasing temperature, as are the Po values of inside-out patches of Chinese hamster ovary cells expressing SK4 (but not SK3). Thus the envelope of evidence indicates that SK4 is the gene that codes for the Gardos channel in human red blood cells. This channel is important pathophysiologically, because it represents the major pathway for cell shrinkage via KCl and water loss that occurs in sickle cell disease.

  20. Crystal Structure of the Agrin-Responsive Immunoglobulin-like Domains 1 and 2 of the Receptor Tyrosine Kinase MuSK

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Stiegler,A.; Burden, S.; Hubbard, S.

    Muscle-specific kinase (MuSK) is a receptor tyrosine kinase expressed exclusively in skeletal muscle, where it is required for formation of the neuromuscular junction. MuSK is activated by agrin, a neuron-derived heparan sulfate proteoglycan. Here, we report the crystal structure of the agrin-responsive first and second immunoglobulin-like domains (Ig1 and Ig2) of the MuSK ectodomain at 2.2 {angstrom} resolution. The structure reveals that MuSK Ig1 and Ig2 are Ig-like domains of the I-set subfamily, which are configured in a linear, semi-rigid arrangement. In addition to the canonical internal disulfide bridge, Ig1 contains a second, solvent-exposed disulfide bridge, which our biochemical datamore » indicate is critical for proper folding of Ig1 and processing of MuSK. Two Ig1-2 molecules form a non-crystallographic dimer that is mediated by a unique hydrophobic patch on the surface of Ig1. Biochemical analyses of MuSK mutants introduced into MuSK{sup -/-} myotubes demonstrate that residues in this hydrophobic patch are critical for agrin-induced MuSK activation.« less

  1. Antioxidant mechanism of mitochondria-targeted plastoquinone SkQ1 is suppressed in aglycemic HepG2 cells dependent on oxidative phosphorylation.

    PubMed

    Ježek, Jan; Engstová, Hana; Ježek, Petr

    2017-09-01

    Previously suggested antioxidant mechanisms for mitochondria-targeted plastoquinone SkQ1 included: i) ion-pairing of cationic SkQ1 + with free fatty acid anions resulting in uncoupling; ii) SkQ1H 2 ability to interact with lipoperoxyl radical; iii) interference with electron flow at the inner ubiquinone (Q) binding site of Complex III (Q i ), involving the reduction of SkQ1 to SkQ1H 2 by ubiquinol. We elucidated SkQ1 antioxidant properties by confocal fluorescence semi-quantification of mitochondrial superoxide (J m ) and cytosolic H 2 O 2 (J c ) release rates in HepG2 cells. Only in glycolytic cells, SkQ1 prevented the rotenone-induced enhancement of J m and J c but not basal releases without rotenone. The effect ceased in glutaminolytic aglycemic cells, in which the redox parameter NAD(P)H/FAD increased after rotenone in contrast to its decrease in glycolytic cells. Autofluorescence decay indicated decreased NADPH/NADH ratios with rotenone in both metabolic modes. SkQ1 did not increase cell respiration and diminished J m established high by antimycin or myxothiazol but not by stigmatellin. The revealed SkQ1 antioxidant modes reflect its reduction to SkQ1H 2 at Complex I I Q or Complex III Q i site. Both reductions diminish electron diversions to oxygen thus attenuating superoxide formation. Resulting SkQ1H 2 oxidizes back to SkQ1at the second (flavin) Complex I site, previously indicated for MitoQ 10 . Regeneration proceeds only at lower NAD(P)H/FAD in glycolytic cells. In contrast, cyclic SkQ1 reduction/SkQ1H 2 oxidation does not substantiate antioxidant activity in intact cells in the absence of oxidative stress (neither pro-oxidant activity, representing a great advantage). A targeted delivery to oxidative-stressed tissues is suggested for the effective antioxidant therapy based on SkQ1. Copyright © 2017 Elsevier B.V. All rights reserved.

  2. The hSK4 (KCNN4) isoform is the Ca2+-activated K+ channel (Gardos channel) in human red blood cells

    PubMed Central

    Hoffman, Joseph F.; Joiner, William; Nehrke, Keith; Potapova, Olga; Foye, Kristen; Wickrema, Amittha

    2003-01-01

    The question is, does the isoform hSK4, also designated KCNN4, represent the small conductance, Ca2+-activated K+ channel (Gardos channel) in human red blood cells? We have analyzed human reticulocyte RNA by RT-PCR, and, of the four isoforms of SK channels known, only SK4 was found. Northern blot analysis of purified and synchronously growing human erythroid progenitor cells, differentiating from erythroblasts to reticulocytes, again showed only the presence of SK4. Western blot analysis, with an anti-SK4 antibody, showed that human erythroid progenitor cells and, importantly, mature human red blood cell ghost membranes, both expressed the SK4 protein. The Gardos channel is known to turn on, given inside Ca2+, in the presence but not the absence of external \\documentclass[12pt]{minimal} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{mathrsfs} \\setlength{\\oddsidemargin}{-69pt} \\begin{document} \\begin{equation*}{\\mathrm{K}}_{{\\mathrm{o}}}^{+}\\end{equation*}\\end{document} and remains refractory to \\documentclass[12pt]{minimal} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{mathrsfs} \\setlength{\\oddsidemargin}{-69pt} \\begin{document} \\begin{equation*}{\\mathrm{K}}_{{\\mathrm{o}}}^{+}\\end{equation*}\\end{document} added after exposure to inside Ca2+. Heterologously expressed SK4, but not SK3, also shows this behavior. In inside–out patches of red cell membranes, the open probability (Po) of the Gardos channel is markedly reduced when the temperature is raised from 27 to 37°C. Net K+ efflux of intact red cells is also reduced by increasing temperature, as are the Po values of inside–out patches of Chinese hamster ovary cells expressing SK4 (but not SK3). Thus the envelope of evidence indicates that SK4 is the gene that codes for the Gardos channel in human red blood cells. This channel is important

  3. 7SK snRNP/P-TEFb couples transcription elongation with alternative splicing and is essential for vertebrate development

    PubMed Central

    Barboric, Matjaz; Lenasi, Tina; Chen, Hui; Johansen, Eric B.; Guo, Su; Peterlin, B. Matija

    2009-01-01

    Eukaryotic gene expression is commonly controlled at the level of RNA polymerase II (RNAPII) pausing subsequent to transcription initiation. Transcription elongation is stimulated by the positive transcription elongation factor b (P-TEFb) kinase, which is suppressed within the 7SK small nuclear ribonucleoprotein (7SK snRNP). However, the biogenesis and functional significance of 7SK snRNP remain poorly understood. Here, we report that LARP7, BCDIN3, and the noncoding 7SK small nuclear RNA (7SK) are vital for the formation and stability of a cell stress-resistant core 7SK snRNP. Our functional studies demonstrate that 7SK snRNP is not only critical for controlling transcription elongation, but also for regulating alternative splicing of pre-mRNAs. Using a transient expression splicing assay, we find that 7SK snRNP disintegration promotes inclusion of an alternative exon via the increased occupancy of P-TEFb, Ser2-phosphorylated (Ser2-P) RNAPII, and the splicing factor SF2/ASF at the minigene. Importantly, knockdown of larp7 or bcdin3 orthologues in zebrafish embryos destabilizes 7SK and causes severe developmental defects and aberrant splicing of analyzed transcripts. These findings reveal a key role for P-TEFb in coupling transcription elongation with alternative splicing, and suggest that maintaining core 7SK snRNP is essential for vertebrate development. PMID:19416841

  4. GSK-3 mediates the okadaic acid-induced modification of collapsin response mediator protein-2 in human SK-N-SH neuroblastoma cells.

    PubMed

    Ni, Mei-Hui; Wu, Chih-Ching; Chan, Wen-Hsiung; Chien, Kun-Yi; Yu, Jau-Song

    2008-04-15

    Collapsin response mediator protein-2 (CRMP-2), a phosphoprotein involved in axonal outgrowth and microtubule dynamics, is aberrantly phosphorylated in Alzheimer's disease (AD) brain. Alteration of glycogen synthase kinase-3 (GSK-3) activity is associated with the pathogenesis of AD. Here, we show that CRMP-2 is one of the major substrates for GSK-3 in pig brain extracts. Both GSK-3alpha and 3beta phosphorylate purified pig brain CRMP-2 and significantly alter its mobility in SDS-gels, resembling the CRMP-2 modification observed in AD brain. Interestingly, this modification can be detected in SK-N-SH neuroblastoma cells treated with a phosphatase inhibitor, okadaic acid (OA), and GSK-3 inhibitors completely block this OA-induced event. Knockdown of both GSK-3alpha and 3beta, but not either kinase alone, impairs OA-induced modification of CRMP-2. Mutation of Ser-518 or Ser-522 of CRMP-2, which are highly phosphorylated in AD brain, to Ala blocks the OA-induced modification of CRMP-2 in SK-N-SH cells. Ser-522 prephosphorylated by Cdk5 is required for subsequent GSK-3alpha-mediated phosphorylation of CRMP-2 in vitro. Collectively, our results demonstrate for the first time that OA can induce phosphorylation of CRMP-2 in SK-N-SH cells at sites aberrantly phosphorylated in AD brain, and both GSK-3alpha and 3beta and Ser-522 kinase(s) are involved in this process.

  5. Distribution, expression and functional effects of small conductance Ca-activated potassium (SK) channels in rat myometrium.

    PubMed

    Noble, Karen; Floyd, Rachel; Shmygol, Andre; Shmygol, Anatoly; Mobasheri, A; Wray, Susan

    2010-01-01

    Calcium-activated potassium channels are important in a variety of smooth muscles, contributing to excitability and contractility. In the myometrium previous work has focussed on the large conductance channels (BK), and the role of small conductance channels (SK) has received scant attention, despite the finding that over-expression of an SK channel isoform (SK3) results in uterine dysfunction and delayed parturition. This study therefore characterises the expression of the three SK channel isoforms (SK1-3) in rat myometrium throughout pregnancy and investigates their effect on cytosolic [Ca] and force and compares this with that of BK channels. Consistent expression of all SK isoform transcripts and clear immunostaining of SK1-3 was found. Inhibition of SK1-3 channels (apamin, scyllatoxin) significantly inhibited outward current, caused membrane depolarisation and elicited action potentials in previously quiescent cells. Apamin or scyllatoxin increased the amplitude of [Ca] and force in spontaneously contracting myometrial strips throughout gestation. The functional effect of SK inhibition was larger than that of BK channel inhibition. Thus we show for the first time that SK1-3 channels are expressed and translated throughout pregnancy and contribute to outward current, regulate membrane potential and hence Ca signals in pregnant rat myometrium. They contribute more to quiescence that BK channels. 2009 Elsevier Ltd. All rights reserved.

  6. hLARP7 C-terminal domain contains an xRRM that binds the 3' hairpin of 7SK RNA

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Eichhorn, Catherine D.; Chug, Rahul; Feigon, Juli

    The 7SK small nuclear ribonucleoprotein (snRNP) sequesters and inactivates the positive transcription elongation factor b (P-TEFb), an essential eukaryotic mRNA transcription factor. The human La-related protein group 7 (hLARP7) is a constitutive component of the 7SK snRNP and localizes to the 3' terminus of the 7SK long noncoding RNA. hLARP7, and in particular its C-terminal domain (CTD), is essential for 7SK RNA stability and assembly with P-TEFb. The hLARP7 N-terminal Lamodule binds and protects the 3' end from degradation, but the structural and functional role of its CTD is unclear.We report the solution NMR structure of the hLARP7 CTD andmore » show that this domain contains an xRRM, a class of atypical RRM first identified in the Tetrahymena thermophila telomerase LARP7 protein p65. The xRRM binds the 3' end of 7SK RNA at the top of stem-loop 4 (SL4) and interacts with both unpaired and base-paired nucleotides. This study thus confirms that the xRRM is general to the LARP7 family of proteins and defines the binding site for hLARP7 on the 7SK RNA, providing insight into function.« less

  7. hLARP7 C-terminal domain contains an xRRM that binds the 3' hairpin of 7SK RNA

    DOE PAGES

    Eichhorn, Catherine D.; Chug, Rahul; Feigon, Juli

    2016-09-26

    The 7SK small nuclear ribonucleoprotein (snRNP) sequesters and inactivates the positive transcription elongation factor b (P-TEFb), an essential eukaryotic mRNA transcription factor. The human La-related protein group 7 (hLARP7) is a constitutive component of the 7SK snRNP and localizes to the 3' terminus of the 7SK long noncoding RNA. hLARP7, and in particular its C-terminal domain (CTD), is essential for 7SK RNA stability and assembly with P-TEFb. The hLARP7 N-terminal Lamodule binds and protects the 3' end from degradation, but the structural and functional role of its CTD is unclear.We report the solution NMR structure of the hLARP7 CTD andmore » show that this domain contains an xRRM, a class of atypical RRM first identified in the Tetrahymena thermophila telomerase LARP7 protein p65. The xRRM binds the 3' end of 7SK RNA at the top of stem-loop 4 (SL4) and interacts with both unpaired and base-paired nucleotides. This study thus confirms that the xRRM is general to the LARP7 family of proteins and defines the binding site for hLARP7 on the 7SK RNA, providing insight into function.« less

  8. The SK-N-AS human neuroblastoma cell line develops osteolytic bone metastases with increased angiogenesis and COX-2 expression

    PubMed Central

    Tsutsumimoto, Takahiro; Williams, Paul; Yoneda, Toshiyuki

    2014-01-01

    Neuroblastoma (NB), which arises from embryonic neural crest cells, is the most common extra-cranial solid tumor of childhood. Approximately half of NB patients manifest bone metastasis accompanied with bone pain, fractures and bone marrow failure, leading to disturbed quality of life and poor survival. To study the mechanism of bone metastasis of NB, we established an animal model in which intracardiac inoculation of the SK-N-AS human NB cells in nude mice developed osteolytic bone metastases with increased osteoclastogenesis. SK-N-AS cells induced the expression of receptor activator of NF-κB ligand and osteoclastogenesis in mouse bone marrow cells in the co-culture. SK-N-AS cells expressed COX-2 mRNA and produced substantial amounts of prostaglandin E2 (PGE2). In contrast, the SK-N-DZ and SK-N-FI human NB cells failed to develop bone metastases, induce osteoclastogenesis, express COX-2 mRNA and produce PGE2. Immunohistochemical examination of SK-N-AS bone metastasis and subcutaneous tumor showed strong expression of COX-2. The selective COX-2 inhibitor NS-398 inhibited PGE2 production and suppressed bone metastases with reduced osteoclastogenesis. NS-398 also inhibited subcutaneous SK-N-AS tumor development with decreased angiogenesis and vascular endothelial growth factor-A expression. Of interest, metastasis to the adrenal gland, a preferential site for NB development, was also diminished by NS-398. Our results suggest that COX2/PGE2 axis plays a critical role in the pathophysiology of osteolytic bone metastases and tumor development of the SK-NS-AS human NB. Inhibition of angiogenesis by suppressing COX-2/PGE2 may be an effective therapeutic approach for children with NB. PMID:26909300

  9. TaSK5, an abiotic stress-inducible GSK3/shaggy-like kinase from wheat, confers salt and drought tolerance in transgenic Arabidopsis.

    PubMed

    Christov, Nikolai Kirilov; Christova, Petya Koeva; Kato, Hideki; Liu, Yuelin; Sasaki, Kentaro; Imai, Ryozo

    2014-11-01

    A novel cold-inducible GSK3/shaggy-like kinase, TaSK5, was isolated from winter wheat using a macroarray-based differential screening approach. TaSK5 showed high similarity to Arabidopsis subgroup I GSK3/shaggy-like kinases ASK-alpha, AtSK-gamma and ASK-epsilon. RNA gel blot analyses revealed TaSK5 induction by cold and NaCl treatments and to a lesser extent by drought treatment. TaSK5 functionally complemented the cold- and salt-sensitive phenotypes of a yeast GSK3/shaggy-like kinase mutant, △mck1. Transgenic Arabidopsis plants overexpressing TaSK5 cDNA showed enhanced tolerance to salt and drought stresses. By contrast, the tolerance of the transgenic plants to freezing stress was not altered. Microarray analysis revealed that a number of abiotic stress-inducible genes were constitutively induced in the transgenic Arabidopsis plants, suggesting that TaSK5 may function in a novel signal transduction pathway that appears to be unrelated to DREB1/CBF regulon and may involve crosstalk between abiotic and hormonal signals. Copyright © 2014 Elsevier Masson SAS. All rights reserved.

  10. RuvBL2 Is Involved in Histone Deacetylase Inhibitor PCI-24781-Induced Cell Death in SK-N-DZ Neuroblastoma Cells

    PubMed Central

    Zhan, Qinglei; Tsai, Sauna; Lu, Yonghai; Wang, Chunmei; Kwan, Yiuwa; Ngai, Saiming

    2013-01-01

    Neuroblastoma is the second most common solid tumor diagnosed during infancy. The survival rate among children with high-risk neuroblastoma is less than 40%, highlighting the urgent needs for new treatment strategies. PCI-24781 is a novel hydroxamic acid-based histone deacetylase (HDAC) inhibitor that has high efficacy and safety for cancer treatment. However, the underlying mechanisms of PCI-24781 are not clearly elucidated in neuroblastoma cells. In the present study, we demonstrated that PCI-24781 treatment significantly inhibited tumor growth at very low doses in neuroblastoma cells SK-N-DZ, not in normal cell line HS-68. However, PCI-24781 caused the accumulation of acetylated histone H3 both in SK-N-DZ and HS-68 cell line. Treatment of SK-N-DZ with PCI-24781 also induced cell cycle arrest in G2/M phase and activated apoptosis signaling pathways via the up-regulation of DR4, p21, p53 and caspase 3. Further proteomic analysis revealed differential protein expression profiles between non-treated and PCI-24781 treated SK-N-DZ cells. Totally 42 differentially expressed proteins were identified by MALDI-TOF MS system. Western blotting confirmed the expression level of five candidate proteins including prohibitin, hHR23a, RuvBL2, TRAP1 and PDCD6IP. Selective knockdown of RuvBL2 rescued cells from PCI-24781-induced cell death, implying that RuvBL2 might play an important role in anti-tumor activity of PCI-24781 in SK-N-DZ cells. The present results provide a new insight into the potential mechanism of PCI-24781 in SK-N-DZ cell line. PMID:23977108

  11. Structural basis for recognition of human 7SK long noncoding RNA by the La-related protein Larp7.

    PubMed

    Eichhorn, Catherine D; Yang, Yuan; Repeta, Lucas; Feigon, Juli

    2018-06-26

    The La and the La-related protein (LARP) superfamily is a diverse class of RNA binding proteins involved in RNA processing, folding, and function. Larp7 binds to the abundant long noncoding 7SK RNA and is required for 7SK ribonucleoprotein (RNP) assembly and function. The 7SK RNP sequesters a pool of the positive transcription elongation factor b (P-TEFb) in an inactive state; on release, P-TEFb phosphorylates RNA Polymerase II to stimulate transcription elongation. Despite its essential role in transcription, limited structural information is available for the 7SK RNP, particularly for protein-RNA interactions. Larp7 contains an N-terminal La module that binds UUU-3'OH and a C-terminal atypical RNA recognition motif (xRRM) required for specific binding to 7SK and P-TEFb assembly. Deletion of the xRRM is linked to gastric cancer in humans. We report the 2.2-Å X-ray crystal structure of the human La-related protein group 7 (hLarp7) xRRM bound to the 7SK stem-loop 4, revealing a unique binding interface. Contributions of observed interactions to binding affinity were investigated by mutagenesis and isothermal titration calorimetry. NMR 13 C spin relaxation data and comparison of free xRRM, RNA, and xRRM-RNA structures show that the xRRM is preordered to bind a flexible loop 4. Combining structures of the hLarp7 La module and the xRRM-7SK complex presented here, we propose a structural model for Larp7 binding to the 7SK 3' end and mechanism for 7SK RNP assembly. This work provides insight into how this domain contributes to 7SK recognition and assembly of the core 7SK RNP.

  12. Application program of CRUST-1 10km continental scientific drilling rig in SK-2 scientific drilling well

    NASA Astrophysics Data System (ADS)

    Sun, Youhong; Gao, Ke; Yu, Ping; Liu, Baochang; Guo, Wei; Ma, Yinlong; Yang, Yang

    2014-05-01

    SK-2 Well is located in DaQing city,where is site of the largest oil field in China,Heilongjiang province, north-east of China.The objective of SK-2 well is to obtain full cores of cretaceous formation in Song Liao basin,and to build the time tunnel of Cretaceous greenhouse climate change,and to clarify the causes,processes and results of the formations of DaQing oil field. This will ensure to achieve our ultimate goals,to test the CRUST-1 drilling rig and improve China's deep scientific drilling technology,to form the scientific drilling technology,method and system with independent intellectual property rights,and to provide technical knowledge and information for China's ten kilometers super-deep scientific drilling technical resources.SK-2 Well is at 6400 meter depth, where the drilling inclination is 90 degree and the continuous coring length is 3535 meter that from 2865 to 6400 meter,the recovery rate of the core is greater or equal to 95 percent with 100 millimeters core diameter and 3.9 degree per 100 meter geothermal gradient.The CRUST-1 rig is designated with special drilling equipment for continental scientific drilling combined to the oil drilling equipment ability with advanced geological drilling technology which is highly automatic and intelligent. CRUST-1 drilling ability is 10000 meter with the maximum hook load 700 tons, the total power is 4610 Kilowatt.CRUST-1 will be integrated with a complete set of automation equipment,including big torque hydraulic top drive,high accuracy automatic drilling rod feeding system, suspended automatic drill string discharge device,hydraulic intelligent iron roughneck,and hydraulic automatic catwalk to fully meet the drilling process requirements of SK-2.Designed with advanced drilling technique for 260 degree in the bottom of SK-2 well and hard rock,including the drilling tools of high temperature hydraulic hammer,high temperature resistance and high strength aluminum drill pipe,high temperature preparation of mud

  13. A dehydrin-dehydrin interaction: the case of SK3 from Opuntia streptacantha

    PubMed Central

    Hernández-Sánchez, Itzell E.; Martynowicz, David M.; Rodríguez-Hernández, Aida A.; Pérez-Morales, Maria B.; Graether, Steffen P.; Jiménez-Bremont, Juan F.

    2014-01-01

    Dehydrins belongs to a large group of highly hydrophilic proteins known as Late Embryogenesis Abundant (LEA) proteins. It is well known that dehydrins are intrinsically disordered plant proteins that accumulate during the late stages of embryogenesis and in response to abiotic stresses; however, the molecular mechanisms by which their functions are carried out are still unclear. We have previously reported that transgenic Arabidopsis plants overexpressing an Opuntia streptacantha SK3 dehydrin (OpsDHN1) show enhanced tolerance to freezing stress. Herein, we show using a split-ubiquitin yeast two-hybrid system that OpsDHN1 dimerizes. We found that the deletion of regions containing K-segments and the histidine-rich region in the OpsDHN1 protein affects dimer formation. Not surprisingly, in silico protein sequence analysis suggests that OpsDHN1 is an intrinsically disordered protein, an observation that was confirmed by circular dichroism and gel filtration of the recombinantly expressed protein. The addition of zinc triggered the association of recombinantly expressed OpsDHN1 protein, likely through its histidine-rich motif. These data brings new insights about the molecular mechanism of the OpsDHN1 SK3-dehydrin. PMID:25346739

  14. Anti-MuSK myasthenia gravis with prolonged remission.

    PubMed

    Bouwyn, Jean Paul; Magnier, Patrick; Bédat-Millet, Anne-Laure; Ahtoy, Patrick; Maltête, David; Lefaucheur, Romain

    2016-07-01

    Myasthenia gravis (MG) with antibodies against muscle-specific tyrosine kinase (MuSK) is a rare disorder of neuromuscular transmission affecting preferentially bulbar, neck and respiratory muscles. We report the case of a 22-year-old man who presented with diplopia on lateral gaze to both sides, facial diplegia, nasal dysarthria and dysphagia. Repetitive nerve stimulation of the trapezius and orbicularis oculi muscles showed amplitude decrements of 19% and 41% respectively supporting the diagnosis of myasthenia gravis. MUsK antibodies were positive. Corticosteroids were introduced and then tapered and discontinued at 6 months after initiation. The patient remained in remission and asymptomatic for 4 years without ongoing treatment or prior treatment with rituximab after this first relapse of MuSK-MG. MuSK- MG is considered a hard-to-treat condition and patients generally remain dependent on immunosuppression or prior treatment with rituximab. Our observation highlights that patients with MuSK-MG can have a benign course and that continued immunosuppressive or immunomodulatory therapy may not always be required. Copyright © 2016 Elsevier B.V. All rights reserved.

  15. Acyclic Cucurbit[n]uril-Type Molecular Container Enables Systemic Delivery of Effective Doses of Albendazole for Treatment of SK-OV-3 Xenograft Tumors.

    PubMed

    Hettiarachchi, Gaya; Samanta, Soumen K; Falcinelli, Shane; Zhang, Ben; Moncelet, Damien; Isaacs, Lyle; Briken, Volker

    2016-03-07

    Approximately, 40-70% of active pharmaceutical ingredients (API) are severely limited by their extremely poor aqueous solubility, and consequently, there is a high demand for excipients that can be used to formulate clinically relevant doses of these drug candidates. Here, proof-of-concept studies demonstrate the potential of our recently discovered acyclic cucurbit[n]uril-type molecular container Motor1 (M1) as a solubilizing agent for insoluble drugs. M1 did not induce significant rates of mutations in various Salmonella typhimurium test strains during the Ames test, suggesting low genotoxicity. M1 also has low risk of causing cardiac toxicity in humans since it did not inhibit the human Ether-à-go-go-Related Gene channel as tested on transfected CHO cell lines via patch clamp analysis. Albendazole (ABZ) is a widely used antihelminthic agent but that has also shown promising efficacy against cancerous cells in vitro. However, due to its low aqueous solubility (2.7 μM) and poor pharmacokinetics, ABZ is clinically limited as an anticancer agent. Here we investigated the potential of M1 as a solubilizing excipient for ABZ formulation. A pharmacokinetic study indicated that ABZ escapes the peritoneal cavity resulting in 78% absolute bioavailability, while its active intermediate metabolite, albendazole sulfoxide, achieved 43% absolute bioavailability. The daily dosing of 681 mg/kg M1 complexed with 3.2 mg/kg of ABZ for 14 days did not result in significant weight loss or pathology in Swiss Webster mice. In vivo efficacy studies using this M1·ABZ inclusion complex showed significant decreases in tumor growth rates and increases in survival of mice bearing SK-OV-3 xenograft tumors. In conclusion, we provide substantial new evidence demonstrating that M1 is a safe and efficient excipient that enables in vivo parenteral delivery of poorly water-soluble APIs.

  16. Calcium-dependent stoichiometries of the KCa2.2 (SK) intracellular domain/calmodulin complex in solution

    PubMed Central

    Halling, D. Brent; Kenrick, Sophia A.; Riggs, Austen F.

    2014-01-01

    Ca2+ activates SK Ca2+-activated K+ channels through the protein Ca2+ sensor, calmodulin (CaM). To understand how SK channels operate, it is necessary to determine how Ca2+ regulates CaM binding to its target on SK. Tagless, recombinant SK peptide (SKp), was purified for binding studies with CaM at low and high Ca2+ concentrations. Composition gradient multi-angle light scattering accurately measures the molar mass, stoichiometry, and affinity of protein complexes. In 2 mM Ca2+, SKp and CaM bind with three different stoichiometries that depend on the molar ratio of SKp:CaM in solution. These complexes include 28 kD 1SKp/1CaM, 39 kD 2SKp/1CaM, and 44 kD 1SKp/2CaM. A 2SKp/2CaM complex, observed in prior crystallographic studies, is absent. At <5 nM Ca2+, 1SKp/1CaM and 2SKp/1CaM were observed; however, 1SKp/2CaM was absent. Analytical ultracentrifugation was used to characterize the physical properties of the three SKp/CaM stoichiometries. In high Ca2+, the sedimentation coefficient is smaller for a 1SKp:1CaM solution than it is for either 2SKp:1CaM or 1SKp:2CaM. At low Ca2+ and at >100 µM protein concentrations, a molar excess of SKp over CaM causes aggregation. Aggregation is not observed in Ca2+ or with CaM in molar excess. In low Ca2+ both 1SKp:1CaM and 1SKp:2CaM solutions have similar sedimentation coefficients, which is consistent with the absence of a 1SKp/2CaM complex in low Ca2+. These results suggest that complexes with stoichiometries other than 2SKp/2CaM are important in gating. PMID:24420768

  17. Calcium-dependent stoichiometries of the KCa2.2 (SK) intracellular domain/calmodulin complex in solution.

    PubMed

    Halling, D Brent; Kenrick, Sophia A; Riggs, Austen F; Aldrich, Richard W

    2014-02-01

    Ca(2+) activates SK Ca(2+)-activated K(+) channels through the protein Ca(2+) sensor, calmodulin (CaM). To understand how SK channels operate, it is necessary to determine how Ca(2+) regulates CaM binding to its target on SK. Tagless, recombinant SK peptide (SKp), was purified for binding studies with CaM at low and high Ca(2+) concentrations. Composition gradient multi-angle light scattering accurately measures the molar mass, stoichiometry, and affinity of protein complexes. In 2 mM Ca(2+), SKp and CaM bind with three different stoichiometries that depend on the molar ratio of SKp:CaM in solution. These complexes include 28 kD 1SKp/1CaM, 39 kD 2SKp/1CaM, and 44 kD 1SKp/2CaM. A 2SKp/2CaM complex, observed in prior crystallographic studies, is absent. At <5 nM Ca(2+), 1SKp/1CaM and 2SKp/1CaM were observed; however, 1SKp/2CaM was absent. Analytical ultracentrifugation was used to characterize the physical properties of the three SKp/CaM stoichiometries. In high Ca(2+), the sedimentation coefficient is smaller for a 1SKp:1CaM solution than it is for either 2SKp:1CaM or 1SKp:2CaM. At low Ca(2+) and at >100 µM protein concentrations, a molar excess of SKp over CaM causes aggregation. Aggregation is not observed in Ca(2+) or with CaM in molar excess. In low Ca(2+) both 1SKp:1CaM and 1SKp:2CaM solutions have similar sedimentation coefficients, which is consistent with the absence of a 1SKp/2CaM complex in low Ca(2+). These results suggest that complexes with stoichiometries other than 2SKp/2CaM are important in gating.

  18. Guidelines for pre-clinical animal and cellular models of MuSK-myasthenia gravis.

    PubMed

    Phillips, W D; Christadoss, P; Losen, M; Punga, A R; Shigemoto, K; Verschuuren, J; Vincent, A

    2015-08-01

    Muscle-specific tyrosine kinase (MuSK) autoantibodies are the hallmark of a form of myasthenia gravis (MG) that can challenge the neurologist and the experimentalist. The clinical disease can be difficult to treat effectively. MuSK autoantibodies affect the neuromuscular junction in several ways. When added to muscle cells in culture, MuSK antibodies disperse acetylcholine receptor clusters. Experimental animals actively immunized with MuSK develop MuSK autoantibodies and muscle weakness. Weakness is associated with reduced postsynaptic acetylcholine receptor numbers, reduced amplitudes of miniature endplate potentials and endplate potentials, and failure of neuromuscular transmission. Similar impairments have been found in mice injected with IgG from MG patients positive for MuSK autoantibody (MuSK-MG). The active and passive models have begun to reveal the mechanisms by which MuSK antibodies disrupt synaptic function at the neuromuscular junction, and should be valuable in developing therapies for MuSK-MG. However, translation into new and improved treatments for patients requires procedures that are not too cumbersome but suitable for examining different aspects of MuSK function and the effects of potential therapies. Study design, conduct and analysis should be carefully considered and transparently reported. Here we review what has been learnt from animal and culture models of MuSK-MG, and offer guidelines for experimental design and conduct of studies, including sample size determination, randomization, outcome parameters and precautions for objective data analysis. These principles may also be relevant to the increasing number of other antibody-mediated diseases that are now recognized. Copyright © 2014 Elsevier Inc. All rights reserved.

  19. Expression of a Diverse Array of Ca2+-Activated K+ Channels (SK1/3, IK1, BK) that Functionally Couple to the Mechanosensitive TRPV4 Channel in the Collecting Duct System of Kidney.

    PubMed

    Li, Yue; Hu, Hongxiang; Butterworth, Michael B; Tian, Jin-Bin; Zhu, Michael X; O'Neil, Roger G

    2016-01-01

    The voltage- and Ca2+-activated, large conductance K+ channel (BK, maxi-K) is expressed in the collecting duct system of kidney where it underlies flow- and Ca2+-dependent K+ excretion. To determine if other Ca2+-activated K+ channels (KCa) may participate in this process, mouse kidney and the K+-secreting mouse cortical collecting duct (CCD) cell line, mCCDcl1, were assessed for TRPV4 and KCa channel expression and cross-talk. qPCR mRNA analysis and immunocytochemical staining demonstrated TRPV4 and KCa expression in mCCDcl1 cells and kidney connecting tubule (CNT) and CCD. Three subfamilies of KCa channels were revealed: the high Ca2+-binding affinity small-conductance SK channels, SK1and SK3, the intermediate conductance channel, IK1, and the low Ca2+-binding affinity, BK channel (BKα subunit). Apparent expression levels varied in CNT/CCD where analysis of CCD principal cells (PC) and intercalated cells (IC) demonstrated differential staining: SK1:PCSK3:PC>IC, IK1:PC>IC, BKα:PC = IC, and TRPV4:PC>IC. Patch clamp analysis and fluorescence Ca2+ imaging of mCCDcl1 cells demonstrated potent TRPV4-mediated Ca2+ entry and strong functional cross-talk between TRPV4 and KCa channels. TRPV4-mediated Ca2+ influx activated each KCa channel, as evidenced by selective inhibition of KCa channels, with each active KCa channel enhancing Ca2+ entry (due to membrane hyperpolarization). Transepithelial electrical resistance (TEER) analysis of confluent mCCDcl1 cells grown on permeable supports further demonstrated this cross-talk where TRPV4 activation induce a decrease in TEER which was partially restored upon selective inhibition of each KCa channel. It is concluded that SK1/SK3 and IK1 are highly expressed along with BKα in CNT and CCD and are closely coupled to TRPV4 activation as observed in mCCDcl1 cells. The data support a model in CNT/CCD segments where strong cross talk between TRPV4-mediated Ca2+ influx and each KCa channel leads to enhance Ca2+ entry which

  20. Global transcriptomic response of Anoxybacillus sp. SK 3-4 to aluminum exposure.

    PubMed

    Lim, Jia Chun; Thevarajoo, Suganthi; Selvaratnam, Chitra; Goh, Kian Mau; Shamsir, Mohd Shahir; Ibrahim, Zaharah; Chong, Chun Shiong

    2017-02-01

    Anoxybacillus sp. SK 3-4 is a Gram-positive, rod-shaped bacterium and a member of family Bacillaceae. We had previously reported that the strain is an aluminum resistant thermophilic bacterium. This is the first report to provide a detailed analysis of the global transcriptional response of Anoxybacillus when the cells were exposed to 600 mg L -1 of aluminum. The transcriptome was sequenced using Illumina MiSeq sequencer. Total of 708 genes were differentially expressed (fold change >2.00) with 316 genes were up-regulated while 347 genes were down-regulated, in comparing to control with no aluminum added in the culture. Based on Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis, the majority of genes encoding for cell metabolism such as glycolysis, sulfur metabolism, cysteine and methionine metabolism were up-regulated; while most of the gene associated with tricarboxylic acid cycle (TCA cycle) and valine, leucine and isoleucine metabolism were down-regulated. In addition, a significant number of the genes encoding ABC transporters, metal ions transporters, and some stress response proteins were also differentially expressed following aluminum exposure. The findings provide further insight and help us to understand on the resistance of Anoxybacillus sp. SK 3-4 toward aluminium. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  1. Characterization of CD4 and CD8 T Cell Responses in MuSK Myasthenia Gravis

    PubMed Central

    Yi, JS; Guidon, A; Sparks, S; Osborne, R; Juel, VC; Massey, JM; Sanders, DB; Weinhold, KJ; Guptill, JT

    2014-01-01

    Muscle specific tyrosine kinase myasthenia gravis (MuSK MG) is a form of autoimmune MG that predominantly affects women and has unique clinical features, including prominent bulbar weakness, muscle atrophy, and excellent response to therapeutic plasma exchange. Patients with MuSK MG have predominantly IgG4 autoantibodies directed against MuSK on the postsynaptic muscle membrane. Lymphocyte functionality has not been reported in this condition. The goal of this study was to characterize T-cell responses in patients with MuSK MG. Intracellular production of IFN-gamma, TNF-alpha, IL-2, IL-17, and IL-21 by CD4+ and CD8+ T-cells was measured by polychromatic flow cytometry in peripheral blood samples from 11 Musk MG patients and 10 healthy controls. Only one MuSK MG patient was not receiving immunosuppressive therapy. Regulatory T-cells (Treg) were also included in our analysis to determine if changes in T cell function were due to altered Treg frequencies. CD8+ T-cells from MuSK MG patients had higher frequencies of polyfunctional responses than controls, and CD4+ T-cells had higher IL-2, TNF-alpha, and IL-17. MuSK MG patients had a higher percentage of CD4+ T-cells producing combinations of IFN-gamma/IL-2/TNF-gamma, TNF-alpha/IL-2, and IFN-gamma/TNF-alpha. Interestingly, Treg numbers and CD39 expression were not different from control values. MuSK MG patients had increased frequencies of Th1 and Th17 cytokines and were primed for polyfunctional proinflammatory responses that cannot be explained by a defect in Treg function or number. PMID:24378287

  2. [Influences of a shiitake (Lentinus edodes)-fructo-oligosaccharide mixture (SK-204) on experimental pulmonary thrombosis in rats].

    PubMed

    Otsuka, M; Shinozuka, K; Hirata, G; Kunitomo, M

    1996-02-01

    Effects of the mixture (SK-204) consisting of dried shiitake mushroom (Lentinus edodes) treated with wet-heating and fructo-oligosaccharides (7:3) on the experimental models of pulmonary thrombosis induced by lactic acidosis in rats were evaluated. Chronic oral administration (10 weeks) of SK-204 significantly prevented the thrombus formation on this thrombosis model. However, decreases in the numbers of platelet and fibrinogen level by lactate were not changed by SK-204. These results suggest that SK-204 have an anti-thrombotic action, which is due to neither the inhibition of platelet aggregation nor coagulation, but probably due to the promotion of fibrinolysis and thrombolysis.

  3. Bioactive products from singlet oxygen photooxygenation of cannabinoids.

    PubMed

    Galal Osman, Ahmed; Elokely, Khaled M; Yadav, Vivek K; Carvalho, Paulo; Radwan, Mohamed; Slade, Desmond; Gul, Waseem; Khan, Shabana; Dale, Olivia R; Husni, Afeef S; Klein, Michael L; Cutler, Stephen J; Ross, Samir A; ElSohly, Mahmoud A

    2018-01-01

    Photooxygenation of Δ 8 tetrahydrocannabinol (Δ 8 -THC), Δ 9 tetrahydrocannabinol (Δ 9 -THC), Δ 9 tetrahydrocannabinolic acid (Δ 9 -THCA) and some derivatives (acetate, tosylate and methyl ether) yielded 24 oxygenated derivatives, 18 of which were new and 6 were previously reported, including allyl alcohols, ethers, quinones, hydroperoxides, and epoxides. Testing these compounds for their modulatory effect on cannabinoid receptors CB 1 and CB 2 led to the identification of 7 and 21 as CB 1 partial agonists with Ki values of 0.043 μM and 0.048 μM, respectively and 23 as a cannabinoid with high binding affinity for CB 2 with Ki value of 0.0095 μM, but much less affinity towards CB 1 (Ki 0.467 μM). The synthesized compounds showed cytotoxic activity against cancer cell lines (SK-MEL, KB, BT-549, and SK-OV-3) with IC 50 values ranging from 4.2 to 8.5 μg/mL. Several of those compounds showed antimicrobial, antimalarial and antileishmanial activities, with compound 14 being the most potent against various pathogens. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

  4. The 7SK snRNP associates with the little elongation complex to promote snRNA gene expression.

    PubMed

    Egloff, Sylvain; Vitali, Patrice; Tellier, Michael; Raffel, Raoul; Murphy, Shona; Kiss, Tamás

    2017-04-03

    The 7SK small nuclear RNP (snRNP), composed of the 7SK small nuclear RNA (snRNA), MePCE, and Larp7, regulates the mRNA elongation capacity of RNA polymerase II (RNAPII) through controlling the nuclear activity of positive transcription elongation factor b (P-TEFb). Here, we demonstrate that the human 7SK snRNP also functions as a canonical transcription factor that, in collaboration with the little elongation complex (LEC) comprising ELL, Ice1, Ice2, and ZC3H8, promotes transcription of RNAPII-specific spliceosomal snRNA and small nucleolar RNA (snoRNA) genes. The 7SK snRNA specifically associates with a fraction of RNAPII hyperphosphorylated at Ser5 and Ser7, which is a hallmark of RNAPII engaged in snRNA synthesis. Chromatin immunoprecipitation (ChIP) and chromatin isolation by RNA purification (ChIRP) experiments revealed enrichments for all components of the 7SK snRNP on RNAPII-specific sn/snoRNA genes. Depletion of 7SK snRNA or Larp7 disrupts LEC integrity, inhibits RNAPII recruitment to RNAPII-specific sn/snoRNA genes, and reduces nascent snRNA and snoRNA synthesis. Thus, through controlling both mRNA elongation and sn/snoRNA synthesis, the 7SK snRNP is a key regulator of nuclear RNA production by RNAPII. © 2017 The Authors.

  5. Sarcoidosis in a case of MuSK-positive myasthenia gravis.

    PubMed

    Spengos, Konstantinos; Vassilopoulou, Sofia; Christou, Yiolanda; Manta, Panagiota

    2008-11-01

    Whereas the coexistence of different autoimmune or rheumatologic diseases with myasthenia gravis is well documented, the combination of myasthenia with sarcoidosis is extremely rare. There very few case reports of patients suffering from these two immune-mediated diseases. Nearly all had acetylcholine receptor antibodies. As far as we know myasthenia gravis with antibodies to muscle-specific tyrosine kinase-MuSK--has not been associated with any form of sarcoidosis. We present probably the first case of MuSK-positive myasthenia gravis with concurrent, asymptomatic pulmonary sarcoidosis.

  6. Endothelial SK(Ca) and IK(Ca) channels regulate brain parenchymal arteriolar diameter and cortical cerebral blood flow.

    PubMed

    Hannah, Rachael M; Dunn, Kathryn M; Bonev, Adrian D; Nelson, Mark T

    2011-05-01

    Calcium-sensitive potassium (K(Ca)) channels have been shown to modulate the diameter of cerebral pial arteries; however, little is known regarding their roles in controlling cerebral parenchymal arterioles (PAs). We explored the function and cellular distribution of small-conductance (SK(Ca)) and intermediate-conductance (IK(Ca)) K(Ca) channels and large-conductance K(Ca) (BK(Ca)) channels in endothelial cells (ECs) and smooth muscle cells (SMCs) of PAs. Both SK(Ca) and IK(Ca) channels conducted the outward current in isolated PA ECs (current densities, ~20 pA/pF and ~28 pA/pF at +40 mV, respectively), but these currents were not detected in PA SMCs. In contrast, BK(Ca) currents were prominent in PA SMCs (~154 pA/pF), but were undetectable in PA ECs. Pressurized PAs constricted to inhibition of SK(Ca) (~16%) and IK(Ca) (~16%) channels, but were only modestly affected by inhibition of BK(Ca) channels (~5%). Blockade of SK(Ca) and IK(Ca) channels decreased resting cortical cerebral blood flow (CBF) by ~15%. NS309 (6,7-dichloro-1H-indole-2,3-dione3-oxime), a SK(Ca)/IK(Ca) channel opener, hyperpolarized PA SMCs by ~27 mV, maximally dilated pressurized PAs, and increased CBF by ~40%. In conclusion, these data show that SK(Ca) and IK(Ca) channels in ECs profoundly modulate PA tone and CBF, whereas BK(Ca) channels in SMCs only modestly influence PA diameter.

  7. Pomegranate inhibits neuroinflammation and amyloidogenesis in IL-1β-stimulated SK-N-SH cells.

    PubMed

    Velagapudi, Ravikanth; Baco, Gina; Khela, Sunjeet; Okorji, Uchechukwu; Olajide, Olumayokun

    2016-06-01

    Pomegranate fruit, Punica granatum L. (Punicaceae), and its constituents have been shown to inhibit inflammation. In this study, we aimed to assess the effects of freeze-dried pomegranate (PWE) on PGE2 production in IL-1β-stimulated SK-N-SH cells. An enzyme immunoassay (EIA) was used to measure prostaglandin E2 (PGE2) production from supernatants of IL-1β-stimulated SK-N-SH cells. Expression of COX-2, phospho-IκB, and phospho-IKK proteins was evaluated, while NF-κB reporter gene assay was carried out in TNFα-stimulated HEK293 cells to determine the effect of PWE on NF-κB transactivation. Levels of BACE-1 and Aβ in SK-N-SH cells stimulated with IL-1β were measured with an in cell ELISA. PWE (25-200 μg/ml) dose dependently reduced COX-2-dependent PGE2 production in SK-N-SH cells stimulated with IL-1β. Phosphorylation of IκB and IKK was significantly (p < 0.001) inhibited by PWE (50-200 μg/ml). Our studies also show that PWE (50-200 μg/ml) significantly (p < 0.01) inhibited NF-κB transactivation in TNFα-stimulated HEK293 cells. Furthermore, PWE inhibited BACE-1 and Aβ expression in SK-N-SH cells treated with IL-1β. Taken together, our study demonstrates that pomegranate inhibits inflammation, as well as amyloidogenesis in IL-1β-stimulated SK-N-SH cells. We propose that pomegranate is a potential nutritional strategy in slowing the progression of neurodegenerative disorders such as Alzheimer's disease.

  8. Improvement of SPM nonlinear limit by chirped duobinary PolSK transmission

    NASA Astrophysics Data System (ADS)

    Yang, Lixiu; Fan, Jiayu; Wang, Lutang; Huang, Zhaoming

    2005-02-01

    In today's terrestrial long-haul optical fiber communication systems, high channel powers are required to obtain a large transmission distance with reasonable optical amplifier spacing. In such systems, however, the presence of nonlinear effects such as the self-phase modulation (SPM) and the fiber dispersion as well as their combined effects, called SPM-induced nonlinear limitation or SPM limit, will seriously degrade the system performances in respect of the effective transmission distance and ultimately become a limiting factor in high-speed, long-haul optical fiber transmission.In this paper, a new transmission format: chirped duobinary PolSK transmission, has been proposed to generate a pre-chirped duobianry signal with fixed polarity (either positive or negative), which is modulated by a PolSK modulator. This format is based on a transmitter setup consisting of a duobinary PolSK Modulation transmitter followed by an additional phase modulator. The chirped duobinary PolSK transmission reduces the signal degradation and spectral broadening in the nonlinear regime significantly. Thus it shifts this SPM nonlinear limit to enable more relaxed dispersion compensation at high optical power compared to the conventional duobinary schemes.The simulation results show chirped duobinary PolSK transmission enlarges the dispersion limited transmission distance, increases the dispersion tolerance and overcome the SPM nonlinear limit.

  9. Effect of cationic plastoquinone SkQ1 on electron transfer reactions in chloroplasts and mitochondria from pea seedlings.

    PubMed

    Samuilov, V D; Kiselevsky, D B

    2015-04-01

    Plastoquinone bound with decyltriphenylphosphonium cation (SkQ1) penetrating through the membrane in nanomolar concentrations inhibited H2O2 generation in cells of epidermis of pea seedling leaves that was detected by the fluorescence of 2',7'-dichlorofluorescein. Photosynthetic electron transfer in chloroplasts isolated from pea leaves is suppressed by SkQ1 at micromolar concentrations: the electron transfer in chloroplasts under the action of photosystem II or I (with silicomolybdate or methyl viologen as electron acceptors, respectively) is more sensitive to SkQ1 than under the action of photosystem II + I (with ferricyanide or p-benzoquinone as electron acceptors). SkQ1 reduced by borohydride is oxidized by ferricyanide, p-benzoquinone, and, to a lesser extent, by silicomolybdate, but not by methyl viologen. SkQ1 is not effective as an electron acceptor supporting O2 evolution from water in illuminated chloroplasts. The data on suppression of photosynthetic O2 evolution or consumption show that SkQ1, similarly to phenazine methosulfate, causes conversion of the chloroplast redox-chain from non-cyclic electron transfer mode to the cyclic mode without O2 evolution. Oxidation of NADH or succinate in mitochondria isolated from pea roots is stimulated by SkQ1.

  10. Mitochondria-targeted antioxidant SkQ1 improves impaired dermal wound healing in old mice.

    PubMed

    Demyanenko, Ilya A; Popova, Ekaterina N; Zakharova, Vlada V; Ilyinskaya, Olga P; Vasilieva, Tamara V; Romashchenko, Valeria P; Fedorov, Artem V; Manskikh, Vasily N; Skulachev, Maxim V; Zinovkin, Roman A; Pletjushkina, Olga Yu; Skulachev, Vladimir P; Chernyak, Boris V

    2015-07-01

    The process of skin wound healing is delayed or impaired in aging animals. To investigate the possible role of mitochondrial reactive oxygen species (mtROS) in cutaneous wound healing of aged mice, we have applied the mitochondria-targeted antioxidant SkQ1. The SkQ1 treatment resulted in accelerated resolution of the inflammatory phase, formation of granulation tissue, vascularization and epithelization of the wounds. The wounds of SkQ1-treated mice contained increased amount of myofibroblasts which produce extracellular matrix proteins and growth factors mediating granulation tissue formation. This effect resembled SkQ1-induced differentiation of fibroblasts to myofibroblast, observed earlierin vitro. The Transforming Growth Factor beta (TGFb) produced by SkQ1-treated fibroblasts was found to stimulated motility of endothelial cells in vitro, an effect which may underlie pro-angiogenic action of SkQ1 in the wounds. In vitro experiments showed that SkQ1 prevented decomposition of VE-cadherin containing contacts and following increase in permeability of endothelial cells monolayer, induced by pro-inflammatory cytokine TNF. Prevention of excessive reaction of endothelium to the pro-inflammatory cytokine(s) might account for anti-inflammatory effect of SkQ1. Our findings point to an important role of mtROS in pathogenesis of age-related chronic wounds.

  11. Mitochondria-targeted antioxidant SkQ1 improves impaired dermal wound healing in old mice

    PubMed Central

    Zakharova, Vlada V.; Ilyinskaya, Olga P.; Vasilieva, Tamara V.; Romashchenko, Valeria P.; Fedorov, Artem V.; Manskikh, Vasily N.; Skulachev, Maxim V.; Zinovkin, Roman A.; Pletjushkina, Olga Yu.; Skulachev, Vladimir P.; Chernyak, Boris V.

    2015-01-01

    The process of skin wound healing is delayed or impaired in aging animals. To investigate the possible role of mitochondrial reactive oxygen species (mtROS) in cutaneous wound healing of aged mice, we have applied the mitochondria-targeted antioxidant SkQ1. The SkQ1 treatment resulted in accelerated resolution of the inflammatory phase, formation of granulation tissue, vascularization and epithelization of the wounds. The wounds of SkQ1-treated mice contained increased amount of myofibroblasts which produce extracellular matrix proteins and growth factors mediating granulation tissue formation. This effect resembled SkQ1-induced differentiation of fibroblasts to myofibroblast, observed earlier in vitro. The Transforming Growth Factor beta (TGFβ)produced by SkQ1-treated fibroblasts was found to stimulated motility of endothelial cells in vitro, an effect which may underlie pro-angiogenic action of SkQ1 in the wounds. In vitro experiments showed that SkQ1 prevented decomposition of VE-cadherin containing contacts and following increase in permeability of endothelial cells monolayer, induced by pro-inflammatory cytokine TNF. Prevention of excessive reaction of endothelium to the pro-inflammatory cytokine(s) might account for anti-inflammatory effect of SkQ1. Our findings point to an important role of mtROS in pathogenesis of age-related chronic wounds. PMID:26187706

  12. Crystal Structure of the Frizzled-Like Cysteine-Rich Domain of the Receptor Tyrosine Kinase MuSK

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Stiegler, A.; Burden, S; Hubbard, S

    Muscle-specific kinase (MuSK) is an essential receptor tyrosine kinase for the establishment and maintenance of the neuromuscular junction (NMJ). Activation of MuSK by agrin, a neuronally derived heparan-sulfate proteoglycan, and LRP4 (low-density lipoprotein receptor-related protein-4), the agrin receptor, leads to clustering of acetylcholine receptors on the postsynaptic side of the NMJ. The ectodomain of MuSK comprises three immunoglobulin-like domains and a cysteine-rich domain (Fz-CRD) related to those in Frizzled proteins, the receptors for Wnts. Here, we report the crystal structure of the MuSK Fz-CRD at 2.1 {angstrom} resolution. The structure reveals a five-disulfide-bridged domain similar to CRDs of Frizzled proteinsmore » but with a divergent C-terminal region. An asymmetric dimer present in the crystal structure implicates surface hydrophobic residues that may function in homotypic or heterotypic interactions to mediate co-clustering of MuSK, rapsyn, and acetylcholine receptors at the NMJ.« less

  13. Bradykinin mediates myogenic differentiation in murine myoblasts through the involvement of SK1/Spns2/S1P2 axis.

    PubMed

    Bruno, Gennaro; Cencetti, Francesca; Bernacchioni, Caterina; Donati, Chiara; Blankenbach, Kira Vanessa; Thomas, Dominique; Meyer Zu Heringdorf, Dagmar; Bruni, Paola

    2018-05-01

    Skeletal muscle tissue retains a remarkable regenerative capacity due to the activation of resident stem cells that in pathological conditions or after tissue damage proliferate and commit themselves into myoblasts. These immature myogenic cells undergo differentiation to generate new myofibers or repair the injured ones, giving a strong contribution to muscle regeneration. Cytokines and growth factors, potently released after tissue injury by leukocytes and macrophages, are not only responsible of the induction of the initial inflammatory response, but can also affect skeletal muscle regeneration. Growth factors exploit sphingosine kinase (SK), the enzyme that catalyzes the production of sphingosine 1-phosphate (S1P), to exert their biological effects in skeletal muscle. In this paper we show for the first time that bradykinin (BK), the leading member of kinin/kallikrein system, is able to induce myogenic differentiation in C2C12 myoblasts. Moreover, evidence is provided that SK1, the specific S1P-transporter spinster homolog 2 (Spns2) and S1P 2 receptor are involved in the action exerted by BK, since pharmacological inhibition/antagonism or specific down-regulation significantly alter BK-induced myogenic differentiation. Moreover, the molecular mechanism initiated by BK involves a rapid translocation of SK1 to plasma membrane, analyzed by time-lapse immunofluorescence analysis. The present study highlights the role of SK1/Spns2/S1P receptor 2 signaling axis in BK-induced myogenic differentiation, thus confirming the crucial involvement of this pathway in skeletal muscle cell biology. Copyright © 2018 Elsevier Inc. All rights reserved.

  14. The mid-face of lower Pleistocene hominins and its bearing on the attribution of SK 847 and StW 53.

    PubMed

    Williams, Frank L'engle; Schroeder, Lauren; Ackermann, Rebecca Rogers

    2012-08-01

    SK 847 and StW 53 have often been cited as evidence for early Homo in South Africa. To examine whether midfacial morphology is in agreement with these attributions, we analyze Euclidean distances calculated from 3-D coordinates on the maxillae of SK 847 and StW 53, as well as Australopithecus africanus (Sts 5, Sts 71), Paranthropus robustus (SK 46, SK 48, SK 52, SK 83), early Homo (KNM-ER 1813, KNM-ER 1805, KNM-ER 3733, KNM-WT 15000), P. boisei (KNM-ER 406, KNM-WT 17000, KNM-WT 17400), Gorilla gorilla (n=116), Homo sapiens (n=342), Pan paniscus (n=21) and P. troglodytes (n=65). Multivariate analyses separate extant hominoids suggesting we have captured taxonomic affinity. With the exception of SK 847 and SK 52, South African fossils tend to cluster together. P. robustus differs substantially from East African robust megadonts. SK 847 and StW 53 resemble the East African Homo specimens that are the most australopith-like, such as KNM-WT 15000 and KNM-ER 1813. The resemblance between StW 53 and Homo is driven partly by similarities in maxillary size. When distances are scaled, StW 53 aligns with A. africanus, while SK 847 clusters primarily with early Homo. Copyright © 2012 Elsevier GmbH. All rights reserved.

  15. Observations of Sk-69 deg 203 and the interstellar extinction towards SN 1987A

    NASA Technical Reports Server (NTRS)

    Fitzpatrick, Edward L.; Walborn, Nolan R.

    1990-01-01

    Optical and UV spectroscopic observations of the Large Magellanic Cloud (LMC) star Sk-69 deg 203 are discussed. The optical data reveal Sk-69 deg 203 to be a BO.7 Ia supergiant with a moderate nitrogen enhancement, and its UV spectrum is consistent with this classification. UV interstellar extinction curves were constructed for the star using, as flux standards, two lightly reddened LMS supergiants, which bracket Sk-69 deg 203's spectral type. The resultant extinction curves are consistent with the extinction law derived previously for the 30 Doradus region, and the results for Sk-69 deg 203 suggest that the general 30 Doradus extinction law is appropriate for dereddening the observed fluxes of SN 1987A. Published H I 21 observations place SN 1987A in a region with a strong E-W gradient in the total hydrogen content. Comparison with the H I column density implied by the reddening indicates that the supernova is imbedded approximately in the middle of the main H I complex.

  16. Solution properties of the capsular polysaccharide produced by Klebsiella pneumoniae SK1.

    PubMed

    Cescutti, P; Paoletti, S; Navarini, L; Flaibani, A

    1993-08-01

    The solution properties of the capsular polysaccharide produced by Klebsiella pneumoniae SK1, SK1-CPS, were investigated by various methods. The SK1-CPS repeating unit is a branched pentasaccharide containing one glucuronic acid as single unit side chain; acetyl groups are present as non-carbohydrate substituents on the uronic acid residue in non-stoichiometric amounts. Chiro-optical, potentiometric, viscometric and rheological measurements have been performed in order to characterize the conformational behaviour of the polymer in water and in aqueous salt solutions. Under the investigated experimental conditions, changes of temperature, ionic strength and pH were shown not to induce any cooperative conformational transition. All the results obtained suggest that the solution conformation of SK1-CPS is a random coil with a certain degree of chain flexibility. The removal of the acetyl substituents apparently does not modify the overall conclusions drawn for the native polymer, except for an incipient tendency to aggregation revealed for high salt conditions.

  17. Cadmium (Cd(2+)) exposure differentially elicits both cell proliferation and cell death related responses in SK-RC-45.

    PubMed

    Sinha, Krishnendu; Pal, Pabitra Bikash; Sil, Parames C

    2014-03-01

    Cadmium (Cd(2+)) is a major nephrotoxic environmental pollutant, affecting mostly proximal convoluted tubule (PCT) cells of the mammalian kidney, while conditionally Cd(2+) could also elicit protective responses with great variety and variability in different systems. The present study was designed to evaluate the molecular mechanism of Cd(2+) toxicity on human PCT derived Renal Cell Carcinoma (RCC), SK-RC-45 and compare its responses with normal human PCT derived cell line, NKE. Exposure of SK-RC-45 cells with different concentrations of CdCl2 (e.g. 0, 10 and 20μM) in serum free medium for 24h generate considerable amount of ROS, accompanied with decreased cell viability and alternations in the cellular and nuclear morphologies, heat shock responses and GCLC mediated protective responses. Also phosphatidylserine externalization, augmentation in the level of caspase-3, PARP, BAD, Apaf1 and cleaved caspase-9 along with decreased expression of Bcl2 and release of cytochrome c confirmed that, Cd(2+) dose dependently induces solely intrinsic pathway of apoptosis in SK-RC-45, independent of JNK. Furthermore, the non-toxic concentration (10μM) of Cd(2+) induced nuclear translocation of Nrf2 and increased expression in the level of HO-1 enzyme suggesting that at the milder concentration, Cd(2+) induces protective signaling pathways. On the other hand, exposure of NKE to different concentrations of CdCl2 (e.g. 0, 10, 20, 30 and 50μM) under the same conditions elevate stronger heat shock and SOD2 mediated protective responses. In contrary to the RCC PCT, the normal PCT derived cell follows JNK dependent and extrinsic pathways of apoptosis. Cumulatively, these results suggest that Cd(2+) exposure dose dependently elicit both cell proliferative and cell death related responses in SK-RC-45 cells and is differentially regulated with respect to normal kidney epithelia derived NKE cells. Copyright © 2013 Elsevier Ltd. All rights reserved.

  18. SkData: data sets and algorithm evaluation protocols in Python

    NASA Astrophysics Data System (ADS)

    Bergstra, James; Pinto, Nicolas; Cox, David D.

    2015-01-01

    Machine learning benchmark data sets come in all shapes and sizes, whereas classification algorithms assume sanitized input, such as (x, y) pairs with vector-valued input x and integer class label y. Researchers and practitioners know all too well how tedious it can be to get from the URL of a new data set to a NumPy ndarray suitable for e.g. pandas or sklearn. The SkData library handles that work for a growing number of benchmark data sets (small and large) so that one-off in-house scripts for downloading and parsing data sets can be replaced with library code that is reliable, community-tested, and documented. The SkData library also introduces an open-ended formalization of training and testing protocols that facilitates direct comparison with published research. This paper describes the usage and architecture of the SkData library.

  19. Structural investigation of the capsular polysaccharide produced by a novel Klebsiella serotype (SK1). Location of O-acetyl substituents using NMR and MS techniques.

    PubMed

    Cescutti, P; Ravenscroft, N; Ng, S; Lam, Z; Dutton, G G

    1993-06-21

    The capsular polysaccharide of Klebsiella SK1 was investigated by methylation analysis, Smith degradation, and 1H NMR spectroscopy. The oligosaccharides (P1 and P2) obtained by bacteriophage phi SK1 degradation of the polymer were studied by methylation analysis, and 1D- and 2D-NMR spectroscopy. The resulting data showed that the parent repeating unit is a branched pentasaccharide having a structure identical to the revised structure recently proposed for Klebsiella serotype K8 capsular polysaccharide. [Formula: see text] The 2D-NMR data showed that one third of the glucuronic acid residues in the SK1 polymer are acetylated at O-2, O-3, or O-4. FABMS studies confirmed the presence of monoacetylated glucuronic acid residues. Thus, the relationship between the Klebsiella K8 and SK1 polymers is akin to that found for Klebsiella polysaccharides K30 and K33, which have been typed as serologically distinct yet their structures differ only in the degree of acetylation.

  20. An efficient synthesis of SK-658 and its analogs as potent histone deacetylase inhibitors.

    PubMed

    Shahidul Islam, Md; Nurul Islam, Md; Ashraful Hoque, Md; Nishino, Norikazu; Kato, Tamaki; Kim, Hyun-Jung; Ito, Akihiro; Yoshida, Minoru

    2015-04-01

    SK-658 is a potent histone deacetylase (HDAC) inhibitor that showed higher activity than SAHA due to the presence of extended hydrophobic group. We designed and synthesized thioester and SS-hybrid bearing SK-658 analogs as HDAC inhibitors. All the compounds were active in nano molar range and showed higher inhibitory activity than SAHA and SK-658. Among these, disulfide compounds showed the highest activity. Copyright © 2015 Elsevier Inc. All rights reserved.

  1. PP2B and PP1α cooperatively disrupt 7SK snRNP to release P-TEFb for transcription in response to Ca2+ signaling

    PubMed Central

    Chen, Ruichuan; Liu, Min; Li, Huan; Xue, Yuhua; Ramey, Wanichaya N.; He, Nanhai; Ai, Nanping; Luo, Haohong; Zhu, Ying; Zhou, Nan; Zhou, Qiang

    2008-01-01

    The positive transcription elongation factor b (P-TEFb), consisting of Cdk9 and cyclin T, stimulates RNA polymerase II elongation and cotranscriptional pre-mRNA processing. To accommodate different growth conditions and transcriptional demands, a reservoir of P-TEFb is kept in an inactive state in the multisubunit 7SK snRNP. Under certain stress or disease conditions, P-TEFb is released to activate transcription, although the signaling pathway(s) that controls this is largely unknown. Here, through analyzing the UV- or hexamethylene bisacetamide (HMBA)-induced release of P-TEFb from 7SK snRNP, an essential role for the calcium ion (Ca2+)–calmodulin–protein phosphatase 2B (PP2B) signaling pathway is revealed. However, Ca2+ signaling alone is insufficient, and PP2B must act sequentially and cooperatively with protein phosphatase 1α (PP1α) to disrupt 7SK snRNP. Activated by UV/HMBA and facilitated by a PP2B-induced conformational change in 7SK snRNP, PP1α releases P-TEFb through dephosphorylating phospho-Thr186 in the Cdk9 T-loop. This event is also necessary for the subsequent recruitment of P-TEFb by the bromodomain protein Brd4 to the preinitiation complex, where Cdk9 remains unphosphorylated and inactive until after the synthesis of a short RNA. Thus, through cooperatively dephosphorylating Cdk9 in response to Ca2+ signaling, PP2B and PP1α alter the P-TEFb functional equilibrium through releasing P-TEFb from 7SK snRNP for transcription. PMID:18483222

  2. Thrombin/Matrix Metalloproteinase-9-Dependent SK-N-SH Cell Migration is Mediated Through a PLC/PKC/MAPKs/NF-κB Cascade.

    PubMed

    Yang, Chien-Chung; Lin, Chih-Chung; Chien, Peter Tzu-Yu; Hsiao, Li-Der; Yang, Chuen-Mao

    2016-11-01

    Thrombin has been known to activate inflammatory genes including matrix metalloproteinases (MMPs). The elevated expression of MMP-9 has been observed in patients with neuroinflammatory diseases and may contribute to the pathology of brain diseases. However, the mechanisms underlying thrombin-induced MMP-9 expression in SK-N-SH cells remain unknown. The effects of thrombin on MMP-9 expression were examined in SK-N-SH cells by gelatin zymography, Western blot, real-time PCR, promoter activity assay, and cell migration assay. The detailed mechanisms were analyzed by using pharmacological inhibitors and small intefering RNA (siRNA) transfection. Here, we demonstrated that thrombin induced the expression of proform MMP-9 and migration of SK-N-SH cells, which were attenuated by pretreatment with the inhibitor of thrombin (PPACK), Gq (GPA2A), PC-PLC (D609), PI-PLC (ET-18-OCH 3 ), nonselective protien kinase C (PKC, GF109203X), PKCα/βII (Gö6983), PKCδ (Rottlerin), p38 mitogen-activated protein kinases (MAPK) (SB202190), JNK1/2 (SP600125), or NF-κB (Bay11-7082 or Helenalin) and transfection with siRNA of Gq, PKCα, PKCβ, PKCδ, p38, JNK1/2, IKKα, IKKβ, or p65. Moreover, thrombin-stimulated PKCα/βII, PKCδ, p38 MAPK, JNK1/2, or p65 phosphorylation was abrogated by their respective inhibitor of PPACK, GPA2A, D609, ET-18-OCH 3 , Gö6983, Rottlerin, SB202190, SP600125, Bay11-7082, or Helenalin. Pretreatment with these inhibitors or transfection with MMP-9 siRNA also blocked thrombin-induced SK-N-SH cell migration. Our results show that thrombin stimulates a Gq/PLC/PKCs/p38 MAPK and JNK1/2 cascade, which in turn triggers NF-κB activation and ultimately induces MMP-9 expression and cell migration in SK-N-SH cells.

  3. Structural basis of agrin-LRP4-MuSK signaling

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Zong, Yinong; Zhang, Bin; Gu, Shenyan

    Synapses are the fundamental units of neural circuits that enable complex behaviors. The neuromuscular junction (NMJ), a synapse formed between a motoneuron and a muscle fiber, has contributed greatly to understanding of the general principles of synaptogenesis as well as of neuromuscular disorders. NMJ formation requires neural agrin, a motoneuron-derived protein, which interacts with LRP4 (low-density lipoprotein receptor-related protein 4) to activate the receptor tyrosine kinase MuSK (muscle-specific kinase). However, little is known of how signals are transduced from agrin to MuSK. Here, we present the first crystal structure of an agrin-LRP4 complex, consisting of two agrin-LRP4 heterodimers. Formation ofmore » the initial binary complex requires the z8 loop that is specifically present in neuronal, but not muscle, agrin and that promotes the synergistic formation of the tetramer through two additional interfaces. We show that the tetrameric complex is essential for neuronal agrin-induced acetylcholine receptor (AChR) clustering. Collectively, these results provide new insight into the agrin-LRP4-MuSK signaling cascade and NMJ formation and represent a novel mechanism for activation of receptor tyrosine kinases.« less

  4. [Construction of a new oncolytic virus oHSV2hGM-CSF and its anti-tumor effects].

    PubMed

    Shi, Gui-Lan; Zhuang, Xiu-Fen; Han, Xiang-Ping; Li, Jie; Zhang, Yu; Zhang, Shu-Ren; Liu, Bin-Lei

    2012-02-01

    The aim of this study was to construct a new oncolytic virus oHSV2hGM-CSF and evaluate its oncolytic activity in vitro and in vivo in parallel with oHSV1hGM-CSF. oHSV2hGM-CSF was a replication-competent, attenuated HSV2 based on the HG52 virus (an HSV2 strain). It was engineered to be specific for cancer by deletion of the viral genes ICP34.5 and ICP47 and insertion of the gene encoding hGM-CSF. To measure the in vitro killing effect of the virus, 15 human tumor cell lines (HeLa, Eca-109, PG, HepG2, SK/FU, CNE-2Z, PC-3, SK-OV3, A-549, 786-0, MCF-7, Hep-2, HT-29, SK-Mel-28, U87-MG) and mouse melanoma (B16R) cell line were seeded into 24-well plates and infected with viruses at MOI = 1 (multiplicity of infection, MOI), or left uninfected. The cells were harvested 24 and 48 hours post infection, and observed under the microscope. For animal studies, the oncolytic viruses were administered intratumorally (at 3-day interval) at a dose of 2.3 x 10(6) PFU (plaque forming unit, PFU) for three times when the tumor volume reached 7-8 mm3. The tumor volume was measured at 3-day intervals and animal survival was recorded. Both oHSV2hCM-CSFand oHSV1hGM-CSF induced widespread cytopathic effects at 24 h after infection. OHSV2hGM-CSF, by contrast, produced more plaques with a syncytial phenotype than oHSV1hGM-CSF. In the in vitro killing experiments for the cell lines HeLa, HepG2, SK-Mel-28, B16R and U87-MG, oHSV2hGM-CSF eradicated significantly more cells than oHSV1hGM-CSF under the same conditions. For the mouse experiments, it was observed that oHSV2hGM-CSF significantly inhibited the tumor growth. At 15 days after B16R tumor cells inoculation, the tumor volumes of the PBS, oHSV1hGCM-CSF and oHSV2hGM-CSF groups were (374.7 +/- 128.24) mm3, (128.23 +/- 45.32) mm3 (P < 0.05, vs. PBS group) or (10.06 +/- 5.1) mm3 (P < 0.01, vs. PBS group), respectively (mean +/- error). The long term therapeutic effect of oHSV2hGM-CSF on the B16R animal model was evaluated by recording animal

  5. SkQ1 Ophthalmic Solution for Dry Eye Treatment: Results of a Phase 2 Safety and Efficacy Clinical Study in the Environment and During Challenge in the Controlled Adverse Environment Model.

    PubMed

    Petrov, Anton; Perekhvatova, Natalia; Skulachev, Maxim; Stein, Linda; Ousler, George

    2016-01-01

    This Phase 2 clinical trial assessed the efficacy and safety of the novel antioxidative, renewable compound SkQ1 for topical treatment of dry eye signs and symptoms. In a single-center, randomized, double-masked, placebo-controlled, 29-day study, 91 subjects with mild to moderate dry eye instilled the study drug twice daily and recorded dry eye symptoms daily. Subjects were randomized 1:1:1 into one of three ophthalmic solution treatment groups: SkQ1 1.55 µg/mL, SkQ1 0.155 µg/mL, or 0.0 µg/mL (placebo). Subjects were exposed to a controlled adverse environment chamber at 3 of the 4 study visits (Day -7, Day 1, and Day 29). Investigator assessments occurred at all study visits. SkQ1 was safe and efficacious in treating dry eye signs and symptoms. Statistically significant improvements with SkQ1 compared to placebo occurred for the dry eye signs of corneal fluorescein staining and lissamine green staining in the central region and lid margin redness, and for the dry eye symptoms of ocular discomfort, dryness, and grittiness. In addition, SkQ1 demonstrated greater efficacy compared to placebo, although the differences were not statistically significant, for corneal fluorescein staining in other regions and/or time points (total staining score, central region, corneal sum score, and temporal region), lissamine green staining for the central and nasal regions, and blink rate scores. This Phase 2 study indicated that SkQ1 is safe and efficacious for the treatment of dry eye signs and symptoms and supported previous study results. Clinicaltrials.gov identifier: NCT02121301. Miotech S.A.

  6. SkICAT: A cataloging and analysis tool for wide field imaging surveys

    NASA Technical Reports Server (NTRS)

    Weir, N.; Fayyad, U. M.; Djorgovski, S. G.; Roden, J.

    1992-01-01

    We describe an integrated system, SkICAT (Sky Image Cataloging and Analysis Tool), for the automated reduction and analysis of the Palomar Observatory-ST ScI Digitized Sky Survey. The Survey will consist of the complete digitization of the photographic Second Palomar Observatory Sky Survey (POSS-II) in three bands, comprising nearly three Terabytes of pixel data. SkICAT applies a combination of existing packages, including FOCAS for basic image detection and measurement and SAS for database management, as well as custom software, to the task of managing this wealth of data. One of the most novel aspects of the system is its method of object classification. Using state-of-theart machine learning classification techniques (GID3* and O-BTree), we have developed a powerful method for automatically distinguishing point sources from non-point sources and artifacts, achieving comparably accurate discrimination a full magnitude fainter than in previous Schmidt plate surveys. The learning algorithms produce decision trees for classification by examining instances of objects classified by eye on both plate and higher quality CCD data. The same techniques will be applied to perform higher-level object classification (e.g., of galaxy morphology) in the near future. Another key feature of the system is the facility to integrate the catalogs from multiple plates (and portions thereof) to construct a single catalog of uniform calibration and quality down to the faintest limits of the survey. SkICAT also provides a variety of data analysis and exploration tools for the scientific utilization of the resulting catalogs. We include initial results of applying this system to measure the counts and distribution of galaxies in two bands down to Bj is approximately 21 mag over an approximate 70 square degree multi-plate field from POSS-II. SkICAT is constructed in a modular and general fashion and should be readily adaptable to other large-scale imaging surveys.

  7. Rituximab as treatment for anti-MuSK myasthenia gravis: Multicenter blinded prospective review.

    PubMed

    Hehir, Michael K; Hobson-Webb, Lisa D; Benatar, Michael; Barnett, Carolina; Silvestri, Nicholas J; Howard, James F; Howard, Diantha; Visser, Amy; Crum, Brian A; Nowak, Richard; Beekman, Rachel; Kumar, Aditya; Ruzhansky, Katherine; Chen, I-Hweii Amy; Pulley, Michael T; LaBoy, Shannon M; Fellman, Melissa A; Greene, Shane M; Pasnoor, Mamatha; Burns, Ted M

    2017-09-05

    To evaluate the efficacy of rituximab in treatment of anti-muscle-specific kinase (MuSK) myasthenia gravis (MG). This was a multicenter, blinded, prospective review, comparing anti-MuSK-positive patients with MG treated with rituximab to those not treated with rituximab. The primary clinical endpoint was the Myasthenia Gravis Status and Treatment Intensity (MGSTI), a novel outcome that combines the Myasthenia Gravis Foundation of America (MGFA) postintervention status (PIS) and the number and dosages of other immunosuppressant therapies used. A priori, an MGSTI of level ≤2 was used to define a favorable outcome. Secondary outcomes included modified MGFA PIS of minimal manifestations or better, mean/median prednisone dose, and mean/median doses of other immunosuppressant drugs. Seventy-seven of 119 patients with anti-MuSK MG evaluated between January 1, 2005, and January 1, 2015, at 10 neuromuscular centers were selected for analysis after review of limited clinical data by a blinded expert panel. An additional 22 patients were excluded due to insufficient follow-up. Baseline characteristics were similar between the rituximab-treated patients (n = 24) and the controls (n = 31). Median follow-up duration was >3.5 years. At last visit, 58% (14/24) of rituximab-treated patients reached the primary outcome compared to 16% (5/31) of controls ( p = 0.002). Number needed to treat for the primary outcome is 2.4. At last visit, 29% of rituximab-treated patients were taking prednisone (mean dose 4.5 mg/day) compared to 74% of controls (mean dose 13 mg/day) ( p = 0.001 and p = 0.005). This study provides Class IV evidence that for patients with anti-MuSK MG, rituximab increased the probability of a favorable outcome. © 2017 American Academy of Neurology.

  8. hnRNP R and its main interactor, the noncoding RNA 7SK, coregulate the axonal transcriptome of motoneurons.

    PubMed

    Briese, Michael; Saal-Bauernschubert, Lena; Ji, Changhe; Moradi, Mehri; Ghanawi, Hanaa; Uhl, Michael; Appenzeller, Silke; Backofen, Rolf; Sendtner, Michael

    2018-03-20

    Disturbed RNA processing and subcellular transport contribute to the pathomechanisms of motoneuron diseases such as amyotrophic lateral sclerosis and spinal muscular atrophy. RNA-binding proteins are involved in these processes, but the mechanisms by which they regulate the subcellular diversity of transcriptomes, particularly in axons, are not understood. Heterogeneous nuclear ribonucleoprotein R (hnRNP R) interacts with several proteins involved in motoneuron diseases. It is located in axons of developing motoneurons, and its depletion causes defects in axon growth. Here, we used individual nucleotide-resolution cross-linking and immunoprecipitation (iCLIP) to determine the RNA interactome of hnRNP R in motoneurons. We identified ∼3,500 RNA targets, predominantly with functions in synaptic transmission and axon guidance. Among the RNA targets identified by iCLIP, the noncoding RNA 7SK was the top interactor of hnRNP R. We detected 7SK in the nucleus and also in the cytosol of motoneurons. In axons, 7SK localized in close proximity to hnRNP R, and depletion of hnRNP R reduced axonal 7SK. Furthermore, suppression of 7SK led to defective axon growth that was accompanied by axonal transcriptome alterations similar to those caused by hnRNP R depletion. Using a series of 7SK-deletion mutants, we show that the function of 7SK in axon elongation depends on its interaction with hnRNP R but not with the PTEF-B complex involved in transcriptional regulation. These results propose a role for 7SK as an essential interactor of hnRNP R to regulate its function in axon maintenance. Copyright © 2018 the Author(s). Published by PNAS.

  9. Current status of SK-Gd project and EGADS

    NASA Astrophysics Data System (ADS)

    Xu, Chenyuan; Super-Kamiokande Collaboration

    2016-05-01

    Supernova Relic Neutrino (SRN) has not been observed yet because of its low event rate and high background. By adding gadolinium into water Cherenkov detector, inverse beta decay will have two signals, the prompt one is positron signal and the delayed one is a ~8 MeV gamma cascade from neutron capture on gadolinium. By this way, background for SRN can be largely reduced by detecting prompt and delayed signals coincidently, and Super-K will also have the ability to distinguish neutrino and anti-neutrino. SK-Gd is a R&D project proposed to dissolve gadolinium into Super-K. As a part of it, EGADS, a 200 ton water Cherenkov detector was built in Kamioka mine. Current status of SK-Gd project and the physics work being performed in EGADS will be presented here.

  10. The Crisis of Education in Arkhangel'sk.

    ERIC Educational Resources Information Center

    Konstantinov, A. S.

    1995-01-01

    Asserts that there is an educational crisis in the Arkhangel'sk area of Russia. Summarizes research studies of students and teachers to illustrate low teacher morale, student dissatisfaction, and low pay for all educators. Argues for financial autonomy for educational institutions. (CFR)

  11. Mitochondria-Targeted Antioxidant SkQ1 Improves Dermal Wound Healing in Genetically Diabetic Mice

    PubMed Central

    Demyanenko, Ilya A.; Zakharova, Vlada V.; Ilyinskaya, Olga P.; Vasilieva, Tamara V.; Fedorov, Artem V.; Skulachev, Vladimir P.

    2017-01-01

    Oxidative stress is widely recognized as an important factor in the delayed wound healing in diabetes. However, the role of mitochondrial reactive oxygen species in this process is unknown. It was assumed that mitochondrial reactive oxygen species are involved in many wound-healing processes in both diabetic humans and animals. We have applied the mitochondria-targeted antioxidant 10-(6′-plastoquinonyl)decyltriphenylphosphonium (SkQ1) to explore the role of mitochondrial reactive oxygen species in the wound healing of genetically diabetic mice. Healing of full-thickness excisional dermal wounds in diabetic C57BL/KsJ-db−/db− mice was significantly enhanced after long-term (12 weeks) administration of SkQ1. SkQ1 accelerated wound closure and stimulated epithelization, granulation tissue formation, and vascularization. On the 7th day after wounding, SkQ1 treatment increased the number of α-smooth muscle actin-positive cells (myofibroblasts), reduced the number of neutrophils, and increased macrophage infiltration. SkQ1 lowered lipid peroxidation level but did not change the level of the circulatory IL-6 and TNF. SkQ1 pretreatment also stimulated cell migration in a scratch-wound assay in vitro under hyperglycemic condition. Thus, a mitochondria-targeted antioxidant normalized both inflammatory and regenerative phases of wound healing in diabetic mice. Our results pointed to nearly all the major steps of wound healing as the target of excessive mitochondrial reactive oxygen species production in type II diabetes. PMID:28761623

  12. Mitochondria-Targeted Antioxidant SkQ1 Improves Dermal Wound Healing in Genetically Diabetic Mice.

    PubMed

    Demyanenko, Ilya A; Zakharova, Vlada V; Ilyinskaya, Olga P; Vasilieva, Tamara V; Fedorov, Artem V; Manskikh, Vasily N; Zinovkin, Roman A; Pletjushkina, Olga Yu; Chernyak, Boris V; Skulachev, Vladimir P; Popova, Ekaterina N

    2017-01-01

    Oxidative stress is widely recognized as an important factor in the delayed wound healing in diabetes. However, the role of mitochondrial reactive oxygen species in this process is unknown. It was assumed that mitochondrial reactive oxygen species are involved in many wound-healing processes in both diabetic humans and animals. We have applied the mitochondria-targeted antioxidant 10-(6'-plastoquinonyl)decyltriphenylphosphonium (SkQ1) to explore the role of mitochondrial reactive oxygen species in the wound healing of genetically diabetic mice. Healing of full-thickness excisional dermal wounds in diabetic C57BL/KsJ-db - /db - mice was significantly enhanced after long-term (12 weeks) administration of SkQ1. SkQ1 accelerated wound closure and stimulated epithelization, granulation tissue formation, and vascularization. On the 7th day after wounding, SkQ1 treatment increased the number of α -smooth muscle actin-positive cells (myofibroblasts), reduced the number of neutrophils, and increased macrophage infiltration. SkQ1 lowered lipid peroxidation level but did not change the level of the circulatory IL-6 and TNF. SkQ1 pretreatment also stimulated cell migration in a scratch-wound assay in vitro under hyperglycemic condition. Thus, a mitochondria-targeted antioxidant normalized both inflammatory and regenerative phases of wound healing in diabetic mice. Our results pointed to nearly all the major steps of wound healing as the target of excessive mitochondrial reactive oxygen species production in type II diabetes.

  13. The Cytoplasmic Adaptor Protein Dok7 Activates the Receptor Tyrosine Kinase MuSK via Dimerization

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Bergamin, E.; Hallock, P; Burden, S

    Formation of the vertebrate neuromuscular junction requires, among others proteins, Agrin, a neuronally derived ligand, and the following muscle proteins: LRP4, the receptor for Agrin; MuSK, a receptor tyrosine kinase (RTK); and Dok7 (or Dok-7), a cytoplasmic adaptor protein. Dok7 comprises a pleckstrin-homology (PH) domain, a phosphotyrosine-binding (PTB) domain, and C-terminal sites of tyrosine phosphorylation. Unique among adaptor proteins recruited to RTKs, Dok7 is not only a substrate of MuSK, but also an activator of MuSK's kinase activity. Here, we present the crystal structure of the Dok7 PH-PTB domains in complex with a phosphopeptide representing the Dok7-binding site on MuSK.more » The structure and biochemical data reveal a dimeric arrangement of Dok7 PH-PTB that facilitates trans-autophosphorylation of the kinase activation loop. The structure provides the molecular basis for MuSK activation by Dok7 and for rationalizing several Dok7 loss-of-function mutations found in patients with congenital myasthenic syndromes.« less

  14. skNAC, a Smyd1-interacting transcription factor, is involved in cardiac development and skeletal muscle growth and regeneration.

    PubMed

    Park, Chong Yon; Pierce, Stephanie A; von Drehle, Morgan; Ivey, Kathryn N; Morgan, Jayson A; Blau, Helen M; Srivastava, Deepak

    2010-11-30

    Cardiac and skeletal muscle development and maintenance require complex interactions between DNA-binding proteins and chromatin remodeling factors. We previously reported that Smyd1, a muscle-restricted histone methyltransferase, is essential for cardiogenesis and functions with a network of cardiac regulatory proteins. Here we show that the muscle-specific transcription factor skNAC is the major binding partner for Smyd1 in the developing heart. Targeted deletion of skNAC in mice resulted in partial embryonic lethality by embryonic day 12.5, with ventricular hypoplasia and decreased cardiomyocyte proliferation that were similar but less severe than in Smyd1 mutants. Expression of Irx4, a ventricle-specific transcription factor down-regulated in hearts lacking Smyd1, also depended on the presence of skNAC. Viable skNAC(-/-) adult mice had reduced postnatal skeletal muscle growth and impaired regenerative capacity after cardiotoxin-induced injury. Satellite cells isolated from skNAC(-/-) mice had impaired survival compared with wild-type littermate satellite cells. Our results indicate that skNAC plays a critical role in ventricular cardiomyocyte expansion and regulates postnatal skeletal muscle growth and regeneration in mice.

  15. Neuroprotective effects of corn silk maysin via inhibition of H2O2-induced apoptotic cell death in SK-N-MC cells.

    PubMed

    Choi, Doo Jin; Kim, Sun-Lim; Choi, Ji Won; Park, Yong Il

    2014-07-25

    Neuroprotective effects of maysin, which is a flavone glycoside that was isolated from the corn silk (CS, Zea mays L.) of a Korean hybrid corn Kwangpyeongok, against oxidative stress (H2O2)-induced apoptotic cell death of human neuroblastoma SK-N-MC cells were investigated. Maysin cytotoxicity was determined by measuring cell viability using MTT and lactate dehydrogenase (LDH) assays. Intracellular reactive oxygen species (ROS) were measured using a 2,7-dichlorofluorescein diacetate (DCF-DA) assay. Apoptotic cell death was monitored by annexin V-FITC/PI double staining and by a TUNEL assay. Antioxidant enzyme mRNA levels were determined by real-time PCR. The cleavage of poly (ADP-ribose) polymerase (PARP) was measured by western blotting. Maysin pretreatment reduced the cytotoxic effect of H2O2 on SK-N-MC cells, as shown by the increase in cell viability and by reduced LDH release. Maysin pretreatment also dose-dependently reduced the intracellular ROS level and inhibited PARP cleavage. In addition, DNA damage and H2O2-induced apoptotic cell death were significantly attenuated by maysin pretreatment. Moreover, maysin pretreatment (5-50 μg/ml) for 2h significantly and dose-dependently increased the mRNA levels of antioxidant enzymes (CAT, GPx-1, SOD-1, SOD-2 and HO-1) in H2O2 (200 μM)-insulted cells. These results suggest that CS maysin has neuroprotective effects against oxidative stress (H2O2)-induced apoptotic death of human brain SK-N-MC cells through its antioxidative action. This report is the first regarding neuroprotective health benefits of corn silk maysin by its anti-apoptotic action and by triggering the expression of intracellular antioxidant enzyme systems in SK-N-MC cells. Copyright © 2014 Elsevier Inc. All rights reserved.

  16. pSK41-Like Plasmid Is Necessary for Inc18-Like vanA Plasmid Transfer from Enterococcus faecalis to Staphylococcus aureus In Vitro

    PubMed Central

    Clark, Nancye; Patel, Jean B.

    2013-01-01

    Vancomycin-resistant Staphylococcus aureus (VRSA) is thought to result from the in vivo conjugative transfer of a vanA plasmid from an Enterococcus sp. to S. aureus. We studied bacterial isolates from VRSA cases that occurred in the United States to identify microbiological factors which may contribute to this plasmid transfer. First, vancomycin-susceptible, methicillin-resistant S. aureus (MRSA) isolates from five VRSA cases were tested for their ability to accept foreign DNA by conjugation in mating experiments with Enterococcus faecalis JH2-2 containing pAM378, a pheromone-response conjugative plasmid. All of the MRSA isolates accepted the plasmid DNA with similar transfer efficiencies (∼10−7/donor CFU) except for one isolate, MRSA8, for which conjugation was not successful. The MRSA isolates were also tested as recipients in mating experiments between an E. faecalis isolate with an Inc18-like vanA plasmid that was isolated from a VRSA case patient. Conjugative transfer was successful for 3/5 MRSA isolates. Successful MRSA recipients carried a pSK41-like plasmid, a staphylococcal conjugative plasmid, whereas the two unsuccessful MRSA recipients did not carry pSK41. The transfer of a pSK41-like plasmid from a successful MRSA recipient to the two unsuccessful recipients resulted in conjugal transfer of the Inc18-like vanA plasmid from E. faecalis at a frequency of 10−7/recipient CFU. In addition, conjugal transfer could be achieved for pSK41-negative MRSA in the presence of a cell-free culture filtrate from S. aureus carrying a pSK41-like plasmid at a frequency of 10−8/recipient CFU. These results indicated that a pSK41-like plasmid can facilitate the transfer of an Inc18-like vanA plasmid from E. faecalis to S. aureus, possibly via an extracellular factor produced by pSK41-carrying isolates. PMID:23089754

  17. In vitro characterization of PlySK1249, a novel phage lysin, and assessment of its antibacterial activity in a mouse model of Streptococcus agalactiae bacteremia.

    PubMed

    Oechslin, Frank; Daraspe, Jean; Giddey, Marlyse; Moreillon, Philippe; Resch, Grégory

    2013-12-01

    Beta-hemolytic Streptococcus agalactiae is the leading cause of bacteremia and invasive infections. These diseases are treated with β-lactams or macrolides, but the emergence of less susceptible and even fully resistant strains is a cause for concern. New bacteriophage lysins could be promising alternatives against such organisms. They hydrolyze the bacterial peptidoglycan at the end of the phage cycle, in order to release the phage progeny. By using a bioinformatic approach to screen several beta-hemolytic streptococci, a gene coding for a lysin was identified on a prophage carried by Streptococcus dysgalactiae subsp. equisimilis SK1249. The gene product, named PlySK1249, harbored an original three-domain structure with a central cell wall-binding domain surrounded by an N-terminal amidase and a C-terminal CHAP domain. Purified PlySK1249 was highly lytic and bactericidal for S. dysgalactiae (2-log10 CFU/ml decrease within 15 min). Moreover, it also efficiently killed S. agalactiae (1.5-log10 CFU/ml decrease within 15 min) but not several streptococcal commensal species. We further investigated the activity of PlySK1249 in a mouse model of S. agalactiae bacteremia. Eighty percent of the animals (n = 10) challenged intraperitoneally with 10(6) CFU of S. agalactiae died within 72 h, whereas repeated injections of PlySK1249 (45 mg/kg 3 times within 24 h) significantly protected the mice (P < 0.01). Thus, PlySK1249, which was isolated from S. dysgalactiae, demonstrated high cross-lytic activity against S. agalactiae both in vitro and in vivo. These encouraging results indicated that PlySK1249 might represent a good candidate to be developed as a new enzybiotic for the treatment of systemic S. agalactiae infections.

  18. Extreme bottom velocities induced by wind wave and currents in the Gulf of Gdańsk

    NASA Astrophysics Data System (ADS)

    Cieślikiewicz, Witold; Dudkowska, Aleksandra; Gic-Grusza, Gabriela; Jędrasik, Jan

    2017-11-01

    The principal goal of this study is to get some preliminary insights about the intensity of water movement generated by wind waves, and due to the currents in the bottom waters of Gulf of Gdańsk, during severe storms. The Gulf of Gdańsk is located in the southern Baltic Sea. This paper presents the results of analysis of wave and current-induced velocities during extreme wind conditions, which are determined based on long-term historical records. The bottom velocity fields originated from wind wave and wind currents, during analysed extreme wind events, are computed independently of each other. The long-term wind wave parameters for the Baltic Sea region are derived from the 44-year hindcast wave database generated in the framework of the project HIPOCAS funded by the European Union. The output from the numerical wave model WAM provides the boundary conditions for the model SWAN operating in high-resolution grid covering the area of the Gulf of Gdańsk. Wind current velocities are calculated with the M3D hydrodynamic model developed in the Institute of Oceanography of the University of Gdańsk based on the POM model. The three dimensional current fields together with trajectories of particle tracers spreading out of bottom boundary layer are modelled, and the calculated fields of bottom velocities are presented in the form of 2D maps. During northerly winds, causing in the Gulf of Gdańsk extreme waves and most significant wind-driven circulation, the wave-induced bottom velocities are greater than velocities due to currents. The current velocities in the bottom layer appeared to be smaller by an order of magnitude than the wave-induced bottom orbital velocities. Namely, during most severe northerly storms analysed, current bottom velocities ranged about 0.1-0.15 m/s, while the root mean square of wave-induced near-seabed velocities reached maximum values of up to 1.4 m/s in the southern part of Gulf of Gdańsk.

  19. Lycopene Inhibits Metastasis of Human Liver Adenocarcinoma SK-Hep-1 Cells by Downregulation of NADPH Oxidase 4 Protein Expression.

    PubMed

    Jhou, Bo-Yi; Song, Tuzz-Ying; Lee, Inn; Hu, Miao-Lin; Yang, Nae-Cherng

    2017-08-16

    NADPH oxidase 4 (NOX4), with the sole function to produce reactive oxygen species (ROS), can be a molecular target for disrupting cancer metastasis. Several studies have indicated that lycopene exhibited anti-metastatic actions in vitro and in vivo. However, the role of NOX4 in the anti-metastatic action of lycopene remains unknown. Herein, we first confirmed the anti-metastatic effect of lycopene (0.1-5 μM) on human liver adenocarcinoma SK-Hep-1 cells. We showed that lycopene significantly inhibited NOX4 protein expression, with the strongest inhibition of 64.3 ± 10.2% (P < 0.05) at 2.5 μM lycopene. Lycopene also significantly inhibited NOX4 mRNA expression, NOX activity, and intracellular ROS levels in SK-Hep-1 cells. We then determined the effects of lycopene on transforming growth factor β (TGF-β)-induced metastasis. We found that TGF-β (5 ng/mL) significantly increased migration, invasion, and adhesion activity, the intracellular ROS level, matrix metalloproteinase 9 (MMP-9) and MMP-2 activities, the level of NOX4 protein expression, and NOX activity. All these TGF-β-induced effects were antagonized by the incubation of SK-Hep-1 cells with lycopene (2.5 μM). Using transient transfection of siRNA against NOX4, we found that the downregulation of NOX4 could mimic lycopene by inhibiting cell migration and the activities of MMP-9 and MMP-2 during the incubation with or without TGF-β on SK-Hep-1 cells. The results demonstrate that the downregulation of NOX4 plays a crucial role in the anti-metastatic action of lycopene in SK-Hep-1 cells.

  20. Genome sequence of Shimia str. SK013, a representative of the Roseobacter group isolated from marine sediment

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Kanukollu, Saranya; Voget, Sonja; Pohlner, Marion

    Shimia strain SK013 is an aerobic, Gram-negative, rod shaped alphaproteobacterium affiliated with the Roseobacter group within the family Rhodobacteraceae. The strain was isolated from surface sediment (0-1 cm) of the Skagerrak at 114 m below sea level. The 4,049,808 bp genome of Shimia str. SK013 comprises 3,981 protein-coding genes and 47 RNA genes. It contains one chromosome and no extrachromosomal elements. The genome analysis revealed the presence of genes for a dimethylsulfoniopropionate lyase, demethylase and the trimethylamine methyltransferase ( mttB) as well as genes for nitrate, nitrite and dimethyl sulfoxide reduction. This indicates that Shimia str. SK013 is able tomore » switch from aerobic to anaerobic metabolism and thus is capable of aerobic and anaerobic sulfur cycling at the seafloor. Among the ability to convert other sulfur compounds it has the genetic capacity to produce climatically active dimethyl sulfide. Growth on glutamate as a sole carbon source results in formation of cell-connecting filaments, a putative phenotypic adaptation of the surface-associated strain to the environmental conditions at the seafloor. Genome analysis revealed the presence of a flagellum ( fla1) and a type IV pilus biogenesis, which is speculated to be a prerequisite for biofilm formation. This is also related to genes responsible for signalling such as N-acyl homoserine lactones, as well as quip-genes responsible for quorum quenching and antibiotic biosynthesis. Pairwise similarities of 16S rRNA genes (98.56 % sequence similarity to the next relative S. haliotis) and the in silico DNA-DNA hybridization (21.20 % sequence similarity to S. haliotis) indicated Shimia str. SK013 to be considered as a new species. In conclusion, the genome analysis of Shimia str. SK013 offered first insights into specific physiological and phenotypic adaptation mechanisms of Roseobacter-affiliated bacteria to the benthic environment.« less

  1. Genome sequence of Shimia str. SK013, a representative of the Roseobacter group isolated from marine sediment

    DOE PAGES

    Kanukollu, Saranya; Voget, Sonja; Pohlner, Marion; ...

    2016-03-12

    Shimia strain SK013 is an aerobic, Gram-negative, rod shaped alphaproteobacterium affiliated with the Roseobacter group within the family Rhodobacteraceae. The strain was isolated from surface sediment (0-1 cm) of the Skagerrak at 114 m below sea level. The 4,049,808 bp genome of Shimia str. SK013 comprises 3,981 protein-coding genes and 47 RNA genes. It contains one chromosome and no extrachromosomal elements. The genome analysis revealed the presence of genes for a dimethylsulfoniopropionate lyase, demethylase and the trimethylamine methyltransferase ( mttB) as well as genes for nitrate, nitrite and dimethyl sulfoxide reduction. This indicates that Shimia str. SK013 is able tomore » switch from aerobic to anaerobic metabolism and thus is capable of aerobic and anaerobic sulfur cycling at the seafloor. Among the ability to convert other sulfur compounds it has the genetic capacity to produce climatically active dimethyl sulfide. Growth on glutamate as a sole carbon source results in formation of cell-connecting filaments, a putative phenotypic adaptation of the surface-associated strain to the environmental conditions at the seafloor. Genome analysis revealed the presence of a flagellum ( fla1) and a type IV pilus biogenesis, which is speculated to be a prerequisite for biofilm formation. This is also related to genes responsible for signalling such as N-acyl homoserine lactones, as well as quip-genes responsible for quorum quenching and antibiotic biosynthesis. Pairwise similarities of 16S rRNA genes (98.56 % sequence similarity to the next relative S. haliotis) and the in silico DNA-DNA hybridization (21.20 % sequence similarity to S. haliotis) indicated Shimia str. SK013 to be considered as a new species. In conclusion, the genome analysis of Shimia str. SK013 offered first insights into specific physiological and phenotypic adaptation mechanisms of Roseobacter-affiliated bacteria to the benthic environment.« less

  2. Optical spectroscopy of the blue supergiant Sk-69° 279 and its circumstellar shell with SALT

    NASA Astrophysics Data System (ADS)

    Gvaramadze, V. V.; Kniazev, A. Y.; Maryeva, O. V.; Berdnikov, L. N.

    2018-02-01

    We report the results of optical spectroscopy of the blue supergiant Sk-69° 279 and its circular shell in the Large Magellanic Cloud (LMC) with the Southern African Large Telescope (SALT). We classify Sk-69° 279 as an O9.2 Iaf star and analyse its spectrum by using the stellar atmosphere code CMFGEN, obtaining a stellar temperature of ≈30 kK, a luminosity of log (L*/ L⊙) = 5.54, a mass-loss rate of log (\\dot{M}/ M_{⊙} yr^{-1}) = -5.26, and a wind velocity of 800km s-1. We found also that Sk-69° 279 possesses an extended atmosphere with an effective temperature of ≈24 kK and that its surface helium and nitrogen abundances are enhanced, respectively, by factors of ≈2 and 20-30. This suggests that either Sk-69° 279 was initially a (single) fast-rotating ( ≳ 400 km s- 1) star, which only recently evolved off the main sequence, or that it is a product of close binary evolution. The long-slit spectroscopy of the shell around Sk-69° 279 revealed that its nitrogen abundance is enhanced by the same factor as the stellar atmosphere, which implies that the shell is composed mostly of the CNO processed material lost by the star. Our findings support previous propositions that some massive stars can produce compact circumstellar shells and, presumably, appear as luminous blue variables while they are still on the main sequence or have only recently left it.

  3. SkZpipe: A Python3 module to produce efficiently PSF-fitting photometry with DAOPHOT, and much more

    NASA Astrophysics Data System (ADS)

    Mauro, F.

    2017-07-01

    In an era characterized by big sky surveys and the availability of large amount of photometric data, it is important for astronomers to have tools to process their data in an efficient, accurate and easy way, minimizing reduction time. We present SkZpipe, a Python3 module designed mainly to process generic data, performing point-spread function (PSF) fitting photometry with the DAOPHOT suite (Stetson 1987). The software has already demonstrated its accuracy and efficiency with the adaptation VVV-SkZ_pipeline (Mauro et al. 2013) for the "VISTA Variables in the Vía Láctea" ESO survey, showing how it can replace the users, avoiding repetitive interaction in all the operations, retaining all of the benefits of the power and accuracy of the DAOPHOT suite, detaching them from the burden of data precessing. This software provides not only a pipeline, but also all the tools to run easily each atomic step of the photometric procedure, to match the results, and to retrieve information from fits headers and the internal instrumental database. We plan to add the support to other photometric softwares in the future.

  4. [The effect of modulators of SK channels on simple spike firing frequency in the discharge of the cerebellar Purkinje cells in laboratory mice].

    PubMed

    Egorova, P A; Karelina, T V; Vlasova, O L; Antonov, S M; Besprozvanny, I B

    2014-01-01

    The effect of CyPPA, a positive modulator of small conductance calcium-activated potassium channels of type 3 and 2 (SK3/SK2), and of NS309, an activator of intermediate and small conductance calcium-activated potassium channels (IK/SK), on the activity of cerebellar Purkinje cells was studied in 2-month-old male mice. The use of 1 mM of CyPPA has led to a decrease of simple spike firing frequency in the discharge of Purkinje cells by 25%, on average, during 1 h after application. At the same time, application of 100 μM of NS309 has promoted a decrease in simple spike firing frequency by 47 %, on average, during 1 h after the beginning of the action. The obtained results confirm the hypothesis that SK channels participate in regulation of simple spike firing frequency in the discharge of Purkinje cells and are responsible for restriction of signal frequency. The effect of NS309 on simple spike firing frequency was more pronounced; therefore, the IK/SK channels may be suggested to play the cardinal role in regulation of spike activity of Purkinje cells. Since increasing simple spike frequency in the discharge of Purkinje cells is observed at many disturbances of motor activity, in particular, at spinocerebellar ataxia, it can be suggested that the studied compounds or substances of similar action are of interest as potential medicinal agents.

  5. Soluble ephrin a1 is necessary for the growth of HeLa and SK-BR3 cells

    PubMed Central

    2010-01-01

    Background Ephrin A1 (EFNA1) is a member of the A-type ephrin family of cell surface proteins that function as ligands for the A-type Eph receptor tyrosine kinase family. In malignancy, the precise role of EFNA1 and its preferred receptor, EPHA2, is controversial. Several studies have found that EFNA1 may suppress EPHA2-mediated oncogenesis, or enhance it, depending on cell type and context. However, little is known about the conditions that influence whether EFNA1 promotes or suppresses tumorigenicity. EFNA1 exists in a soluble form as well as a glycophosphatidylinositol (GPI) membrane attached form. We investigated whether the contradictory roles of EFNA1 in malignancy might in part be related to the existence of both soluble and membrane attached forms of EFNA1 and potential differences in the manner in which they interact with EPHA2. Results Using a RNAi strategy to reduce the expression of endogenous EFNA1 and EPHA2, we found that both EFNA1 and EPHA2 are required for growth of HeLa and SK-BR3 cells. The growth defects could be rescued by conditioned media from cells overexpressing soluble EFNA1. Interestingly, we found that overexpression of the membrane attached form of EFNA1 suppresses growth of HeLa cells in 3D but not 2D. Knockdown of endogenous EFNA1, or overexpression of full-length EFNA1, resulted in relocalization of EPHA2 from the cell surface to sites of cell-cell contact. Overexpression of soluble EFNA1 however resulted in more EPHA2 distributed on the cell surface, away from cell-cell contacts, and promoted the growth of HeLa cells. Conclusions We conclude that soluble EFNA1 is necessary for the transformation of HeLa and SK-BR3 cells and participates in the relocalization of EPHA2 away from sites of cell-cell contact during transformation. PMID:20979646

  6. Rb-Raf-1 interaction disruptor RRD-251 induces apoptosis in metastatic melanoma cells and synergizes with dacarbazine

    PubMed Central

    Singh, Sandeep; Davis, Rebecca; Alamanda, Vignesh; Pireddu, Roberta; Pernazza, Daniel; Sebti, Said; Lawrence, Nicholas; Chellappan, Srikumar

    2010-01-01

    Metastatic melanoma is an aggressive cancer with very low response rate against conventional chemotherapeutic agents such as dacarbazine (DTIC). Inhibitor of Rb-Raf-1 interaction (RRD-251) was tested against the melanoma cell lines SK-MEL-28, SK-MEL-5 and SK-MEL-2. RRD-251 was found to be a potent inhibitor of melanoma cell proliferation, irrespective of V600E B-Raf mutation status of the cell lines. In a SK-MEL-28 xenograft experiment, RRD-251 exerted a significant suppression of tumor growth compared to vehicle (p=0.003). Similar to in vitro effects, tumors from RRD-251 treated animals showed decreased Rb-Raf-1 interaction in vivo. Growth suppressive effects of RRD-251 were associated with induction of apoptosis as well as a G1 arrest, with an accompanying decrease in S-phase cells. RRD-251 inhibited Rb phosphorylation, and downregulated E2F1 protein levels in these cells. Real-time PCR analysis showed that RRD-251 caused downregulation of cell cycle regulatory genes thymidylate synthase (TS) and cdc6 as well as anti-apoptotic gene Mcl-1. Combinatorial treatment of RRD-251 and DTIC resulted in a significantly higher apoptosis in DTIC resistant cell lines SK-MEL-28 and SK-MEL-5, as revealed by increased Caspase-3 activity and PARP cleavage. Since aberrant Rb/E2F pathway is associated with melanoma progression and resistance to apoptosis, these results suggest that the Rb-Raf-1 inhibitor could be an effective agent for melanoma treatment, either alone or in combination with DTIC. PMID:21139044

  7. Rb-Raf-1 interaction disruptor RRD-251 induces apoptosis in metastatic melanoma cells and synergizes with dacarbazine.

    PubMed

    Singh, Sandeep; Davis, Rebecca; Alamanda, Vignesh; Pireddu, Roberta; Pernazza, Daniel; Sebti, Said; Lawrence, Nicholas; Chellappan, Srikumar

    2010-12-01

    Metastatic melanoma is an aggressive cancer with very low response rate against conventional chemotherapeutic agents such as dacarbazine (DTIC). Inhibitor of Rb-Raf-1 interaction RRD-251 was tested against the melanoma cell lines SK-MEL-28, SK-MEL-5, and SK-MEL-2. RRD-251 was found to be a potent inhibitor of melanoma cell proliferation, irrespective of V600E B-Raf mutation status of the cell lines. In a SK-MEL-28 xenograft experiment, RRD-251 exerted a significant suppression of tumor growth compared with vehicle (P = 0.003). Similar to in vitro effects, tumors from RRD-251-treated animals showed decreased Rb-Raf-1 interaction in vivo. Growth suppressive effects of RRD-251 were associated with induction of apoptosis as well as a G(1) arrest, with an accompanying decrease in S-phase cells. RRD-251 inhibited Rb phosphorylation and downregulated E2F1 protein levels in these cells. Real-time PCR analysis showed that RRD-251 caused downregulation of cell-cycle regulatory genes thymidylate synthase (TS) and cdc6 as well as the antiapoptotic gene Mcl-1. Combinatorial treatment of RRD-251 and DTIC resulted in a significantly higher apoptosis in DTIC resistant cell lines SK-MEL-28 and SK-MEL-5, as revealed by increased caspase-3 activity and PARP cleavage. Because aberrant Rb/E2F pathway is associated with melanoma progression and resistance to apoptosis, these results suggest that the Rb-Raf-1 inhibitor could be an effective agent for melanoma treatment, either alone or in combination with DTIC. ©2010 AACR.

  8. Preliminary study on the L ate Cretaceous ostracods from continental scientific drilling SK1 in the Songliao Basin, NE China

    NASA Astrophysics Data System (ADS)

    Xi, Dangpeng; Qu, Haiying; Shi, Zhongye; Wan, Xiaoqiao

    2017-04-01

    Songliao Basin is one of the biggest lacustrine systems in Asia during Cretaceous age. Widespread deposits in the basin are mainly composed of clastic sediments which contain abundant fossils including gastropod, bivalves, ostracods, vertebrates and others. These well preserved ostracod fossils provide us valuable information about past climate changes and biotic responses in a greenhouse environment.The Cretaceous Continental Scientific Drilling in the Songliao Basin (SK1) offers a rare opportunity to study Late Cretaceous non-marine ostracod. The SK1 was drilled separately in two boreholes: the lower 959.55-meter-thick south core (SK1(s)), and the upper 1636.72-meter-thick north core (SK1 (n)), containing the Upper Quantou, Qingshankou, Yaojia, Nenjiang Formation, Sifangtai, Mingshui and lower Taikang formations. Here we establish high-resolution non-marine ostracod biostratigraphy based on SK1. 80 species belonging to 12 genera in the SK1(S) and 45 species assigned to 20 genera in the SK1(n) have been recovered. Nineteen ostracod assemblage zones have been recognized: 1. Mongolocypris longicaudata-Cypridea Assemblage Zone, 2.Triangulicypris torsuosus-Triangulicypris torsuosus. nota Assemblage Zone, 3. Cypridea dekhoinensis-Cypridea gibbosa Assemblage Zone, 4.Cypridea nota-Sunliavia tumida Assemblage Zone, 5.Cypridea edentula-Lycopterocypris grandis Assemblage Zone, 6.Cypridea fuyuensis-Triangulicypris symmetrica Assemblage Zone, 7.Triangulicypris vestilus-Triangulicypris fusiformis-Triangulicypris pumilis Assemblage Zone, 8.Cypridea panda-Mongolocypris obscura Assemblage Zone, 9. Cypridea exornata-Cypridea dongfangensis Assemblage Zone, 10.Cypridea favosa-Mongolocypris tabulata Assemblage Zone, 11.Cypridea formosa-Cypridea sunghuajiangensis Assemblage Zone, 12. Cypridea anonyma-Candona fabiforma Assemblage Zone, 13.Cypridea gracila-Cypridea gunsulinensis Assemblage Zone, 14.Mongolocypris magna-Mongolocypris heiluntszianensis Assemblage Zone, 15.Cypridea

  9. SK-N-MC cell death occurs by distinct molecular mechanisms in response to hydrogen peroxide and superoxide anions: involvements of JAK2-STAT3, JNK, and p38 MAP kinases pathways.

    PubMed

    Moslehi, Maryam; Yazdanparast, Razieh

    2013-07-01

    Oxidative stress plays a vital role in the pathogenesis of neurodegenerative diseases. Nerve cells are incessantly exposed to environmental stresses leading to overproduction of some harmful species like reactive oxygen species (ROS). ROS including hydrogen peroxide and superoxide anion are potent inducers of various signaling pathways encompassing MAPKs and JAK-STAT pathways. In the current study, we scrutinized the effects of hydrogen peroxide and/or menadione (superoxide anion generator) on JNK/p38-MAPKs and JAK2-STAT3 pathways to elucidate the mechanism(s) by which each oxidant modulated the above-mentioned pathways leading to SK-N-MC cell death. Our results delineated that hydrogen peroxide and superoxide anion radical induced distinct responses as we showed that STAT3 and p38 were activated in response to hydrogen peroxide, but not superoxide anion radicals indicating the specificity in ROS-induced signaling pathways activations and behaviors. We also observed that menadione induced JNK-dependent p53 expression and apoptotic death in SK-N-MC cells while H2O2-induced JNK activation was p53 independent. Thus, we declare that ROS type has a key role in selective instigation of JNK/p38-MAPKs and JAK2-STAT3 pathways in SK-N-MC cells. Identifying these differential behaviors and mechanisms of hydrogen peroxide and superoxide anion functions illuminates the possible therapeutic targets in the prevention or treatment of ROS-induced neurodegenerative diseases such as Alzheimer's disease.

  10. Production of coagulation factor VII in human cell lines Sk-Hep-1 and HKB-11.

    PubMed

    Corrêa de Freitas, Marcela Cristina; Bomfim, Aline de Sousa; Mizukami, Amanda; Picanço-Castro, Virgínia; Swiech, Kamilla; Covas, Dimas Tadeu

    2017-09-01

    Recombinant factor VII (rFVII) is the main therapeutic choice for hemophilia patients who have developed inhibitory antibodies against conventional treatments (FVIII and FIX). Because of the post-translational modifications, rFVII needs to be produced in mammalian cell lines. In this study, for the first time, we have shown efficient rFVII production in HepG2, Sk-Hep-1, and HKB-11 cell lines. Experiments in static conditions for a period of 96 h showed that HepG2-FVII produced the highest amounts of rhFVII, with an average of 1843 ng/mL. Sk-hep-1-FVII cells reached a maximum protein production of 1432 ng/mL and HKB-11-FVII cells reached 1468 ng/mL. Sk-Hep-1-rFVII and HKB-11-rFVII were selected for the first step of scale-up. Over 10 days of spinner flask culture, HKB-11 and SK-Hep-1 cells showed a cumulative production of rFVII of 152 μg and 202.6 μg in 50 mL, respectively. Thus, these human cell lines can be used for an efficient production of recombinant FVII. With more investment in basic research, human cell lines can be optimized for the commercial production of different bio therapeutic proteins. Copyright © 2017 Elsevier Inc. All rights reserved.

  11. Preliminary approach to elucidate the role of pigment as a binding site for drugs and chemicals in anagen hair: differential uptake of 3H-haloperidol by pigment-producing compared to non-pigment-producing cell lines.

    PubMed

    Pötsch, L; Emmerich, P; Skopp, G

    2002-02-01

    A striking difference was observed for cellular-bound drug in HaCaT and Sk-Mel-1 cells for a fixed drug exposure time of 72 h and varying 3H-haloperidol concentrations in the culture media. Drug uptake was dependent on drug concentration and linearly correlated for both the non-pigment- and the pigment-producing cells which however was different in magnitude. In an additional investigation the time course of drug uptake during 3H-haloperidol exposure (400 pmol/ml; 28 days) revealed increasing drug concentrations in the Sk-Mel-1 population, whereas drug concentrations in the keratinocytes reached a plateau within a short time period. In contrast to the HaCaT cells no tendency to saturation was observed for the pigment-producing cell line. At the end of the experiments 3H-haloperidol concentrations in Sk-Mel-1 were found to be approximately tenfold higher than in HaCaT.

  12. HSP-70 mitigates LPS/SKI-induced cell damage by increasing sphingosine kinase 1 (SK1).

    PubMed

    Ding, Xuan Z; Feng, Xiao R; Borschel, Richard H; Nikolich, Mikeljon P; Feng, Jie; Li, Yan S; Hoover, David L

    2010-06-01

    Heat shock proteins (HSPs) are potent protectors of cellular integrity against environmental stresses, including toxic microbial products. To investigate the mechanism of HSP-70 cell protection against bacterial lipopolysaccharide (LPS), we established a stable HSP-70 gene-transfected RAW 264.7 murine macrophage model of LPS-induced cell death. Bacterial LPS increases the activity of sphingosine kinase 1 (SK1), which catalyzes formation of sphingosine-1-phosphate (S1P). S1P functions as a critical signal for initiation and maintenance of diverse aspects of immune cell activation and function. When mouse macrophages were incubated with Escherichia coli LPS (1 microg/ml) and sphingosine kinase inhibitor (SKI, 5 microM), 90% of cells died. Neither LPS nor SKI alone at these doses damaged the cells. The LPS/SKI-induced cell death was partially reversed by overexpression of HSP-70 in gene-transfected macrophages. The specificity of HSP-70 in this reversal was demonstrated by transfection of HSP-70-specific siRNA. Down-regulation of HSP-70 expression after transfection of siRNA specific for HSP-70 was associated with increased LPS/SKI-induced cell damage. Overexpression of human or murine HSP-70 (HSPA1A and Hspa1a, respectively) increased both cellular SK1 mRNA and protein levels. Cellular heat shock also increased SK1 protein. These studies confirm the importance of SK1 as a protective moiety in LPS-induced cell injury and demonstrate that HSP-70-mediated protection from cells treated with LPS/SKI is accompanied by upregulating expression of SK1. HSP-70-mediated increases in SK1 and consequent increased levels of S1P may also play a role in protection of cells from other processes that lead to programmed cell death. Published by Elsevier Inc.

  13. Survivin knockdown increased anti-cancer effects of (-)-epigallocatechin-3-gallate in human malignant neuroblastoma SK-N-BE2 and SH-SY5Y cells.

    PubMed

    Hossain, Md Motarab; Banik, Naren L; Ray, Swapan K

    2012-08-01

    Neuroblastoma is a solid tumor that mostly occurs in children. Malignant neuroblastomas have poor prognosis because conventional chemotherapeutic agents are hardly effective. Survivin, which is highly expressed in some malignant neuroblastomas, plays a significant role in inhibiting differentiation and apoptosis and promoting cell proliferation, invasion, and angiogenesis. We examined consequences of survivin knockdown by survivin short hairpin RNA (shRNA) plasmid and then treatment with (-)-epigallocatechin-3-gallate (EGCG), a green tea flavonoid, in malignant neuroblastoma cells. Our Western blotting and laser scanning confocal immunofluorescence microscopy showed that survivin was highly expressed in malignant neuroblastoma SK-N-BE2 and SH-SY5Y cell lines and slightly in SK-N-DZ cell line. Expression of survivin was very faint in malignant neuroblastoma IMR32 cell line. We transfected SK-N-BE2 and SH-SY-5Y cells with survivin shRNA, treated with EGCG, and confirmed knockdown of survivin at mRNA and protein levels. Survivin knockdown induced morphological features of neuronal differentiation, as we observed following in situ methylene blue staining. Combination of survivin shRNA and EGCG promoted neuronal differentiation biochemically by increases in the expression of NFP, NSE, and e-cadherin and also decreases in the expression of Notch-1, ID2, hTERT, and PCNA. Our in situ Wright staining and Annexin V-FITC/PI staining showed that combination therapy was highly effective in inducing, respectively, morphological and biochemical features of apoptosis. Apoptosis occurred with activation of caspase-8 and cleavage of Bid to tBid, increase in Bax:Bcl-2 ratio, mitochondrial release of cytochrome c, and increases in the expression and activity of calpain and caspase-3. Combination therapy decreased migration of cells through matrigel and inhibited proliferative (p-Akt and NF-κB), invasive (MMP-2 and MMP-9), and angiogenic (VEGF and b-FGF) factors. Also, in vitro

  14. HEXIM1 is a promiscuous double-stranded RNA-binding protein and interacts with RNAs in addition to 7SK in cultured cells

    PubMed Central

    Li, Qintong; Cooper, Jeffrey J.; Altwerger, Gary H.; Feldkamp, Michael D.; Shea, Madeline A.; Price, David H.

    2007-01-01

    P-TEFb regulates eukaryotic gene expression at the level of transcription elongation, and is itself controlled by the reversible association of 7SK RNA and an RNA-binding protein HEXIM1 or HEXIM2. In an effort to determine the minimal region of 7SK needed to interact with HEXIM1 in vitro, we found that an oligo comprised of nucleotides 10–48 sufficed. A bid to further narrow down the minimal region of 7SK led to a surprising finding that HEXIM1 binds to double-stranded RNA in a sequence-independent manner. Both dsRNA and 7SK (10–48), but not dsDNA, competed efficiently with full-length 7SK for HEXIM1 binding in vitro. Upon binding dsRNA, a large conformational change was observed in HEXIM1 that allowed the recruitment and inhibition of P-TEFb. Both subcellular fractionation and immunofluorescence demonstrated that, while most HEXIM1 is found in the nucleus, a significant fraction is found in the cytoplasm. Immunoprecipitation experiments demonstrated that both nuclear and cytoplasmic HEXIM1 is associated with RNA. Interestingly, the one microRNA examined (mir-16) was found in HEXIM1 immunoprecipitates, while the small nuclear RNAs, U6 and U2, were not. Our study illuminates novel properties of HEXIM1 both in vitro and in vivo, and suggests that HEXIM1 may be involved in other nuclear and cytoplasmic processes besides controlling P-TEFb. PMID:17395637

  15. Mitochondria-targeted plastoquinone derivatives as tools to interrupt execution of the aging program. 4. Age-related eye disease. SkQ1 returns vision to blind animals.

    PubMed

    Neroev, V V; Archipova, M M; Bakeeva, L E; Fursova, A Zh; Grigorian, E N; Grishanova, A Yu; Iomdina, E N; Ivashchenko, Zh N; Katargina, L A; Khoroshilova-Maslova, I P; Kilina, O V; Kolosova, N G; Kopenkin, E P; Korshunov, S S; Kovaleva, N A; Novikova, Yu P; Philippov, P P; Pilipenko, D I; Robustova, O V; Saprunova, V B; Senin, I I; Skulachev, M V; Sotnikova, L F; Stefanova, N A; Tikhomirova, N K; Tsapenko, I V; Shchipanova, A I; Zinovkin, R A; Skulachev, V P

    2008-12-01

    Mitochondria-targeted cationic plastoquinone derivative SkQ1 (10-(6'-plastoquinonyl) decyltriphenylphosphonium) has been investigated as a potential tool for treating a number of ROS-related ocular diseases. In OXYS rats suffering from a ROS-induced progeria, very small amounts of SkQ1 (50 nmol/kg per day) added to food were found to prevent development of age-induced cataract and retinopathies of the eye, lipid peroxidation and protein carbonylation in skeletal muscles, as well as a decrease in bone mineralization. Instillation of drops of 250 nM SkQ1 reversed cataract and retinopathies in 3-12-month-old (but not in 24-month-old) OXYS rats. In rabbits, experimental uveitis and glaucoma were induced by immunization with arrestin and injections of hydroxypropyl methyl cellulose to the eye anterior sector, respectively. Uveitis was found to be prevented or reversed by instillation of 250 nM SkQ1 drops (four drops per day). Development of glaucoma was retarded by drops of 5 microM SkQ1 (one drop daily). SkQ1 was tested in veterinarian practice. A totally of 271 animals (dogs, cats, and horses) suffering from retinopathies, uveitis, conjunctivitis, and cornea diseases were treated with drops of 250 nM SkQ1. In 242 cases, positive therapeutic effect was obvious. Among animals suffering from retinopathies, 89 were blind. In 67 cases, vision returned after SkQ1 treatment. In ex vivo studies of cultivated posterior retina sector, it was found that 20 nM SkQ1 strongly decreased macrophagal transformation of the retinal pigmented epithelial cells, an effect which might explain some of the above SkQ1 activities. It is concluded that low concentrations of SkQ1 are promising in treating retinopathies, cataract, uveitis, glaucoma, and some other ocular diseases.

  16. SK3/TRPC1/Orai1 complex regulates SOCE-dependent colon cancer cell migration: a novel opportunity to modulate anti-EGFR mAb action by the alkyl-lipid Ohmline

    PubMed Central

    Guéguinou, Maxime; Harnois, Thomas; Crottes, David; Uguen, Arnaud; Deliot, Nadine; Gambade, Audrey; Chantôme, Aurélie; Haelters, Jean Pierre; Jaffrès, Paul Alain; Jourdan, Marie Lise; Weber, Günther; Soriani, Olivier; Bougnoux, Philippe; Mignen, Olivier; Bourmeyster, Nicolas; Constantin, Bruno; Lecomte, Thierry

    2016-01-01

    Background Barely 10-20% of patients with metastatic colorectal cancer (mCRC) receive a clinical benefit from the use of anti-EGFR monoclonal antibodies (mAbs). We hypothesized that this could depends on their efficiency to reduce Store Operated Calcium Entry (SOCE) that are known to enhance cancer cells. Results In the present study, we demonstrate that SOCE promotes migration of colon cancer cell following the formation of a lipid raft ion channel complex composed of TRPC1/Orai1 and SK3 channels. Formation of this complex is stimulated by the phosphorylation of the reticular protein STIM1 by EGF and activation of the Akt pathway. Our data show that, in a positive feedback loop SOCE activates both Akt pathway and SK3 channel activity which lead to SOCE amplification. This amplification occurs through the activation of Rac1/Calpain mediated by Akt. We also show that Anti-EGFR mAbs can modulate SOCE and cancer cell migration through the Akt pathway. Interestingly, the alkyl-lipid Ohmline, which we previously showed to be an inhibitor of SK3 channel, can dissociated the lipid raft ion channel complex through decreased phosphorylation of Akt and modulation of mAbs action. Conclusions This study demonstrates that the inhibition of the SOCE-dependent colon cancer cell migration trough SK3/TRPC1/Orai1 channel complex by the alkyl-lipid Ohmline may be a novel strategy to modulate Anti-EGFR mAb action in mCRC. PMID:27102434

  17. A High Molecular-Mass Anoxybacillus sp. SK3-4 Amylopullulanase: Characterization and Its Relationship in Carbohydrate Utilization

    PubMed Central

    Kahar, Ummirul Mukminin; Chan, Kok-Gan; Salleh, Madihah Md.; Hii, Siew Mee; Goh, Kian Mau

    2013-01-01

    An amylopullulanase of the thermophilic Anoxybacillus sp. SK3-4 (ApuASK) was purified to homogeneity and characterized. Though amylopullulanases larger than 200 kDa are rare, the molecular mass of purified ApuASK appears to be approximately 225 kDa, on both SDS-PAGE analyses and native-PAGE analyses. ApuASK was stable between pH 6.0 and pH 8.0 and exhibited optimal activity at pH 7.5. The optimal temperature for ApuASK enzyme activity was 60 °C, and it retained 54% of its total activity for 240 min at 65 °C. ApuASK reacts with pullulan, starch, glycogen, and dextrin, yielding glucose, maltose, and maltotriose. Interestingly, most of the previously described amylopullulanases are unable to produce glucose and maltose from these substrates. Thus, ApuASK is a novel, high molecular-mass amylopullulanase able to produce glucose, maltose, and maltotriose from pullulan and starch. Based on whole genome sequencing data, ApuASK appeared to be the largest protein present in Anoxybacillus sp. SK3-4. The α-amylase catalytic domain present in all of the amylase superfamily members is present in ApuASK, located between the cyclodextrin (CD)-pullulan-degrading N-terminus and the α-amylase catalytic C-terminus (amyC) domains. In addition, the existence of a S-layer homology (SLH) domain indicates that ApuASK might function as a cell-anchoring enzyme and be important for carbohydrate utilization in a streaming hot spring. PMID:23759984

  18. Development and preclinical studies of 64Cu-NOTA-pertuzumab F(ab′)2 for imaging changes in tumor HER2 expression associated with response to trastuzumab by PET/CT

    PubMed Central

    Lam, Karen; Chan, Conrad; Reilly, Raymond M.

    2017-01-01

    ABSTRACT We previously reported that microSPECT/CT imaging with 111In-labeled pertuzumab detected decreased HER2 expression in human breast cancer (BC) xenografts in athymic mice associated with response to treatment with trastuzumab (Herceptin). Our aim was to extend these results to PET/CT by constructing F(ab′)2 of pertuzumab modified with NOTA chelators for complexing 64Cu. The effect of the administered mass (5–200 µg) of 64Cu-NOTA-pertuzumab F(ab′)2 was studied in NOD/SCID mice engrafted with HER2-positive SK-OV-3 human ovarian cancer xenografts. Biodistribution studies were performed in non-tumor bearing Balb/c mice to predict radiation doses to normal organs in humans. Serial PET/CT imaging was conducted on mice engrafted with HER2-positive and trastuzumab-sensitive BT-474 or trastuzumab-insensitive SK-OV-3 xenografted mice treated with weekly doses of trastuzumab. There were no significant effects of the administered mass of 64Cu-NOTA-pertuzumab F(ab′)2 on tumor or normal tissue uptake. The predicted total body dose in humans was 0.015 mSv/MBq, a 3.3-fold reduction compared to 111In-labeled pertuzumab. MicroPET/CT images revealed specific tumor uptake of 64Cu-NOTA-pertuzumab F(ab′)2 at 24 or 48 h post-injection in mice with SK-OV-3 tumors. Image analysis of mice treated with trastuzumab showed 2-fold reduced uptake of 64Cu-NOTA-pertuzumab F(ab′)2 in BT-474 tumors after 1 week of trastuzumab normalized to baseline, and 1.9-fold increased uptake in SK-OV-3 tumors after 3 weeks of trastuzumab, consistent with tumor response and resistance, respectively. We conclude that PET/CT imaging with 64Cu-NOTA-pertuzumab F(ab′)2 detected changes in HER2 expression in response to trastuzumab while delivering a lower total body radiation dose compared to 111In-labeled pertuzumab. PMID:27813707

  19. Survivin knockdown increased anti-cancer effects of (−)-epigallocatechin-3-gallate in human malignant neuroblastoma SK-N- BE2 and SH-SY5Y cells

    PubMed Central

    Hossain, Md. Motarab; Banik, Naren L.; Ray, Swapan K.

    2012-01-01

    Neuroblastoma is a solid tumor that mostly occurs in children. Malignant neuroblastomas have poor prognosis because conventional chemotherapeutic agents are hardly effective. Survivin, which is highly expressed in some malignant neuroblastomas, plays a significant role in inhibiting differentiation and apoptosis and promoting cell proliferation, invasion, and angiogenesis. We examined consequences of survivin knockdown by survivin short hairpin RNA (shRNA) plasmid and then treatment with (−)-epigallocatechin-3-gallate (EGCG), a green tea flavonoid, in malignant neuroblastoma cells. Our Western blotting and laser scanning confocal immunofluorescence microscopy showed that survivin was highly expressed in malignant neuroblastoma SK-N-BE2 and SH-SY5Y cell lines and slightly in SK-N-DZ cell line. Expression of survivin was very faint in malignant neuroblastoma IMR32 cell line. We transfected SK-N-BE2 and SH-SY-5Y cells with survivin shRNA, treated with EGCG, and confirmed knockdown of survivin at mRNA and protein levels. Survivin knockdown induced morphological features of neuronal differentiation, as we observed following in situ methylene blue staining. Combination of survivin shRNA and EGCG promoted neuronal differentiation biochemically by increases in expression of NFP, NSE, and e-cadherin and also decreases in expression of Notch-1, ID2, hTERT, and PCNA. Our in situ Wright staining and Annexin V-FITC/PI staining showed that combination therapy was highly effective in inducing, respectively, morphological and biochemical features of apoptosis. Apoptosis occurred with activation of caspase-8 and cleavage of Bid to tBid, increase in Bax:Bcl-2 ratio, mitochondrial release of cytochrome c, and increases in expression and activity of calpain and caspase-3. Combination therapy decreased migration of cells through matrigel and inhibited proliferative (p-Akt and NF-κB), invasive (MMP-2 and MMP-9), and angiogenic (VEGF and b-FGF) factors. Also, in vitro network

  20. High-quality draft genome sequence of Effusibacillus lacus strain skLN1T, facultative anaerobic spore-former isolated from freshwater lake sediment.

    PubMed

    Watanabe, Miho; Tokizawa, Riho; Kojima, Hisaya; Fukui, Manabu

    2017-01-01

    10.1601/nm.25721 strain skLN1 T is the type strain of the type species in the genus 10.1601/nm.25720 which is the one of the genera in the family 10.1601/nm.5070 within the phylum 10.1601/nm.3874. 10.1601/nm.25721 strain skLN1 T is a Gram-positive, spore-forming thermophilic neutrophile isolated from freshwater lake sediment. Here, we present the draft genome sequence of strain skLN1 T , which consists of 3,902,380 bp with a G + C content of 50.38%.

  1. Arsenic trioxide induces cell cycle arrest and affects Trk receptor expression in human neuroblastoma SK-N-SH cells.

    PubMed

    Xiong, Xilin; Li, Yang; Liu, Ling; Qi, Kai; Zhang, Chi; Chen, Yueqin; Fang, Jianpei

    2018-06-13

    Arsenic trioxide (As 2 O 3 ), a drug that has been used in China for approximately two thousand years, induces cell death in a variety of cancer cell types, including neuroblastoma (NB). The tyrosine kinase receptor (Trk) family comprises three members, namely TrkA, TrkB and TrkC. Various studies have confirmed that TrkA and TrkC expression is associated with a good prognosis in NB, while TrkB overexpression can lead to tumor cell growth and invasive metastasis. Previous studies have shown that As 2 O 3 can inhibit the growth and proliferation of a human NB cell line and can also affect the N-Myc mRNA expression. It remains unclear whether As 2 O 3 regulates Trks for the purposes of treating NB. The aim of the present study was to investigate the effect of As 2 O 3 on Trk expression in NB cell lines and its potential therapeutic efficacy. SK-N-SH cells were grown with increasing doses of As 2 O 3 at different time points. We cultured SK-N-SH cells, which were treated with increasing doses of As 2 O 3 at different time points. Trk expression in the NB samples was quantified by immunohistochemistry, and the cell cycle was analyzed by flow cytometry. TrkA, TrkB and TrkC mRNA expression was evaluated by real-time PCR analysis. Immunohistochemical and real-time PCR analyses indicated that TrkA and TrkC were over-expressed in NB, and specifically during stages 1, 2 and 4S of the disease progression. TrkB expression was increased in stage 3 and 4 NB. As 2 O 3 significantly arrested SK-N-SH cells in the G2/M phase. In addition, TrkA, TrkB and TrkC expression levels were significantly upregulated by higher concentrations of As 2 O 3 treatment, notably in the 48-h treatment period. Our findings suggested that to achieve the maximum effect and appropriate regulation of Trk expression in NB stages 1, 2 and 4S, As 2 O 3 treatment should be at relatively higher concentrations for longer delivery times;however, for NB stages 3 and 4, an appropriate concentration and infusion time

  2. LRP4 third β-propeller domain mutations cause novel congenital myasthenia by compromising agrin-mediated MuSK signaling in a position-specific manner

    PubMed Central

    Ohkawara, Bisei; Cabrera-Serrano, Macarena; Nakata, Tomohiko; Milone, Margherita; Asai, Nobuyuki; Ito, Kenyu; Ito, Mikako; Masuda, Akio; Ito, Yasutomo; Engel, Andrew G.; Ohno, Kinji

    2014-01-01

    Congenital myasthenic syndromes (CMS) are heterogeneous disorders in which the safety margin of neuromuscular transmission is compromised by one or more specific mechanisms. Using Sanger and exome sequencing in a CMS patient, we identified two heteroallelic mutations, p.Glu1233Lys and p.Arg1277His, in LRP4 coding for the postsynaptic low-density lipoprotein receptor-related protein 4. LRP4, expressed on the surface of the postsynaptic membrane of the neuromuscular junction, is a receptor for neurally secreted agrin, and LRP4 bound by agrin activates MuSK. Activated MuSK in concert with Dok-7 stimulates rapsyn to concentrate and anchor AChR on the postsynaptic membrane and interacts with other proteins implicated in the assembly and maintenance of the neuromuscular junction. LRP4 also functions as an inhibitor of Wnt/beta-catenin signaling. The identified mutations in LRP4 are located at the edge of its 3rd beta-propeller domain and decrease binding affinity of LRP4 for both MuSK and agrin. Mutations in the LRP4 3rd beta-propeller domain were previously reported to impair Wnt signaling and cause bone diseases including Cenani–Lenz syndactyly syndrome and sclerosteosis-2. By analyzing naturally occurring and artificially introduced mutations in the LRP4 3rd beta-propeller domain, we show that the edge of the domain regulates the MuSK signaling whereas its central cavity governs Wnt signaling. We conclude that LRP4 is a new CMS disease gene and that the 3rd beta propeller domain of LRP4 mediates the two signaling pathways in a position-specific manner. PMID:24234652

  3. Mitochondria-targeted antioxidant SkQ1 reduces age-related alterations in the ultrastructure of the lacrimal gland

    PubMed Central

    Bakeeva, Lora E.; Eldarov, Chupalav M.; Vangely, Irina M.; Kolosova, Nataliya G.; Vays, Valeriya B.

    2016-01-01

    Dry eye syndrome is an eye disorder affecting many people at an old age. Because dry eye syndrome is accelerated by aging, a useful approach to the prevention of this syndrome may be an intervention into the aging process. Previously, we showed that the mitochondria-targeted antioxidant SkQ1 delays manifestations of aging and inhibits the development of age-related diseases including dry eye syndrome. Nevertheless, the link between SkQ1's effects and its suppression of age-related changes in the lacrimal gland remains unclear. Here we demonstrated that dietary supplementation with SkQ1 (250 nmol/[kg body weight] daily) starting at age 1.5 months significantly alleviated the pathological changes in lacrimal glands of Wistar rats by age 24 months. By this age, lacrimal glands underwent dramatic deterioration of the ultrastructure that was indicative of irreversible disturbances in these glands' functioning. In contrast, in SkQ1-treated rats, the ultrastructure of the lacrimal gland was similar to that in much younger rats. Morphometric analysis of electron-microscopic specimens of lacrimal glands revealed the presence of numerous secretory granules in acinar cells and a significant increase in the number of operating intercalary ducts. Our results confirm that dietary supplementation with SkQ1 is a promising approach to healthy ageing and to prevention of aberrations in the lacrimal gland that underlie dry eye syndrome. PMID:27852065

  4. Exogenous melatonin administration modifies cutaneous vasoconstrictor response to whole body skin cooling in humans.

    PubMed

    Aoki, Ken; Zhao, Kun; Yamazaki, Fumio; Sone, Ryoko; Alvarez, Guy E; Kosiba, Wojciech A; Johnson, John M

    2008-03-01

    Humans and other diurnal species experience a fall in internal temperature (T(int)) at night, accompanied by increased melatonin and altered thermoregulatory control of skin blood flow (SkBF). Also, exogenous melatonin induces a fall in T(int), an increase in distal skin temperatures and altered control of the cutaneous active vasodilator system, suggesting an effect of melatonin on the control of SkBF. To test whether exogenous melatonin also affects the more tonically active vasoconstrictor system in glabrous and nonglabrous skin during cooling, healthy males (n = 9) underwent afternoon sessions of whole body skin temperature (T(sk)) cooling (water-perfused suits) after oral melatonin (Mel; 3 mg) or placebo (Cont). Cutaneous vascular conductance (CVC) was calculated from SkBF (laser Doppler flowmetry) and non-invasive blood pressure. Baseline T(int) was lower in Mel than in Cont (P < 0.01). During progressive reduction of T(sk) from 35 degrees C to 32 degrees C, forearm CVC was first significantly reduced at T(sk) of 34.33 +/- 0.01 degrees C (P < 0.05) in Cont. In contrast, CVC in Mel was not significantly reduced until T(sk) reached 33.33 +/- 0.01 degrees C (P < 0.01). The decrease in forearm CVC in Mel was significantly less than in Cont at T(sk) of 32.66 +/- 0.01 degrees C and lower (P < 0.05). In Mel, palmar CVC was significantly higher than in Cont above T(sk) of 33.33 +/- 0.01 degrees C, but not below. Thus exogenous melatonin blunts reflex vasoconstriction in nonglabrous skin and shifts vasoconstrictor system control to lower T(int). It provokes vasodilation in glabrous skin but does not suppress the sensitivity to falling T(sk). These findings suggest that by affecting the vasoconstrictor system, melatonin has a causal role in the nocturnal changes in body temperature and its control.

  5. Comparative Phosphoproteomics Reveals the Role of AmpC β-lactamase Phosphorylation in the Clinical Imipenem-resistant Strain Acinetobacter baumannii SK17*

    PubMed Central

    Lai, Juo-Hsin; Yang, Jhih-Tian; Chern, Jeffy; Chen, Te-Li; Wu, Wan-Ling; Liao, Jiahn-Haur; Tsai, Shih-Feng; Liang, Suh-Yuen; Chou, Chi-Chi

    2016-01-01

    Nosocomial infectious outbreaks caused by multidrug-resistant Acinetobacter baumannii have emerged as a serious threat to human health. Phosphoproteomics of pathogenic bacteria has been used to identify the mechanisms of bacterial virulence and antimicrobial resistance. In this study, we used a shotgun strategy combined with high-accuracy mass spectrometry to analyze the phosphoproteomics of the imipenem-susceptible strain SK17-S and -resistant strain SK17-R. We identified 410 phosphosites on 248 unique phosphoproteins in SK17-S and 285 phosphosites on 211 unique phosphoproteins in SK17-R. The distributions of the Ser/Thr/Tyr/Asp/His phosphosites in SK17-S and SK17-R were 47.0%/27.6%/12.4%/8.0%/4.9% versus 41.4%/29.5%/17.5%/6.7%/4.9%, respectively. The Ser-90 phosphosite, located on the catalytic motif S88VS90K of the AmpC β-lactamase, was first identified in SK17-S. Based on site-directed mutagenesis, the nonphosphorylatable mutant S90A was found to be more resistant to imipenem, whereas the phosphorylation-simulated mutant S90D was sensitive to imipenem. Additionally, the S90A mutant protein exhibited higher β-lactamase activity and conferred greater bacterial protection against imipenem in SK17-S compared with the wild-type. In sum, our results revealed that in A. baumannii, Ser-90 phosphorylation of AmpC negatively regulates both β-lactamase activity and the ability to counteract the antibiotic effects of imipenem. These findings highlight the impact of phosphorylation-mediated regulation in antibiotic-resistant bacteria on future drug design and new therapies. PMID:26499836

  6. A Long-Gap Peripheral Nerve Injury Therapy Using Human Skeletal Muscle-Derived Stem Cells (Sk-SCs): An Achievement of Significant Morphological, Numerical and Functional Recovery

    PubMed Central

    Hirata, Maki; Nakajima, Nobuyuki; Saito, Kosuke; Hashimoto, Hiroyuki; Soeda, Shuichi; Uchiyama, Yoshiyasu; Watanabe, Masahiko

    2016-01-01

    Losses in vital functions of the somatic motor and sensory nervous system are induced by severe long-gap peripheral nerve transection injury. In such cases, autologous nerve grafts are the gold standard treatment, despite the unavoidable sacrifice of other healthy functions, whereas the prognosis is not always favorable. Here, we use human skeletal muscle-derived stem cells (Sk-SCs) to reconstitute the function after long nerve-gap injury. Muscles samples were obtained from the amputated legs from 9 patients following unforeseen accidents. The Sk-SCs were isolated using conditioned collagenase solution, and sorted as CD34+/45- (Sk-34) and CD34-/45-/29+ (Sk-DN/29+) cells. Cells were separately cultured/expanded under optimal conditions for 2 weeks, then injected into the athymic nude mice sciatic nerve long-gap model (7-mm) bridging an acellular conduit. After 8–12 weeks, active cell engraftment was observed only in the Sk-34 cell transplanted group, showing preferential differentiation into Schwann cells and perineurial/endoneurial cells, as well as formation of the myelin sheath and perineurium/endoneurium surrounding regenerated axons, resulted in 87% of numerical recovery. Differentiation into vascular cell lineage (pericyte and endothelial cells) were also observed. A significant tetanic tension recovery (over 90%) of downstream muscles following electrical stimulation of the sciatic nerve (at upper portion of the gap) was also achieved. In contrast, Sk-DN/29+ cells were completely eliminated during the first 4 weeks, but relatively higher numerical (83% vs. 41% in axon) and functional (80% vs. 60% in tetanus) recovery than control were observed. Noteworthy, significant increase in the formation of vascular networks in the conduit during the early stage (first 2 weeks) of recovery was observed in both groups with the expression of key factors (mRNA and protein levels), suggesting the paracrine effects to angiogenesis. These results suggested that the human Sk

  7. A Long-Gap Peripheral Nerve Injury Therapy Using Human Skeletal Muscle-Derived Stem Cells (Sk-SCs): An Achievement of Significant Morphological, Numerical and Functional Recovery.

    PubMed

    Tamaki, Tetsuro; Hirata, Maki; Nakajima, Nobuyuki; Saito, Kosuke; Hashimoto, Hiroyuki; Soeda, Shuichi; Uchiyama, Yoshiyasu; Watanabe, Masahiko

    2016-01-01

    Losses in vital functions of the somatic motor and sensory nervous system are induced by severe long-gap peripheral nerve transection injury. In such cases, autologous nerve grafts are the gold standard treatment, despite the unavoidable sacrifice of other healthy functions, whereas the prognosis is not always favorable. Here, we use human skeletal muscle-derived stem cells (Sk-SCs) to reconstitute the function after long nerve-gap injury. Muscles samples were obtained from the amputated legs from 9 patients following unforeseen accidents. The Sk-SCs were isolated using conditioned collagenase solution, and sorted as CD34+/45- (Sk-34) and CD34-/45-/29+ (Sk-DN/29+) cells. Cells were separately cultured/expanded under optimal conditions for 2 weeks, then injected into the athymic nude mice sciatic nerve long-gap model (7-mm) bridging an acellular conduit. After 8-12 weeks, active cell engraftment was observed only in the Sk-34 cell transplanted group, showing preferential differentiation into Schwann cells and perineurial/endoneurial cells, as well as formation of the myelin sheath and perineurium/endoneurium surrounding regenerated axons, resulted in 87% of numerical recovery. Differentiation into vascular cell lineage (pericyte and endothelial cells) were also observed. A significant tetanic tension recovery (over 90%) of downstream muscles following electrical stimulation of the sciatic nerve (at upper portion of the gap) was also achieved. In contrast, Sk-DN/29+ cells were completely eliminated during the first 4 weeks, but relatively higher numerical (83% vs. 41% in axon) and functional (80% vs. 60% in tetanus) recovery than control were observed. Noteworthy, significant increase in the formation of vascular networks in the conduit during the early stage (first 2 weeks) of recovery was observed in both groups with the expression of key factors (mRNA and protein levels), suggesting the paracrine effects to angiogenesis. These results suggested that the human Sk

  8. Comparative Phosphoproteomics Reveals the Role of AmpC β-lactamase Phosphorylation in the Clinical Imipenem-resistant Strain Acinetobacter baumannii SK17.

    PubMed

    Lai, Juo-Hsin; Yang, Jhih-Tian; Chern, Jeffy; Chen, Te-Li; Wu, Wan-Ling; Liao, Jiahn-Haur; Tsai, Shih-Feng; Liang, Suh-Yuen; Chou, Chi-Chi; Wu, Shih-Hsiung

    2016-01-01

    Nosocomial infectious outbreaks caused by multidrug-resistant Acinetobacter baumannii have emerged as a serious threat to human health. Phosphoproteomics of pathogenic bacteria has been used to identify the mechanisms of bacterial virulence and antimicrobial resistance. In this study, we used a shotgun strategy combined with high-accuracy mass spectrometry to analyze the phosphoproteomics of the imipenem-susceptible strain SK17-S and -resistant strain SK17-R. We identified 410 phosphosites on 248 unique phosphoproteins in SK17-S and 285 phosphosites on 211 unique phosphoproteins in SK17-R. The distributions of the Ser/Thr/Tyr/Asp/His phosphosites in SK17-S and SK17-R were 47.0%/27.6%/12.4%/8.0%/4.9% versus 41.4%/29.5%/17.5%/6.7%/4.9%, respectively. The Ser-90 phosphosite, located on the catalytic motif S(88)VS(90)K of the AmpC β-lactamase, was first identified in SK17-S. Based on site-directed mutagenesis, the nonphosphorylatable mutant S90A was found to be more resistant to imipenem, whereas the phosphorylation-simulated mutant S90D was sensitive to imipenem. Additionally, the S90A mutant protein exhibited higher β-lactamase activity and conferred greater bacterial protection against imipenem in SK17-S compared with the wild-type. In sum, our results revealed that in A. baumannii, Ser-90 phosphorylation of AmpC negatively regulates both β-lactamase activity and the ability to counteract the antibiotic effects of imipenem. These findings highlight the impact of phosphorylation-mediated regulation in antibiotic-resistant bacteria on future drug design and new therapies. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

  9. Role of degassing of the Noril’sk nickel deposits in the Permian–Triassic mass extinction event

    PubMed Central

    Barnes, Stephen J.; Mungall, James E.

    2017-01-01

    The largest mass extinction event in Earth's history marks the boundary between the Permian and Triassic Periods at circa 252 Ma and has been linked with the eruption of the basaltic Siberian Traps large igneous province (SLIP). One of the kill mechanisms that has been suggested is a biogenic methane burst triggered by the release of vast amounts of nickel into the atmosphere. A proposed Ni source lies within the huge Noril’sk nickel ore deposits, which formed in magmatic conduits widely believed to have fed the eruption of the SLIP basalts. However, nickel is a nonvolatile element, assumed to be largely sequestered at depth in dense sulfide liquids that formed the orebodies, preventing its release into the atmosphere and oceans. Flotation of sulfide liquid droplets by surface attachment to gas bubbles has been suggested as a mechanism to overcome this problem and allow introduction of Ni into the atmosphere during eruption of the SLIP lavas. Here we use 2D and 3D X-ray imagery on Noril’sk nickel sulfide, combined with simple thermodynamic models, to show that the Noril’sk ores were degassing while they were forming. Consequent “bubble riding” by sulfide droplets, followed by degassing of the shallow, sulfide-saturated, and exceptionally volatile and Cl-rich SLIP lavas, permitted a massive release of nickel-rich volcanic gas and subsequent global dispersal of nickel released from this gas as aerosol particles. PMID:28223492

  10. A Model for Spheroid versus Monolayer Response of SK-N-SH Neuroblastoma Cells to Treatment with 15-Deoxy-PGJ 2

    PubMed Central

    Dunham, Ann; Chen, Paula X.; Chen, Michelle; Huynh, Milan; Rheingold, Evan; Prosper, Olivia

    2016-01-01

    Researchers have observed that response of tumor cells to treatment varies depending on whether the cells are grown in monolayer, as in vitro spheroids or in vivo. This study uses data from the literature on monolayer treatment of SK-N-SH neuroblastoma cells with 15-deoxy-PGJ 2 and couples it with data on growth rates for untreated SK-N-SH neuroblastoma cells grown as multicellular spheroids. A linear model is constructed for untreated and treated monolayer data sets, which is tuned to growth, death, and cell cycle data for the monolayer case for both control and treatment with 15-deoxy-PGJ 2. The monolayer model is extended to a five-dimensional nonlinear model of in vitro tumor spheroid growth and treatment that includes compartments of the cell cycle (G 1, S, G 2/M) as well as quiescent (Q) and necrotic (N) cells. Monolayer treatment data for 15-deoxy-PGJ 2 is used to derive a prediction of spheroid response under similar treatments. For short periods of treatment, spheroid response is less pronounced than monolayer response. The simulations suggest that the difference in response to treatment of monolayer versus spheroid cultures observed in laboratory studies is a natural consequence of tumor spheroid physiology rather than any special resistance to treatment. PMID:28044089

  11. Neuroglobin overexpression plays a pivotal role in neuroprotection through mitochondrial raft-like microdomains in neuroblastoma SK-N-BE2 cells.

    PubMed

    Garofalo, Tina; Ferri, Alberto; Sorice, Maurizio; Azmoon, Pardis; Grasso, Maria; Mattei, Vincenzo; Capozzi, Antonella; Manganelli, Valeria; Misasi, Roberta

    2018-04-01

    Since stressing conditions induce a relocalization of endogenous human neuroglobin (NGB) to mitochondria, this research is aimed to evaluate the protective role of NGB overexpression against neurotoxic stimuli, through mitochondrial lipid raft-associated complexes. To this purpose, we built a neuronal model of oxidative stress by the use of human dopaminergic neuroblastoma cells, SK-N-BE2, stably overexpressing NGB by transfection and treated with 1-methyl-4-phenylpyridinium ion (MPP+). We preliminary observed the redistribution of NGB to mitochondria following MPP+ treatment. The analysis of mitochondrial raft-like microdomains revealed that, following MPP+ treatment, NGB translocated to raft fractions (Triton X-100-insoluble), where it interacts with ganglioside GD3. Interestingly, the administration of agents capable of perturbating microdomain before MPP+ treatment, significantly affected viability in SK-N-BE2-NGB cells. The overexpression of NGB was able to abrogate the mitochondrial injuries on complex IV activity or mitochondrial morphology induced by MPP+ administration. The protective action of NGB on mitochondria only takes place if the mitochondrial lipid(s) rafts-like microdomains are intact, indeed NGB fails to protect complex IV activity when purified mitochondria were treated with the lipid rafts disruptor methyl-β-cyclodextrin. Thus, our unique in vitro model of stably transfected cells overexpressing endogenous NGB allowed us to suggest that the role in neuroprotection played by NGB is reliable only through interaction with mitochondrial lipid raft-associated complexes. Copyright © 2018 Elsevier Inc. All rights reserved.

  12. Survivin knockdown increased anti-cancer effects of (-)-epigallocatechin-3-gallate in human malignant neuroblastoma SK-N-BE2 and SH-SY5Y cells

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Hossain, Md. Motarab; Banik, Naren L.; Ray, Swapan K., E-mail: swapan.ray@uscmed.sc.edu

    Neuroblastoma is a solid tumor that mostly occurs in children. Malignant neuroblastomas have poor prognosis because conventional chemotherapeutic agents are hardly effective. Survivin, which is highly expressed in some malignant neuroblastomas, plays a significant role in inhibiting differentiation and apoptosis and promoting cell proliferation, invasion, and angiogenesis. We examined consequences of survivin knockdown by survivin short hairpin RNA (shRNA) plasmid and then treatment with (-)-epigallocatechin-3-gallate (EGCG), a green tea flavonoid, in malignant neuroblastoma cells. Our Western blotting and laser scanning confocal immunofluorescence microscopy showed that survivin was highly expressed in malignant neuroblastoma SK-N-BE2 and SH-SY5Y cell lines and slightly inmore » SK-N-DZ cell line. Expression of survivin was very faint in malignant neuroblastoma IMR32 cell line. We transfected SK-N-BE2 and SH-SY-5Y cells with survivin shRNA, treated with EGCG, and confirmed knockdown of survivin at mRNA and protein levels. Survivin knockdown induced morphological features of neuronal differentiation, as we observed following in situ methylene blue staining. Combination of survivin shRNA and EGCG promoted neuronal differentiation biochemically by increases in the expression of NFP, NSE, and e-cadherin and also decreases in the expression of Notch-1, ID2, hTERT, and PCNA. Our in situ Wright staining and Annexin V-FITC/PI staining showed that combination therapy was highly effective in inducing, respectively, morphological and biochemical features of apoptosis. Apoptosis occurred with activation of caspase-8 and cleavage of Bid to tBid, increase in Bax:Bcl-2 ratio, mitochondrial release of cytochrome c, and increases in the expression and activity of calpain and caspase-3. Combination therapy decreased migration of cells through matrigel and inhibited proliferative (p-Akt and NF-{kappa}B), invasive (MMP-2 and MMP-9), and angiogenic (VEGF and b-FGF) factors. Also, in

  13. Arabidopsis mutant sk156 reveals complex regulation of SPL15 in a miR156-controlled gene network.

    PubMed

    Wei, Shu; Gruber, Margaret Y; Yu, Bianyun; Gao, Ming-Jun; Khachatourians, George G; Hegedus, Dwayne D; Parkin, Isobel A P; Hannoufa, Abdelali

    2012-09-18

    The Arabidopsis microRNA156 (miR156) regulates 11 members of the SQUAMOSA PROMOTER BINDING PROTEIN LIKE (SPL) family by base pairing to complementary target mRNAs. Each SPL gene further regulates a set of other genes; thus, miR156 controls numerous genes through a complex gene regulation network. Increased axillary branching occurs in transgenic Arabidopsis overexpressing miR156b, similar to that observed in loss-of-function max3 and max4 mutants with lesions in carotenoid cleavage dioxygenases. Arabidopsis miR156b was found to enhance carotenoid levels and reproductive shoot branching when expressed in Brassica napus, suggesting a link between miR156b expression and carotenoid metabolism. However, details of the miR156 regulatory network of SPL genes related to carotenoid metabolism are not known. In this study, an Arabidopsis T-DNA enhancer mutant, sk156, was identified due to its altered branching and trichome morphology and increased seed carotenoid levels compared to wild type (WT) ecovar Columbia. Enhanced miR156b expression due to the 35S enhancers present on the T-DNA insert was responsible for these phenotypes. Constitutive and leaf primodium-specific expression of a miR156-insensitive (mutated) SPL15 (SPL15m) largely restored WT seed carotenoid levels and plant morphology when expressed in sk156. The Arabidopsis native miR156-sensitive SPL15 (SPL15n) and SPL15m driven by a native SPL15 promoter did not restore the WT phenotype in sk156. Our findings suggest that SPL15 function is somewhat redundant with other SPL family members, which collectively affect plant phenotypes. Moreover, substantially decreased miR156b transcript levels in sk156 expressing SPL15m, together with the presence of multiple repeats of SPL-binding GTAC core sequence close to the miR156b transcription start site, suggested feedback regulation of miR156b expression by SPL15. This was supported by the demonstration of specific in vitro interaction between DNA-binding SBP domain of SPL15

  14. Croton lechleri sap and isolated alkaloid taspine exhibit inhibition against human melanoma SK23 and colon cancer HT29 cell lines.

    PubMed

    Montopoli, Monica; Bertin, Riccardo; Chen, Zheng; Bolcato, Jenny; Caparrotta, Laura; Froldi, Guglielmina

    2012-12-18

    Croton lechleri Mull. Arg. (Euphorbiaceae) is a traditional medicinal plant which produces a red sap, traditionally known as "Sangre de Drago"; it is used in folk medicine externally for wounds, fractures, and haemorrhoids, internally for intestinal and stomach ulcers and also for the empirical cure of cancers. We investigated the effects of Croton lechleri sap and taspine in comparison with taxol and vinblastine on the growth of human cancer cell lines of SK23 (melanoma), LoVo and HT29 (colorectal cancer) using MTT and Trypan blue assays. Further, we studied cell cycle by flow cytometry and detected acetylated-α-tubulin by confocal microscope. Croton lechleri inhibited cell proliferation starting from 1 μg/mL in SK23 cells, whereas 10 times higher concentrations were required for growth inhibition of HT-29 and LoVo cell lines. Also taspine (0.1 μg/mL) inhibited the SK23 and HT29 cell proliferation. Further, assay was assessed on SK23 and HT29 cell lines with 24-48 h treatment with sap and taspine. Both sap and taspine inhibited cancer cell proliferation; taspine showed higher activity on SK23 cells, which was significantly increased after 48 h of SK23 treatment. Using confocal microscopy we observed that Croton lechleri (1 μg/mL) caused a loss of microtubule structure, whereas taspine (0.5 μg/mL) caused an increase in acetylated α-tubulin and a modification of cellular morphology, mainly in SK23 cells. Croton lechleri sap 10 and 50 μg/mL influence cell cycle; 50 μg/mL sap caused a dramatic reduction of cells in G(1)/G(0) and S phases with a great increase of subG(0) cells. The data showed that Croton lechleri and taspine could inhibit cell proliferation with higher potency against melanoma SK23 cells, supporting the empirical use of the sap as anticancer in ethnomedicine and taspine as a possible anticancer agent. Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.

  15. Anti-MuSK myasthenia gravis presenting with Epstein-Barr virus-associated mononucleosis and immune-mediated diabetes mellitus.

    PubMed

    Bhibhatbhan, Arunee; Kline, Gregory; Vincent, Angela; Toth, Cory

    2007-08-01

    We report a young woman with the abrupt onset of infectious mononucleosis due to Epstein-Barr virus associated with a subsequent autoimmune form of diabetes mellitus and myasthenia gravis with anti-muscle-specific kinase (MuSK) antibodies. The simultaneous onset of these two autoimmune diseases preceded by a systemic viral illness supports a causal relationship between MuSK antibodies and myasthenia gravis and suggests the possibility of a viral trigger in some cases.

  16. Intracellular synthesis of glutamic acid in Bacillus methylotrophicus SK19.001, a glutamate-independent poly(γ-glutamic acid)-producing strain.

    PubMed

    Peng, Yingyun; Zhang, Tao; Mu, Wanmeng; Miao, Ming; Jiang, Bo

    2016-01-15

    Bacillus methylotrophicus SK19.001 is a glutamate-independent strain that produces poly(γ-glutamic acid) (γ-PGA), a polymer of D- and L-glutamic acids that possesses applications in food, the environment, agriculture, etc. This study was undertaken to explore the synthetic pathway of intracellular L- and D-glutamic acid in SK19.001 by investigating the effects of tricarboxylic acid cycle intermediates and different amino acids as metabolic precursors on the production of γ-PGA and analyzing the activities of the enzymes involved in the synthesis of L- and D-glutamate. Tricarboxylic acid cycle intermediates and amino acids could participate in the synthesis of γ-PGA via independent pathways in SK19.001. L-Aspartate aminotransferase, L-glutaminase and L-glutamate synthase were the enzymatic sources of L-glutamate. Glutamate racemase was responsible for the formation of D-glutamate for the synthesis of γ-PGA, and the synthetase had stereoselectivity for glutamate substrate. The enzymatic sources of L-glutamate were investigated for the first time in the glutamate-independent γ-PGA-producing strain, and multiple enzymatic sources of L-glutamate were verified in SK19.001, which will benefit efforts to improve production of γ-PGA with metabolic engineering strategies. © 2015 Society of Chemical Industry.

  17. On the specificity of a bacteriophage-borne endoglycanase for the native capsular polysaccharide produced by Klebsiella pneumoniae SK1 and its derived polymers.

    PubMed

    Cescutti, P; Paoletti, S

    1994-02-15

    The specificity of the endoglycanase associated with the bacteriophage phi SK1 particles was tested on the native capsular polysaccharide produced by Klebsiella pneumoniae serotype SK1 and on three chemically modified polymers derived from it. The primary structure of the SK1 capsular polysaccharide is: [formula: see text] and the beta 1-3 linkage between the glucose and the galactose residues is the one cleaved by the phage enzyme. The enzyme activity was assayed on the deacetylated polysaccharide and on two derivatives obtained by removal of both the side-chain sugars and of only the alpha-D-galactosyl unit, respectively. The endoglycanase was more active on the deacetylated polysaccharide than on the native one, suggesting that the presence of the acetyl groups interferes with the enzyme-polysaccharide interaction. A possible role of the acetyl groups in the control of the polysaccharide chain length and hence on the rheological behaviour of the capsule cannot be ruled out, as already indicated for other bacterial polysaccharides. On the contrary, the removal of the side chains, either complete or selective, caused the modification of the recognition site in such a way that the enzymatic depolymerization no longer occurred. Therefore, it can be inferred that the phi SK1 endoglycanase requires the presence of both the side chain sugars to exhibit its cleaving activity, although this latter is in the main chain.

  18. Solar Neutrino flare detection in Hyperkamiokande and SK

    NASA Astrophysics Data System (ADS)

    Fargion, Daniele

    2016-07-01

    The possible buid and near activity of a Megaton neutrino detection in HyperKamiokande and the older SK implementation by Gadolinium liqid might open to future detection of largest solar flare (pion trace at tens MeV) electron neutrino and antineutrino. The multiwave detection of X-gamma and neutrino event might offer a deep view of such solar acelleration and of neutrino flavor mix along its flight. The possoble near future discover of such events will open a third neutrino astronomy windows after rarest SN 1987A and persistent Solar nuclear signals.

  19. Detection of large color variation in the potentially hazardous asteroid (297274) 1996 SK

    NASA Astrophysics Data System (ADS)

    Lin, Chien-Hsien; Ip, Wing-Huen; Lin, Zhong-Yi; Yoshida, Fumi; Cheng, Yu-Chi

    2014-03-01

    Low-inclination near-earth asteroid (NEA) (297274) 1996 SK, which is also classified as a potentially hazardous asteroid, has a highly eccentric orbit. It was studied by multi-wavelength photometry within the framework of an NEA color survey at Lulin Observatory. Here, we report the finding of large color variation across the surface of (297274) 1996 SK within one asteroidal rotation period of 4.656 ± 0.122 hours and classify it as an S-type asteroid according to its average colors of B — V = 0.767 ± 0.033, V — R = 0.482 ± 0.021, V — I = 0.801 ± 0.025 and the corresponding relative reflectance spectrum. These results might be indicative of differential space weathering or compositional inhomogeneity in the surface materials.

  20. The Chinese Cretaceous Continental Scientific Drilling Project in the Songliao Basin, NE China: Organic-rich source rock evaluation with geophysical logs from Borehole SK-2

    NASA Astrophysics Data System (ADS)

    Zhang, X.; Zou, C.

    2017-12-01

    The Cretaceous strata have been recognized as an important target of oil or gas exploration in the Songliao Basin, northeast China. The second borehole (SK-2) of the Chinese Cretaceous Continental Scientific Drilling Project in the Songliao Basin (CCSD-SK) is the first one to drill through the Cretaceous continental strata in the frame of ICDP. It was designed not only to solve multiple scientific problems (including the Cretaceous paleoenvironment and paleoclimate, as well as deep resources exploration of the Songliao Basin), but also to expect to achieve new breakthroughs in oil and gas exploration. Based on the project, various geophysical log data (including gamma, sonic, resistivity, density etc.) and core samples have been collected from Borehole SK-2. We do research on organic-rich source rocks estimation using various geophysical log data. Firstly, we comprehensively analyzed organic-rich source rocks' geophysical log response characteristics. Then, source rock's identification methods were constructed to identify organic-rich source rocks with geophysical logs. The main identification methods include cross-plot, multiple overlap and Decision Tree method. Finally, the technique and the CARBOLOG method were applied to evaluate total organic carbon (TOC) content from geophysical logs which provide continuous vertical profile estimations (Passey, 1990; Carpentier et al., 1991). The results show that source rocks are widely distributed in Borehole SK-2, over a large depth strata (985 5700m), including Nenjiang, Qingshankou, Denglouku, Yingcheng, Shahezi Formations. The organic-rich source rocks with higher TOC content occur in the Qingshankou (1647 1650m), Denglouku (2534 2887m) and Shahezi (3367 5697m) Formations. The highest TOC content in these formations can reach 10.31%, 6.58%, 12.79% respectively. The bed thickness of organic-rich source rocks in the these formations are totally up to 7.88m, 74.34m, 276.60m respectively. These organic-rich rocks in the

  1. Azelaic acid was sensitizing effect in the chemotherapeutic treatment of several melanoma cell lines.

    PubMed

    Rodriguez-Vicente, J; Vicente-Ortega, V; Canteras-Jordana

    1996-12-01

    Chemotherapy for melanoma results in low response and must be reinforced with sensitizer compounds. We believed that azelaic acid (AZA) could modulate melanomas' resistance to antineoplastics. Therefore we tried to compare in vitro treatment with antineoplastics alone versus AZA treatment followed by antineoplastics. We carried out MTT assays to evaluate the cytotoxicity of melphalan, lomustine (CCNU), fotemustine, and 4-Hydroxyanisole (4-HA) on three melanoma lines (B16F10, SK-MEL-28, and SK-MEL-1), and the modulating effect of pretreatment with AZA (1 mM). AZA showed a dose-dependent antineoplastic activity on the three lines. Melphalan was the most active drug followed by CCNU, fotemustine, and 4-HA. The most sensitive line was B16F10 and the least sensitive was SK-meL-1. Previous treatment with AZA of B16F10 reinforced the effect of melphalan (2.5 times), CCNU (10 times), and fotemustine (14 times); whereas for SK-MEL-28 and SK-MEL-1, only the cytotoxicity of CCNU and fotemustine increased. An antagonist effect was produced by 4-HA on all three lines. We concluded that AZA enhances in vitro cytotoxicity of CCNU and fotemustine.

  2. Polyploidization on SK-N-MC human neuroblastoma cells infected with herpes simplex virus 1.

    PubMed

    Karalyan, Zaven; Izmailyan, Roza; Karalova, Elena; Abroyan, Liana; Hakobyan, Lina; Avetisyan, Aida; Semerjyan, Zara

    2016-01-01

    Polyploidization is one of the most dramatic changes occurring within cell genome owing to various reasons including under many viral infections. We examined the impact of herpes simplex virus-1 (HSV-1) on SK-N-MC human neuroblastoma cell line. The infected cells were followed from 6 hours up to 96 hours post infection (hpi). A large number of polyploid cells with giant nuclei was observed under the influence of HSV-1 at 24 hpi with the DNA content of 32c to 64c or more, in comparison with control SK-N-MC cells that were characterized by relatively moderate values of ploidy, i.e. 8с to 16с (where 1c is the haploid amount of nuclear DNA found in normal diploid populations in G0/G1). After 48-96 hpi, the population of polyploid cells with giant nuclei decreased to the benchmark level. The SK-NMC cells infected with HSV-1 for 24 hours were stained with gallocyanine and monitored for cytological features. The infected cells underwent virus induced cellcell and nuclei fusion with the formation of dense nuclei syncytium. The metabolic activity of HSV-1 infected cells was higher in both nuclei and nucleoli when compared to control cells.

  3. Inhibition of the CSF-1 receptor sensitizes ovarian cancer cells to cisplatin.

    PubMed

    Yu, Rong; Jin, Hao; Jin, Congcong; Huang, Xuefeng; Lin, Jinju; Teng, Yili

    2018-03-01

    Ovarian cancer is one of the most common female malignancies, and cisplatin-based chemotherapy is routinely used in locally advanced ovarian cancer patients. Acquired or de novo cisplatin resistance remains the barrier to patient survival, and the mechanisms of cisplatin resistance are still not well understood. In the current study, we found that colony-stimulating-factor-1 receptor (CSF-1R) was upregulated in cisplatin-resistant SK-OV-3 and CaoV-3 cells. Colony-stimulating-factor-1 receptor knockdown suppressed proliferation and enhanced apoptosis in cisplatin-resistant SK-OV-3 and CaoV-3 cells. However, CSF-1R overexpression had inverse effects. While parental SK-OV-3 and CaoV-3 cells were more resistant to cisplatin after CSF-1R overexpression, CSF-1R knockdown in SK-OV-3 and CaoV-3 cells promoted cisplatin sensitivity. Overexpression and knockdown studies also showed that CSF-1R significantly promoted active AKT and ERK1/2 signalling pathways in cisplatin-resistant cells. Furthermore, a combination of cisplatin and CSF-1R inhibitor effectively inhibited tumour growth in xenografts. Taken together, our results provide the first evidence that CSF-1R inhibition can sensitize cisplatin-refractory ovarian cancer cells. This study may help to increase understanding of the molecular mechanisms underlying cisplatin resistance in tumours. Copyright © 2018 John Wiley & Sons, Ltd.

  4. Effect of the oncolytic ECHO-7 virus Rigvir® on the viability of cell lines of human origin in vitro.

    PubMed

    Tilgase, Andra; Patetko, Liene; Blāķe, Ilze; Ramata-Stunda, Anna; Borodušķis, Mārtiņš; Alberts, Pēteris

    2018-01-01

    Background: The role of oncolytic viruses in cancer treatment is increasingly studied. The first oncolytic virus (Rigvir®, ECHO-7) was registered in Latvia over a decade ago. In a recent retrospective study Rigvir® decreased mortality 4.39-6.57-fold in stage IB-IIC melanoma patients. The aims of the present study are to test the effect of Rigvir® on cell line viability in vitro and to visualize the cellular presence of Rigvir® by immunocytochemistry. Methods: The cytolytic effect of Rigvir® on the viability of FM-9, RD, AGS, A549, HDFa, HPAF‑II, MSC, MCF7, HaCaT, and Sk-Mel-28 cell lines was measured using live cell imaging. PBMC viability was measured using flow cytometry. The presence of ECHO-7 virus was visualized using immunocytochemistry. Statistical difference between treatment groups was calculated using two-way ANOVA. Results: Rigvir® (10%, volume/volume) reduced cell viability in FM-9, RD, AGS, A549, HDFa, HPAF‑II and MSC cell lines by 67-100%. HaCaT cell viability was partly affected while Rigvir® had no effect on MCF7, Sk-Mel-28 and PBMC viability. Detection of ECHO-7 by immunocytochemistry in FM-9, RD, AGS, A549, HDFa, HPAF-II and Sk-Mel-28 cell lines suggests that the presence of Rigvir® in the cells preceded or coincided with the time of reduction of cell viability. Rigvir® (10%) had no effect on live PBMC count. Conclusions: The results suggest that Rigvir® in vitro reduces the viability of cells of human melanoma, rhabdomyosarcoma, gastric adenocarcinoma, lung carcinoma, pancreas adenocarcinoma but not in PBMC. The presence of Rigvir® in the sensitive cells was confirmed using anti-ECHO-7 antibodies. The present results suggest that a mechanism of action for the clinical benefit of Rigvir® is its cytolytic properties. The present results suggest that the effect of Rigvir® could be tested in other cancers besides melanoma. Further studies of possible Rigvir® entry receptors are needed.

  5. Improved health-span and lifespan in mtDNA mutator mice treated with the mitochondrially targeted antioxidant SkQ1

    PubMed Central

    Shabalina, Irina G.; Vyssokikh, Mikhail Yu.; Gibanova, Natalia; Csikasz, Robert I.; Edgar, Daniel; Hallden-Waldemarson, Anne; Rozhdestvenskaya, Zinaida; Bakeeva, Lora E.; Vays, Valeria B.; Pustovidko, Antonina V.; Skulachev, Maxim V.; Cannon, Barbara; Skulachev, Vladimir P.; Nedergaard, Jan

    2017-01-01

    MtDNA mutator mice exhibit marked features of premature aging. We find that these mice treated from age of ≈100 days with the mitochondria-targeted antioxidant SkQ1 showed a delayed appearance of traits of aging such as kyphosis, alopecia, lowering of body temperature, body weight loss, as well as ameliorated heart, kidney and liver pathologies. These effects of SkQ1 are suggested to be related to an alleviation of the effects of an enhanced reactive oxygen species (ROS) level in mtDNA mutator mice: the increased mitochondrial ROS released due to mitochondrial mutations probably interact with polyunsaturated fatty acids in cardiolipin, releasing malondialdehyde and 4-hydroxynonenal that form protein adducts and thus diminishes mitochondrial functions. SkQ1 counteracts this as it scavenges mitochondrial ROS. As the results, the normal mitochondrial ultrastructure is preserved in liver and heart; the phosphorylation capacity of skeletal muscle mitochondria as well as the thermogenic capacity of brown adipose tissue is also improved. The SkQ1-treated mice live significantly longer (335 versus 290 days). These data may be relevant in relation to treatment of mitochondrial diseases particularly and the process of aging in general. PMID:28209927

  6. The sigma-1 receptor modulates NMDA receptor synaptic transmission and plasticity via SK channels in rat hippocampus

    PubMed Central

    Martina, Marzia; Turcotte, Marie-Eve B; Halman, Samantha; Bergeron, Richard

    2007-01-01

    The sigma receptor (σR), once considered a subtype of the opioid receptor, is now described as a distinct pharmacological entity. Modulation of N-methyl-d-aspartate receptor (NMDAR) functions by σR-1 ligands is well documented; however, its mechanism is not fully understood. Using patch-clamp whole-cell recordings in CA1 pyramidal cells of rat hippocampus and (+)pentazocine, a high-affinity σR-1 agonist, we found that σR-1 activation potentiates NMDAR responses and long-term potentiation (LTP) by preventing a small conductance Ca2+-activated K+ current (SK channels), known to shunt NMDAR responses, to open. Therefore, the block of SK channels and the resulting increased Ca2+ influx through the NMDAR enhances NMDAR responses and LTP. These results emphasize the importance of the σR-1 as postsynaptic regulator of synaptic transmission. PMID:17068104

  7. The sigma-1 receptor modulates NMDA receptor synaptic transmission and plasticity via SK channels in rat hippocampus.

    PubMed

    Martina, Marzia; Turcotte, Marie-Eve B; Halman, Samantha; Bergeron, Richard

    2007-01-01

    The sigma receptor (sigmaR), once considered a subtype of the opioid receptor, is now described as a distinct pharmacological entity. Modulation of N-methyl-D-aspartate receptor (NMDAR) functions by sigmaR-1 ligands is well documented; however, its mechanism is not fully understood. Using patch-clamp whole-cell recordings in CA1 pyramidal cells of rat hippocampus and (+)pentazocine, a high-affinity sigmaR-1 agonist, we found that sigmaR-1 activation potentiates NMDAR responses and long-term potentiation (LTP) by preventing a small conductance Ca2+-activated K+ current (SK channels), known to shunt NMDAR responses, to open. Therefore, the block of SK channels and the resulting increased Ca2+ influx through the NMDAR enhances NMDAR responses and LTP. These results emphasize the importance of the sigmaR-1 as postsynaptic regulator of synaptic transmission.

  8. Effects of Mitochondrial Antioxidant SkQ1 on Biochemical and Behavioral Parameters in a Parkinsonism Model in Mice.

    PubMed

    Pavshintsev, V V; Podshivalova, L S; Frolova, O Y; Belopolskaya, M V; Averina, O A; Kushnir, E A; Marmiy, N V; Lovat, M L

    2017-12-01

    According to one hypothesis, Parkinson's disease pathogenesis is largely caused by dopamine catabolism that is catalyzed on mitochondrial membranes by monoamine oxidase. Reactive oxygen species are formed as a byproduct of these reactions, which can lead to mitochondrial damage followed by cell degeneration and death. In this study, we investigated the effects of administration of the mitochondrial antioxidant SkQ1 on biochemical, immunohistochemical, and behavioral parameters in a Parkinson-like condition caused by protoxin MPTP injections in C57BL/6 mice. SkQ1 administration increased dopamine quantity and decreased signs of sensory-motor deficiency as well as destruction of dopaminergic neurons in the substantia nigra and ventral tegmental area in mice with the Parkinson-like condition.

  9. Mechanisms underlying the cardiac pacemaker: the role of SK4 calcium-activated potassium channels

    PubMed Central

    Weisbrod, David; Khun, Shiraz Haron; Bueno, Hanna; Peretz, Asher; Attali, Bernard

    2016-01-01

    The proper expression and function of the cardiac pacemaker is a critical feature of heart physiology. The sinoatrial node (SAN) in human right atrium generates an electrical stimulation approximately 70 times per minute, which propagates from a conductive network to the myocardium leading to chamber contractions during the systoles. Although the SAN and other nodal conductive structures were identified more than a century ago, the mechanisms involved in the generation of cardiac automaticity remain highly debated. In this short review, we survey the current data related to the development of the human cardiac conduction system and the various mechanisms that have been proposed to underlie the pacemaker activity. We also present the human embryonic stem cell-derived cardiomyocyte system, which is used as a model for studying the pacemaker. Finally, we describe our latest characterization of the previously unrecognized role of the SK4 Ca2+-activated K+ channel conductance in pacemaker cells. By exquisitely balancing the inward currents during the diastolic depolarization, the SK4 channels appear to play a crucial role in human cardiac automaticity. PMID:26725737

  10. Mechanisms underlying the cardiac pacemaker: the role of SK4 calcium-activated potassium channels.

    PubMed

    Weisbrod, David; Khun, Shiraz Haron; Bueno, Hanna; Peretz, Asher; Attali, Bernard

    2016-01-01

    The proper expression and function of the cardiac pacemaker is a critical feature of heart physiology. The sinoatrial node (SAN) in human right atrium generates an electrical stimulation approximately 70 times per minute, which propagates from a conductive network to the myocardium leading to chamber contractions during the systoles. Although the SAN and other nodal conductive structures were identified more than a century ago, the mechanisms involved in the generation of cardiac automaticity remain highly debated. In this short review, we survey the current data related to the development of the human cardiac conduction system and the various mechanisms that have been proposed to underlie the pacemaker activity. We also present the human embryonic stem cell-derived cardiomyocyte system, which is used as a model for studying the pacemaker. Finally, we describe our latest characterization of the previously unrecognized role of the SK4 Ca(2+)-activated K(+) channel conductance in pacemaker cells. By exquisitely balancing the inward currents during the diastolic depolarization, the SK4 channels appear to play a crucial role in human cardiac automaticity.

  11. Recessive variants of MuSK are associated with late onset CMS and predominant limb girdle weakness.

    PubMed

    Owen, David; Töpf, Ana; Preethish-Kumar, Veeramani; Lorenzoni, Paulo José; Vroling, Bas; Scola, Rosana Herminia; Dias-Tosta, Elza; Geraldo, Argemiro; Polavarapu, Kiran; Nashi, Saraswati; Cox, Daniel; Evangelista, Teresinha; Dawson, John; Thompson, Rachel; Senderek, Jan; Laurie, Steven; Beltran, Sergi; Gut, Marta; Gut, Ivo; Nalini, Atchayaram; Lochmüller, Hanns

    2018-04-28

    Congenital myasthenic syndrome (CMS) is a heterogeneous disorder that causes fatigable muscle weakness. CMS has been associated with variants in the MuSK gene and, to date, 16 patients have been reported. MuSK-CMS patients present a different phenotypic pattern of limb girdle weakness. Here, we describe four additional patients and discuss the phenotypic and clinical relationship with those previously reported. Two novel damaging missense variants are described: c.1742T > A; p.I581N found in homozygosis, and c.1634T > C; p.L545P found in compound heterozygosis with p.R166*. The reported patients had predominant limb girdle weakness with symptom onset at 12, 17, 18, and 30 years of age, and the majority exhibited a good clinical response to Salbutamol therapy, but not to esterase inhibitors. Meta-analysis including previously reported variants revealed an increased likelihood of a severe, respiratory phenotype with null alleles. Missense variants exclusively affecting the kinase domain, but not the catalytic site, are associated with late onset. These data refine the phenotype associated with MuSK-related CMS. © 2018 Wiley Periodicals, Inc.

  12. Cations SkQ1 and MitoQ accumulated in mitochondria delay opening of ascorbate/FeSO4-induced nonspecific pore in the inner mitochondrial membrane.

    PubMed

    Khailova, L S; Dedukhova, V I; Mokhova, E N

    2008-10-01

    It is known that an addition of FeSO4 in the presence of ascorbic acid to cells or mitochondria can injure energy coupling and some other functions in mitochondria. The present study demonstrates that decrease in ascorbate concentration from 4 to 0.2 mM in the presence of the same low concentrations of FeSO4 accelerates the nonspecific pore opening, while cyclosporin A prevents and under some conditions reverses the pore opening. Hydrophobic cations SkQ1 and MitoQ (structural analogs of plastoquinone and coenzyme Q(10), respectively) delay pore opening, SkQ1 being more efficient. It is known that an increase in matrix ADP concentration delays pore opening, while an addition of carboxyatractylate to mitochondria accelerates the beginning of pore opening. Preliminary addition of SkQ1 into a mitochondrial suspension increased the effect of ADP and decreased the effect of carboxyatractylate. These results suggest that under the conditions used SkQ1 protects mitochondria from oxidative damage as an antioxidant when added at extremely low concentrations.

  13. Expression of Meiotic Drive Elements Spore Killer-2 and Spore Killer-3 in Asci of Neurospora Tetrasperma

    PubMed Central

    Raju, N. B.; Perkins, D. D.

    1991-01-01

    It was shown previously that when a chromosomal Spore killer factor is heterozygous in Neurospora species with eight-spored asci, the four sensitive ascospores in each ascus die and the four survivors are all killers. Sk-2(K) and Sk-3(K) are nonrecombining haplotypes that segregate with the centromere of linkage group III. No killing occurs when either one of these killers is homozygous, but each is sensitive to killing by the other in crosses of Sk-2(K) X Sk-3(K). In the present study, Sk-2(K) and Sk-3(K) were transferred by recurrent backcrosses from the eight-spored species Neurospora crassa into Neurospora tetrasperma, a pseudohomothallic species which normally makes asci with four large spores, each heterokaryotic for mating type and for any other centromere-linked genes that are heterozygous in the cross. The action of Sk-2(K) and Sk-3(K) in N. tetrasperma is that predicted from their behavior in eight-spored species. A sensitive nucleus is protected from killing if it is enclosed in the same ascospore with a killer nucleus. Crosses of Sk-2(K) X Sk-2(S), Sk-3(K) X Sk-3(S), and Sk-2(K) X Sk-3(K) all produce four-spored asci that are wild type in appearance, with the ascospores heterokaryotic and viable. The Eight-spore gene E, which shows variable penetrance, was used to obtain N. tetrasperma asci in which two to eight spores are small and homokaryotic. When killer and sensitive alleles are segregating in the presence of E, only those ascospores that contain a killer allele survive. Half of the small ascospores are killed. In crosses of Sk-2(K) X Sk-3(K) (with E heterozygous), effectively all small ascospores are killed. The ability of N. tetrasperma to carry killer elements in cryptic condition suggests a possible role for Spore killers in the origin of pseudohomothallism, with adoption of the four-spored mode restoring ascospore viability of crosses in which killing would otherwise occur. PMID:1834522

  14. Complementary functions of SK and Kv7/M potassium channels in excitability control and synaptic integration in rat hippocampal dentate granule cells

    PubMed Central

    Mateos-Aparicio, Pedro; Murphy, Ricardo; Storm, Johan F

    2014-01-01

    The dentate granule cells (DGCs) form the most numerous neuron population of the hippocampal memory system, and its gateway for cortical input. Yet, we have only limited knowledge of the intrinsic membrane properties that shape their responses. Since SK and Kv7/M potassium channels are key mechanisms of neuronal spiking and excitability control, afterhyperpolarizations (AHPs) and synaptic integration, we studied their functions in DGCs. The specific SK channel blockers apamin or scyllatoxin increased spike frequency (excitability), reduced early spike frequency adaptation, fully blocked the medium-duration AHP (mAHP) after a single spike or spike train, and increased postsynaptic EPSP summation after spiking, but had no effect on input resistance (Rinput) or spike threshold. In contrast, blockade of Kv7/M channels by XE991 increased Rinput, lowered the spike threshold, and increased excitability, postsynaptic EPSP summation, and EPSP–spike coupling, but only slightly reduced mAHP after spike trains (and not after single spikes). The SK and Kv7/M channel openers 1-EBIO and retigabine, respectively, had effects opposite to the blockers. Computational modelling reproduced many of these effects. We conclude that SK and Kv7/M channels have complementary roles in DGCs. These mechanisms may be important for the dentate network function, as CA3 neurons can be activated or inhibition recruited depending on DGC firing rate. PMID:24366266

  15. Ultrananocrystalline diamond-coated nanoporous membranes support SK-N-SH neuroblastoma endothelial cell attachment.

    PubMed

    Yang, Kai-Hung; Nguyen, Alexander K; Goering, Peter L; Sumant, Anirudha V; Narayan, Roger J

    2018-06-06

    Ultrananocrystalline diamond (UNCD) has been demonstrated to have attractive features for biomedical applications and can be combined with nanoporous membranes for applications in drug delivery systems, biosensing, immunoisolation and single molecule analysis. In this study, free-standing nanoporous UNCD membranes with pore sizes of 100 or 400 nm were fabricated by directly depositing ultrathin UNCD films on nanoporous silicon nitride membranes and then etching away silicon nitride using reactive ion etching. Successful deposition of UNCD on the substrate with a novel process was confirmed with Raman spectroscopy, X-ray photoelectron spectroscopy, cross-section scanning electron microscopy (SEM) and transmission electron microscopy. Both sample types exhibited uniform geometry and maintained a clear hexagonal pore arrangement. Cellular attachment of SK-N-SH neuroblastoma endothelial cells was examined using confocal microscopy and SEM. Attachment of SK-N-SH cells onto UNCD membranes on both porous regions and solid surfaces was shown, indicating the potential use of UNCD membranes in biomedical applications such as biosensors and tissue engineering scaffolds.

  16. A numerical investigation of phytoplankton and Pseudocalanus elongatus dynamics in the spring bloom time in the Gdańsk Gulf

    NASA Astrophysics Data System (ADS)

    Dzierzbicka-Głowacka, Lidia

    2005-01-01

    A nutrient-phytoplankton-zooplankton-detritus (1D-NPZD) `phytoplankton {Phyt} and Pseudocalanus elongatus {Zoop} dynamics in the spring bloom time in the Gdańsk Gulf. The 1D-NPZD model consists of three coupled, partial second-order differential equations of the diffusion type for phytoplankton {Phyt}, zooplankton {Zoop}, nutrients {Nutr} and one ordinary first-order differential equation for benthic detritus pool {Detr}, together with initial and boundary conditions. In this model, the {Zoop} is presented by only one species of copepod ( P. elongatus) and {Zoop} is composed of six cohorts of copepods with weights ( Wi) and numbers ( Zi); where Zoop= limit∑i=16W iZ i. The calculations were made for 90 days (March, April, May) for two stations at Gdańsk Gulf with a vertical space step of 0.5m and a time step of 900 s. The flow field and water temperature used as the inputs in the biological model 1D-NPZD were reproduced by the prognostic numerical simulation technique using hydrographic climatological data. The results of the numerical investigations described here were compared with the mean observed values of surface chlorophyll- a and depth integrated P. elongatus biomass for 10 years, 1980-1990. The slight differences between the calculated and mean observed values of surface chlorophyll- a and zooplankton biomass are ca. 10-60 mg C m -3 and ca. 5-23 mg C m -2, respectively, depending on the location of the hydrographic station. The 1D-NPZD model with a high-resolution zooplankton module for P. elongatus can be used to describe the temporal patterns for phytoplankton biomass and P. elongatus in the centre of the Gdańsk Gulf.

  17. Protective Effect of Coriolus versicolor Cultivated in Citrus Extract Against Nitric Oxide-Induced Apoptosis in Human Neuroblastoma SK-N-MC Cells

    PubMed Central

    Kim, Byung-Chul; Kim, Youn-Sub; Lee, Jin-Woo; Seo, Jin-Hee; Ji, Eun-Sang; Lee, Hyejung; Park, Yong-Il

    2011-01-01

    Nitric oxide (NO) is a reactive free radical and a messenger molecule in many physiological functions. However, excessive NO is believed to be a mediator of neurotoxicity. The medicinal plant Coriolus versicolor is known to possess anti-tumor and immune-potentiating activities. In this study, we investigated whether Coriolus versicolor possesses a protective effect against NO donor sodium nitroprusside (SNP)-induced apoptosis in the human neuroblastoma cell line SK-N-MC. We utilized 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, flow cytometry, 4,6-diamidino-2-phenylindole (DAPI) staining, terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick end labeling (TUNEL) assay, DNA fragmentation assay, reverse transcription-polymerase chain reaction (RT-PCR), Western blot analysis, and caspase-3 enzyme activity assay in SK-N-MC cells. MTT assay showed that SNP treatment significantly reduces the viability of cells, and the viabilities of cells pre-treated with the aqueous extract of Coriolus versicolor cultivated in citrus extract (CVEcitrus) was increased. However, aqueous extract of Coriolus versicolor cultivated in synthetic medium (CVEsynthetic) showed no protective effect and aqueous citrus extract (CE) had a little protective effect. The cell treated with SNP exhibited several apoptotic features, while those pre-treated for 1 h with CVEcitrus prior to SNP expose showed reduced apoptotic features. The cells pre-treated for 1 h with CVEcitrus prior to SNP expose inhibited p53 and Bax expressions and caspase-3 enzyme activity up-regulated by SNP. We showed that CVEcitrus exerts a protective effect against SNP-induced apoptosis in SK-N-MC cells. Our study suggests that CVEcitrus has therapeutic value in the treatment of a variety of NO-induced brain diseases. PMID:22110367

  18. Protective Effect of Coriolus versicolor Cultivated in Citrus Extract Against Nitric Oxide-Induced Apoptosis in Human Neuroblastoma SK-N-MC Cells.

    PubMed

    Kim, Byung-Chul; Kim, Youn-Sub; Lee, Jin-Woo; Seo, Jin-Hee; Ji, Eun-Sang; Lee, Hyejung; Park, Yong-Il; Kim, Chang-Ju

    2011-06-01

    Nitric oxide (NO) is a reactive free radical and a messenger molecule in many physiological functions. However, excessive NO is believed to be a mediator of neurotoxicity. The medicinal plant Coriolus versicolor is known to possess anti-tumor and immune-potentiating activities. In this study, we investigated whether Coriolus versicolor possesses a protective effect against NO donor sodium nitroprusside (SNP)-induced apoptosis in the human neuroblastoma cell line SK-N-MC. We utilized 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, flow cytometry, 4,6-diamidino-2-phenylindole (DAPI) staining, terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick end labeling (TUNEL) assay, DNA fragmentation assay, reverse transcription-polymerase chain reaction (RT-PCR), Western blot analysis, and caspase-3 enzyme activity assay in SK-N-MC cells. MTT assay showed that SNP treatment significantly reduces the viability of cells, and the viabilities of cells pre-treated with the aqueous extract of Coriolus versicolor cultivated in citrus extract (CVE(citrus)) was increased. However, aqueous extract of Coriolus versicolor cultivated in synthetic medium (CVE(synthetic)) showed no protective effect and aqueous citrus extract (CE) had a little protective effect. The cell treated with SNP exhibited several apoptotic features, while those pre-treated for 1 h with CVE(citrus) prior to SNP expose showed reduced apoptotic features. The cells pre-treated for 1 h with CVE(citrus) prior to SNP expose inhibited p53 and Bax expressions and caspase-3 enzyme activity up-regulated by SNP. We showed that CVE(citrus) exerts a protective effect against SNP-induced apoptosis in SK-N-MC cells. Our study suggests that CVE(citrus) has therapeutic value in the treatment of a variety of NO-induced brain diseases.

  19. [Temporal dynamics of ethnic assortativeness in the Tsivil'sk region of Chuvashia].

    PubMed

    El'chinova, G I; Znichenko, R A; Ginter, E K

    2003-04-01

    Based on marriage records for 1950-1959, 1970-1979, and 1990-1999, ethnic assortativeness of Chuvash and Russian inhabitants of Tsivil'sk raion of the Chuvash Republic was calculated. It was shown that in half a century ethnic assortativeness of Chuvashes did not change (1.18, 1.15, and 1.15), while in Russians a clear downward trend was observed (4.20, 2.18, and 1.28). The data on ethnic assortativeness obtained were compared with those for the same populations from other regions of the Chuvashia examined earlier.

  20. The 5K70SK automatically tuned, high power, S-band klystron

    NASA Technical Reports Server (NTRS)

    Goldfinger, A.

    1977-01-01

    Primary objectives include delivery of 44 5K70SK klystron amplifier tubes and 26 remote tuner assemblies with spare parts kits. Results of a reliability demonstration on a klystron test cavity are discussed, along with reliability tests performed on a remote tuning unit. Production problems and one design modification are reported and discussed. Results of PAT and DVT are included.

  1. Relationship between 4-hydroxyanisole toxicity and dopa oxidase activity for three melanoma cell lines.

    PubMed

    Rodriguez-Vicente, J; Vicente-Ortega, V; Canteras-Jordana, M; Calderon-Rubiales, F

    1997-10-01

    We studied the response of mouse B16F10 and SK-MEL-28 and SK-MEL-1 human melanoma cell lines to treatment with 4-hydroxyanisole (4-HA), and attempted to relate the response to the dopa oxidase levels and the morphological characteristics of each cell line. Clear dose-response curves were observed after 24 h of treatment in each cell line, the 4-HA being more toxic to the B16F10 cells, with an ID50 value of 215 microM. This was much lower than that observed for the SK-MEL-28 and SK-MEL-1 cell lines (ID50 of 5.98 mM and 7.17 mM, respectively). There was a direct relationship between toxicity levels and dopa oxidase activity, since the highest specific activity was obtained for B16F10 (15.9 mU), while lower activity was registered for SK-MEL-28 (4.59 mU) and SK-MEL-1 (1.24 mU), which also showed lower 4-HA toxicity. Morphologically, we observed the typical characteristics of cellular injury, with swelling and dilation of the internal membranes and organelles, an increased number of vacuoles, and an increased number of abnormal multilamellar melanosomes or thick clumps of irregularly distributed melanin. On the other hand, we observed that the two cell lines with the lowest dopa oxidase activity contained more mature fully melanized melanosomes than B16F10, pointing to possible alterations in the melanosome transference mechanism and lower enzymatic activity in the mature melanosomes of these two human cell lines.

  2. Response of the diatom flora in the Gdańsk Basin (southern Baltic Sea) to eutrophication in the last century

    NASA Astrophysics Data System (ADS)

    Witak, Małgorzata

    2016-04-01

    The diatom flora in six short cores (length of 10-20 cm) taken from Gdańsk Basin, southern Baltic Sea, was analyzed in order to response of diatoms to environmental changes over the past 100 years. This study is a part of CLISED project (Climate Change Impact on Ecosystem Health - Marine Sediments Indicators) funded from Norway Grants in the Polish-Norwegian Research Programme. Three cores P116, M1 and P1 were retrieved from the Gdańsk Deep region, whereas P104, BMPK10 and P110 were drilled from the shallower part - Gulf of Gdańsk. The sediment cores were dated by 210Pb and represented time interval of AD 1895-2015. In this period an artificial mouth of the Vistula River near Świbno was excavated and riverine waters flowed directly to the Gulf of Gdańsk. Urbanization, progressive industrialization and an increase in the inflow of waste water had a strong influence on ecosystem in the last 120 years. The transport of large amounts of nutrients, organic matter and pollutants by the rivers resulted in environmental degradation of the Gdańsk Basin and its progressive eutrophication in particular. Diatoms, being the major component of algal assemblages, are excellent bioindicators for assessing the state of basin ecosystems. They are usually abundant in marine sediments, diverse and sensitive to numerous environmental variables, including physical (e.g. light, temperature, water transparency) and chemical (e.g. pH, alkalinity, nutrient availability) factors. Changes in nutrients concentrations may lead to substantial transformations in the structure of diatom assemblages. Samples for diatom analyses (ca. 0.3-2.0 g dry sediment) were prepared following the standard procedure for diatom observation under light microscope (Battarbee, 1986). To estimate the concentration of siliceous microfossils per unit weight of dry sediment (absolute abundance), a random settling technique was used (Bodén, 1991). Permanent diatom preparations were mounted in Naphrax® (refractive

  3. Loss of function mutation in LARP7, chaperone of 7SK ncRNA, causes a syndrome of facial dysmorphism, intellectual disability, and primordial dwarfism.

    PubMed

    Alazami, Anas M; Al-Owain, Mohammad; Alzahrani, Fatema; Shuaib, Taghreed; Al-Shamrani, Hussain; Al-Falki, Yahya H; Al-Qahtani, Saleh M; Alsheddi, Tarfa; Colak, Dilek; Alkuraya, Fowzan S

    2012-10-01

    Primordial dwarfism (PD) is a clinically and genetically heterogeneous condition. Various molecular mechanisms are known to underlie the disease including impaired mitotic mechanics, abnormal IGF2 expression, perturbed DNA damage response, defective spliceosomal machinery, and abnormal replication licensing. Here, we describe a syndromic form of PD associated with severe intellectual disability and distinct facial features in a large multiplex Saudi family. Analysis reveals a novel underlying mechanism for PD involving depletion of 7SK, an abundant cellular noncoding RNA (ncRNA), due to mutation of its chaperone LARP7. We show that 7SK levels are tightly linked to LARP7 expression across cell lines, and that this chaperone is ubiquitously expressed in the mouse embryo. The 7SK is known to influence the expression of a wide array of genes through its inhibitory effect on the positive transcription elongation factor b (P-TEFb) as well as its competing role in HMGA1-mediated transcriptional regulation. This study documents a critical role played by ncRNA in human development and adds to the growing list of molecular mechanisms that, when perturbed, converge on the PD phenotype. © 2012 Wiley Periodicals, Inc.

  4. Protection of SK-N-MC cells against β-amyloid peptide-induced degeneration using neuron growth factor-loaded liposomes with surface lactoferrin.

    PubMed

    Kuo, Yung-Chih; Wang, Cheng-Ting

    2014-07-01

    A liposomal system with surface lactoferrin (Lf) was developed for delivering neuron growth factor (NGF) across the blood-brain barrier (BBB) and improving the viability of neuron-like SK-N-MC cells with deposited β-amyloid peptide (Aβ). The Lf-grafted liposomes carrying NGF (Lf/NGF-liposomes) were applied to a monolayer of human brain-microvascular endothelial cells (HBMECs) regulated by human astrocytes (HAs) and to fibrillar Aβ1-42-insulted SK-N-MC cells. An increase in cholesterol mole percentage enhanced the particle size, absolute value of zeta potential, and physical stability, however, reduced the entrapment efficiency and release rate of NGF. In addition, an increase in Lf concentration increased the particle size, surface nitrogen percentage, NGF permeability across the BBB, and viability of HBMECs, HAs, and SK-N-MC cells, however, decreased the absolute value of zeta potential, surface phosphorus percentage, and loading efficiency of Lf. After treating with Lf/NGF-liposomes, a higher Aβ concentration yielded a lower survival of SK-N-MC cells. The current Lf/NGF-liposomes are efficacious drug carriers to target the BBB and inhibit the Aβ-induced neurotoxicity as potential pharmacotherapy for Alzheimer's disease. Copyright © 2014 Elsevier Ltd. All rights reserved.

  5. Modulation of notch signaling pathway to prevent H2O2/menadione-induced SK-N-MC cells death by EUK134.

    PubMed

    Kamarehei, Maryam; Yazdanparast, Razieh

    2014-10-01

    The brain in Alzheimer's disease is under increased oxidative stress, and this may have a role in the pathogenesis and neural death in this disorder. It has been verified that numerous signaling pathways involved in neurodegenerative disorders are activated in response to reactive oxygen species (ROS). EUK134, a synthetic salen-manganese antioxidant complex, has been found to possess many interesting pharmacological activities awaiting exploration. The present study is to characterize the role of Notch signaling in apoptotic cell death of SK-N-MC cells. The cells were treated with hydrogen peroxide (H2O2) or menadione to induce oxidative stress. The free-radical scavenging capabilities of EUK134 were studied through the MTT assay, glutathione peroxidase (GPx) enzyme activity assay, and glutathione (GSH) Levels. The extents of lipid peroxidation, protein carbonyl formation, and intracellular ROS levels, as markers of oxidative stress, were also studied. Our results showed that H2O2/menadione reduced GSH levels and GPx activity. However, EUK134 protected cells against ROS-induced cell death by down-regulation of lipid peroxidation and protein carbonyl formation as well as restoration of antioxidant enzymes activity. ROS induced apoptosis and increased NICD and HES1 expression. Inhibition of NICD production proved that Notch signaling is involved in apoptosis through p53 activation. Moreover, H2O2/menadione led to Numb protein down-regulation which upon EUK134 pretreatment, its level increased and subsequently prevented Notch pathway activation. We indicated that EUK134 can be a promising candidate in designing natural-based drugs for ROS-induced neurodegenerative diseases. Collectively, ROS activated Notch signaling in SK-N-MC cells leading to cell apoptosis.

  6. Constant envelope OFDM scheme for 6PolSK-QPSK

    NASA Astrophysics Data System (ADS)

    Li, Yupeng; Ding, Ding

    2018-03-01

    A constant envelope OFDM scheme with phase modulator (PM-CE-OFDM) for 6PolSK-QPSK modulation was demonstrated. Performance under large fiber launch power is measured to check its advantages in counteracting fiber nonlinear impairments. In our simulation, PM-CE-OFDM, RF-assisted constant envelope OFDM (RF-CE-OFDM) and conventional OFDM (Con-OFDM) are transmitted through 80 km standard single mode fiber (SSMF) single channel and WDM system. Simulation results confirm that PM-CE-OFDM has best performance in resisting fiber nonlinearity. In addition, benefiting from the simple system structure, the complexity and cost of PM-CE-OFDM system could be reduced effectively.

  7. Modification of cutaneous vasodilator response to heat stress by daytime exogenous melatonin administration.

    PubMed

    Aoki, Ken; Stephens, Dan P; Zhao, Kun; Kosiba, Wojciech A; Johnson, John M

    2006-09-01

    In humans, the nocturnal fall in internal temperature is associated with increased endogenous melatonin and with a shift in the thermoregulatory control of skin blood flow (SkBF), suggesting a role for melatonin in the control of SkBF. The purpose of this study was to test whether daytime exogenous melatonin would shift control of SkBF to lower internal temperatures during heat stress, as is seen at night. Healthy male subjects (n = 8) underwent body heating with melatonin administration (Mel) or without (control), in random order at least 1 wk apart. SkBF was monitored at sites pretreated with bretylium to block vasoconstrictor nerve function and at untreated sites. Cutaneous vascular conductance, calculated from SkBF and arterial pressure, sweating rate (SR), and heart rate (HR) were monitored. Skin temperature was elevated to 38 degrees C for 35-50 min. Baseline esophageal temperature (Tes) was lower in Mel than in control (P < 0.01). The Tes threshold for cutaneous vasodilation and the slope of cutaneous vascular conductance with respect to Tes were also lower in Mel at both untreated and bretylium-treated sites (P < 0.05). The Tes threshold for the onset of sweating and the Tes for a standard HR were reduced in Mel. The slope of the relationship of HR, but not SR, to Tes was lower in Mel (P < 0.05). These findings suggest that melatonin affects the thermoregulatory control of SkBF during hyperthermia via the cutaneous active vasodilator system. Because control of SR and HR are also modified, a central action of melatonin is suggested.

  8. The sanitary state of Pomeranian Bay and Gulf of Gdańsk waters during the flood of 1997

    NASA Astrophysics Data System (ADS)

    Michalska, Małgorzata; Bartoszewicz, Maria

    1998-06-01

    Due to the 1997 summer flood, the Pomeranian and Gdańsk Bays were polluted with a significant amount of bacteriologically contaminated Oder and Vistula rivers waters. The purpose of this study was to determine the sanitary state of both bays during the flood and directly after it, the range of direction of distribution of the bacteriological contamination brought in by Vistula and Oder waters. On the basis of examination results, the sanitary state of the bays waters was found to be worse, particularly with regard to indicators: total bacterial number (at 20 °C and 37 °C) and Most Probable Number (MPN) of coliform and faecal coliform bacteria. Bacteria of the Pseudomonas group (Pseudomonas aeruginosa, Pseudomonas fluorescens), Enterobacteriaceae group (m.in. Proteus vulgaris), as well as enterococci (Streptoccus) and staphylococci (Staphylococcus) were found in water samples from Pomeranian and Gdańsk Bays. In the Pomeranian Bay the waters spread to the west (due to weather conditions), whereas in Gdańsk Bay they remained mainly in the coastal zone and then moved to the east. An improvement in the sanitary state of both bay waters was observed at the end of August.

  9. Structural elucidation of polysaccharide fractions from the brown alga Coccophora langsdorfii and in vitro investigation of their anticancer activity.

    PubMed

    Imbs, Tatiana I; Ermakova, Svetlana P; Malyarenko Vishchuk, Olesya S; Isakov, Vladimir V; Zvyagintseva, Tatiana N

    2016-01-01

    Laminaran, fucoidan, and alginate were isolated from the brown alga Coccophora langsdorfii collected in the Japan Sea. The structural characteristics of polysaccharides were investigated by NMR spectroscopy. The laminaran was determined as β-d-glucan, which consisted of 80% of 1,3- and 20% of 1,6-linked residues and was terminated with mannitol. The alginate was a guluronic acid-rich polysaccharide (M/G=0.85). Fucoidan, sulfated α-l-fucan, contained a linear backbone of alternating (1→3)- and (1→4)- linked α-l-fucopyranose residues with sulfate at C2 and C4 of (1→3)-α-l-fucopyranose residues. Anticancer activity of this fucoidan was investigated in comparison with activity of fucoidan having similar linear backbone from the brown alga Fucus evanescens. The fucoidan from C. langsdorfii significantly inhibited colony formation of SK-MEL-5 and SK-MEL-28 melanoma cells (the percentage of inhibition was 28 and 76, respectively) and weakly inhibited colony formation of breast adenocarcinoma cells MDA-MB-231 (the percentage of inhibition was about 5). Similar results were obtained for fucoidan from F. evanescens; the percentage of inhibition of colony formation of SK-MEL-5 and SK-MEL-28 melanoma cells was 54 and 56, respectively. The inhibition of colony formation of breast adenocarcinoma cells MDA-MB-231 was weak. We suppose that other sulfated and partially acetylated fucoidans consisting of (1→3)- and (1→4)-linked α-l-fucopyranose residues may suppress progression of melanoma cell colony formation similar to fucoidans of C. langsdorfii and F. evanescens. Copyright © 2015 Elsevier Ltd. All rights reserved.

  10. Ethanolic Extract of Agaricus blazei Fermentation Product Inhibits the Growth and Invasion of Human Hepatoma HA22T/VGH and SK-Hep-1 Cells

    PubMed Central

    Tung, Yen-Chen; Su, Zheng-Yuan; Kuo, Min-Liang; Sheen, Lee-Yan

    2012-01-01

    Hepatoma is a leading cause of death in the world. SK-Hep-1 and HA22T/VGH cells are poorly differentiated human hepatocellular carcinoma cell lines with invasive and migratory abilities. Agaricus blazei (AB) is a mushroom with many biological effects and active ingredients, and the ethanolic extract of AB fermentation product (AB-pE) was demonstrated to inhibit the growth of hepatoma Hep3B and HepG2 cells in our previous study. In this study, we further investigated the anticancer and anti-invasive abctivities of the AB-pE. Results showed that the AB-pE inhibited the growth of SK-Hep1 and HA22T/VGH cells (with IC50 values of 26.8 and 28.7 μg/mL, respectively) and led cells toward apoptosis after 48 h of treatment. Activation of caspase-3 by AB-pE (12.5~200 μg/mL) in a dose-dependent manner was observed in both cell lines using fluorescence microscopy and flow cytometry. The apoptosis triggered by the AB-pE was regulated by the increased expression of Bax, the activation of caspase-3, caspase-9, and PARP, and the decreased expression of Bcl-2. Additionally, the AB-pE showed the potential ability to inhibit invasion of SK-Hep1 and HA22T/VGH cells according to the results of a Matrigel invasion assay. Our results suggested that the AB-pE may be a further developed for its potential against hepatoma due to its antiproliferative (via apoptosis) and anti-invasive activities in hepatoma cells. PMID:24716127

  11. Neuroprotection against apoptosis of SK-N-MC cells using RMP-7- and lactoferrin-grafted liposomes carrying quercetin

    PubMed Central

    Kuo, Yung-Chih; Tsao, Chien-Wei

    2017-01-01

    A drug delivery system of quercetin (QU)-encapsulated liposomes (LS) grafted with RMP-7, a bradykinin analog, and lactoferrin (Lf) was developed to permeate the blood–brain barrier (BBB) and rescue degenerated neurons, acting as an Alzheimer’s disease (AD) pharmacotherapy. This colloidal formulation of QU-encapsulated LS grafted with RMP-7 and Lf (RMP-7-Lf-QU-LS) was used to traverse human brain microvascular endothelial cells (HBMECs) regulated by human astrocytes (HAs) and to treat SK-N-MC cells after an insult with cytotoxic β-amyloid (Aβ) fibrils. We found that surface RMP-7 and Lf enhanced the ability of QU to cross the BBB without inducing strong toxicity and damaging the tight junction. In addition, RMP-7-Lf-QU-LS significantly reduced Aβ-induced neurotoxicity and improved the viability of SK-N-MC cells. Compared with free QU, RMP-7-Lf-QU-LS could also significantly inhibit the expression of phosphorylated c-Jun N terminal kinase, phosphorylated p38, and phosphorylated tau protein at serine 202 by SK-N-MC cells, indicating an important role of RMP-7, Lf, and LS in protecting neurons against apoptosis. RMP-7-Lf-QU-LS is a promising carrier targeting the BBB to prevent Aβ-insulted neurodegeneration and may have potential in managing AD in future clinical applications. PMID:28435263

  12. Dissipation processes in the insulating skyrmion compound Cu2OSeO3

    NASA Astrophysics Data System (ADS)

    Levatić, I.; Šurija, V.; Berger, H.; Živković, I.

    2014-12-01

    We present a detailed study of the phase diagram surrounding the skyrmion lattice (SkL) phase of Cu2OSe2O3 using high-precision magnetic ac susceptibility measurements. An extensive investigation of transition dynamics around the SkL phase using the imaginary component of the susceptibility revealed that at the conical-to-SkL transition a broad dissipation region exists with a complex frequency dependence. The analysis of the observed behavior within the SkL phase indicates a distribution of relaxation times intrinsically related to SkL. At the SkL-to-paramagnet transition a narrow first-order peak is found that exhibits a strong frequency and magnetic field dependence. Surprisingly, very similar dependence has been discovered for the first-order transition below the SkL phase, i.e., where the system enters the helical and conical state(s), indicating similar processes across the order-disorder transition.

  13. [Maria Skłodowska-Curie--her chemistry at the centenary of the second Nobel Prize].

    PubMed

    Zagórski, Zbigniew Paweł; Kornacka, Ewa Maria

    2012-01-01

    The article presents from the perspective of one hundred years the work of Maria Curie-Skłodowska, which in many cases was ahead of the state of knowledge of the time. It opened new horizons and for this reason we made many digressions. The fact of awarding her the Nobel Prize twice is a sensation enough to present the values of careful activity of the Nobel Prize Committee that emphasizes the importance of Maria's achievements. A significant element of Maria Skłodowska-Curie's achievements was still mysterious character of the radiation in her time, and only chemical approach made it possible to organise the phenomena and explain the origin of the radiation. The essence of the research was an arduous separation of components following the track of growing radiation of successive fractions of preparations. This research was a start of the technology of educement of dispersed elements in great mass of materials. We underline the paramount role of the chemical research Maria Skłodowska conducted while still in Warsaw in the laboratories of the Museum of Industry and Agriculture under the guidance of an excellent chemist Józef Jerzy Boguski. Her research in Paris was the origin of the semi-commercial scale in chemistry and setting aside a special shed outside the university building was the beginning of the institutes that now function beyond universities and are key element of scientific and technical progress. Technology of splitting developed by Maria Skłodowska-Curie was applied also by other radiochemists, e.g. By Otto Hahn. Lively movement in radiochemistry of her lifetime resulted in Maria's disputes with e.g. German chemist Marckwald, who questioned the originality of polonium. The scientific disputes like this one Maria won triumphantly although in several others she had to accept opponents' argument, as in the case of radon. Her experiments were planned with utmost rationality as it was with the rejection of the hypothesis saying that radioactivity was

  14. Under the glacier, the groundwater - the case of Skálafell area, Iceland

    NASA Astrophysics Data System (ADS)

    Vincent, Aude; Hart, Jane

    2017-04-01

    The research addressing glaciers evolution under climate change is well developed, and is now looking not only at their mass balance, but also at the associated subsurface hydrology and downstream hydrology. However, the groundwater component is rarely considered, even though it will be required to forecast the evolution of water resources and of water linked hazards under climate change. The few available studies demonstrate the existence of sub-or pro-glacial aquifers. Some of them suggest strong coupling between rivers and the aquifer, observe the flooding due to water table rising following enhanced glacier melting, or expect stronger recharge in the future due to glacier melting. The present study is the first step of a wider project, GlacAq, aiming at filling this knowledge gap, by characterizing the particular hydrogeology encountered under and downstream of glaciers of alpine type, i.e. sub-, pro- and periglacial hydrogeology, and its sensibility to climate change, in order to provide operational management directions. Skálafell glacier area (Iceland) has been chosen as it has already been followed for climatic, glaciological, and surface hydrology data (Hart et al. (2015), Young et al. (2015)). The present work will use those data, as well as topographic and surface data from the National Land Survey of Iceland, and geological data, to run a comprehensive numerical modelling. The work conducted on the Skálafell site will lead both to the achievement of an operational understanding of a poorly known underground system, and to the anticipation of its hydrodynamic response to climate change. The foreseen mechanisms include an enhanced sub-glacial aquifer recharge, intense surface water bodies-aquifer exchanges, and the aquifer discharge either through springs, or to an offshore system. Those offshore stocks are being increasingly recognised, but their origins are still only guessed at. Skálafell site allows the exploration of the potential role of the

  15. High-sensitive LC-MS/MS method for the simultaneous determination of mirodenafil and its major metabolite, SK-3541, in human plasma: application to microdose clinical trials of mirodenafil.

    PubMed

    Cho, Doo-Yeoun; Bae, Soo Hyeon; Shon, Ji-Hong; Bae, Soo Kyung

    2013-03-01

    A high-sensitivity LC/MS/MS method was developed and validated for the simultaneous determination of mirodenafil and its major metabolite, SK-3541, in human plasma. Mirodenafil, SK-3541, and udenafil as an internal standard were extracted from plasma samples with methyl tert-butyl ether. Chromatographic separation was performed on a Luna phenyl-hexyl column (100 × 2.0 mm) with an isocratic mobile phase consisting of 5 mM ammonium formate and ACN (23:77, v/v) at a flow rate of 0.35 mL/min. Detection and quantification were performed using a mass spectrometer in selected reaction monitoring mode with positive ESI at m/z 532.3 → 296.1 for mirodenafil, m/z 488.1 → 296.1 for SK-3541, and m/z 517.3 → 283.2 for udenafil. The calibration curves were linear over a concentration range of 2-500 pg/mL using 0.5 mL plasma for the microdose of mirodenafil (100 μg). Analytical method validation of the clinical dose (100 mg), with a calibration curve range of 2-500 ng/mL using 0.025-mL plasma, was also conducted. The other LC-MS/MS conditions were similar to those used for the microdosing. Each method was applied successfully to pharmacokinetic studies after a microdose or clinical dose of mirodenafil to six healthy Korean male volunteers. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  16. Atmospheric deposition and riverine load of (90)Sr and (137)Cs to the Gulf of Gdańsk (southern Baltic Sea) in the period 2005-2011.

    PubMed

    Saniewski, Michał; Zalewska, Tamara

    2016-01-01

    In the period 2005-2011 total atmospheric fallout and the riverine input to the Gulf of Gdańsk was 1168.8 GBq of (90)Sr and 424.1 GBq (137)Cs. The major source of both radionuclides to the Gulf of Gdańsk is the Vistula river; its contribution reached 99.7% in the case of (90)Sr and 95.8% regarding (137)Cs. The atmospheric load of (137)Cs, 18.1 GBq, was nearly 4 times bigger than in the case of (90)Sr (3.75 GBq). In the period 2005-2010, the average annual atmospheric load were at the levels 2-3 GBq for (137)Cs and 0.4-0.6 GBq for (90)Sr, while in 2011, due to the Fukuchima Dai-ichi power plant break down, an increase of annual atmospheric loads was noted to 5.3 GBq of (137)Cs and to 0.87 GBq of (90)Sr. The additional loads did not have an increasing effect on the activity concentrations of (137)Cs and (90)Sr in seawater of the Gulf of Gdańsk, where mean activity concentrations in seawater were equal to 31.1 Bq m(-3) and 7.6 Bq m(-3) in the case of (137)Cs and (90)Sr respectively. Copyright © 2015 Elsevier Ltd. All rights reserved.

  17. The effects of different antineoplastic agents and of pretreatment by modulators on three melanoma lines.

    PubMed

    Rodriguez-Vicente, J; Vicente-Ortega, V; Canteras-Jordana, M

    1998-02-01

    The chemotherapy of melanoma patients must be improved because of the naturally poor response and acquired resistance of this disease. The authors used mouse (B16F10) and human (SK-MEL-28 and SK-MEL-1) melanoma lines for in vitro treatment with melphalan, lomustine, fotemustine, and 4-hydroxyanisole (4-HA) alone, combined and after pretreatment with buthionine sulfoximine (BSO), ethacrynic acid (EA), and azelaic acid (AZA). Melphalan was the most effective individual drug, followed by lomustine, fotemustine, and 4-HA. The simultaneous administration of two agents was disappointing, although some combinations slightly improved the response compared with the individual treatments. Pretreatment with BSO enhanced the cytotoxicity of melphalan and lomustine 10-fold in B16F10 and 7.5-fold in SK-MEL-28, increasing the toxicity of fotemustine in all 3 lines. EA potentiated lomustine and fotemustine 9-fold and melphalan 5-fold in B16F10 and SK-MEL-28. AZA increased the effectiveness of lomustine and fotemustine in B16F10 and to a lower degree in the two human lines. 4-HA was the poorest drug for sensitization; only B16F10 BSO followed by 4-HA treatment demonstrated increased toxicity, and all other combinations with 4-HA were negative or antagonistic. There was a strong relationship between dopa oxidase activity and the toxicity of 4-HA. B16F10 was the most sensitive to all treatments and SK-MEL-1 the most resistant. Melphalan was the most active individual drug and 4-HA the least. Combinations of two drugs did not result in improved activity compared with drugs administered alone. Pretreatment with modulator seems to be a potential method for enhancing some treatments.

  18. P2Y receptor-mediated transient relaxation of rat longitudinal ileum preparations involves phospholipase C activation, intracellular Ca(2+) release and SK channel activation.

    PubMed

    Mader, Felix; Krause, Ludwig; Tokay, Tursonjan; Hakenberg, Oliver W; Köhling, Rüdiger; Kirschstein, Timo

    2016-05-01

    Purinergic signaling plays a major role in the enteric nervous system, where it governs gut motility through a number of P2X and P2Y receptors. The aim of this study was to investigate the P2Y receptor-mediated motility in rat longitudinal ileum preparations. Ileum smooth muscle strips were prepared from rats, and fixed in an organ bath. Isometric contraction and relaxation responses of the muscle strips were measured with force transducers. Drugs were applied by adding of stock solutions to the organ bath to yield the individual final concentrations. Application of the non-hydrolyzable P2 receptor agonists α,β-Me-ATP or 2-Me-S-ADP (10, 100 μmol/L) dose-dependently elicited a transient relaxation response followed by a sustained contraction. The relaxation response was largely blocked by SK channel blockers apamin (500 nmol/L) and UCL1684 (10 μmol/L), PLC inhibitor U73122 (100 μmol/L), IP3 receptor blocker 2-APB (100 μmol/L) or sarcoendoplasmic Ca(2+) ATPase inhibitor thapsigargin (1 μmol/L), but not affected by atropine, NO synthase blocker L-NAME or tetrodotoxin. Furthermore, α,β-Me-ATP-induced relaxation was suppressed by P2Y1 receptor antagonist MRS2179 (50 μmol/L) or P2Y13 receptor antagonist MRS2211 (100 μmol/L), and was abolished by co-application of the two antagonists, whereas 2-Me-S-ADP-induced relaxation was abolished by P2Y6 receptor antagonist MRS2578 (50 μmol/L). In addition, P2Y1 receptor antagonist MRS2500 (1 μmol/L) not only abolished α,β-Me-ATP-induced relaxation, but also suppressed 2-Me-S-ADP-induced relaxation. P2Y receptor agonist-induced transient relaxation of rat ileum smooth muscle strips is mediated predominantly by P2Y1 receptor, but also by P2Y6 and P2Y13 receptors, and involves PLC, IP3, Ca(2+) release and SK channel activation, but is independent of acetylcholine and NO release.

  19. Neuroprotective effects of phytosterols and flavonoids from Cirsium setidens and Aster scaber in human brain neuroblastoma SK-N-SH cells.

    PubMed

    Chung, Mi Ja; Lee, Sanghyun; Park, Yong Il; Lee, Jisun; Kwon, Ki Han

    2016-03-01

    We investigated the neuroprotective effects and action mechanism of three major compounds [daucosterol (Dau), pectolinarin (Pec), and astragalin (Ast)] isolated from edible plants against H2O2-induced cell death of human brain neuroblastoma SK-N-SH cells. Cytotoxicity was determined by MTT and lactate dehydrogenase (LDH) assays. Apoptotic cell death was monitored by annexin V-FITC/PI double staining and by TUNEL assay. The formation of reactive oxygen species (ROS), expression of antioxidant enzymes and phosphorylation of mitogen-activated protein kinase (MAPK) were determined by 2,7-dichlorofluorescein diacetate (DCF-DA) assay, RT-PCR, and western blotting, respectively. The ethyl acetate fractions from Cirsium setidens (CSEA) and Aster scaber (ASEA) showed neuroprotective effects in SK-N-SH cells. The phytochemicals were isolated from CSEA and ASEA and identified by spectral analyses, as β-sitosterol, Dau, Pec, Ast, or isoquercitrin. Pretreatment with Dau, Pec, or Ast showed protective effects against H2O2-induced cell death and inhibited ROS generation by oxidative stress. HO-1 mRNA and protein levels were increased by the presence of H2O2 and were further elevated by pretreatment with Dau and Ast. Dau pretreatment resulted in further increases of H2O2-induced enhancement in levels of CAT and SOD2. Pretreatment with Dau, Pec, and Ast inhibited phosphorylation of MAPK, such as extracellular protein regulated protein kinase, p38, and c-Jun N-terminal kinase by H2O2. Dau exerts its neuroprotective effects by down regulation of MAPK pathways and upregulation of the HO-1, CAT and SOD2 antioxidant genes and is associated with reduced oxidative stress in SK-N-SH cells. Copyright © 2016 Elsevier Inc. All rights reserved.

  20. [Prevalence of the Diabetic Retinopathy and Genetic Factors Significance in the Development of Diabetic Retinopathy in Patients with Diabetes Mellitus type I and II in Slovakia (DIARET SK study). Overview of Actual Findings and Design of the Epidemiological DIARET SK Study].

    PubMed

    Krásnik, V; Štefaničková, J; Fabková, J; Bucková, D; Helbich, M

    2015-09-01

    Diabetic retinopathy (DR) is the second most common microvascular complication and the most common cause of blindness in patients with diabetes mellitus (DM). Despite the ongoing research, the findings of diabetic retinopathy epidemiological and risk factors are, until now, not consistent. More finding may be revealed by epidemiological studies, consistently mapping DR epidemiology under the current possibilities of investigations and treatment of the DM. DIARET SK Study, with 5 000 enrolled patients with diabetes mellitus in the Slovak Republic, is, until now, the largest epidemiological study to set the prevalence of diabetic retinopathy. The primary aim is to establish the prevalence of diabetic retinopathy in patients with diabetes mellitus type I and II, according to the duration of the disease. The secondary aim is to establish prevalence of the different stages of the DR and diabetic macular edema (DME) and analysis of the risk factors influence. Included are patients with DM type I and II regardless to the ocular complications history and the period of DM duration. Each enrolled patient has both complex diabetic and ophthalmic examinations.Projects to establish DR prevalence: Tens of projects concerned with diabetic retinopathy epidemiology with different approaches to establish the prevalence and with different patients population. Results from different studies vary significantly (from 12.3 % to 66.9 %). The results depend on the design of the study and the patients recruitment, used examination methods, specific patients population with regard to the geography, prevalence of risk factors, period of diabetes duration, glycated hemoglobin (HbA1C) level, blood pressure, and is higher in type I diabetic patients. The most accurate results are from population epidemiological studies with well-controlled patient recruitment and uniform complex examination that are similar to the DIARET SK study. The DIARET SK study represents the largest epidemiological study

  1. Zircon and baddeleyite from the economic ultramafic-mafic Noril'sk-1 intrusion (Russia): Hf-isotope constraints on source composition

    NASA Astrophysics Data System (ADS)

    Malitch, K. N.; Belousova, E. A.; Badanina, I. Yu.; Griffin, W. L.

    2012-04-01

    The ultramafic-mafic Noril'sk-1 intrusion in the northwestern part of the Siberian Craton (Russia) represents one of three known Noril'sk-type, ore-bearing intrusions, which host one of the world's major economic sulphide platinum-group-element (PGE)-Cu-Ni deposits. Zircon and baddeleyite dated previously both by SHRIMP (i.e. 248.0 ± 3.7 Ma, Campbell et al. 1992) and ID-TIMS (251.1 ± 3.6 Ma, Kamo et al. 1996) have been restricted to one lithology (e.g. leucogabbro) of the Noril'sk-1 intrusion. To better constrain the age of igneous event and sources involved in its generation our multi-technique study utilized ten rock samples characteristic of unmineralized and mineralized lithologies. The rocks investigated comprise (from top to bottom) gabbro-diorite (sample N1-1), leucogabbro (N1-3), olivine-free gabbro (N1-2 and N1-4), olivine-bearing gabbro (N1-5), olivine gabbro (N1-6), plagiowehrlite and plagiodunite (N1-7), taxitic-textured rocks comprising melanotroctolite, olivine gabbro with relics of ultramafic rocks (N1-8, N1-9) and contact fine-grained gabbro (N1-10). Sulphide PGE-Cu-Ni ores occur in ultramafic (N1-7) and taxitic-textured rocks (N1-8 and N1-9), which have thickness of about 17 m, whereas the low-sulphide horizon of about 1 m thick occurs in the upper part of intrusion (N1-3). In situ U-Pb analyses of zircon from these rocks, combined with detailed study of crystal morphology and internal structure, identify four zircon populations (Malitch et al. 2012). The U-Pb ages of baddeleyite and the defined zircon populations cover a significant time span, from Late Paleozoic to Early Mesozoic (e.g., 290 ± 2.8; 261.3 ± 1.6; 245.7 ± 1.1; 236.5 ± 1.8 and 226.7 ± 0.9 Ma). The established distribution of U-Pb ages implies that crystallization of baddeleyite and zircon corresponds to several stages of protracted evolution of ultramafic-mafic magmas at deep-seated staging chambers and/or probably characterizes interaction of distinct magmas during formation

  2. Immunization of mice with LRP4 induces myasthenia similar to MuSK-associated myasthenia gravis.

    PubMed

    Mori, Shuuichi; Motohashi, Norio; Takashima, Rumi; Kishi, Masahiko; Nishimune, Hiroshi; Shigemoto, Kazuhiro

    2017-11-01

    Since the first report of experimental animal models of myasthenia gravis (MG) with autoantibodies against low-density lipoprotein receptor-related protein 4 (LRP4), there have not been any major reports replicating the pathogenicity of anti-LRP4 antibodies (Abs). Recent clinical studies have cast doubt on the specificity and pathogenicity of anti-LRP4 antibodies for MG, highlighting the need for further research. In this study, we purified antigens corresponding to the extracellular region of human LRP4 stably expressed with chaperones in 293 cells and used these antigens to immunize female A/J mice. Immunization with LRP4 protein caused mice to develop myasthenia having similar electrophysiological and histological features as are observed in MG patients with circulating Abs against muscle-specific kinase (MuSK). Our results clearly demonstrate that active immunization of mice with LRP4 proteins causes myasthenia similar to the MG induced by anti-MuSK Abs. Further experimental and clinical studies are required to prove the pathogenicity of anti-LRP4 Abs in MG patients. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

  3. Application of geochemical logging for palaeoenvironmental research in the Late Cretaceous Qingshankou Formation from the Chinese Continental Scientific Drilling Project-SK-2e, Songliao Basin, NE China

    NASA Astrophysics Data System (ADS)

    Peng, Cheng; Zou, Changchun; Pan, Li; Niu, Yixiong

    2017-08-01

    The Chinese Continental Scientific Drilling Project of the Cretaceous Songliao Basin (CCSD-SK) provides an excellent opportunity to understand the response of terrestrial environments to greenhouse climate change in the Cretaceous. We conducted a palaeoenvironmental study of the Late Cretaceous Qingshankou Formation (K2qn) based on geochemical log data from the SK-2 east borehole. According to the characteristic of Ti mainly from terrigenous minerals, the content of authigenic elements was calculated. Correlation space was proposed to study the variation of the correlation between two log curves along the depth. Palaeoenvironmental proxies were selected from log data to study the evolution of the climate and lake, productivity of the paleolake, and organic matter deposition. The results demonstrate that the productivity of the paleolake was driven by chemical weathering in K2qn, in which the first section of the Qingshankou Formation (K2qn1) has higher productivity than the second and third sections of the Qingshankou Formation (K2qn2+3). The high content of pyrite in several thin layers reveals lake water of high sulfate concentration. This may have been caused by acid rain related to large volcanic activity. In K2qn2+3, several periods of high productivity without the formation of source rocks and high organic matter content were identified. This may show that organic matter deposition was limited by low accommodation space or oxidation environment. Therefore, the preservation condition is suggested as the main controlling factor of organic matter deposition in K2qn.

  4. KLF4 overexpression and apigenin treatment down regulated anti-apoptotic Bcl-2 proteins and matrix metalloproteinases to control growth of human malignant neuroblastoma SK-N-DZ and IMR-32 cells.

    PubMed

    Mohan, Nishant; Ai, Walden; Chakrabarti, Mrinmay; Banik, Naren L; Ray, Swapan K

    2013-06-01

    Neuroblastoma is a childhood tumor that arises from immature neuroblasts of the sympathetic nervous system. Krüpple-like factor 4 (KLF4) is a transcription factor, the precise function of which in neuroblastoma is unclear. We examined the effects of KLF4 overexpression and apigenin (APG) treatment in human malignant neuroblastoma SK-N-DZ and IMR-32 cell lines. KLF4 overexpression in both SK-N-DZ and IMR-32 cell lines was confirmed by laser scanning immunofluorescent confocal microscopy and Western blotting. We found that 100 nM KLF4 plasmid and 25 μM APG synergistically inhibited the growth of SK-N-DZ and IMR-32 cells. We also found increase in KLF4 expression in response to treatment with various concentrations of APG. Combination of KLF4 plasmid and APG treatment significantly increased the amounts of apoptosis in both cell lines when compared with control vector or single treatment. We also noticed that the combination therapy decreased expression of the anti-apoptotic proteins Bcl-2 and Mcl-1, increased expression of the pro-apoptotic proteins Bax, Noxa, and Puma, upregulated p53, and caused activation of caspase-3 for cleavage of the inhibitor of caspase-activated DNase (ICAD) leading to completion of apoptosis machinery. Further, combination of KLF4 overexpression and APG treatment was highly effective in inhibiting migration of both neuroblastoma cell lines and was associated with down regulation of matrix metalloproteinases (MMPs) such as MMP-2 and MMP-9. Collectively, our results from this investigation strongly suggest that KLF4 functions as a tumor suppressor and potentiates the anti-cancer activities of APG in two different human malignant neuroblastoma cell lines. Copyright © 2012 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

  5. Draft Genome Sequence of Cryophilic Basidiomycetous Yeast Mrakia blollopis SK-4, Isolated from an Algal Mat of Naga-ike Lake in the Skarvsnes Ice-Free Area, East Antarctica.

    PubMed

    Tsuji, Masaharu; Kudoh, Sakae; Hoshino, Tamotsu

    2015-01-22

    Mrakia blollopis strain SK-4 was isolated from an algal mat of Naga-ike, a lake in Skarvsnes, East Antarctica. Here, we report the draft genome sequence of M. blollopis SK-4. This is the first report on the genome sequence of any cold-adapted fungal species. Copyright © 2015 Tsuji et al.

  6. Genetic Pd, Pt, Au, Ag, and Rh mineralogy in Noril'sk sulfide ores

    NASA Astrophysics Data System (ADS)

    Spiridonov, E. M.; Kulagov, E. A.; Serova, A. A.; Kulikova, I. M.; Korotaeva, N. N.; Sereda, E. V.; Tushentsova, I. N.; Belyakov, S. N.; Zhukov, N. N.

    2015-09-01

    The undeformed ore-bearing intrusions of the Noril'sk ore field (NOF) cut through volcanic rocks of the Late Permian-Early Triassic trap association folded in brachysynclines. Due to the nonuniform load on the roof of intrusive bodies, most sulfide melts were squeezed, up to the tops of ore-bearing intrusions; readily fusible Ni-Fe-Cu sulfide melts were almost completely squeezed. In our opinion, not only one but two stages of mineralization developed at the Noril'sk deposits: (i) syntrap magmatic and (ii) epigenetic post-trap metamorphic-hydrothermal. All platinum-group minerals (PGM) and minerals of gold are metasomatic in the Noril'sk ores. They replaced sulfide solid solutions and exsolution structures. All types of PGM and Au minerals occur in the ores, varying in composition from pyrrhotite to chalcopyrite, talnakhite, mooihoekite, and rich in galena; they are localized in the inner and outer contact zones and differ only in the quantitative proportions of ore minerals. The aureoles of PGM and Au-Ag minerals are wider than the contours of sulfide bodies and coincide with halos of fluid impact on orebodies and adjacent host rocks. The pneumatolytic PGM and Au-Ag minerals are correlated in abundance with the dimensions of sulfide bodies. Their amounts are maximal in veins of late fusible ore composed of eutectic PbS ss and iss intergrowths, as well as at their contacts. The Pd and Pt contents in eutectic sulfide ores of NOF are the world's highest. In the process of noble-metal mineral formation, the fluids supply Pd, Pt, Au, As, Sb, Sn, Bi, and a part of Te, whereas Fe, Ni, Cu, Pb, Ag, Rh, a part of Te and Pd are leached from the replaced sulfide minerals. The pneumatolytic PGM of the early stage comprises Pd and Pt intermetallic compounds enriched in Au along with Pd-Pt-Fe-Ni-Cu-Sn-Pb(As) and (Pd,Pt,Au)(Sn,Sb,Bi,Te,As) solid solutions. Pneumatolytic PGM and Au minerals of the middle stage are products of solid-phase transformation and recrystallization of

  7. FOXP3 over-expression inhibits melanoma tumorigenesis via effects on proliferation and apoptosis.

    PubMed Central

    Tan, BeeShin; Anaka, Matthew; Deb, Siddhartha; Freyer, Claudia; Ebert, Lisa M.; Chueh, Anderly C.; Al-Obaidi, Sheren; Behren, Andreas; Jayachandran, Aparna; Cebon, Jonathan; Chen, Weisan; Mariadason, John M.

    2014-01-01

    The Forkhead box P3 (FOXP3) transcription factor is the key driver of regulatory T cell (Treg cells) differentiation and immunosuppressive function. In addition, FOXP3 has been reported to be expressed in many tumors, including melanoma. However, its role in tumorigenesis is conficting, with both tumor suppressive and tumor promoting functions described. The aim of the current study was to characterize the expression and function of FOXP3 in melanoma. FOXP3 expression was detected by immunohistochemistry (IHC) in 12% (18/146) of stage III and IV melanomas. However expression was confined to fewer than 1% of cells in these tumors. Stable over-expression of FOXP3 in the SK-MEL-28 melanoma cell line reduced cell proliferation and clonogenicity in vitro, and reduced xenograft growth in vivo. FOXP3 over-expression also increased pigmentation and the rate of apoptosis of SK-MEL-28 cells. Based on its infrequent expression in human melanoma, and its growth inhibitory and pro-apoptotic effect in over-expressing melanoma cells, we conclude that FOXP3 is not likely to be a key tumor suppressor or promoter in melanoma. PMID:24406338

  8. FOXP3 over-expression inhibits melanoma tumorigenesis via effects on proliferation and apoptosis.

    PubMed

    Tan, BeeShin; Anaka, Matthew; Deb, Siddhartha; Freyer, Claudia; Ebert, Lisa M; Chueh, Anderly C; Al-Obaidi, Sheren; Behren, Andreas; Jayachandran, Aparna; Cebon, Jonathan; Chen, Weisan; Mariadason, John M

    2014-01-15

    The Forkhead box P3 (FOXP3) transcription factor is the key driver of regulatory T cell (Treg cells) differentiation and immunosuppressive function. In addition, FOXP3 has been reported to be expressed in many tumors, including melanoma. However, its role in tumorigenesis is conflicting, with both tumor suppressive and tumor promoting functions described. The aim of the current study was to characterize the expression and function of FOXP3 in melanoma. FOXP3 expression was detected by immunohistochemistry (IHC) in 12% (18/146) of stage III and IV melanomas. However expression was confined to fewer than 1% of cells in these tumors. Stable over-expression of FOXP3 in the SK-MEL-28 melanoma cell line reduced cell proliferation and clonogenicity in vitro, and reduced xenograft growth in vivo. FOXP3 over-expression also increased pigmentation and the rate of apoptosis of SK-MEL-28 cells. Based on its infrequent expression in human melanoma, and its growth inhibitory and pro-apoptotic effect in over-expressing melanoma cells, we conclude that FOXP3 is not likely to be a key tumor suppressor or promoter in melanoma.

  9. Effects of the Hematoregulatory Peptide SK&F 107647 Alone and in Combination with Amphotericin B against Disseminated Candidiasis in Persistently Neutropenic Rabbits

    PubMed Central

    Lyman, Caron A.; Gonzalez, Corina; Schneider, Mark; Lee, James; Walsh, Thomas J.

    1999-01-01

    The effects of the hematoregulatory peptide SK&F 107647 were examined in a persistently and profoundly neutropenic rabbit model of disseminated candidiasis in order to determine its potential to enhance resistance against infection and its role as an adjunct to conventional antifungal chemotherapy. In healthy animals, SK&F 107647 elicited a time-dependent increase in CD11b-positive monocytes and neutrophils. When administered to neutropenic rabbits infected with Candida albicans, no significant differences in the number of CFU per gram in any of the tissues tested compared with the number in untreated control rabbits were detected. However, when SK&F 107647 was administered in combination with low doses of amphotericin B, there was a significant reduction in organism burden in the lungs, liver, spleen, and kidneys compared with the burdens in the organs of untreated control animals and in the lungs and kidneys compared with the burdens in the lungs and kidneys of animals treated with amphotericin B alone. These data suggest a potential role for this peptide as adjunctive therapy in combination with conventional antifungal agents in the treatment of disseminated candidiasis in the setting of profound and persistent neutropenia. PMID:10471559

  10. Alterporriol-Type Dimers from the Mangrove Endophytic Fungus, Alternaria sp. (SK11), and Their MptpB Inhibitions

    PubMed Central

    Xia, Guoping; Li, Jia; Li, Hanxiang; Long, Yuhua; Lin, Shao’e; Lu, Yongjun; He, Lei; Lin, Yongcheng; Liu, Lan; She, Zhigang

    2014-01-01

    A new alterporriol-type anthranoid dimer, alterporriol S (1), along with seven known anthraquinone derivatives, (+)-aS-alterporriol C (2), hydroxybostrycin (3), halorosellinia A (4), tetrahydrobostrycin (5), 9α-hydroxydihydrodesoxybostrycin (6), austrocortinin (7) and 6-methylquinizarin (8), were isolated from the culture broth of the mangrove fungus, Alternaria sp. (SK11), from the South China Sea. Their structures and the relative configurations were elucidated using comprehensive spectroscopic methods, including 1D and 2D NMR spectra. The absolute configurations of 1 and the axial configuration of 2 were defined by experimental and theoretical ECD spectroscopy. 1 was identified as the first member of alterporriols consisting of a unique C-10−C-2′ linkage. Atropisomer 2 exhibited strong inhibitory activity against Mycobacterium tuberculosis protein tyrosine phosphatase B (MptpB) with an IC50 value 8.70 μM. PMID:24840716

  11. Analysis and Risk Evaluation on the Case of Alteration, Revitalization and Conversion of a Historic Building in Gdańsk

    NASA Astrophysics Data System (ADS)

    Grzyl, Beata; Kristowski, Adam; Miszewska-Urbańska, Emilia

    2017-10-01

    Each investment plan, including the one concerning a building, is exposed to the consequences of various types of threats taking place. Therefore, in the case of some large-scale, atypical and complicated building ventures, some actions included in the procedure of risk management should be taken (identifications, analysis, measurements, control and supervision of the risk). This will allow for the risk to be eliminated or limited. While preparing a building venture, an investor does not possess full information about the course of events on each stage of investment completion. The identification of the above-mentioned unknowns, subjecting them to quantification and specifying the method of dealing with them, allows an investor to increase the effectiveness of the intended plan. The enterprise discussed in this article and analyzed in the context of risk, concerns alteration, revitalization and conversion for office purposes of two buildings located in Gdańsk at 1 and 2 Lastadia Street. These buildings are situated on the area of historical urban layout of Gdańsk, in the northern-eastern part of Stare Przedmieście District (Old Suburb), about 800 meters south from Dlugi Targ Street and 200 meters west from The Old Motława River. The investor is “Gdańskie Melioracje Ltd.”, a limited liability company, which belongs to the Council of Gdańsk. In order to increase the effectiveness of the intended investment venture, while organizing the investment process, the investor commissioned preparation of an analysis and risk evaluation connected with the above-mentioned intention. Based on an on-site visit, the opinions of experts, who have been involved in the process of the preparation of the investment, studies of the available monographies about the technical condition of the buildings at 1 and 2 Lastadia Street and their own experiences, the authors identified 54 types of relevant risks, which have been systematized into 10 subject groups (among others

  12. Kv7/KCNQ/M and HCN/h, but not KCa2/SK channels, contribute to the somatic medium after-hyperpolarization and excitability control in CA1 hippocampal pyramidal cells

    PubMed Central

    Gu, Ning; Vervaeke, Koen; Hu, Hua; Storm, Johan F

    2005-01-01

    In hippocampal pyramidal cells, a single action potential (AP) or a burst of APs is followed by a medium afterhyperpolarization (mAHP, lasting ∼0.1 s). The currents underlying the mAHP are considered to regulate excitability and cause early spike frequency adaptation, thus dampening the response to sustained excitatory input relative to responses to abrupt excitation. The mAHP was originally suggested to be primarily caused by M-channels (at depolarized potentials) and h-channels (at more negative potentials), but not SK channels. In recent reports, however, the mAHP was suggested to be generated mainly by SK channels or only by h-channels. We have now re-examined the mechanisms underlying the mAHP and early spike frequency adaptation in CA1 pyramidal cells by using sharp electrode and whole-cell recording in rat hippocampal slices. The specific M-channel blocker XE991 (10 μm) suppressed the mAHP following 1–5 APs evoked by current injection at −60 mV. XE991 also enhanced the excitability of the cell, i.e. increased the number of APs evoked by a constant depolarizing current pulse, reduced their rate of adaptation, enhanced the afterdepolarization and promoted bursting. Conversely, the M-channel opener retigabine reduced excitability. The h-channel blocker ZD7288 (4-ethylphenylamino-1,2-dimethyl-6-methylaminopyrimidinium chloride; 10 μm) fully suppressed the mAHP at −80 mV, but had little effect at −60 mV, whereas XE991 did not measurably affect the mAHP at −80 mV. Likewise, ZD7288 had little or no effect on excitability or adaptation during current pulses injected from −60 mV, but changed the initial discharge during depolarizing pulses injected from −80 mV. In contrast to previous reports, we found that blockade of Ca2+-activated K+ channels of the SK/KCa type by apamin (100–400 nm) failed to affect the mAHP or adaptation. A computational model of a CA1 pyramidal cell predicted that M- and h-channels will generate mAHPs in a voltage

  13. In situ S-K XANES study of polymer electrolyte fuel cells: changes in the chemical states of sulfonic groups depending on humidity.

    PubMed

    Isegawa, Kazuhisa; Nagami, Tetsuo; Jomori, Shinji; Yoshida, Masaaki; Kondoh, Hiroshi

    2016-09-14

    Changes in the chemical states of sulfonic groups of Nafion in polymer electrolyte fuel cells (PEFCs) under gas-flowing conditions were studied using in situ S-K XANES spectroscopy. The applied potential to the electrodes and the humidity of the cell were changed under flowing H 2 gas in the anode and He gas in the cathode. While the potential shows no significant effect on the S-K XANES spectra, the humidity is found to induce reversible changes in the spectra. Comparison of the spectral changes with simulations based on the density functional theory calculations indicates that the humidity influences the chemical state of the sulfonic group; under wet conditions the sulfonic group is in the form of a sulfonate ion. By drying treatment the sulfonate ion binds to hydrogen and becomes sulfonic acid. Furthermore, a small fraction of the sulfonic acid irreversibly decomposes to atomic sulfur. The peak energy of the atomic sulfur suggests that the generated atomic sulfur is adsorbed on the Pt catalyst surfaces.

  14. Noril'sk/Siberian plateau basalts and Bahama hot spot: Impact triggered?

    NASA Technical Reports Server (NTRS)

    Deitz, R. S.; Mchone, J. F.

    1992-01-01

    Twenty-eight years after one of us argued that Sudbury was an astrobleme, this interpretation has only recently attained wide acceptance; not so for the view that the Sudbury Cu/Ni sulfide ores are cosmogenic. Other research has provided the triggering of plateau basalts by super-large impacts a modicum of respectability. The recent apparent successful tying in of the K/T extinctions to the Chicxulub astrobleme in the Yucatan encourages the search for an impact event that may have caused the other two major post-Paleozoic extinctions (P/Tr, Tr/J). This gives us heart to offer two further outrageous hypotheses. The cosmogenic concept for the Sudbury ore deposite remains viable because it is giant, nonultramafic, and unique (except for Noril'sk). The Triassic/Jurassic boundary catastrophic extinctions have been attributed to the Manicouagan asteroidal impact, but recent radiometric dating indicates these events are diachronous (Manicouagan astrobleme 212 +/- 2 Ma and Tr/J boundary 200 Ma).

  15. Induction of apoptosis in PA-1 ovarian cancer cells by vitamin K2 is associated with an increase in the level of TR3/Nur77 and its accumulation in mitochondria and nuclei.

    PubMed

    Sibayama-Imazu, Toshiko; Fujisawa, Yukari; Masuda, Yutaka; Aiuchi, Toshihiro; Nakajo, Shigeo; Itabe, Hiroyuki; Nakaya, Kazuyasu

    2008-07-01

    We examined the growth-inhibitory and apoptosis-inducing effects of vitamin K(2) (VK(2); menaquinone-4) on various lines of human ovarian cancer cells to study the mechanism of induction of apoptosis by VK(2). Cell proliferation was determined by XTT method, and apoptotic cells were detected by Hoechst staining. TR3, also known as Nur77 and NGFI-B, was detected by immunoblotting and immunofluorescence analysis. Role of TR3 on induction of apoptosis was examined by a siRNA experiment. We found that PA-1 cells were the most sensitive to VK(2) (IC(50) = 5.0 +/- 0.7 microM), while SK-OV-3 cells were resistant to VK(2). Immunoblotting and immunofluorescence analyses indicated that levels of TR3 were elevated in cell lysates 48 h after the start of treatment with 30 microM VK(2). In the VK(2)-treated cells, TR3 accumulated at significant levels in mitochondria, as well as in the nuclei of PA-1 cells. No similar changes were observed in SK-OV-3 cells under the same conditions. Treatment of PA-1 cells with small interfering RNA (siRNA) directed against TR3, and with cycloheximide or SP600125 (an inhibitor of c-jun N-terminal kinase; JNK), separately, inhibited the VK(2)-induced synthesis of TR3 and apoptosis. From these results, we can conclude that an increase in the synthesis of TR3 and the accumulation of TR3 in mitochondria and in nuclei might be involved in the induction of apoptosis by VK(2) and that the synthesis of TR3 might be regulated through a JNK signaling pathway.

  16. Production of Mannitol from a High Concentration of Glucose by Candida parapsilosis SK26.001.

    PubMed

    Meng, Qing; Zhang, Tao; Wei, Wenting; Mu, Wanmeng; Miao, Ming

    2017-01-01

    A novel strain, SK26.001, which can produce mannitol from a high concentration of glucose without the addition of fructose, was isolated from sugarcane juice. This strain was identified as Candida parapsilosis based on 18S ribosomal RNA (rRNA) sequence analysis and the morphological and physiological-biochemical characteristics of the strain. Under optimized fermentation conditions, the mannitol concentration in shake flasks reached 68.5 g/L. When batch fermentation was performed, the fed glucose was completely consumed after 72 h, resulting in a final mannitol concentration of 80.3 g/L. Fed-batch fermentation was then performed with glucose feed. During the fed-batch process, ammonia water was added to maintain the pH at 4.0. The mannitol concentration in the fermenter reached 97.1 g/L after 120 h, with a total glucose consumption of 284 g/L.

  17. Fractional crystallization-induced variations in sulfides from the Noril’sk-Talnakh mining district (polar Siberia, Russia)

    USGS Publications Warehouse

    Duran, C.J.; Barnes, S-J.; Pleše, P.; Prašek, M. Kudrna; Zientek, Michael L.; Pagé, P.

    2017-01-01

    The distribution of platinum-group elements (PGE) within zoned magmatic ore bodies has been extensively studied and appears to be controlled by the partitioning behavior of the PGE during fractional crystallization of magmatic sulfide liquids. However, other chalcophile elements, especially TABS (Te, As, Bi, Sb, and Sn) have been neglected despite their critical role in forming platinum-group minerals (PGM). TABS are volatile trace elements that are considered to be mobile so investigating their primary distribution may be challenging in magmatic ore bodies that have been somewhat altered. Magmatic sulfide ore bodies from the Noril’sk-Talnakh mining district (polar Siberia, Russia) offer an exceptional opportunity to investigate the behavior of TABS during fractional crystallization of sulfide liquids and PGM formation as the primary features of the ore bodies have been relatively well preserved. In this study, new petrographic (2D and 3D) and whole-rock geochemical data from Cu-poor to Cu-rich sulfide ores of the Noril’sk-Talnakh mining district are integrated with published data to consider the role of fractional crystallization in generating mineralogical and geochemical variations across the different ore types (disseminated to massive). Despite textural variations in Cu-rich massive sulfides (lenses, veins, and breccias), these sulfides have similar chemical compositions, which suggests that Cu-rich veins and breccias formed from fractionated sulfide liquids that were injected into the surrounding rocks. Numerical modeling using the median disseminated sulfide composition as the initial sulfide liquid composition and recent DMSS/liq and DISS/liq predicts the compositional variations observed in the massive sulfides, especially in terms of Pt, Pd, and TABS. Therefore, distribution of these elements in the massive sulfides was likely controlled by their partitioning behavior during sulfide liquid fractional crystallization, prior to PGM formation. Our

  18. [¹²³I]ICF01012 melanoma imaging and [¹³¹I]ICF01012 dosimetry allow adapted internal targeted radiotherapy in preclinical melanoma models.

    PubMed

    Viallard, Claire; Perrot, Yann; Boudhraa, Zied; Jouberton, Elodie; Miot-Noirault, Elisabeth; Bonnet, Mathilde; Besse, Sophie; Mishellany, Florence; Cayre, Anne; Maigne, Lydia; Rbah-Vidal, Latifa; D'Incan, Michel; Cachin, Florent; Chezal, Jean-Michel; Degoul, Françoise

    2015-01-01

    Melanin-targeting radiotracers are interesting tools for imaging and treatment of pigmented melanoma metastases. However, variation of the pigment concentration may alter the efficiency of such targeting. A clear assessment of both tumor melanin status and dosimetry are therefore prerequisites for internal radiotherapy of disseminated melanoma. The melanin tracer ICF01012 was labelled with iodine-123 for melanoma imaging in pigmented murine B16F0 and human SK-Mel 3 melanomas. In vivo imaging showed that the uptake of [(123)I]ICF01012 to melanomas correlated significantly with melanin content. Schedule treatment of 3 × 25 MBq [(131)I]ICF01012 significantly reduced SK-Mel 3 tumor growth and significantly increased the median survival in treated mice. For this protocol, the calculated delivered dose was 53.2 Gy. Radio-iodinated ICF01012 is a good candidate for both imaging and therapeutic purposes for patients with metastatic pigmented melanomas.

  19. Spectral Knowledge (SK-UTALCA): Software for Exploratory Analysis of High-Resolution Spectral Reflectance Data on Plant Breeding.

    PubMed

    Lobos, Gustavo A; Poblete-Echeverría, Carlos

    2016-01-01

    This article describes public, free software that provides efficient exploratory analysis of high-resolution spectral reflectance data. Spectral reflectance data can suffer from problems such as poor signal to noise ratios in various wavebands or invalid measurements due to changes in incoming solar radiation or operator fatigue leading to poor orientation of sensors. Thus, exploratory data analysis is essential to identify appropriate data for further analyses. This software overcomes the problem that analysis tools such as Excel are cumbersome to use for the high number of wavelengths and samples typically acquired in these studies. The software, Spectral Knowledge (SK-UTALCA), was initially developed for plant breeding, but it is also suitable for other studies such as precision agriculture, crop protection, ecophysiology plant nutrition, and soil fertility. Various spectral reflectance indices (SRIs) are often used to relate crop characteristics to spectral data and the software is loaded with 255 SRIs which can be applied quickly to the data. This article describes the architecture and functions of SK-UTALCA and the features of the data that led to the development of each of its modules.

  20. Peginterferon Beta-1a Shows Antitumor Activity as a Single Agent and Enhances Efficacy of Standard of Care Cancer Therapeutics in Human Melanoma, Breast, Renal, and Colon Xenograft Models.

    PubMed

    Boccia, Antonio; Virata, Cyrus; Lindner, Daniel; English, Nicki; Pathan, Nuzhat; Brickelmaier, Margot; Hu, Xiao; Gardner, Jennifer L; Peng, Liaomin; Wang, Xinzhong; Zhang, Xiamei; Yang, Lu; Perron, Keli; Yco, Grace; Kelly, Rebecca; Gamez, James; Scripps, Thomas; Bennett, Donald; Joseph, Ingrid B; Baker, Darren P

    2017-01-01

    Because of its tumor-suppressive effect, interferon-based therapy has been used for the treatment of melanoma. However, limited data are available regarding the antitumor effects of pegylated interferons, either alone or in combination with approved anticancer drugs. We report that treatment of human WM-266-4 melanoma cells with peginterferon beta-1a induced apoptotic markers. Additionally, peginterferon beta-1a significantly inhibited the growth of human SK-MEL-1, A-375, and WM-266-4 melanoma xenografts established in immunocompromised mice. Peginterferon beta-1a regressed large, established WM-266-4 xenografts in nude mice. Treatment of SK-MEL-1 tumor-bearing mice with a combination of peginterferon beta-1a and the MEK inhibitor PD325901 ((R)-N-(2,3-dihydroxypropoxy)-3,4-difluoro-2-(2-fluoro-4-iodophenylamino)benzamide) significantly improved tumor growth inhibition compared with either agent alone. Examination of the antitumor activity of peginterferon beta-1a in combination with approved anticancer drugs in breast and renal carcinomas revealed improved antitumor activity in these preclinical xenograft models, as did the combination of peginterferon beta-1a and bevacizumab in a colon carcinoma xenograft model.

  1. [Heat-shock protein HSP70 protects neuroblastoma cells SK-N-SH from the neurotoxic effects hydrogen peroxide and the β-amyloid peptide].

    PubMed

    Yurinskaya, M M; Mit'kevich, V A; Barykin, E P; Garbuz, D G; Evgen'ev, M B; Makarov, A A; Vinokurov, M G

    2015-01-01

    Neuronal cell death in Alzheimer's disease is associated with the development of oxidative stress caused by the reactive oxygen species (ROS), which can be generated as a result of the effect of beta-amyloid peptides. One of the sources of ROS is hydrogen peroxide, inducing the apoptosis and necrosis of neural tissue cells. The mechanism of hydrogen peroxide apoptotic action includes launching signaling pathways that involve protein kinases PI3K, p38MAPK, JNK and ERK. Oxidative stress leads to increased synthesis of heat-shock proteins in the cells including HSP70. It was shown that the exogenous HSP70 could reduce generation of ROS in cells. In this study, we determined how HSP70 affected apoptosis and necrosis in human neuroblastoma cells SK-N-SH, induced by hydrogen peroxide and β-amyloid peptide Aβ(1-42). It was shown that HSP70 reduces the cytotoxic effects of hydrogen peroxide and beta-amyloid, and protein kinases PI3K and JNK play an important role in the mechanism of HSP70 protective effect on the peroxide induced apoptosis in SK-N-SH cells.

  2. Loss of inhibitor of apoptosis proteins as a determinant of polyamine analog-induced apoptosis in human melanoma cells.

    PubMed

    Chen, Ying; Kramer, Debora L; Li, Fengzhi; Porter, Carl W

    2003-08-07

    We have previously shown that the clinically relevant polyamine analog N1,N11-diethylnorspermine (DENSPM) causes rapid apoptosis in human melanoma SK-MEL-28 cells via a series of events that include mitochondrial release of cytochrome c and activation of the caspase cascade. Upstream to these events, DENSPM downregulates polyamine biosynthesis and potently upregulates polyamine catabolism at the level of spermidine/spermine N1-acetyltransferase (SSAT). In searching for downstream effectors that either contribute to or abrogate the apoptotic response, we observed that DENSPM treatment of SK-MEL-28 cells for 30 h led to cytosolic release of Smac/Diablo, a mitochondrial protein known to bind and inhibit the function of inhibitor of apoptosis proteins (IAPs). Subsequently, we found that DENSPM markedly lowered survivin and ML-IAP protein (but not XIAP) levels by 18 h via an apparently Smac/Diablo-independent pathway. Proteasome inhibitors fully prevented survivin and ML-IAP protein loss as well as apoptosis, suggesting that the proteasome-mediated degradation of survivin and ML-IAP is causally linked to the cellular outcome. We also observed that structural analogs of DENSPM which differentially induced SSAT and apoptosis lowered survivin and ML-IAP levels in a manner that correlated with enzyme activity. The linkage between IAPs and SSAT was more directly established by the finding that selective prevention of SSAT induction by small interfering RNA prevented survivin and ML-IAP loss as well as apoptosis during DENSPM treatment. Among the melanoma cell lines (SK-MEL-28, MALME-3M, A375 and LOX), survivin degradation correlated temporally with the onset of DENSPM induced apoptosis or growth inhibition. By contrast, ML-IAP degradation occurred only during rapid apoptosis seen in SK-MEL-28 cells. These data suggest a sequence of events whereby DENSPM induction of SSAT leads to loss of IAP proteins and a more fulminate apoptotic response. The findings implicate survivin

  3. Neuronal uptake and neuroprotective effect of curcumin-loaded PLGA nanoparticles on the human SK-N-SH cell line.

    PubMed

    Doggui, Sihem; Sahni, Jasjeet Kaur; Arseneault, Madeleine; Dao, Lé; Ramassamy, Charles

    2012-01-01

    Curcumin, a natural polyphenolic pigment present in the spice turmeric (Curcuma longa), is known to possess a pleiotropic activity such as antioxidant, anti-inflammatory, and anti-amyloid-β activities. However, these benefits of curcumin are limited by its poor aqueous solubility and oral bioavailability. In the present study, a polymer-based nanoparticle approach has been utilized to deliver drugs to neuronal cells. Curcumin was encapsulated in biodegradable poly (lactide-co-glycolide) (PLGA) based-nanoparticulate formulation (Nps-Cur). Dynamic laser light scattering and transmission electronic microscopy analysis indicated a particle diameter ranging from 80 to 120 nm. The entrapment efficiency was 31% with 15% drug-loading. In vitro release kinetics of curcumin from Nps-Cur revealed a biphasic pattern with an initial exponential phase followed by a slow release phase. Cellular internalization of Nps-Cur was confirmed by fluorescence and confocal microscopy with a wide distribution of the fluorescence in the cytoplasm and within the nucleus. The prepared nanoformulation was characterized for cellular toxicity and biological activity. Cytotoxicity assays showed that void PLGA-nanoparticles (Nps) and curcumin-loaded PLGA nanoparticles (Nps-Cur) were nontoxic to human neuroblastoma SK-N-SH cells. Moreover, Nps-Cur was able to protect SK-N-SH cells against H2O2 and prevent the elevation of reactive oxygen species and the consumption of glutathione induced by H2O2. Interestingly, Nps-Cur was also able to prevent the induction of the redox-sensitive transcription factor Nrf2 in the presence of H2O2. Taken together, these results suggest that Nps-Cur could be a promising drug delivery strategy to protect neurons against oxidative damage as observed in Alzheimer's disease.

  4. Differential targeting of the cyclin-dependent kinase inhibitor, p21CIP1/WAF1, by chelators with anti-proliferative activity in a range of tumor cell-types

    PubMed Central

    Moussa, Rayan S.; Kovacevic, Zaklina; Richardson, Des R.

    2015-01-01

    Chelators such as 2-hydroxy-1-napthylaldehyde isonicotinoyl hydrazone (311) and di-2-pyridylketone-4,4-dimethyl-3-thiosemicarbazone (Dp44mT) target tumor cell iron pools and inhibit proliferation. These agents also modulate multiple targets, one of which is the cyclin-dependent kinase inhibitor, p21. Hence, this investigation examined the mechanism of action of these compounds in targeting p21. All the chelators up-regulated p21 mRNA in the five tumor cell-types assessed. In contrast, examining their effect on total p21 protein levels, these agents induced either: (1) down-regulation in MCF-7 cells; (2) up-regulation in SK-MEL-28 and CFPAC-1 cells; or (3) had no effect in LNCaP and SK-N-MC cells. The nuclear localization of p21 was also differentially affected by the ligands depending upon the cell-type, with it being decreased in MCF-7 cells, but increased in SK-MEL-28 and CFPAC-1 cells. Further studies assessing the mechanisms responsible for these effects demonstrated that p21 expression was not correlated with p53 status, suggesting a p53-independent mechanism. Considering this, we examined proteins that modulate p21 independently of p53, namely NDRG1, MDM2 and ΔNp63. These studies demonstrated that a dominant negative MDM2 isoform (p75MDM2) closely resembled p21 expression in response to chelation in three cell lines. These data suggest MDM2 may be involved in the regulation of p21 by chelators. PMID:26335183

  5. New magnetostratigraphy data and correlation of the traps of Noril'sk and Maymecha-Kotuy regions of the Siberian platform

    NASA Astrophysics Data System (ADS)

    Fetisova, A. M.; Pavlov, V.; Veselovskiy, R. V.

    2013-12-01

    The Siberian traps are one of the largest igneous provinces on the Earth and trap volcanism has been considered as a possible trigger for the end-Permian mass extinction. Duration of volcanic activity and volume of erupted material are the critical characteristics for determining of the Siberian traps' disastrous effect. These characteristics can be obtained from the correlation of the most complete trap cross-sections. Studied sections are located close to the Norilsk city and in the Maymecha-Kotuy region in the northern part of the Siberian platform. Some possible correlations of these trap sections have been suggested. All of them are based on geochronological, geochemical, paleontological and paleomagnetic data and more or less controversial. In this study we re-analyzed the possible schemes of correlation of these trap sections using new magnetostratigraphic data on these regions. Our analysis allows us to suggest a partial overlapping of the Noril'sk and Maymecha-Kotuy trap sections. Comparison of the virtual geomagnetic poles (VGP) supports a correlation of Arydzhangsky and lower part of Onkuchaksky formations (lower part of Maymecha-Kotuy section) with Morongovsky and Mokulaevsky formations (middle part of Noril'sk section). We also discuss an important question about correlation the designed magnetostratigraphic scale of the northern Siberian traps with the latest version of a global scale of the magnetic polarity (Hounslow et al., 2010). Using two groups of high-quality isotopic ages for the northern Siberia volcanic sections, obtained by Sandra Kamo (Kamo et al., 2003) and Sam Bowring and Seth Burgess (personal comm.), we conclude that the Noril'sk and Maymecha-Kotuy trap sequences could be referred to the lower Induan stage or to the lower Olenekian stage of the Early Triassic. This study was funded by grants RFBR 12-05-31149, 13-05-12030, 13-05-00235 and EAR-0807585 (NSF).

  6. Spectral Knowledge (SK-UTALCA): Software for Exploratory Analysis of High-Resolution Spectral Reflectance Data on Plant Breeding

    PubMed Central

    Lobos, Gustavo A.; Poblete-Echeverría, Carlos

    2017-01-01

    This article describes public, free software that provides efficient exploratory analysis of high-resolution spectral reflectance data. Spectral reflectance data can suffer from problems such as poor signal to noise ratios in various wavebands or invalid measurements due to changes in incoming solar radiation or operator fatigue leading to poor orientation of sensors. Thus, exploratory data analysis is essential to identify appropriate data for further analyses. This software overcomes the problem that analysis tools such as Excel are cumbersome to use for the high number of wavelengths and samples typically acquired in these studies. The software, Spectral Knowledge (SK-UTALCA), was initially developed for plant breeding, but it is also suitable for other studies such as precision agriculture, crop protection, ecophysiology plant nutrition, and soil fertility. Various spectral reflectance indices (SRIs) are often used to relate crop characteristics to spectral data and the software is loaded with 255 SRIs which can be applied quickly to the data. This article describes the architecture and functions of SK-UTALCA and the features of the data that led to the development of each of its modules. PMID:28119705

  7. The Staphylococcus aureus pSK41 plasmid-encoded ArtA protein is a master regulator of plasmid transmission genes and contains a RHH motif used in alternate DNA-binding modes.

    PubMed

    Ni, Lisheng; Jensen, Slade O; Ky Tonthat, Nam; Berg, Tracey; Kwong, Stephen M; Guan, Fiona H X; Brown, Melissa H; Skurray, Ronald A; Firth, Neville; Schumacher, Maria A

    2009-11-01

    Plasmids harbored by Staphylococcus aureus are a major contributor to the spread of bacterial multi-drug resistance. Plasmid conjugation and partition are critical to the dissemination and inheritance of such plasmids. Here, we demonstrate that the ArtA protein encoded by the S. aureus multi-resistance plasmid pSK41 is a global transcriptional regulator of pSK41 genes, including those involved in conjugation and segregation. ArtA shows no sequence homology to any structurally characterized DNA-binding protein. To elucidate the mechanism by which it specifically recognizes its DNA site, we obtained the structure of ArtA bound to its cognate operator, ACATGACATG. The structure reveals that ArtA is representative of a new family of ribbon-helix-helix (RHH) DNA-binding proteins that contain extended, N-terminal basic motifs. Strikingly, unlike most well-studied RHH proteins ArtA binds its cognate operators as a dimer. However, we demonstrate that it is also able to recognize an atypical operator site by binding as a dimer-of-dimers and the extended N-terminal regions of ArtA were shown to be essential for this dimer-of-dimer binding mode. Thus, these data indicate that ArtA is a master regulator of genes critical for both horizontal and vertical transmission of pSK41 and that it can recognize DNA utilizing alternate binding modes.

  8. The Staphylococcus aureus pSK41 plasmid-encoded ArtA protein is a master regulator of plasmid transmission genes and contains a RHH motif used in alternate DNA-binding modes

    PubMed Central

    Ni, Lisheng; Jensen, Slade O.; Ky Tonthat, Nam; Berg, Tracey; Kwong, Stephen M.; Guan, Fiona H. X.; Brown, Melissa H.; Skurray, Ronald A.; Firth, Neville; Schumacher, Maria A.

    2009-01-01

    Plasmids harbored by Staphylococcus aureus are a major contributor to the spread of bacterial multi-drug resistance. Plasmid conjugation and partition are critical to the dissemination and inheritance of such plasmids. Here, we demonstrate that the ArtA protein encoded by the S. aureus multi-resistance plasmid pSK41 is a global transcriptional regulator of pSK41 genes, including those involved in conjugation and segregation. ArtA shows no sequence homology to any structurally characterized DNA-binding protein. To elucidate the mechanism by which it specifically recognizes its DNA site, we obtained the structure of ArtA bound to its cognate operator, ACATGACATG. The structure reveals that ArtA is representative of a new family of ribbon–helix–helix (RHH) DNA-binding proteins that contain extended, N-terminal basic motifs. Strikingly, unlike most well-studied RHH proteins ArtA binds its cognate operators as a dimer. However, we demonstrate that it is also able to recognize an atypical operator site by binding as a dimer-of-dimers and the extended N-terminal regions of ArtA were shown to be essential for this dimer-of-dimer binding mode. Thus, these data indicate that ArtA is a master regulator of genes critical for both horizontal and vertical transmission of pSK41 and that it can recognize DNA utilizing alternate binding modes. PMID:19759211

  9. Re-Os isotopic evidence for an enriched-mantle source for the Noril'sk-type, ore-bearing intrusions, Siberia

    USGS Publications Warehouse

    Walker, R.J.; Morgan, J.W.; Horan, M.F.; Czamanske, G.K.; Krogstad, E.J.; Fedorenko, V.A.; Kunilov, V.E.

    1994-01-01

    Magmatic Cu-Ni sulfide ores and spatially associated ultramafic and mafic rocks from the Noril'sk I, Talnakh, and Kharaelakh intrusions are examined for Re-Os isotopic systematics. Neodymium and lead isotopic data also are reported for the ultramafic and mafic rocks. The Re-Os data for most samples indicate closed-system behavior since the ca. 250 Ma igneous crystallization age of the intrusions. There are small but significant differences in the initial osmium isotopic compositions of samples from the three intrusions. Ores from the Noril'sk I intrusion have ??Os values that vary from +0.4 to +8.8, but average +5.8. Ores from the Talnakh intrusion have ??Os values that range from +6.7 to +8.2, averaging +7.7. Ores from the Kharaelakh intrusion have ??Os values that range from +7.8 to +12.9, with an average value of +10.4. The osmium isotopic compositions of the ore samples from the Main Kharaelakh orebody exhibit minimal overlap with those for the Noril'sk I and Talnakh intrusions, indicating that these Kharaelakh ores were derived from a more radiogenic source of osmium than the other ores. Combined osmium and lead data for major orebodies in the three intrusions plot in three distinct fields, indicating derivation of osmium and lead from at least three isotopically distinct sources. Some of the variation in lead isotopic compositions may be the result of minor lower-crustal contamination. However, in contrast to most other isotopic and trace element data, Os-Pb variations are generally inconsistent with significant crustal contamination or interaction with the subcontinental lithosphere. Thus, the osmium and lead isotopic compositions of these intrusions probably reflect quite closely the compositions of their mantle source, and suggest that these two isotope systems were insensitive to lithospheric interaction. Ultramafic and mafic rocks have osmium and lead isotopic compositions that range only slightly beyond the compositions of the ores. These rocks also

  10. Innocence Is No Defense: Politicized Childhood in Antonio Skármeta's "La Composición/The Composition"

    ERIC Educational Resources Information Center

    Nance-Carroll, Niall

    2014-01-01

    Chilean author Antonio Skármeta's picture book "The Composition" (2000), illustrated by Alfonso Ruano, focuses on the political involvement of 9-year-old Pedro. This essay brings together Maria Nikolajeva's concept of aetonormativity and Elisabeth Young-Bruehl's concept of childism to examine how "The…

  11. Identity and function of a cardiac mitochondrial small conductance Ca2+-activated K+ channel splice variant

    PubMed Central

    Yang, MeiYing; Camara, Amadou K.S.; Aldakkak, Mohammed; Kwok, Wai-Meng; Stowe, David F.

    2017-01-01

    We provide evidence for location and function of a small conductance, Ca2+-activated K+ (SKCa) channel isoform 3 (SK3) in mitochondria (m) of guinea pig, rat and human ventricular myocytes. SKCa agonists protected isolated hearts and mitochondria against ischemia/reperfusion (IR) injury; SKCa antagonists worsened IR injury. Intravenous infusion of a SKCa channel agonist/antagonist, respectively, in intact rats was effective in reducing/enhancing regional infarct size induced by coronary artery occlusion. Localization of SK3 in mitochondria was evidenced by Western blot of inner mitochondrial membrane, immunocytochemical staining of cardiomyocytes, and immunogold labeling of isolated mitochondria. We identified a SK3 splice variant in guinea pig (SK3.1, aka SK3a) and human ventricular cells (SK3.2) by amplifying mRNA, and show mitochondrial expression in mouse atrial tumor cells (HL-1) by transfection with full length and truncated SK3.1 protein. We found that the Nterminus is not required for mitochondrial trafficking but the C-terminus beyond the Ca2+ calmodulin binding domain is required for Ca2+ sensing to induce mK+ influx and/or promote mitochondrial localization. In isolated guinea pig mitochondria and in SK3 overexpressed HL-1 cells, mK+ influx was driven by adding CaCl2. Moreover, there was a greater fall in membrane potential (ΔΨm), and enhanced cell death with simulated cell injury after silencing SK3.1 with siRNA. Although SKCa channel opening protects the heart and mitochondria against IR injury, the mechanism for favorable bioenergetics effects resulting from SKCa channel opening remains unclear. SKCa channels could play an essential role in restraining cardiac mitochondria from inducing oxidative stress-induced injury resulting from mCa2+ overload. PMID:28342809

  12. Antioxidant activity and oxidative stress protection of duck proteins hydrolysates in SK-N-SH cells.

    PubMed

    Guo, Yuxing; Pan, Daodong; Wu, Zhen; Zhao, Chuanchuan; Cao, Jinxuan

    2013-02-26

    Studies have found that natural antioxidants, which are free-radical scavengers, can reduce the risk of diseases caused by free radicals. This work investigated the antioxidant properties of duck proteins hydrolysates. The free-radical scavenging function of CP-1 (M(r) > 10 kDa), CP-2 (5 kDa < M(r) < 10 kDa) and CP-3 (M(r) < 5 kDa), obtained through ultrafiltration and gel filtration were evaluated. The results showed that the lower molecular weight fraction exhibited a stronger free-radical scavenging ability. The highest free-radical scavenging activity was detected in the fraction of p4 purified from CP-3 using Sephadex G-15 column chromatography. The 50% inhibitory value (IC(50)) of p4 for scavenging radicals of superoxide, hydroxyl and 1,1-diphenyl-2-pycrylhydrazyl (DPPH) were, respectively, 0.97 mg mL(-1), 0.84 mg mL(-1) and 1.84 mg mL(-1). Furthermore, the p4 fraction at a concentration of 10 μg mL(-1) increased cell viability from 84.8% to 94% under antioxidative stress in neuroblastoma SK-N-SH cells.

  13. Synthesis and activity of novel 16-dehydropregnenolone acetate derivatives as inhibitors of type 1 5α-reductase and on cancer cell line SK-LU-1.

    PubMed

    Silva-Ortiz, Aylin Viviana; Bratoeff, Eugene; Ramírez-Apan, Teresa; Heuze, Yvonne; Sánchez, Araceli; Soriano, Juan; Cabeza, Marisa

    2015-12-15

    Testosterone (T) plays a crucial role in prostate growth. In androgen-dependent tissues T is reduced to dihydrotestosterone (DHT) because of the presence of the 5α-reductase enzyme. This androgen is more active than T, since it has a higher affinity for the androgen receptor (AR). When this mechanism is altered, androgen-dependent diseases, including prostate cancer, could result. The aim of this study was to synthesize several 16-dehydropregnenolone acetate derivatives containing a triazole ring at C-21 and a linear or alicyclic ester moiety at C-3 of the steroidal skeleton. These steroids were designed as potential inhibitors of the activity of both types (1 and 2) of 5α-reductase. The cytotoxic activity of these compounds was also evaluated on a panel of PC-3, MCF7, and SK-LU-1 human cancer cell lines. The results from this study showed that with the exception of steroids 20-oxo-21-(1H-1,2,4-triazole-1-yl)pregna-5,16-dien-3β-yl-propionate and 20-oxo-21-(1H-1,2,4-triazole-1-yl)pregna-5,16-dien-3β-yl-pentanoate, the compounds exhibit a lower inhibitory activity for both isoenzymes of 5α-reductase than finasteride. Furthermore the 3β-hydroxy-21-(1H-1,2,4-triazole-1-yl)pregna-5,16-dien-20-one and 20-oxo-21-(1H-1,2,4-triazole-1-yl)pregna-5,16-dien-3β-yl-acetate derivatives display 80% cytotoxic activity on the SK-LU-1 cell line. These results also indicated that the triazole derivatives, which have a hydroxyl or acetoxy group at C-3, could have an anticancer effect, whereas the derivatives with a alicyclic ester group at C-3 do not show biological activity. Copyright © 2015 Elsevier Ltd. All rights reserved.

  14. Bioavailability of angiotensin I-converting enzyme (ACE) inhibitory peptides derived from Virgibacillus halodenitrificans SK1-3-7 proteinases hydrolyzed tilapia muscle proteins.

    PubMed

    Toopcham, Tidarat; Mes, Jurriaan J; Wichers, Harry J; Roytrakul, Sittiruk; Yongsawatdigul, Jirawat

    2017-04-01

    The angiotensin I-converting enzyme (ACE) inhibitory activity of protein hydrolysates from tilapia muscle fractions, namely mince (M), washed mince (WM), and sarcoplasmic protein (SP), were investigated. Each fraction was hydrolyzed by Virgibacillus halodenitrificans SK1-3-7 proteinases for up to 24h. After 8h of hydrolysis, the M hydrolysate (48% degree of hydrolysis (DH)) showed the highest ACE inhibitory activity, with an IC 50 value of 0.54mg/ml, while the SP hydrolysate exhibited the lowest DH and ACE inhibition. In vitro gastrointestinal digestion reduced the ACE inhibitory activity of the M hydrolysate but enhanced its transport across Caco-2 cell monolayers. The transported peptides were found to contain 3-4 amino acid residues showing strong ACE inhibition. The novel ACE inhibitory peptide with the highest inhibition was found to be MCS, with an IC 50 value of 0.29μM. Therefore, tilapia mince hydrolyzed by V. halodenitrificans proteinases contained ACE inhibitory peptides that are potentially bioavailable. Copyright © 2016 Elsevier Ltd. All rights reserved.

  15. Developments in damage assessment by Marie Skłodowska-Curie TRUSS ITN project

    NASA Astrophysics Data System (ADS)

    González, A.

    2017-05-01

    The growth of cities, the impacts of climate change and the massive cost of providing new infrastructure provide the impetus for TRUSS (Training in Reducing Uncertainty in Structural Safety), a3.7 million Marie Skłodowska-Curie Action Innovative Training Network project funded by EU’s Horizon 2020 programme, which aims to maximize the potential of infrastructure that already exists (http://trussitn.eu). For that purpose, TRUSS brings together an international, inter-sectoral and multidisciplinary collaboration between five academic and eleven industry institutions from five European countries. The project covers rail and road infrastructure, buildings and energy and marine infrastructure. This paper reports progress in fields such as advanced sensor-based structural health monitoring solutions - unmanned aerial vehicles, optical backscatter reflectometry, monitoring sensors mounted on vehicles, … - and innovative algorithms for structural designs and short- and long-term assessments of buildings, bridges, pavements, ships, ship unloaders, nuclear components and wind turbine towers that will support infrastructure operators and owners in managing their assets.

  16. [The Outpatient Clinic for Adolescent Risk-taking and Self-harm behaviors (AtR!Sk) - A Pioneering Approach of Outpatient Early Detection and Intervention of Borderline Personality Disorder].

    PubMed

    Kaess, Michael; Ghinea, Denisa; Fischer-Waldschmidt, Gloria; Resch, Franz

    2017-07-01

    The Outpatient Clinic for Adolescent Risk-taking and Self-harm behaviors (AtR!Sk) - A Pioneering Approach of Outpatient Early Detection and Intervention of Borderline Personality Disorder Self-harm and risk-taking behaviors are frequently occurring problems in adolescents' everyday life and commonly challenge the present child and adolescent health-care system. Those behaviors are typical features of borderline-personality disorder (BPD), a severe mental disorder that is associated with immense psychological strain, increased risk of mortality and poor psychosocial functioning. Despite controversy in the past, recent evidence shows that BPD is a valid and reliable diagnosis in adolescence. Consequently, specified and effective intervention programs for this age group are necessary. Instead, present health-care services for children and adolescents in Germany are usually characterized by long waiting periods for specialized outpatient treatments and unnecessary expanses of unnecessary long inpatient stays. Alternatively, and in order to prevent serious long-term consequences, new concepts - integrated in an ambulatory setting and close to patients' daily routines - should focus on early detection and treatment of adolescents at-risk presenting with borderline personality features. The specialist outpatient clinic AtR!Sk at the University Hospital Heidelberg ensures a low-threshold initial contact service, comprehensive and accurate diagnosis of BPD features, and rapidly "tailored" therapy for young people presenting with any types of risk-taking and self-harm behavior. AtR!Sk - as a pioneering approach in south Germany - provides evidence-based early detection and intervention for adolescent BPD.

  17. Mild uncoupling of respiration and phosphorylation as a mechanism providing nephro- and neuroprotective effects of penetrating cations of the SkQ family.

    PubMed

    Plotnikov, E Y; Silachev, D N; Jankauskas, S S; Rokitskaya, T I; Chupyrkina, A A; Pevzner, I B; Zorova, L D; Isaev, N K; Antonenko, Y N; Skulachev, V P; Zorov, D B

    2012-09-01

    It is generally accepted that mitochondrial production of reactive oxygen species is nonlinearly related to the value of the mitochondrial membrane potential with significant increment at values exceeding 150 mV. Due to this, high values of the membrane potential are highly dangerous, specifically under pathological conditions associated with oxidative stress. Mild uncoupling of oxidative phosphorylation is an approach to preventing hyperpolarization of the mitochondrial membrane. We confirmed data obtained earlier in our group that dodecylrhodamine 19 (C(12)R1) (a penetrating cation from SkQ family not possessing a plastoquinone group) has uncoupling properties, this fact making it highly potent for use in prevention of pathologies associated with oxidative stress induced by mitochondrial hyperpolarization. Further experiments showed that C(12)R1 provided nephroprotection under ischemia/reperfusion of the kidney as well as under rhabdomyolysis through diminishing of renal dysfunction manifested by elevated level of blood creatinine and urea. Similar nephroprotective properties were observed for low doses (275 nmol/kg) of the conventional uncoupler 2,4-dinitrophenol. Another penetrating cation that did not demonstrate protonophorous activity (SkQR4) had no effect on renal dysfunction. In experiments with induced ischemic stroke, C(12)R1 did not have any effect on the area of ischemic damage, but it significantly lowered neurological deficit. We conclude that beneficial effects of penetrating cation derivatives of rhodamine 19 in renal pathologies and brain ischemia may be at least partially explained by uncoupling of oxidation and phosphorylation.

  18. Influence of Melanosome Dynamics on Melanoma Drug Sensitivity

    PubMed Central

    Chen, Kevin G.; Leapman, Richard D.; Zhang, Guofeng; Lai, Barry; Valencia, Julio C.; Cardarelli, Carol O.; Vieira, Wilfred D.; Hearing, Vincent J.

    2009-01-01

    Background Malignant melanomas are intrinsically resistant to many conventional treatments, such as radiation and chemotherapy, for reasons that are poorly understood. Here we propose and test a model that explains drug resistance or sensitivity in terms of melanosome dynamics. Methods The growth and sensitivity to cisplatin of MNT-1 cells, which are melanotic and enriched with mature stage III and IV melanosomes, and SK-MEL-28 cells, which have only immature stage I and II melanosomes, were compared using clonogenic assays. Differences in pigmentation, melanosome stages, melanosome number, and cellular structures in different cell lines in response to various treatments were examined by electron microscopy. The relative numbers of melanosomes of different stages were compared after treatment with 1-phenyl-2-thiourea. The relationship between drug transporter function and endogenous melanogenic toxicity was assessed by treating cells with the cyclosporin analog PSC-833 and by assessing vacuole formation and cell growth inhibition. All statistical tests were two-sided. Results Endogenous melanogenic cytotoxicity, produced by damaged melanosomes, resulted in pronounced cell growth inhibition in MNT-1 cells compared with amelanotic SK-MEL-28 cells. The sensitivity to CDDP of MNT-1 cells was 3.8-fold higher than that of SK-MEL-28 cells (mean IC50 for SK-MEL-28 and MNT-1 = 2.13 μM and 0.56 μM, respectively; difference = 1.57 μM, 95% confidence interval = 1.45 to 1.69; P = .0017). After treatment with 6.7 μM CDDP for 72 hours, the number of stage II-III melanosomes in surviving MNT-1 cells was 6.8-fold that of untreated cells. Modulation of MNT-1 cells to earlier-stage (II, II-III, III) melanosomes by treatment with the tyrosinase inhibitor 1-phenyl-2-thiourea dramatically increased CDDP resistance. Furthermore, PSC-833 principally suppressed MNT-1 melanotic cell growth via an elevation of autophagosome-like vacuolar structures, possibly by inhibiting melanosome

  19. Influence of melanosome dynamics on melanoma drug sensitivity.

    PubMed

    Chen, Kevin G; Leapman, Richard D; Zhang, Guofeng; Lai, Barry; Valencia, Julio C; Cardarelli, Carol O; Vieira, Wilfred D; Hearing, Vincent J; Gottesman, Michael M

    2009-09-16

    Malignant melanomas are intrinsically resistant to many conventional treatments, such as radiation and chemotherapy, for reasons that are poorly understood. Here we propose and test a model that explains drug resistance or sensitivity in terms of melanosome dynamics. The growth and sensitivity to cisplatin of MNT-1 cells, which are melanotic and enriched with mature stage III and IV melanosomes, and SK-MEL-28 cells, which have only immature stage I and II melanosomes, were compared using clonogenic assays. Differences in pigmentation, melanosome stages, melanosome number, and cellular structures in different cell lines in response to various treatments were examined by electron microscopy. The relative numbers of melanosomes of different stages were compared after treatment with 1-phenyl-2-thiourea. The relationship between drug transporter function and endogenous melanogenic toxicity was assessed by treating cells with the cyclosporin analog PSC-833 and by assessing vacuole formation and cell growth inhibition. All statistical tests were two-sided. Endogenous melanogenic cytotoxicity, produced by damaged melanosomes, resulted in pronounced cell growth inhibition in MNT-1 cells compared with amelanotic SK-MEL-28 cells. The sensitivity to CDDP of MNT-1 cells was 3.8-fold higher than that of SK-MEL-28 cells (mean IC(50) for SK-MEL-28 and MNT-1 = 2.13 microM and 0.56 microM, respectively; difference = 1.57 microM, 95% confidence interval = 1.45 to 1.69; P = .0017). After treatment with 6.7 microM CDDP for 72 hours, the number of stage II-III melanosomes in surviving MNT-1 cells was 6.8-fold that of untreated cells. Modulation of MNT-1 cells to earlier-stage (II, II-III, III) melanosomes by treatment with the tyrosinase inhibitor 1-phenyl-2-thiourea dramatically increased CDDP resistance. Furthermore, PSC-833 principally suppressed MNT-1 melanotic cell growth via an elevation of autophagosome-like vacuolar structures, possibly by inhibiting melanosome membrane

  20. A standardized crisis management model for self-harming and suicidal individuals with three or more diagnostic criteria of borderline personality disorder: The Brief Admission Skåne randomized controlled trial protocol (BASRCT).

    PubMed

    Liljedahl, Sophie I; Helleman, Marjolein; Daukantaité, Daiva; Westrin, Åsa; Westling, Sofie

    2017-06-15

    Brief Admission is a crisis and risk management strategy in which self-harming and suicidal individuals with three or more diagnostic criteria of borderline personality disorder self-admit to hospital at times of increasing risk when other efforts to stay safe are failing. Standardized in the current randomized controlled trial, the intensity of Brief Admission Skåne is implemented in durations of three days, with a maximum frequency of three times a month. Brief Admission is integrated into existing treatment plans in advance of crises to prevent reliance on general psychiatric admissions for risk management, as these may be lengthy, unstructured, and of uncertain therapeutic value. The overall objective of the Brief Admission Skåne randomized controlled trial is to determine if Brief Admission can replace general psychiatric admission for self-harming and suicidal individuals with complex mental illness at times of escalating risk. Other objectives of the study are to evaluate whether Brief Admission increases daily functioning and enhances coping, reduces psychiatric symptoms including frequency and severity of self-harm and suicidal behaviours. A final objective is to determine if Brief Admission is an effective crisis management model for this population. Participants are randomized at an individual level to either Brief Admission Skåne plus Treatment as Usual or Treatment As Usual. Based on a priori power analyses, N = 124 participants will be recruited to the study. Data collection is in progress, and will continue until June 2018. All participant data are single-blinded and will be handled with intention-to-treat analysis. Based on the combined clinical experience of our international research group, the Brief Admission Skåne randomized controlled trial upon which the current protocol is based represents the first initiative to standardize, implement and evaluate Brief Admission amongst self-harming and suicidal individuals, including those with

  1. 10Be exposure age chronology of the last glaciation of the Roháčská Valley in the Western Tatra Mountains, central Europe

    NASA Astrophysics Data System (ADS)

    Engel, Zbyněk; Mentlík, Pavel; Braucher, Régis; Křížek, Marek; Pluháčková, Markéta; Arnold, Maurice; Aumaître, Georges; Bourlès, Didier; Keddadouche, Karim; Aster Team; Arnold, Maurice; Aumaître, Georges; Bourlès, Didier; Keddadouche, Karim

    2017-09-01

    10Be exposure ages from moraines and bedrock sites in the Roháčská Valley provide chronology of the last glaciation in the largest valley of the Western Tatra Mts., the Western Carpathians. The minimum apparent exposure age of 19.4 ± 2.1 ka obtained for the oldest sampled boulder and the mean age of 18.0 ± 0.8 ka calculated for the terminal moraine indicate that the oldest preserved moraine was probably deposited at the time of the global Last Glacial Maximum (LGM). The age of this moraine coincides with the termination of the maximum glacier expansion in other central European ranges, including the adjacent High Tatra Mts. and the Alps. The equilibrium line altitude (ELA) of the LGM glacier in the Roháčská Valley, estimated at 1400-1410 m a.s.l., was 50-80 m lower than in the eastern part of the range, indicating a positive ELA gradient from west to east among the north-facing glaciers in the Tatra Mts. Lateglacial glacier expansion occurred no later than 13.4 ± 0.5 ka and 11.9 ± 0.5 ka, as indicated by the mean exposure ages calculated for re-advance moraines. This timing is consistent with the exposure age chronology of the last Lateglacial re-advance in the High Tatra Mts., Alps and lower mountain ranges in central Europe. The ELA in the Roháčská Valley estimated at 1690-1770 m a.s.l. in this period was located 130-300 m lower than in the north-facing valleys in the High Tatra Mts. 10Be exposure ages obtained for a rock glacier constrains the timing of this landform stabilization in the Salatínska Valley and provides the first chronological evidence for the Lateglacial activity of rock glaciers in the Carpathians.

  2. Elementary analysis and energetic potential of the municipal sewage sludges from the Gdańsk and Kościerzyna WWTPs

    NASA Astrophysics Data System (ADS)

    Ostojski, Arkadiusz

    2018-01-01

    This paper aims to present municipal sewage sludge (MSS) elementary analysis and energetic potential based on measurement of heat of combustion (higher heating value HHV) and calculation of calorific values (lower heating value LHV). The analysis takes into the consideration water content in sewage sludge, at different utilization stages, in wastewater treatment plants in Gdańsk Wschód and Kościerzyna - Pomeranian Voivodeship. The study yielded the following results (in % dry matter): ash 19÷31 %, C - 31÷36 %, H - 5÷6 %, N - 4÷6 %, O - 28÷32 %, S - 1 %. Calorific value of stabilized sludges in Gdańsk was on average 13.8÷15 MJ/kg. In case of sludges not undergoing digestion from Kościerzyna WWTP, the calorific value was at the level of 17.5 MJ/kg. Thus, sewage sludges are good energy carriers. High water content though is the problem, as it lowers the useful effect of heat. There is no alternative for thermal sewage sludge neutralization, which is in conformity with valid Polish National Waste Management Plan (KPGO 2022).

  3. Late Permian volcanic dykes in the crystalline basement of the Považský Inovec Mts. (Western Carpathians): U-Th-Pb zircon SHRIMP and monazite chemical dating

    NASA Astrophysics Data System (ADS)

    Pelech, Ondrej; Vozárová, Anna; Uher, Pavel; Petrík, Igor; Plašienka, Dušan; Šarinová, Katarína; Rodionov, Nikolay

    2017-08-01

    This paper presents geochronological data for the volcanic dykes located in the northern Považský Inovec Mts. The dykes are up to 5 m thick and tens to hundreds of metres long. They comprise variously inclined and oriented lenses, composed of strongly altered grey-green alkali basalts. Their age was variously interpreted and discussed in the past. Dykes were emplaced into the Tatricum metamorphic rocks, mostly consisting of mica schists and gneisses of the Variscan (early Carboniferous) age. Two different methods, zircon SHRIMP and monazite chemical dating, were applied to determine the age of these dykes. U-Pb SHRIMP dating of magmatic zircons yielded the concordia age of 260.2 ± 1.4 Ma. The Th-U-Pb monazite dating of the same dyke gave the CHIME age of 259 ± 3Ma. Both ages confirm the magmatic crystallization at the boundary of the latest Middle Permian to the Late Permian. Dyke emplacement was coeval with development of the Late Paleozoic sedimentary basin known in the northern Považský Inovec Mts. and could be correlated with other pre-Mesozoic Tethyan regions especially in the Southern Alps.

  4. The effect of differentiation agents on inflammatory and oxidative responses of the human neuroblastoma cell line SK-N-SH.

    PubMed

    Niewiarowska-Sendo, Anna; Patrzalek, Katarzyna; Kozik, Andrzej; Guevara-Lora, Ibeth

    2015-01-01

    Obtaining a suitable experimental cellular model is a major problem for neuroscience studies. Neuroblastoma cell lines have been often applied in studies related to pathological disorders of nervous system. However, in the search for an ideal model, these cells must be differentiated to cancel their tumor character. The subsequent reactions that are caused by differentiation are not always indifferent to the same model. We evaluated the effect of two well known substances, used for SH-N-SK cell line differentiation, retinoic acid (RA) and phorbol-12-myristate-13-acetate (PMA), on the induction of pro-inflammatory and pro-oxidative reactions in these cells. Cells differentiated with PMA were able to produce significantly higher amounts of pro-inflammatory cytokines whereas the release of nitric oxide radicals was similar to that in undifferentiated cells. On the contrary, in RA-differentiated cells no significant changes in cytokine production were observed and the nitric oxide release was decreased. Additionally, the RA-differentiated neuronal model was more sensible to lipopolysaccharide stimulation, producing pro-inflammatory cytokines abundantly. These results suggest that RA-differentiated SH-N-SK cells provide a more suitable experimental model for the study of molecular and cellular mechanisms of the inflammation and oxidative stress in neuronal cells.

  5. Elevated STAT3 Signaling-Mediated Upregulation of MMP-2/9 Confers Enhanced Invasion Ability in Multidrug-Resistant Breast Cancer Cells

    PubMed Central

    Zhang, Fei; Wang, Zhiyong; Fan, Yanling; Xu, Qiao; Ji, Wei; Tian, Ran; Niu, Ruifang

    2015-01-01

    The development of multidrug resistance greatly impedes effective cancer therapy. Recent advances in cancer research have demonstrated that acquisition of multidrug resistance by cancer cells is usually accompanied by enhanced cell invasiveness. Several lines of evidence indicated that cross activation of other signaling pathways during development of drug resistance may increase invasive potential of multidrug-resistant (MDR) cancer cells. However, the accurate mechanism of this process is largely undefined. In this study, to better understand the associated molecular pathways responsible for cancer progression induced by drug resistance, a MDR human breast cancer cell line SK-BR-3/EPR with P-glycoprotein overexpression was established using stepwise long-term exposure to increasing concentration of epirubicin. The SK-BR-3/EPR cell line exhibited decreased cell proliferative activity, but enhanced cell invasive capacity. We showed that the expression of metastasis-related matrix metalloproteinase (MMP)-2/9 was elevated in SK-BR-3/EPR cells. Moreover, SK-BR-3/EPR cells showed elevated activation of STAT3. Activation of STAT3 signaling is responsible for enhanced invasiveness of SK-BR-3/EPR cells through upregulation of MMP-2/9. STAT3 is a well-known oncogene and is frequently implicated in tumorigenesis and chemotherapeutic resistance. Our findings augment insight into the mechanism underlying the functional association between MDR and cancer invasiveness. PMID:26501276

  6. Petrographic and Geochemical Characterization of Ore-Bearing Intrusions of the Noril'sk type, Siberia; With Discussion of Their Origin, Including Additional Datasets and Core Logs

    USGS Publications Warehouse

    Czamanske, Gerald K.

    2002-01-01

    The Noril'sk I, Talnakh, and Kharaelakh intrusions of the Noril'sk district host one of the outstanding metal concentrations in the world; contained Cu-Ni resources are comparable to the deposits at Sudbury, Ontario and the platinum group element (PGE) resource is second only to that of the Bushveld Complex. Our opportunity to cooperatively sample and study this district in Siberian Russia arose in 1990 through a memorandum of understanding between the U.S. Geological Survey and the former Ministry of Geology of the U.S.S.R. The world-class significance of these deposits and the possibility that understanding their geologic context, including construction of a credible 'ore-deposit model,' will lead to discovery of similar deposits elsewhere, inspired extensive studies of the ores, the mafic-intrusions which host them, and associated flood basalts.

  7. Analysis of CPolSK-based FSO system working in space-to-ground channel

    NASA Astrophysics Data System (ADS)

    Su, Yuwei; Sato, Takuro

    2018-03-01

    In this article, the transmission performance of a circle polarization shift keying (CPolSK)-based free space optical (FSO) system working in space-to-ground channel is analyzed. Formulas describing the optical polarization distortion caused by the atmospheric turbulence and the communication qualities in terms of signal-to-noise-ratio (SNR), bit-error-ratio (BER) and outage probability of the proposed system are derived. Based on the Stokes parameters data measured by a Japanese optical communication satellite, we evaluate the space-to-ground FSO link and simulate the system performance under a varying regime of turbulence strength. The proposed system provides a more efficient way to compensate scintillation effects in a comparison with the on-off-keying (OOK)-based FSO system. These results are useful to the designing and evaluating of a deep space FSO communication system.

  8. Mild aerobic training with blood flow restriction increases the hypertrophy index and MuSK in both slow and fast muscles of old rats: Role of PGC-1α.

    PubMed

    Bahreinipour, Mohammad-Ali; Joukar, Siyavash; Hovanloo, Fariborz; Najafipour, Hamid; Naderi, Vida; Rajiamirhasani, Alireza; Esmaeili-Mahani, Saeed

    2018-06-01

    Existing evidence emphasize the role of mitochondrial dysfunction in sarcopenia which is revealed as loss of skeletal muscle mass and neuromuscular junction remodeling. We assessed the effect of low-intensity aerobic training along with blood flow restriction on muscle hypertrophy index, muscle-specific kinase (MuSK), a pivotal protein of the neuromuscular junction and Peroxisome proliferator-activated receptor gamma co-activator 1-alpha (PGC-1α) in aged male rats. Animals groups were control (CTL), sham (Sh), leg blood flow restriction (BFR), exercise (Ex), sham + exercise (Sh + Ex), and BFR plus exercise (BFR + Ex) groups. The exercise groups were trained with low intensity exercise for 10 weeks. 48 h after the last training session, animals were sacrificed under anesthesia. Soleus and EDL muscles were isolated, hypertrophy index was estimated and MuSK and PGC-1α were measured by western blot method. Hypertrophy index enhanced in soleus and Extensor digitorum longus (EDL) muscles of BFR + Ex group (P < 0.01 versus CTL and Sh groups, and P < 0.001 versus other groups). The MuSK protein of soleus and EDL muscles increased in BFR + Ex group (P < 0.01 and P < 0.001, respectively) in comparison with CTL and Sh groups. In BFR + Ex group, the PGC-1α protein increased in both soleus and EDL (P < 0.001 compared to other groups). Also the PGC-1α of soleus muscle was higher in Ex and Sh + Ex groups versus CTL and Sh groups (P < 0.05). Findings suggest that low endurance exercise plus BFR improves the MuSK and hypertrophy index of both slow and fast muscles of elderly rats probably through the rise of PGC-1α expression. Copyright © 2018. Published by Elsevier Inc.

  9. Maria Skłodowska-Curie, her life and work - the 150 anniversary of her birthday

    PubMed

    Dobrzyńska, Małgorzata M.

    Maria Skłodowska was born on November 7, 1867 in Warsaw (Poland). Her parents were teachers. Maria’s mother has died in 1878 of tuberculosis. In 1893 and 1894, respectively, Maria was awarded master’s degrees in physics and in mathematics from the Sorbonne University. In 1895 Maria married Pierre Curie. In 1897 their daughter Irene was born. Maria investigated rays emitted by uranium salts. She hypothesized that the radiation come from atom and called this phenomenon “radioactivity”. In 1898, Maria and Pierre discovered new radioactive elements polonium and radium. In 1902 she isolated pure radium chloride and defined radium atomic mass. In June 1903, Maria supervised by Professor Lippmann was awarded her doctorate in physics from the Sorbonne University of Paris after presentation of the thesis “Investigation of radioactive bodies”. In December 1903, Maria was awarded the Nobel Prize in Physics, along with her husband Pierre and Henri Becquerel, for their work on radioactivity. In 1904, the daughter Eve was born. On 19 April 1906, Pierre was killed in a road accident in Paris. In 1910 Maria isolated radium as a pure metal. She also defined an international standard for radioactive emissions (curie), published her fundamental results on radioactivity and textbook of radiology. She also defined the international pattern of radium. In 1911, she won her second Nobel Prize, this time in Chemistry, for her discovery of radium and polonium. In 1914 she was appointed director in the Radium Institute in Paris. During World War I, Maria became the director of the Red Cross Radiology Service and set up France’s first military radiology centre. In May 1932 she has attended the official opening ceremony of the Radium Institute in Warsaw. On 4 July 1934, Maria Skłodowska-Curie has died aged 66 years in Sancellemoz sanatorium (France) of aplastic anemia.

  10. Cellular uptake and anticancer activity of carboxylated gallium corroles.

    PubMed

    Pribisko, Melanie; Palmer, Joshua; Grubbs, Robert H; Gray, Harry B; Termini, John; Lim, Punnajit

    2016-04-19

    We report derivatives of gallium(III) tris(pentafluorophenyl)corrole, 1 [Ga(tpfc)], with either sulfonic (2) or carboxylic acids (3, 4) as macrocyclic ring substituents: the aminocaproate derivative, 3 [Ga(ACtpfc)], demonstrated high cytotoxic activity against all NCI60 cell lines derived from nine tumor types and confirmed very high toxicity against melanoma cells, specifically the LOX IMVI and SK-MEL-28 cell lines. The toxicities of 1, 2, 3, and 4 [Ga(3-ctpfc)] toward prostate (DU-145), melanoma (SK-MEL-28), breast (MDA-MB-231), and ovarian (OVCAR-3) cancer cells revealed a dependence on the ring substituent: IC50values ranged from 4.8 to >200 µM; and they correlated with the rates of uptake, extent of intracellular accumulation, and lipophilicity. Carboxylated corroles 3 and 4, which exhibited about 10-fold lower IC50values (<20 µM) relative to previous analogs against all four cancer cell lines, displayed high efficacy (Emax= 0). Confocal fluorescence imaging revealed facile uptake of functionalized gallium corroles by all human cancer cells that followed the order: 4 > 3 > 2 > 1 (intracellular accumulation of gallium corroles was fastest in melanoma cells). We conclude that carboxylated gallium corroles are promising chemotherapeutics with the advantage that they also can be used for tumor imaging.

  11. Cellular uptake and anticancer activity of carboxylated gallium corroles

    PubMed Central

    Pribisko, Melanie; Palmer, Joshua; Grubbs, Robert H.; Gray, Harry B.; Termini, John; Lim, Punnajit

    2016-01-01

    We report derivatives of gallium(III) tris(pentafluorophenyl)corrole, 1 [Ga(tpfc)], with either sulfonic (2) or carboxylic acids (3, 4) as macrocyclic ring substituents: the aminocaproate derivative, 3 [Ga(ACtpfc)], demonstrated high cytotoxic activity against all NCI60 cell lines derived from nine tumor types and confirmed very high toxicity against melanoma cells, specifically the LOX IMVI and SK-MEL-28 cell lines. The toxicities of 1, 2, 3, and 4 [Ga(3-ctpfc)] toward prostate (DU-145), melanoma (SK-MEL-28), breast (MDA-MB-231), and ovarian (OVCAR-3) cancer cells revealed a dependence on the ring substituent: IC50 values ranged from 4.8 to >200 µM; and they correlated with the rates of uptake, extent of intracellular accumulation, and lipophilicity. Carboxylated corroles 3 and 4, which exhibited about 10-fold lower IC50 values (<20 µM) relative to previous analogs against all four cancer cell lines, displayed high efficacy (Emax = 0). Confocal fluorescence imaging revealed facile uptake of functionalized gallium corroles by all human cancer cells that followed the order: 4 >> 3 > 2 >> 1 (intracellular accumulation of gallium corroles was fastest in melanoma cells). We conclude that carboxylated gallium corroles are promising chemotherapeutics with the advantage that they also can be used for tumor imaging. PMID:27044076

  12. [Analytical chemistry in works of Maria Skłodowska-Curie].

    PubMed

    Hulanicki, Adam

    2012-01-01

    Maria Skłodowska-Curie--a Nobel Prize winner in chemistry--the elements of learning of chemistry gained just by a dint of work of more than ten months in Warsaw in the Institute of Industry and Agriculture Museum. The Nobel Prize concerned a contribution to the progress of chemistry through the discovery of radium and polonium, separation of radium and study of properties of this amazing element. It was awarded for an extremely arduous work, during which the chemical reactions being the principles of analytical chemistry were realized. Unlike to a typical analytical procedure, an initial attempt here was the thousands of kilograms of uranium ore: pitchblende. The final effect was small amounts of new elements: polonium and radium. Both the knowledge and the intuition of the researcher let her have a triumph. The difficulties she experienced because the properties of the searched chemical elements could only be evaluated thanks to the knowledge on other chemical elements. A significant achievement was the determination of the samples by means of radioactivity measurement, which gave rise to radiochemical analytical methods. An extreme analytical precision was demanded in multiple processes of fractional crystallization and precipitation which finally led to the calculation of the atomic mass of radium.

  13. Cellular uptake and retention measurements of alkylphosphocholines in the SK-BR-3 breast cancer and Molt-4 leukemia cell line using capillary gas chromatography.

    PubMed

    Brochez, V; Van Heuverswyn, D; Diniz, J A; De Potter, C R; Van den Eeckhout, E G

    1999-05-01

    The determination of cellular content of octadecylphosphocholine (D-19391) and hexadecylphosphocholine (HePC, D-18506), two anticancer agents of the alkylphosphocholine group, using capillary gas chromatography is described. The compounds' cytotoxicity was first determined by the MTT [3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyltetrazolium] assay, being indicative for the concentration used in the uptake and retention measurements. D-19391 was added to the SK-BR-3 breast cancer cell line and HePC to the Molt-4 leukemia cell line in concentrations of, respectively, 18.6 and 15.0 microM, during a 36-h incubation period at 37 degrees C, 5% CO2. HePC uptake in the leukemia cells was followed by a 24-h reversibility test in drug-free medium. Subsequently, sample clean-up was performed on a weak cation-exchange column. For the quantitative analysis, HePC was used as internal standard for the D-19391 measurements and vice versa. Derivatization of the samples with trimethylsilylbromide was followed by capillary gas chromatographic analysis. From these data we conclude that our uptake results are quite similar with those of a previous study of HePC cellular uptake in the more resistant Caco-2T colon cancer cell line. Without having investigated the mechanism that underlies the cellular uptake results obtained, our study points to no direct correlation between the compounds' cellular uptake and their cytotoxic effects.

  14. Determination of In-situ Rock Thermal Properties from Geophysical Log Data of SK-2 East Borehole, Continental Scientific Drilling Project of Songliao Basin, NE China

    NASA Astrophysics Data System (ADS)

    Zou, C.; Zhao, J.; Zhang, X.; Peng, C.; Zhang, S.

    2017-12-01

    Continental Scientific Drilling Project of Songliao Basin is a drilling project under the framework of ICDP. It aims at detecting Cretaceous environmental/climate changes and exploring potential resources near or beneath the base of the basin. The main hole, SK-2 East Borehole, has been drilled to penetrate through the Cretaceous formation. A variety of geophysical log data were collected from the borehole, which provide a great opportunity to analyze thermal properties of in-situ rock surrounding the borehole.The geothermal gradients were derived directly from temperature logs recorded 41 days after shut-in. The matrix and bulk thermal conductivity of rock were calculated with the geometric-mean model, in which mineral/rock contents and porosity were required as inputs (Fuchs et. al., 2014). Accurate mineral contents were available from the elemental capture spectroscopy logs and porosity data were derived from conventional logs (density, neutron and sonic). The heat production data were calculated by means of the concentrations of uranium, thorium and potassium determined from natural gamma-ray spectroscopy logs. Then, the heat flow was determined by using the values of geothermal gradients and thermal conductivity.The thermal parameters of in-situ rock over the depth interval of 0 4500m in the borehole were derived from geophysical logs. Statistically, the numerical ranges of thermal parameters are in good agreement with the measured values from both laboratory and field in this area. The results show that high geothermal gradient and heat flow exist over the whole Cretaceous formation, with anomalously high values in the Qingshankou formation (1372.0 1671.7m) and the Quantou formation (1671.7 2533.5m). It is meaningful for characterization of geothermal regime and exploration of geothermal resources in the basin. Acknowledgment: This work was supported by the "China Continental Scientific Drilling Program of Cretaceous Songliao Basin (CCSD-SK)" of China

  15. Partial apamin sensitivity of human small conductance Ca2+-activated K+ channels stably expressed in Chinese hamster ovary cells.

    PubMed

    Dale, T J; Cryan, J E; Chen, M X; Trezise, D J

    2002-11-01

    The bee venom toxin apamin is an important drug tool for characterising small conductance Ca(2+)-activated K(+) channels (SK channels). In recombinant expression systems both rSK2 and rSK3 channels are potently blocked by apamin, whilst the sensitivity of SK1 channels is somewhat less clear. In the present study we have conducted a detailed analysis by patch clamp electrophysiology of the effects of apamin on human SK channels (SK1, SK2 and SK3) stably expressed in Chinese hamster ovary (CHO-K1) cells. CHO-K1 cell lines expressing either hSK1, 2 or 3 channels were first validated using specific antibodies and Western blotting. Specific protein bands of a size corresponding to the predicted channel tetramer (approximately 250-290 kDa) were detected. In each cell line, but not wild-type untransfected cells, large, time-independent inwardly rectifying Ca(2+)-dependent K(+) currents were observed under voltage-clamp. In CHO-hSK1, this current was markedly reduced by apamin (IC(50) value 8 nM), however, a significant fraction of the current remained unblocked (39+/-5%), even at saturating concentrations (1 microM apamin). The apamin-sensitive and -insensitive currents possess very similar biophysical and pharmacological properties. Each are Ca(2+)-dependent, inwardly rectify and have relative ionic permeabilities of K(+)>Cs(+)>Li(+)=Na(+). Both components were resistant to block by charybdotoxin and iberiotoxin, known IK and BK channel blockers, but were attenuated by the tricyclic antidepressant cyproheptadine (>95% block at 1 mM). The SK channel opener 1-EBIO could still produce channel activation in the presence of apamin. Importantly, hSK2 and hSK3 channels also exhibit partial apamin sensitivity in our experimental paradigm (IC(50) values of 0.14 nM and 1.1 nM, respectively, and maximal percentage inhibition values of 47+/-7% and 58+/-9%, respectively). Our data indicate that, at least in a recombinant expression system, all three SK channels can be partially

  16. High glucose upregulates BACE1-mediated Aβ production through ROS-dependent HIF-1α and LXRα/ABCA1-regulated lipid raft reorganization in SK-N-MC cells

    PubMed Central

    Lee, Hyun Jik; Ryu, Jung Min; Jung, Young Hyun; Lee, Sei-Jung; Kim, Jeong Yeon; Lee, Sang Hun; Hwang, In Koo; Seong, Je Kyung; Han, Ho Jae

    2016-01-01

    There is an accumulation of evidence indicating that the risk of Alzheimer’s disease is associated with diabetes mellitus, an indicator of high glucose concentrations in blood plasma. This study investigated the effect of high glucose on BACE1 expression and amyloidogenesis in vivo, and we present details of the mechanism associated with those effects. Our results, using ZLC and ZDF rat models, showed that ZDF rats have high levels of amyloid-beta (Aβ), phosphorylated tau, BACE1, and APP-C99. In vitro result with mouse hippocampal neuron and SK-N-MC, high glucose stimulated Aβ secretion and apoptosis in a dose-dependent manner. In addition, high glucose increased BACE1 and APP-C99 expressions, which were reversed by a reactive oxygen species (ROS) scavenger. Indeed, high glucose increased intracellular ROS levels and HIF-1α expression, associated with regulation of BACE1 and Liver X Receptor α (LXRα). In addition, high glucose induced ATP-binding cassette transporter A1 (ABCA1) down-regulation, was associated with LXR-induced lipid raft reorganization and BACE1 localization on the lipid raft. Furthermore, silencing of BACE1 expression was shown to regulate Aβ secretion and apoptosis of SK-N-MC. In conclusion, high glucose upregulates BACE1 expression and activity through HIF-1α and LXRα/ABCA1-regulated lipid raft reorganization, leading to Aβ production and apoptosis of SK-N-MC. PMID:27829662

  17. Developing translational medicine professionals: the Marie Skłodowska-Curie action model.

    PubMed

    Petrelli, Alessandra; Prakken, Berent J; Rosenblum, Norman D

    2016-11-29

    End goal of translational medicine is to combine disciplines and expertise to eventually promote improvement of the global healthcare system by delivering effective therapies to individuals and society. Well-trained experts of the translational medicine process endowed with profound knowledge of biomedical technology, ethical and clinical issues, as well as leadership and teamwork abilities are essential for the effective development of tangible therapeutic products for patients. In this article we focus on education and, in particular, we discuss how programs providing training on the broad spectrum of the translational medicine continuum have still a limited degree of diffusion and do not provide professional support and mentorship in the long-term, resulting in the lack of well established professionals of translational medicine (TMPs) in the scientific community. Here, we describe the Marie Skłodowska-Curie Actions program ITN-EUtrain (EUropean Translational tRaining for Autoimmunity & Immune manipulation Network) where training on the Translational Medicine machinery was integrated with education on professional and personal skills, mentoring, and a long-lasting network of TMPs.

  18. Cytotoxicity and apoptosis of ovarian and breast cancer cell lines induced by OVS1 monoclonal antibody and paclitaxel.

    PubMed

    Moongkarndi, Primchanien; Kaslungka, Sineenart; Kosem, Nuttavut; Junnu, Sarawut; Jongsomboonkusol, Suna; Theptaranon, Yodsaward; Neungton, Neelobol

    2003-03-01

    OVS1 monoclonal antibody (MAb) produced against ovarian cancer is currently used to identify mucinous cystadenocarcinoma antigen as a tumor marker secreted in serum. The potential of OVS1 MAb in ovarian cancer treatment was studied by evaluating the induction of cytotoxicity and apoptosis of SKOV3 ovarian cancer and BT549 breast cancer cell lines induced by OVS1. Paclitaxel, an antitumor drug, was used as positive control and applied as a combined drug together with OVS1 MAb. OVS1 MAb and paclitaxel were found by MTT assay to induce cytotoxicity against both cell lines. The ED50 of OVS1 MAb were 26.25 and 25.00 microg/ml and of paclitaxel were 21.88 and 9.20 nM against SKOV3 and BT549 cell lines, respectively. The quantitative amount of cells determined by fluorimetric assay was correlated to the results of the MTT assay. The combined application of OVS1 MAb and paclitaxel on these two cell lines resulted in a greater cytotoxicity than observed by either agent alone. OVS1 MAb and paclitaxel applied against both cell lines induced the morphological changes of apoptotic cell death at 24 hours visualized by two color fluorescence dyes, Ho33342 and propidium iodide. Combination of the two substances enhanced the rate of apoptosis compared to either OVS1 MAb or paclitaxel given alone. DNA fragmentation was detected in an agarose gel electrophoresis after treating cells with OVS1 MAb and paclitaxel at 24 hours. These findings on the induction of cytotoxicity and apoptosis by OVS1 MAb on cancer cell lines have implications on the potential application of OVS1 MAb for clinical therapy.

  19. The ethanol extract of Scutellaria baicalensis and the active compounds induce cell cycle arrest and apoptosis including upregulation of p53 and Bax in human lung cancer cells

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Gao Jiayu; Morgan, Winston A.; Sanchez-Medina, Alberto

    2011-08-01

    Despite a lack of scientific authentication, Scutellaria baicalensis is clinically used in Chinese medicine as a traditional adjuvant to chemotherapy of lung cancer. In this study, cytotoxicity assays demonstrated that crude ethanolic extracts of S. baicalensis were selectively toxic to human lung cancer cell lines A549, SK-LU-1 and SK-MES-1 compared with normal human lung fibroblasts. The active compounds baicalin, baicalein and wogonin did not exhibit such selectivity. Following exposure to the crude extracts, cellular protein expression in the cancer cell lines was assessed using 2D gel electrophoresis coupled with MALDI-TOF-MS/Protein Fingerprinting. The altered protein expression indicated that cell growth arrestmore » and apoptosis were potential mechanisms of cytotoxicity. These observations were supported by PI staining cell cycle analysis using flow cytometry and Annexin-V apoptotic analysis by fluorescence microscopy of cancer cells treated with the crude extract and pure active compounds. Moreover, specific immunoblotting identification showed the decreased expression of cyclin A results in the S phase arrest of A549 whereas the G{sub 0}/G{sub 1} phase arrest in SK-MES-1 cells results from the decreased expression of cyclin D1. Following treatment, increased expression in the cancer cells of key proteins related to the enhancement of apoptosis was observed for p53 and Bax. These results provide further insight into the molecular mechanisms underlying the clinical use of this herb as an adjuvant to lung cancer therapy. - Research Highlights: > Scutellaria baicalensis is a clinical adjuvant to lung cancer chemotherapy in China. > Scutellaria ethanol extracts selectively toxic to A549, SK-LU-1 and SK-MES-1. > Baicalin, baicalein and wogonin were toxic to all lung cancer cell lines. > Proteomics identified increased p53 and BAX in response to Scutellaria extracts.« less

  20. Effect of a membrane-targeted sphingosine kinase 1 on cell proliferation and survival

    PubMed Central

    2005-01-01

    Numerous extracellular stimuli activate SK1 (sphingosine kinase type 1) to catalyse the production of sphingosine 1-phosphate, a bioactive lipid that functions as both an extracellular ligand for a family of G-protein-linked receptors and as a putative intracellular messenger. Phorbol esters, calcium or immunoglobulin receptors stimulate SK1 by promoting its translocation to the plasma membrane, which brings it into proximity both to its substrate (i.e. sphingosine) and to activating acidic phospholipids (e.g. phosphatidylserine). To evaluate the consequence of SK translocation, we generated an SK1-derivative tagged with a myristoylation sequence (Myr-SK1) on its N-terminus and overexpressed the construct in 3T3-L1 fibroblasts using recombinant retrovirus. Myr-SK1 overexpression increased SK activity by more than 50-fold in crude membranes, while only stimulating cytoplasmic SK activity by 4-fold. In contrast, the overexpression of WT-SK1 (wild-type SK1), as well as that of a construct containing a false myristoylation sequence (A2-Myr-SK1), markedly increased SK activity in both membrane and cytoplasmic compartments. Immunofluorescence confirmed that Myr-SK1 preferentially localized at the plasma membrane, whereas WT-SK1 and A2-Myr-SK1 partitioned in cytoplasmic/perinuclear cellular regions. Surprisingly, Myr-SK1 overexpression significantly decreased the rates of cell proliferation by delaying exit from G0/G1 phase. Moreover, expression of Myr-SK1 but not WT-SK1 or A2-Myr-SK1 protected cells from apoptosis induced by serum withdrawal. Collectively, these findings reveal that altering the subcellular location of SK1 has marked effects on cell function, with plasma membrane-associated SK having a potent inhibitory effect on the G1–S phase transition. PMID:15693752

  1. Felicjan Sławoj-Składkowski (1885-1962): The only known doctor-soldier-politician with an outdoor-wooden-toilet named after him.

    PubMed

    Ohry, Avi

    2017-05-01

    Felicjan Sławoj-Składkowski was a Polish physician, General and politician who served as Polish Minister of Internal Affairs and was the last Prime Minister of Poland before the Second World War. The lack of basic sanitation in many of Poland's villages caused him to issue a decree that every household in Poland must have a latrine in working order. Wooden sheds were built in the backyards, subsequently named 'sławojkis'.

  2. Observations of the anisotropy in the cosmic microwave background by the FIRS, SK93, and MSAM-I experiments

    NASA Technical Reports Server (NTRS)

    Kowitt, Matt; Cheng, Ed; Silverberg, Bob; Ganga, Ken; Page, Lyman; Jarosik, Norm; Netterfield, Barth; Wilkinson, Dave; Meyer, Stephan; Inman, Casey; hide

    1994-01-01

    The observations and results from the FIRS, SK93, and MSAM-1, experiments are discussed. These experiments search for anisotropy in the cosmic microwave background over a range in angular scale from 180 deg to 0.5 deg and a range in frequency from 26 to 680 GHz. Emphasis is placed on the observing strategy and potential systematic errors. Contamination of the data by galactic sources is addressed. Future directions are indicated. The results for all three experiments, as found by us and others, are given in the context of the standard CDM model, Q(sub CDM), and the model-independent band-power estimates.

  3. Biodistribution of iodine-125 and indium-111 labeled OV-TL 3 intact antibodies and F(ab')2 fragments in tumor-bearing athymic mice.

    PubMed

    Massuger, L F; Boerman, O C; Corstens, F H; Verheijen, R H; Claessens, R A; Poels, L G; van den Broek, W J; Kenemans, P

    1991-01-01

    The monoclonal antibody OV-TL 3, directed against an ovarian carcinoma-associated antigenic determinant, was tested as a vehicle for radioimmunolocalization of ovarian carcinomas in athymic mice bearing NIH:OVCAR-3 xenografts. The biodistribution of intact. OV-TL 3 was compared with the distribution of OC 125. Tumor uptake with OV-TL 3 was significantly higher than with OC 125, and almost 7 times higher than with a non-specific control antibody (OV-TL 19). Administration of a mixture of intact OV-TL 3 and OC 125 did not improve tumor uptake in comparison with OV-TL 3 alone. Subsequently, intact OV-TL 3 and its F(ab')2 fragments were labeled with either 111In or 125I. The highest tumor uptake was obtained with 111In-labeled intact OV-TL 3 (14.7% ID/g, 48 hr p.i.). For both antibody forms uptake of 111In in liver, spleen and kidneys was very high. Furthermore, 111In cleared more slowly from most tissues than 125I. As a result, tumor/tissue ratios with 111In-labeled OV-TL 3 were lower than with 125I-labeled OV-TL 3. The highest tumor/tissue ratios (6.9 to 53) were obtained with 125I-labeled OV-TL 3 F(ab')2 fragments, 48 hr post injection. 111In-labeled OV-TL 3 F(ab')2 has already been shown to be a clinically useful label for the detection of ovarian cancer. The results of our comparative animal study suggest that these clinical results may even be improved by using 123I-labeled OV-TL 3 F(ab')2.

  4. The Tanytarsini (Diptera: Chironomidae) in the collection of the Museum of Amber Inclusions, University of Gdańsk.

    PubMed

    Zakrzewska, Marta; Giłka, Wojciech

    2015-04-10

    Non-biting midges of the tribe Tanytarsini collected in the Museum of Amber Inclusions, University of Gdańsk, Poland, are reviewed. Among over 1500 chironomid specimens examined (inclusions in Baltic amber), 44 Tanytarsini individuals were found, of which 27 well preserved specimens were determined to 7 species, including 3 species described as new. Stempellinella electra sp. nov. (male) displays morphological hypopygial characters unique for the genus, and the antenna composed of 13 flagellomeres. A new checklist of fossil and extant species of this genus is also given, including Stempellinella sofiae (Fusari et Lamas, 2014) comb. nov. Tanytarsus glaesarius sp. nov. (male) is the only Eocene species of the genus with a reduced number of antennal flagellomeres. Tanytarsus protogregarius sp. nov. (male) is the oldest known representative of the gregarius species group. Notes on phylogenetic relations of the new species with their extant congeners are also provided.

  5. Whitening and anti-wrinkle activities of ferulic acid isolated from Tetragonia tetragonioides in B16F10 melanoma and CCD-986sk fibroblast cells.

    PubMed

    Park, Hye-Jin; Cho, Jun-Hyo; Hong, Shin-Hyub; Kim, Dong-Hee; Jung, Hee-Young; Kang, In-Kyu; Cho, Young-Je

    2018-01-01

    Ferulic acid isolated from Tetragonia tetragonioides was tested for its whitening effect on the B16F10 mouse melanoma cell line and its anti-wrinkle activity on the CCD-986sk human dermal fibroblast cell line. Ferulic acid, one of the primary phenolic compounds that can be isolated from T. tetragonioides, has been reported to show potential as a functional food, for its whitening effect and anti-wrinkle activity. To measure its whitening and anti-wrinkle activities, cells were treated with ferulic acid isolated from T. tetragonioides at concentrations between 5 and 20 μM. Ferulic acid showed no cytotoxicity at concentrations up to 20 μM. Ferulic acid inhibited melanin synthesis, tyrosinase expression, and microphthalmia transcription factor expression in B16F10 cells stimulated with α-melanocyte stimulating hormone. Ferulic acid induced procollagen synthesis, hyaluronic acid synthesis, tissue inhibitor of metalloproteinase synthesis, and inhibited matrix metalloproteinase (MMP)-1 and MMP-9 expression in CCD-986sk cells stimulated with UV-B. On the basis of these results, we conclude that ferulic acid isolated from T. tetragonioides shows potential for use as a functional food, with whitening and anti-wrinkle activities.

  6. Two homolog wheat Glycogen Synthase Kinase 3/SHAGGY--like kinases are involved in brassinosteroid signaling.

    PubMed

    Bittner, Thomas; Nadler, Sabine; Schulze, Eija; Fischer-Iglesias, Christiane

    2015-10-13

    Glycogen Synthase Kinase 3/SHAGGY-like kinases (GSKs) are multifunctional non-receptor ser/thr kinases. Plant GSKs are involved in hormonal signaling networks and are required for growth, development, light as well as stress responses. So far, most studies have been carried out on Arabidopsis or on other eudicotyledon GSKs. Here, we evaluated the role of TaSK1 and TaSK2, two homolog wheat (Triticum aestivum) GSKs, in brassinosteroid signaling. We explored in addition the physiological effects of brassinosteroids on wheat growth and development. A bin2-1 like gain-of-function mutation has been inserted respectively in one of the homoeologous gene copies of TaSK1 (TaSK1-A.2-1) and in one of the homoeologous gene copies of TaSK2 (TaSK2-A.2-1). Arabidopsis plants were transformed with these mutated gene copies. Severe dwarf phenotypes were obtained closely resembling those of Arabidopsis bin2-1 lines and Arabidopsis BR-deficient or BR-signaling mutants. Expression of BR downstream genes, SAUR-AC1, CPD and BAS1 was deregulated in TaSK1.2-1 and TaSK2.2-1 transgenic lines. Severe dwarf lines were partially rescued by Bikinin beforehand shown to inhibit TaSK kinase activity. This rescue was accompanied with changes in BR downstream gene expression levels. Wheat embryos and seedlings were treated with compounds interfering with BR signaling or modifying BR levels to gain insight into the role of brassinosteroids in wheat development. Embryonic axis and scutellum differentiation were impaired, and seedling growth responses were affected when embryos were treated with Epibrassinolides, Propiconazole, and Bikinin. In view of our findings, TaSKs are proposed to be involved in BR signaling and to be orthologous of Arabidopsis Clade II GSK3/SHAGGY-like kinases. Observed effects of Epibrassinolide, Propiconazole and Bikinin treatments on wheat embryos and seedlings indicate a role for BR signaling in embryonic patterning and seedling growth.

  7. Liquefied Natural Gas Storage of Variable Composition / Magazynowanie Skroplonego Gazu Ziemnego O Zmiennym Składzie

    NASA Astrophysics Data System (ADS)

    Łaciak, Mariusz

    2015-03-01

    Thanks to the increasing diversification of LNG supply sources, being a result of the growing number of LNG liquefaction installations over the World, increase of short-term trade contracts and general trend to globally liberalize gas markets, reception terminals have to cope with the broad range of qualitatively diversified LNG deliveries from various sources. Different LNG deliveries potentially have different density caused by different gas composition. Although the LNG composition depends on LNG source, it mainly consists of methane, ethane, propane, butane and trace nitrogen. When a new supply of LNG is transported to the tank, the LNG composition and temperature in the tank can be different from LNG as delivered. This may lead to the liquid stratification in the tank, and consequently the rollover. As a result, LNG rapidly evaporates and the pressure in the tank increases. More and more restrictive safety regulations require fuller understanding of the formation and evolution of layers. The paper is focused on the analysis of liquid stratification in the tank which may take place when storing LNG, and which process leads to the rapid evaporation of considerable quantities of LNG. The aim was to attempt modeling of the process of liquid stratification in an LNG tank. The paper is closed with the results of modelling. Dzięki rosnącej dywersyfikacji źródeł dostaw LNG, spowodowanej zwiększającą się liczbą instalacji skraplania gazu na całym świecie, wzrostem ilości kontraktów krótkoterminowych w handlu i ogólnej tendencji do globalnej liberalizacja rynków gazu, terminale do odbioru muszą radzić sobie z coraz większą gamą różnych jakościowo dostaw LNG z różnych źródeł. Różne dostawy LNG mają potencjalnie inną gęstość dzięki różnym składom gazu. Chociaż kompozycja LNG zależy od źródła, to przede wszystkim składa się z metanu, etanu, propanu, butanu i w śladowych ilościach z azotu. Gdy nowa

  8. Rice MEL2, the RNA recognition motif (RRM) protein, binds in vitro to meiosis-expressed genes containing U-rich RNA consensus sequences in the 3'-UTR.

    PubMed

    Miyazaki, Saori; Sato, Yutaka; Asano, Tomoya; Nagamura, Yoshiaki; Nonomura, Ken-Ichi

    2015-10-01

    Post-transcriptional gene regulation by RNA recognition motif (RRM) proteins through binding to cis-elements in the 3'-untranslated region (3'-UTR) is widely used in eukaryotes to complete various biological processes. Rice MEIOSIS ARRESTED AT LEPTOTENE2 (MEL2) is the RRM protein that functions in the transition to meiosis in proper timing. The MEL2 RRM preferentially associated with the U-rich RNA consensus, UUAGUU[U/A][U/G][A/U/G]U, dependently on sequences and proportionally to MEL2 protein amounts in vitro. The consensus sequences were located in the putative looped structures of the RNA ligand. A genome-wide survey revealed a tendency of MEL2-binding consensus appearing in 3'-UTR of rice genes. Of 249 genes that conserved the consensus in their 3'-UTR, 13 genes spatiotemporally co-expressed with MEL2 in meiotic flowers, and included several genes whose function was supposed in meiosis; such as Replication protein A and OsMADS3. The proteome analysis revealed that the amounts of small ubiquitin-related modifier-like protein and eukaryotic translation initiation factor3-like protein were dramatically altered in mel2 mutant anthers. Taken together with transcriptome and gene ontology results, we propose that the rice MEL2 is involved in the translational regulation of key meiotic genes on 3'-UTRs to achieve the faithful transition of germ cells to meiosis.

  9. Evaluation of ultrasound in assessing body composition of high school wrestlers.

    PubMed

    Utter, Alan C; Hager, Marion E

    2008-05-01

    To evaluate the accuracy of ultrasound (ULTRA) in assessing fat-free mass (FFM) in comparison with hydrostatic weighing (HW) and skinfolds (SK) in high school wrestlers in a hydrated state. Body composition was determined by ULTRA, HW, and three-site SK in 70 high school wrestlers (mean +/- SD: age, 15.5 +/- 1.5; height, 1.60 +/- 0.08 m; body mass, 65.8 +/- 12.7 kg). For all methods, body density (Db) was converted to percent body fat (%BF) using the Brozek equation. Hydration state was quantified by evaluating urine specific gravity. There were no significant differences for estimated FFM between ULTRA (57.2 +/- 9.7 kg) and HW (57.0 +/- 9.9 kg); however, SK (54.9 +/- 8.8 kg) were significantly different from HW. The standard errors of estimate for FFM with HW as the reference method were 2.40 kg for ULTRA and 2.74 kg for SK. Significant correlations were found for FFM between HW and ULTRA (r = 0.97, P < 0.001) and between HW and SK (r = 0.96, P < 0.001). A systematic bias was found for SK, as the difference between SK and HW significantly correlated with the FFM average of the two methods (r = -0.38, P < 0.001). This systematic bias was not found for ULTRA (r = - 0.07). This study demonstrates that ULTRA provides similar estimates of FFM when compared with HW in a heterogeneous high school wrestling population during a hydrated state. ULTRA should be considered as an alternative field-based method of estimating the FFM of high school wrestlers.

  10. Lycopene inhibits reactive oxygen species production in SK-Hep-1 cells and attenuates acetaminophen-induced liver injury in C57BL/6 mice.

    PubMed

    Bandeira, Ana Carla Balthar; da Silva, Talita Prato; de Araujo, Glaucy Rodrigues; Araujo, Carolina Morais; da Silva, Rafaella Cecília; Lima, Wanderson Geraldo; Bezerra, Frank Silva; Costa, Daniela Caldeira

    2017-02-01

    Our aim was to investigate the antioxidant potential of lycopene in different experimental liver models: in vitro, to evaluate the influence of lycopene on reactive oxygen species (ROS) production mediated by the PKC pathway and in vivo, to evaluate the protective effects of lycopene in an experimental model of hepatotoxicity. The in vitro study assessed the lycopene antioxidant potential by the quantification of ROS production in SK-Hep-1 cells unstimulated or stimulated by an activator of the PKC pathway. The role of NADPH oxidase was evaluated by measuring its inhibition potential using an inhibitor of this enzyme. In the in vivo study, male C57BL/6 mice received lycopene (10 or 100 mg/kg by oral gavage) and 1 h later, acetaminophen (APAP) (500 mg/kg) was administrated. Lycopene decreased ROS production in SK-Hep-1 cells through inhibition of NADPH oxidase, brought about in the PKC pathway. Lycopene improved hepatotoxicity acting as an antioxidant, reduced GSSG and regulated tGSH and CAT levels, reduced oxidative damage primarily by decreasing protein carbonylation, promoted the downregulation of MMP-2 and reduced areas of necrosis improving the general appearance of the lesion in C57BL/6 mice. Lycopene is a natural compound that was able to inhibit the production of ROS in vitro and mitigate the damage caused by APAP overdose in vivo. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

  11. Multiepitope HER2 targeting enhances photoimmunotherapy of HER2-overexpressing cancer cells with pyropheophorbide-a immunoconjugates.

    PubMed

    Savellano, Mark D; Pogue, Brian W; Hoopes, P Jack; Vitetta, Ellen S; Paulsen, Keith D

    2005-07-15

    Multi-targeting strategies improve the efficacy of antibody and immunotoxin therapies but have not yet been thoroughly explored for HER2-based cancer treatments. We investigated multi-epitope HER2 targeting to boost photosensitizer immunoconjugate uptake as a way of enhancing photoimmunotherapy. Photoimmunotherapy may allow targeted photodynamic destruction of malignancies and may also potentiate anticancer antibodies. However, one obstacle preventing its clinical use is the delivery of enough photosensitizer immunoconjugates to target cells. Anti-HER2 photosensitizer immunoconjugates were constructed from two monoclonal antibodies (mAb), HER50 and HER66, using a novel method originally developed to label photosensitizer immunoconjugates with the photosensitizer, benzoporphyrin derivative verteporfin. Photosensitizer immunoconjugates were labeled instead with a promising alternative photosensitizer, pyropheophorbide-a (PPa), which required only minor changes to the conjugation procedure. Uptake and phototoxicity experiments using human cancer cells were conducted with the photosensitizer immunoconjugates and, for comparison, with free PPa. SK-BR-3 and SK-OV-3 cells served as HER2-overexpressing target cells. MDA-MB-468 cells served as HER2-nonexpressing control cells. Photosensitizer immunoconjugates with PPa/mAb molar ratios up to approximately 10 specifically targeted and photodynamically killed HER2-overexpressing cells. On a per mole basis, photosensitizer immunoconjugates were less phototoxic than free PPa, but photosensitizer immunoconjugates were selective for target cells whereas free PPa was not. Multiepitope targeted photoimmunotherapy with a HER50 and HER66 photosensitizer immunoconjugate mixture was significantly more effective than single-epitope targeted photoimmunotherapy with a single anti-HER2 photosensitizer immunoconjugate, provided photosensitizer immunoconjugate binding was saturated. This study shows that multiepitope targeting enhances HER2

  12. [Medicine and politics in the lives of Kazimierz Noiszewski, Józef Piłsudski, Bolesław Wieniawa-Długoszowski and Felicjan Sławoj Składkowski].

    PubMed

    Polak, Agnieszka; Rejdak, Robert; Czepita, Damian; Rekas, Marek; Wróbel, Andrzej

    2015-01-01

    Changes in the economy in Poland in the years 1864-1914 generated changes in the structure of society. A new social stratum, the intelligentsia, was formed from the educated children of rich landowners. To meet social needs Polish universities in Cracow and Lviv opened new departments of medicine. The article presents fragments of the biographies of 4 Poles: Kazimierz Noiszewski, Józef Piłsudski, Bolesław Wieniawa-Długoszowski and Stanislaw Sławoj Składkowski to illustrate how their study of medicine affected their further activity as patriots and politicians. Kazimierz Noiszewski was an emigrant who lived and worked as an ophthalmologist in Russia. He tried to support the Polish community and Polish medical knowledge. For Józef Pilsudski and Stanisław Sławoj Składkowski medical studies gave freedom of thought and the opportunity to meet other patriots. Boleslaw Wieniawa-Długoszowski was a man of many interests, and after graduating from a medical university in Lviv, and a short stay in Paris, he became engaged in military service for his country.

  13. Tyrosinase overexpression promotes ATM-dependent p53 phosphorylation by quercetin and sensitizes melanoma cells to dacarbazine.

    PubMed

    Thangasamy, Thilakavathy; Sittadjody, Sivanandane; Limesand, Kirsten H; Burd, Randy

    2008-01-01

    Dacarbazine (DTIC) has been used for the treatment of melanoma for decades. However, monotherapy with this chemotherapeutic agent results only in moderate response rates. To improve tumor response to DTIC current clinical trials in melanoma focus on combining a novel targeted agent with chemotherapy. Here, we demonstrate that tyrosinase which is commonly overexpressed in melanoma activates the bioflavonoid quercetin (Qct) and promotes an ataxia telangiectasia mutated (ATM)-dependent DNA damage response. This response sensitizes melanoma cells that overexpress tyrosinase to DTIC. In DB-1 melanoma cells that overexpress tyrosinase (Tyr(+) cells), the threshold for phosphorylation of ATM and p53 at serine 15 was observed at a low dose of Qct (25 microM) when compared to the mock transfected pcDNA3 cells, which required a higher dose (75 microM). Both pcDNA3 and Tyr(+) DB-1 cells demonstrated similar increases in phosphorylation of p53 at other serine sites, but in the Tyr(+) cells, DNApk expression was found to be reduced compared to control cells, indicating a shift towards an ATM-mediated response. The DB-1 control cells were resistant to DTIC, but were sensitized to apoptosis with high dose Qct, while Tyr(+) cells were sensitized to DTIC with low or high dose Qct. Qct also sensitized SK Mel 5 (p53 wildtype) and 28 (p53 mutant) cells to DTIC. However, when SK Mel 5 cells were transiently transfected with tyrosinase and treated with Qct plus DTIC, SK Mel 5 cells demonstrated a more than additive induction of apoptosis. Therefore, this study demonstrates that tyrosinase overexpression promotes an ATM-dependent p53 phosphorylation by Qct treatment and sensitizes melanoma cells to dacarbazine. In conclusion, these results suggest that Qct or Qct analogues may significantly improve DTIC response rates in tumors that express tyrosinase.

  14. Tyrosinase Overexpression Promotes ATM-Dependent p53 Phosphorylation by Quercetin and Sensitizes Melanoma Cells to Dacarbazine

    PubMed Central

    Thangasamy, Thilakavathy; Sittadjody, Sivanandane; H. Limesand, Kirsten; Burd, Randy

    2008-01-01

    Dacarbazine (DTIC) has been used for the treatment of melanoma for decades. However, monotherapy with this chemotherapeutic agent results only in moderate response rates. To improve tumor response to DTIC current clinical trials in melanoma focus on combining a novel targeted agent with chemotherapy. Here, we demonstrate that tyrosinase which is commonly overexpressed in melanoma activates the bioflavonoid quercetin (Qct) and promotes an ataxia telangiectasia mutated (ATM)-dependent DNA damage response. This response sensitizes melanoma cells that overexpress tyrosinase to DTIC. In DB-1 melanoma cells that overexpress tyrosinase (Tyr cells), the threshold for phosphorylation of ATM and p53 at serine 15 was observed at a low dose of Qct (25 μM) when compared to the mock transfected pcDNA3 cells, which required a higher dose (75 μM). Both pcDNA3 and Tyr DB-1 cells demonstrated similar increases in phosphorylation of p53 at other serine sites, but in the Tyr cells, DNApk expression was found to be reduced compared to control cells, indicating a shift towards an ATM-mediated response. The DB-1 control cells were resistant to DTIC, but were sensitized to apoptosis with high dose Qct, while Tyr cells were sensitized to DTIC with low or high dose Qct. Qct also sensitized SK Mel 5 (p53 wildtype) and 28 (p53 mutant) cells to DTIC. However, when SK Mel 5 cells were transiently transfected with tyrosinase and treated with Qct plus DTIC, SK Mel 5 cells demonstrated a more than additive induction of apoptosis. Therefore, this study demonstrates that tyrosinase overexpression promotes an ATM-dependent p53 phosphorylation by Qct treatment and sensitizes melanoma cells to dacarbazine. In conclusion, these results suggest that Qct or Qct analogues may significantly improve DTIC response rates in tumors that express tyrosinase. PMID:18791269

  15. The opposing effects of calmodulin, adenosine 5 prime -triphosphate, and pertussis toxin on phorbol ester induced inhibition of atrial natriuretic factor stimulated guanylate cyclase in SK-NEP-1 cells

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Sekiya, M.; Frohlich, E.D.; Cole, F.E.

    1991-01-01

    In the present study, we investigated the effects of calmodulin, adenosine 5{prime}-triphosphate (ATP) and pertussis toxin (PT) on phorbol ester (PMA) induced inhibition of ANF-stimulated cyclic GMP formation in cells from the human renal cell line, SK-NEP-1. PMA inhibited ANF-stimulated guanylate cyclase activity in particulate membranes by about 65%. Calmodulin reversed this inhibition in a dose dependent manner. ATP potentiated Mg++ but not Mn++ supported guanylate cyclase activity. In PMA treated membranes, ATP potentiating effects were abolished. PMA also inhibited ANF-stimulated cGMP accumulation, but pretreatment with PT prevented this PMA inhibition. PT did not affect basal or ANF-stimulated cGMP accumulation.more » In conclusion, these results demonstrated that PMA inhibited ANF stimulation of particulate guanylate cyclase in opposition to the activating effects of calmodulin or ATP in SK-NEP-1 cells. The protein kinase C inhibitory effects appeared to be mediated via a PT-sensitive G protein.« less

  16. Structural and computational characterization of 4‧,4‧,6‧,6‧-tetrachloro-3-(2-methoxyethyl)-3H,4H-spiro-1,3,2-benzoxaza phosphinine-2,2‧- [1,3,5,2,4,6] triazatriphosphinine

    NASA Astrophysics Data System (ADS)

    Işıklan, Muhammet; Yıldırım, Erdem Kamil; Atiş, Murat; Sonkaya, Ömer; Çoşut, Bünyemin

    2016-08-01

    In this study a new monospirocyclic phosphazene derivative, 4‧,4‧,6‧,6‧-tetrachloro-3-(2-methoxyethyl)-3H,4H-spiro [1,3,2-benzoxazaphosphinine-2,2‧- [1,3,5,2,4,6] triazatriphosphinine] (SP1) was synthesized from the reaction of hexachlorocyclotriphosphazene (N3P3Cl6) with N/O donor-type, 2-{[(2-Metoxyethyl) amino]methyl}phenol. The structural investigations of the compound were verified by elemental analyses, MS, FTIR, 1H, 13C, 31P NMR spectroscopy and the single crystal X-ray diffraction analysis. The structural and spectroscopic data of the molecule in the ground state were calculated by using density functional method (DFT) using 6-311++G (d, p) basis set. The complete assignments of all vibrational modes were performed on the basis of the total energy distributions (TED). Isotropic chemical shifts (31P, 1H and 13C NMR) were calculated using the gauge-invariant atomic orbital (GIAO) method. Theoretical calculations of bond parameters, harmonic vibration frequencies and nuclear magnetic resonance are in good agreement with experimental results. The electrophilic and nucleophilic attack centers in SP1 were predicted with the local softness values (sk+, and sk-) of individual atoms and it is confirmed that P atoms of the PCl2 groups are nucleophilic attack centers.

  17. Immunoscintigraphy of ovarian cancer with indium-111-labeled OV-TL 3 F(ab')2 monoclonal antibody.

    PubMed

    Massuger, L F; Kenemans, P; Claessens, R A; Verheijen, R H; Schijf, C P; Strijk, S P; Poels, L G; van Hoesel, R G; Corstens, F H

    1990-11-01

    The safety and diagnostic accuracy of immunoscintigraphy with the indium-111-labeled monoclonal antibody OV-TL 3 F(ab')2(111In-OV-TL 3 F(ab')2) for diagnosis and follow-up of ovarian cancer was prospectively studied in 31 patients. Planar and SPECT scintigraphy were performed up to 4 days after i.v. injection of 140 MBq 111In-OV-TL 3 F(ab')2. Surgical evaluation was possible in 22 out of 31 patients. Imaging results were compared with X-ray computed tomography, ultrasound, and CA 125 serum level using the histologically confirmed surgical findings as a "gold standard." Apart from a transient rash observed in two patients, no other immediate or delayed adverse reactions were observed. Within the surgically evaluated group, ovarian cancer lesions were detected in 16 out of 17 patients (94%). Of 45 distinct tumor deposits found at operation, 67% were detected and localized with immunoscintigraphy while X-ray computed tomography and ultrasound visualized 53% and 23%, respectively.

  18. Identification and characterization of two wheat Glycogen Synthase Kinase 3/ SHAGGY-like kinases.

    PubMed

    Bittner, Thomas; Campagne, Sarah; Neuhaus, Gunther; Rensing, Stefan A; Fischer-Iglesias, Christiane

    2013-04-18

    Plant Glycogen Synthase Kinase 3/ SHAGGY-like kinases (GSKs) have been implicated in numerous biological processes ranging from embryonic, flower, stomata development to stress and wound responses. They are key regulators of brassinosteroid signaling and are also involved in the cross-talk between auxin and brassinosteroid pathways. In contrast to the human genome that contains two genes, plant GSKs are encoded by a multigene family. Little is known about Liliopsida resp. Poaceae in comparison to Brassicaceae GSKs. Here, we report the identification and structural characterization of two GSK homologs named TaSK1 and TaSK2 in the hexaploid wheat genome as well as a widespread phylogenetic analysis of land plant GSKs. Genomic and cDNA sequence alignments as well as chromosome localization using nullisomic-tetrasomic lines provided strong evidence for three expressed gene copies located on homoeolog chromosomes for TaSK1 as well as for TaSK2. Predicted proteins displayed a clear GSK signature. In vitro kinase assays showed that TaSK1 and TaSK2 possessed kinase activity. A phylogenetic analysis of land plant GSKs indicated that TaSK1 and TaSK2 belong to clade II of plant GSKs, the Arabidopsis members of which are all involved in Brassinosteroid signaling. Based on a single ancestral gene in the last common ancestor of all land plants, paralogs were acquired and retained through paleopolyploidization events, resulting in six to eight genes in angiosperms. More recent duplication events have increased the number up to ten in some lineages. To account for plant diversity in terms of functionality, morphology and development, attention has to be devoted to Liliopsida resp Poaceae GSKs in addition to Arabidopsis GSKs. In this study, molecular characterization, chromosome localization, kinase activity test and phylogenetic analysis (1) clarified the homologous/paralogous versus homoeologous status of TaSK sequences, (2) pointed out their affiliation to the GSK multigene

  19. Ganglioside GM2 mediates migration of tumor cells by interacting with integrin and modulating the downstream signaling pathway.

    PubMed

    Kundu, Manjari; Mahata, Barun; Banerjee, Avisek; Chakraborty, Sohini; Debnath, Shibjyoti; Ray, Sougata Sinha; Ghosh, Zhumur; Biswas, Kaushik

    2016-07-01

    The definitive role of ganglioside GM2 in mediating tumor-induced growth and progression is still unknown. Here we report a novel role of ganglioside GM2 in mediating tumor cell migration and uncovered its mechanism. Data shows differential expression levels of GM2-synthase as well as GM2 in different human cancer cells. siRNA mediated knockdown of GM2-synthase in CCF52, A549 and SK-RC-26B cells resulted in significant inhibition of tumor cell migration as well as invasion in vitro without affecting cellular proliferation. Over-expression of GM2-synthase in low-GM2 expressing SK-RC-45 cells resulted in a consequent increase in migration thus confirming the potential role GM2 and its downstream partners play in tumor cell migration and motility. Further, treatment of SK-RC-45 cells with exogenous GM2 resulted in a dramatic increase in migratory and invasive capacity with no change in proliferative capacity, thereby confirming the role of GM2 in tumorigenesis specifically by mediating tumor migration and invasion. Gene expression profiling of GM2-synthase silenced cells revealed altered expression of several genes involved in cell migration primarily those controlling the integrin mediated signaling. GM2-synthase knockdown resulted in decreased phosphorylation of FAK, Src as well as Erk, while over-expression and/or exogenous GM2 treatment caused increased FAK and Erk phosphorylation respectively. Again, GM2 mediated invasion and Erk phosphorylation is blocked in integrin knockdown SK-RC-45 cells, thus confirming that GM2 mediated migration and phosphorylation of Erk is integrin dependent. Finally, confocal microscopy suggested co-localization while co-immunoprecipitation and surface plasmon resonance (SPR) confirmed direct interaction of membrane bound ganglioside, GM2 with the integrin receptor. Copyright © 2016 Elsevier B.V. All rights reserved.

  20. Next-generation Sequencing (NGS) Analysis on Single Circulating Tumor Cells (CTCs) with No Need of Whole-genome Amplification (WGA).

    PubMed

    Palmirotta, Raffaele; Lovero, Domenica; Silvestris, Erica; Felici, Claudia; Quaresmini, Davide; Cafforio, Paola; Silvestris, Franco

    2017-01-01

    Isolation and genotyping of circulating tumor cells (CTCs) is gaining an increasing interest by clinical researchers in oncology not only for investigative purposes, but also for concrete application in clinical practice in terms of diagnosis, prognosis and decision treatment with targeted therapies. For the mutational analysis of single CTCs, the most advanced biotechnology methodology currently available includes the combination of whole genome amplification (WGA) followed by next-generation sequencing (NGS). However, the sequence of these molecular techniques is time-consuming and may also favor operator-dependent errors, related to the procedures themselves that, as in the case of the WGA technique, might affect downstream molecular analyses. A preliminary approach of molecular analysis by NGS on a model of CTCs without previous WGA procedural step was performed. We set-up an artificial sample obtained by spiking the SK-MEL-28 melanoma cell line in normal donor peripheral whole blood. Melanoma cells were first enriched using an AutoMACS® (Miltenyi) cell separator and then isolated as single and pooled CTCs by DEPArray™ System (Silicon Biosystems). NGS analysis, using the Ion AmpliSeq™ Cancer Hotspot Panel v2 (Life Technologies) with the Ion Torrent PGM™ system (Life Technologies), was performed on the SK-MEL-28 cell pellet, a single CTC previously processed with WGA and on 1, 2, 4 and 8 recovered CTCs without WGA pre-amplification. NGS directly carried out on CTCs without WGA showed the same mutations identified in SK-MEL-28 cell line pellet, with a considerable efficiency and avoiding the errors induced by the WGA procedure. We identified a cost-effective, time-saving and reliable methodological approach that could improve the analytical accuracy of the liquid biopsy and appears promising in studying CTCs from cancer patients for both research and clinical purposes. Copyright© 2017, International Institute of Anticancer Research (Dr. George J

  1. Agrin-LRP4-MuSK signaling as a therapeutic target for myasthenia gravis and other neuromuscular disorders.

    PubMed

    Ohno, Kinji; Ohkawara, Bisei; Ito, Mikako

    2017-10-01

    Signal transduction at the neuromuscular junction (NMJ) is compromised in a diverse array of diseases including myasthenia gravis, Lambert-Eaton myasthenic syndrome, Isaacs' syndrome, congenital myasthenic syndromes, Fukuyama-type congenital muscular dystrophy, amyotrophic lateral sclerosis, and sarcopenia. Except for sarcopenia, all are orphan diseases. In addition, the NMJ signal transduction is impaired by tetanus, botulinum, curare, α-bungarotoxin, conotoxins, organophosphate, sarin, VX, and soman to name a few. Areas covered: This review covers the agrin-LRP4-MuSK signaling pathway, which drives clustering of acetylcholine receptors (AChRs) and ensures efficient signal transduction at the NMJ. We also address diseases caused by autoantibodies against the NMJ molecules and by germline mutations in genes encoding the NMJ molecules. Expert opinion: Representative small compounds to treat the defective NMJ signal transduction are cholinesterase inhibitors, which exert their effects by increasing the amount of acetylcholine at the synaptic space. Another possible therapeutic strategy to enhance the NMJ signal transduction is to increase the number of AChRs, but no currently available drug has this functionality.

  2. Impact of the initial classic section during a simulated cross-country skiing skiathlon on the cardiopulmonary responses during the subsequent period of skate skiing.

    PubMed

    Mourot, Laurent; Fabre, Nicolas; Andersson, Erik; Willis, Sarah J; Hébert-Losier, Kim; Holmberg, Hans-Christer

    2014-08-01

    The aim of this study was to assess potential changes in the performance and cardiorespiratory responses of elite cross-country skiers following transition from the classic (CL) to the skating (SK) technique during a simulated skiathlon. Eight elite male skiers performed two 6 km (2 × 3 km) roller-skiing time trials on a treadmill at racing speed: one starting with the classic and switching to the skating technique (CL1-SK2) and another employing the skating technique throughout (SK1-SK2), with continuous monitoring of gas exchanges, heart rates, and kinematics (video). The overall performance times in the CL1-SK2 (21:12 ± 1:24) and SK1-SK2 (20:48 ± 2:00) trials were similar, and during the second section of each performance times and overall cardiopulmonary responses were also comparable. However, in comparison with SK1-SK2, the CL1-SK2 trial involved significantly higher increases in minute ventilation (V̇E, 89.8 ± 26.8 vs. 106.8 ± 17.6 L·min(-1)) and oxygen uptake (V̇O2; 3.1 ± 0.8 vs 3.5 ± 0.5 L·min(-1)) 2 min after the transition as well as longer time constants for V̇E, V̇O2, and heart rate during the first 3 min after the transition. This higher cardiopulmonary exertion was associated with ∼3% faster cycle rates. In conclusion, overall performance during the 2 time trials did not differ. The similar performance times during the second sections were achieved with comparable mean cardiopulmonary responses. However, the observation that during the initial 3-min post-transition following classic skiing cardiopulmonary responses and cycle rates were slightly higher supports the conclusion that an initial section of classic skiing exerts an impact on performance during a subsequent section of skate skiing.

  3. Ferrotochilinite, 6FeS · 5Fe(OH)2, a new mineral from the Oktyabr'sky deposit, Noril'sk district, Siberia, Russia

    NASA Astrophysics Data System (ADS)

    Pekov, I. V.; Sereda, E. V.; Polekhovsky, Yu. S.; Britvin, S. N.; Chukanov, N. V.; Yapaskurt, V. O.; Bryzgalov, I. A.

    2013-12-01

    A new mineral, ferrotochilinite, ideally 6FeS · 5Fe(OH)2, was found at the Oktyabr'sky Mine, Oktyabr'skoe Cu-Ni deposit, Noril'sk, Krasnoyarsk krai, Siberia, Russia. It is associated with ferrovalleriite, magnetite and Fe-rich, chlorite-like phyllosilicate in the cavities of pentlandite-mooihoekite-cubanite ore with subordinate magnetite and chalcopyrite. Ferrotochilinite occurs as flattened on [001], prismatic to elongated lamellar crystals up to 0.1 × 0.5 × 3.2 mm, typically split and curved. Aggregates (up to 6.5 mm in size) are fanlike, rosette-like, or chaotic. Ferrotochilinite is dark bronze. The streak is black. The luster is moderately metallic. The Mohs' hardness is ca. 1; VHN is 13 kg/mm2. Cleavage is {001} perfect, micalike. Individuals are flexible, inelastic. D(calc) = 3.467 g/cm3. In reflected light, ferrotochilinite is gray, with the hue changing from pale beige to bluish; bireflectance is distinct. Anisotropy is distinct, with gray bluish to yellowish beige rotation colors. No internal reflections. Reflectance values [ R min- R max, % (λ, nm)] are: 11.6-11.4 (470), 11.2-12.4 (546), 11.1-13.6 (589), 11.0-15.5 (650). The IR spectrum shows the presence of (OH) groups bonded with Fe cations and the absence of H2O molecules. Chemical composition (wt %; electron probe; H content is calculated) is as follows: 0.02 Mg, 61.92 Fe, 0.03 Ni, 0.09 Cu, 19.45 S, 16.3 O, 1.03 H calc; the total is 98.84. The empirical formula calculated on the basis of 6 S atoms is: Mg0.01Fe10.96Ni0.005Cu0.015S6(OH)10.07 = (Fe5.98Cu0.0015Ni0.005)Σ6S6(OH)9.80(Fe{4.89/2+}Mg0.01)Σ4.90(OH)9.80Fe{0.09/3+}(OH)0.27. Ferrotochilinite is monoclinic, space group is C2/ m, Cm or C2, the unit-cell dimensions are: a = 5.463(5), b = 15.865(17), c = 10.825(12) Å, β = 93.7(1)°, V = 936(3) Å3, Z = 2. The strongest reflections in the X-ray powder diffraction pattern ( d, Å- I[ hkl]) are: 10.83-13[001], 5.392-100[002], 3.281-7[023], 2.777-7[150], 2.696-12[004, ], 2.524-12[, ], 2

  4. Protein disulfide isomerases: Impact of thapsigargin treatment on their expression in melanoma cell lines.

    PubMed

    Silva, Zélia; Veríssimo, Teresa; Videira, Paula A; Novo, Carlos

    2015-08-01

    Anti-cancer treatments usually elevate the content of unfolded or misfolded proteins in the endoplasmic reticulum (ER). Here we aimed to get insights into the relation between sensitivity of melanoma cell lines to the ER stress inducer thapsigargin (THG) and the genetic expression of protein disulfide isomerase family members (PDIs). The expression of PDIs was analysed by flow cytometry and real-time PCR. The results showed that SK-MEL-30, the less THG sensitive cell line, displays higher basal PDIs' expression levels and the sensitivity is increased by the PDIs inhibitor bacitracin. While SK-MEL-30 PDIs' expression is not THG dose-dependent, an increase in glucose related protein 78 (GRP78), PDIA5, PDIA6, and thioredoxin-related-transmembrane proteins' (TMX3 and TMX4) expression, in response to higher drug concentrations, was observed in MNT-1. The differences in PDIs' gene expression in MNT-1 suggest a different response to ER stress compared to the other cell lines and highlight the importance of understanding the diversity among cancer cells. Copyright © 2015 Elsevier B.V. All rights reserved.

  5. Small and intermediate conductance Ca(2+)-activated K+ channels confer distinctive patterns of distribution in human tissues and differential cellular localisation in the colon and corpus cavernosum.

    PubMed

    Chen, Mao Xiang; Gorman, Shelby A; Benson, Bill; Singh, Kuljit; Hieble, J Paul; Michel, Martin C; Tate, Simon N; Trezise, Derek J

    2004-06-01

    The SK/IK family of small and intermediate conductance calcium-activated potassium channels contains four members, SK1, SK2, SK3 and IK1, and is important for the regulation of a variety of neuronal and non-neuronal functions. In this study we have analysed the distribution of these channels in human tissues and their cellular localisation in samples of colon and corpus cavernosum. SK1 mRNA was detected almost exclusively in neuronal tissues. SK2 mRNA distribution was restricted but more widespread than SK1, and was detected in adrenal gland, brain, prostate, bladder, liver and heart. SK3 mRNA was detected in almost every tissue examined. It was highly expressed in brain and in smooth muscle-rich tissues including the clitoris and the corpus cavernosum, and expression in the corpus cavernosum was upregulated up to 5-fold in patients undergoing sex-change operations. IK1 mRNA was present in surface-rich, secretory and inflammatory cell-rich tissues, highest in the trachea, prostate, placenta and salivary glands. In detailed immunohistochemical studies of the colon and the corpus cavernosum, SK1-like immunoreactivity was observed in the enteric neurons. SK3-like immunoreactivity was observed strongly in smooth muscle and vascular endothelium. IK1-like immunoreactivity was mainly observed in inflammatory cells and enteric neurons of the colon, but absent in corpus cavernosum. These distinctive patterns of distribution suggest that these channels are likely to have different biological functions and could be specifically targeted for a number of human diseases, such as irritable bowel syndrome, hypertension and erectile dysfunction.

  6. Seven diverse human embryonic stem cell-derived chondrogenic clonal embryonic progenitor cell lines display site-specific cell fates.

    PubMed

    Sternberg, Hal; Kidd, Jennifer; Murai, James T; Jiang, Jianjie; Rinon, Ariel; Erickson, Isaac E; Funk, Walter D; Wang, Qian; Chapman, Karen B; Vangsness, C Thomas; West, Michael D

    2013-03-01

    The transcriptomes of seven diverse clonal human embryonic progenitor cell lines with chondrogenic potential were compared with that of bone marrow-derived mesenchymal stem cells (MSCs). The cell lines 4D20.8, 7PEND24, 7SMOO32, E15, MEL2, SK11 and SM30 were compared with MSCs using immunohistochemical methods, gene expression microarrays and quantitative real-time PCR. In the undifferentiated progenitor state, each line displayed unique combinations of site-specific markers, including AJAP1, ALDH1A2, BMP5, BARX1, HAND2, HOXB2, LHX1, LHX8, PITX1, TBX15 and ZIC2, but none of the lines expressed the MSC marker CD74. The lines showed diverse responses when differentiated in the presence of combinations of TGF-β3, BMP2, 4, 6 and 7 and GDF5, with the lines 4D20.8, SK11, SM30 and MEL2 showing osteogenic markers in some differentiation conditions. The line 7PEND24 showed evidence of regenerating articular cartilage and, in some conditions, markers of tendon differentiation. The scalability of site-specific clonal human embryonic stem cell-derived embryonic progenitor cell lines may provide novel models for the study of differentiation and methods for preparing purified and identified cells types for use in therapy.

  7. Suppressive effects of chlorophyllin on mutagen-induced umu C gene expression in Salmonella typhimurium (TA 1535/pSK 1002) and tumor promoter-dependent ornithine decarboxylase induction in BALB/c 3T3 fibroblast cells.

    PubMed

    Okai, Y; Higashi-Okai, K; Yano, Y; Otani, S

    1996-08-01

    The potentially protective role of chlorophyllin, the sodium and copper salt of chlorophyll a against the initiation and promotion stages in carcinogenesis was studied by in vitro short-term assays. Chlorophyllin showed a dose-dependent suppressive effect on 3-amino-1,4-dimethyl-5H-pyrido[4,3-b]indol (Trp-P-1)-induced umu C gene expression of Salmonella typhimurium (TA 1535/pSK 1002) in the presence of metabolizing enzyme mixture. The similar inhibitory effect of chlorophyllin was detected in mitomycin C (MMC)-dependent umu C gene expression in the absence of metabolizing enzyme mixture. Furthermore chlorophyllin also exhibited a dose-dependent inhibition on 12-O-tetradecanoyl-phorbol-13-acetate (TPA)-induced ornithine decarboxylase (ODC) activity of 3T3 fibroblast cells at the same concentrations. However, when chlorophyll a isolated from Japanese tea leaves was applied on the same assay systems as a comparative experiment, chlorophyll a showed much weaker activity compared with that of chlorophyllin. The significance of this finding is discussed from the viewpoint of the protective role of chlorophyllin against carcinogenesis.

  8. Synthesis, structure-activity relationship of novel substituted 4H-chromen-1,2,3,4-tetrahydropyrimidine-5-carboxylates as potential anti-mycobacterial and anticancer agents.

    PubMed

    Raju, B China; Rao, R Nageswara; Suman, P; Yogeeswari, P; Sriram, D; Shaik, Thokhir Basha; Kalivendi, Shasi Vardhan

    2011-05-15

    Series of 4H-chromen-1,2,3,4-tetrahydropyrimidine-5-carboxylate derivatives 7a-7zb, 8a-8d and 9a-9d were synthesized and screened for their in vitro anti-mycobacterial activity against Mycobacterium tuberculosis H(37)Rv (MTB) and cytotoxicity against three human cancer cell lines including A549, SK-N-SH and HeLa. The results indicate that six compounds are more potent and 7za is most effective anti-mycobacterial derivative compared to the standard drugs Ethambutol and Ciprofloxacin. However, 12 compounds exhibited cytotoxicity against human neuroblastoma cell line; amongst them the compound 7v is most effective compared to the standard drug Doxorubicin. This is the first report assigning in vitro anti-mycobacterial, anticancer and structure-activity relationship for this new class of 4H-chromen-1,2,3,4-tetrahydropyrimidine-5-carboxylates. Copyright © 2011 Elsevier Ltd. All rights reserved.

  9. Aging as Evolution-Facilitating Program and a Biochemical Approach to Switch It Off

    NASA Astrophysics Data System (ADS)

    Skulachev, Vladimir P.

    A concept is presented considering aging of living organisms as a final step of their ontogenetic program. It is assumed that such an aging program was invented by biological evolution to facilitate the evolutionary process. Indications are summarized suggesting that controlled production of toxic forms of oxygen (so called reactive oxygen species) by respiring intracellular organelles (mitochondria) is an obligatory component of the aging program. First results of a research project devoted to an attempt to interrupt aging program by antioxidants specifically addressed to mitochondria have been described. Within the framework of the project, antioxidants of a new type (SkQ) were synthesized. SkQs are composed of (i) plastoquinone (an antioxidant moiety), (ii) a penetrating cation, and (iii) a decane or pentane linker. Using planar bilayer phospholipid membranes, we selected SkQ derivatives of the highest penetrability, namely plastoquinonyl decyl triphenylphosphonium (SkQ1), plastoquinonyl decyl rhodamine 19 (SkQR1), and methylplastoquinonyl decyl triphenylphosphonium (SkQ3). Anti- and prooxidant properties of these substances and also of ubiquinonyl-decyl-triphenylphosphonium (MitoQ) were tested in isolated mitochondria. Micromolar concentrations of cationic quinones are found to be very strong prooxidants, but in the lower (sub-micromolar) concentrations they display antioxidant activity which decreases in the series SkQ1 = SkQR1 > SkQ3 > MitoQ. Thus, the window between the anti- and prooxidant effects is the smallest for MitoQ and the largest for SkQ1 and SkQR1. SkQ1 is rapidly reduced by complex III of the mitochondrial respiratory chain, i.e. it is a rechargeable antioxidant. Extremely low concentrations of SkQ1 and SkQR1 completely arrest the H2O2-induced apoptosis in human fibroblasts and HeLa cells (for SkQ1, C 1/2 = 8 · 10-9M). Higher concentrations of SkQ1 are required to block necrosis initiated by reactive oxygen species (ROS). In mice, SkQ1

  10. Baccharis trimera inhibits reactive oxygen species production through PKC and down-regulation p47 phox phosphorylation of NADPH oxidase in SK Hep-1 cells.

    PubMed

    de Araújo, Glaucy Rodrigues; Rabelo, Ana Carolina Silveira; Meira, Janaína Serenato; Rossoni-Júnior, Joamyr Victor; Castro-Borges, William de; Guerra-Sá, Renata; Batista, Maurício Azevedo; Silveira-Lemos, Denise da; Souza, Gustavo Henrique Bianco de; Brandão, Geraldo Célio; Chaves, Míriam Martins; Costa, Daniela Caldeira

    2017-02-01

    Baccharis trimera, popularly known as "carqueja", is a native South-American plant possessing a high concentration of polyphenolic compounds and therefore high antioxidant potential. Despite the antioxidant potential described for B. trimera, there are no reports concerning the signaling pathways involved in this process. So, the aim of the present study was to assess the influence of B. trimera on the modulation of PKC signaling pathway and to characterize the effect of the nicotinamide adenine dinucleotide phosphate oxidase enzyme (NOX) on the generation of reactive oxygen species in SK Hep-1 cells. SK-Hep 1 cells were treated with B. trimera, quercetin, or rutin and then stimulated or not with PMA/ionomycin and labeled with carboxy H 2 DCFDA for detection of reactive oxygen species by flow cytometer. The PKC expression by Western blot and enzyme activity was performed to evaluate the influence of B. trimera and quercetin on PKC signaling pathway. p47 phox and p47 phox phosphorylated expression was performed by Western blot to evaluate the influence of B. trimera on p47 phox phosphorylation. The results showed that cells stimulated with PMA/ionomycin (activators of PKC) showed significantly increased reactive oxygen species production, and this production returned to baseline levels after treatment with DPI (NOX inhibitor). Both B. trimera and quercetin modulated reactive oxygen species production through the inhibition of PKC protein expression and enzymatic activity, also with inhibition of p47 phox phosphorylation. Taken together, these results suggest that B. trimera has a potential mechanism for inhibiting reactive oxygen species production through the PKC signaling pathway and inhibition subunit p47 phox phosphorylation of nicotinamide adenine dinucleotide phosphate oxidase.

  11. Polonium 210Po activities in human blood of patients with ischaemic heart disease from Gdańsk in Poland.

    PubMed

    Boryło, Alicja; Skwarzec, Bogdan; Romańczyk, Grzegorz; Siebert, Janusz

    The determination of polonium 210 Po in human blood samples is presented and discussed in this paper. The human blood samples were collected from patients of Medical University of Gdańsk with ischaemic heart disease ( morbus ischaemicus cordis , MIC ). The polonium concentrations in analyzed human blood samples are very differentiated. 210 Po is of particular interest in public health and although is present in the environment in extremely low amounts, it is easily bioaccumulated to the human body. The study shows that the amount of 210 Po that is incorporated into the human body depends on the food habits and some difference in its levels could be observed between smokers and non-smokers.

  12. Recombinant streptokinase vs phenylephrine-based suppositories in acute hemorrhoids, randomized, controlled trial (THERESA-3)

    PubMed Central

    Hernández-Bernal, Francisco; Castellanos-Sierra, Georgina; Valenzuela-Silva, Carmen M; Catasús-Álvarez, Karem M; Valle-Cabrera, Roselin; Aguilera-Barreto, Ana; López-Saura, Pedro A

    2014-01-01

    AIM: To compare the efficacy and safety of recombinant streptokinase (rSK) and phenylephrine-based suppositories in acute hemorrhoidal disease. METHODS: A multicenter (14 sites), randomized (1:1), open, parallel groups, active controlled trial was done. After inclusion, subjects with acute symptoms of hemorrhoids, who gave their written, informed consent to participate, were centrally randomized to receive, as outpatients, rSK (200000 IU) or 0.25% phenylephrine suppositories, which had different organoleptic characteristics. Treatment was administered by the rectal route, one unit every 6 h during 48 h for rSK, and up to a maximum of 5 d (20 suppositories) for phenylephrine. Evaluations were performed at 3, 5 and 10 d post-inclusion. The main end-point was the 5th-day complete clinical response (disappearance of pain and edema, and ≥ 70% reduction of the lesion size). Time to response and need for thrombectomy were secondary efficacy variables. Adverse events were evaluated too. RESULTS: 5th day complete response rates were 83/110 (75.5%) and 36/110 (32.7%) with rSK and phenylephrine suppositories, respectively. This 42.7% difference (95%CI: 30.5-54.2) was highly significant (P < 0.001). The advantage was detected since the early 3rd day evaluation (37.3% vs 6.4% for the rSK and active control groups, respectively; P < 0.001) and was kept even at the late 10th day assessment (83.6% vs 58.2% for rSK and phenylephrine, respectively; P < 0.001). Time for complete response was significantly shorter (P = 0.031; log-rank test) in the rSK group (median: 4.9 d; 95%CI: 4.8-5.0) with respect to the active control (median: 9.8 d; 95%CI: 9.8-10.0). Thrombectomy was necessary in 1/59 and 8/57 patients with baseline thrombosis in the rSK and phenylephrine groups, respectively (P = 0.016). There were no adverse events attributable to the experimental treatment. CONCLUSION: rSK suppositories showed a significant advantage over a widely used over-the-counter phenylephrine

  13. [Criminal methods in archaeology--collaboration between Forensic Medicine Department, Pomeranian Medical University and the Archaeological Museum in Gdańsk].

    PubMed

    Kempińska-Podhorodecka, Agnieszka; Knap, Oktawian; Parafiniuk, Mirosław

    2007-01-01

    During excavation works carried in the Old Town by the Archaeological Museum in Gdańsk, human remains were found which date back to the turn of the 12th and 13th centuries. On the basis of Gdańsk townsmen's skulls, Forensic Medicine Department, Pomeranian Medical University (PAM) performed the skull based face reconstruction of 8 individuals. In this study, we wanted to present possibilities of using Gierasimow reconstruction method for museum goals. Reconstruction is an anthropological method which aims at reconstructing bony elements of a skull and head soft tissue. The most commonly employed modern way of reconstruction is Gierasimow's method which is based on the observation of soft tissue thickness and its dependence on the form and level of development of different skull areas. Standards for tissue thickness were elaborated for various points (along the profile and transverse sections); they were based on the examination of soft tissue thickness performed on the corpse (for each sex separately). Deviations from the standards result from racial affiliation, age, and the level of development of adequate skull areas. The research scheme includes determination of sex and age, and collection of the detailed craniometrical and cranioscopic data with comprehensive description of the features which can affect the appearance of soft parts. After relevant measurements are done, the muscles are modeled. During the following stage, soft tissue thickness is marked in particular points as stalks and ridges. Next they are joined together to achieve the final effect of reconstruction. From this moment, finishing works are continued by a sculptor in cooperation with an anthropologist. The results of research conducted by anthropologists, anatomists, morphologists, physicians and criminologists are of great importance and they convey both cognitive and practical meaning. Reconstructions appeal to human imagination, and for that reason they are also addressed to non

  14. STS-30 MS Cleave uses camcorder to record FEA-2 crystal growth

    NASA Image and Video Library

    1989-05-08

    STS030-10-002 (8 May 1989) --- STS-30 Mission Specialist Mary L. Cleave operates 8mm video camcorder at Fluids Experiment Apparatus 2 (FEA-2) (SK73-000102) unit located in aft middeck locker onboard Atlantis, Orbiter Vehicle (OV) 103. Two 8mm video camcorders are positioned above FEA-2 unit to record experiment titled "Floating Zone Crystal Growth and Purification". Rockwell International (RI) through its Space Transportation Systems Division, Downey, California, is engaged in a joint endeavor agreement (JEA) with NASA's Office of Commercial Programs in the field for floating zone crystal growth research. Utah State University Aggies decal appears on aft bulkhead above FEA-2 unit.

  15. Acylated iridoids from the roots of Valeriana officinalis var. latifolia.

    PubMed

    Han, Zhu-zhen; Yan, Zhao-hui; Liu, Qing-xin; Hu, Xian-qing; Ye, Ji; Li, Hui-liang; Zhang, Wei-dong

    2012-10-01

    Phytochemical investigation of the roots of Valeriana officinalis var. latifolia resulted in the isolation and characterization of six new acylated iridoids, (5S,7S,8S,9S)-7-hydroxy-8-isovaleroyloxy-Δ⁴,¹¹-dihyronepetalactone (1), (5S,7S,8S,9S)-7-hydroxy-10-isovaleroyloxy-Δ⁴,¹¹-dihyronepetalactone (2), (5S,8S,9S)-10-isovaleroyloxy-Δ⁴,¹¹-dihyronepetalactone (3), (5S,6S,8S,9R)-6-isovaleroyloxy-Δ⁴,¹¹-1,3-diol (4), (5S,6S,8S,9R)-1,3-isovaleroxy-Δ4,11-1,3-diol (5), and (5S,6S,8S,9R)-3-isovaleroxy-6-isovaleroyloxy-Δ⁴,¹¹-1,3-diol (6). Their structures were determined mainly by 1D and 2D NMR spectroscopic techniques. We also report herein for the first time the single crystal X-ray structure of compound 1. In addition, the cytotoxic activities of compounds 1-6 were evaluated against A549 (human lung adenocarcinoma), HCT116 (human colon carcinoma), SK-BR-3 (human breast carcinoma), and HepG2 (human hepatoma) cell lines. Compound 6 showed weak cell growth inhibition of A549, HCT116, SK-BR-3, and HepG2 cells. Georg Thieme Verlag KG Stuttgart · New York.

  16. Purification of a phospholipase A2 from Daboia russelii siamensis venom with anticancer effects

    PubMed Central

    Khunsap, Suchitra; Pakmanee, Narumol; Khow, Orawan; Chanhome, Lawan; Sitprija, Visith; Suntravat, Montamas; Lucena, Sara E; Perez, John C; Sánchez, Elda E

    2011-01-01

    Venom phospholipases A2 (PLA2) are associated with neurotoxic, myotoxic, cardiotoxic, platelet aggregation, and edema activities. A PLA2 (Drs-PLA2) was purified from Daboia russelii siamensis venom by a two-step purification procedure consisting of size-exclusion, followed by anion exchange high performance liquid chromatography (HPLC). The molecular weight of the Drs-PLA2 was 13,679Da, which was determined by MALDI-TOF mass spectrometry. Its N-terminal amino acid sequence was homologous to basic PLA2s of viperid snake venoms. The Drs-PLA2 had indirect hemolytic and anticoagulant activities, cytotoxic activity with a CC50 of 65.8nM, and inhibited SK-MEL-28 cell migration with an IC50 of 25.6nM. In addition, the Drs-PLA2 inhibited the colonization of B16F10 cells in lungs of BALB/c mice by ∼65%. PMID:22091349

  17. LL-37 Recruits Immunosuppressive Regulatory T Cells to Ovarian Tumors

    DTIC Science & Technology

    2009-11-01

    receptor. Western blot analysis of MSC lysates showed that ERK-1 and -2 are robustly phosphorylated beginning 10 minutes after LL-37 treatment and...Carretero, Escamez et al. 2008; von Haussen, Koczulla et al. 2008). Western blot analysis of LL-37-treated SK-OV-3 cell lysates showed the robust...mesenchymal stem cells in the treatment of gliomas ." Cancer Res 65(8): 3307-18. Studeny, M., F. C. Marini, et al. (2004). "Mesenchymal stem cells: potential

  18. Bisphenol A, 4-tert-octylphenol, and 4-nonylphenol in the Gulf of Gdańsk (Southern Baltic).

    PubMed

    Staniszewska, Marta; Falkowska, Lucyna; Grabowski, Paweł; Kwaśniak, Justyna; Mudrak-Cegiołka, Stella; Reindl, Andrzej R; Sokołowski, Adam; Szumiło, Emilia; Zgrundo, Aleksandra

    2014-10-01

    The organic derivatives of phenol are classed as dangerous compounds, and their presence has been detected in surface water, bottom water, phytoplankton, zooplankton, and mussel as well as liver and muscle of fish from the Gulf of Gdańsk and in liver, muscle, and guano of gulls residing in the coastal zone of this basin. The greatest sources of bisphenol A (BPA), 4-tert-octylphenol (OP), and 4-nonylphenol (NP) were found to be the Vistula River and the water purification plant in Dębogórze. In living organisms, concentrations of BPA, OP, and NP ranged between the limit of quantification and several hundred ng g(-1) dry weight (dw), and the highest concentrations were found for BPA. Prolonged alimentary exposure to BPA, OP, and NP in fish and birds was indicated by liver/muscle concentration ratios generally >1. The most influential factors on BPA and alkylphenol concentrations in the tissues of fishes and gulls were mainly diet and habitat. The study confirmed possible bioaccumulation in the food web. High BPA and NP concentrations in guano (≤2,700 and ≤300 ng g(-1) dw, respectively) indicated the ability of birds to detoxify and signalled the reintroduction of these compounds to seawater. Herring, flounder, and cod from the Southern Baltic are a safe food source for human consumption because their BPA and alkylphenol contents are low.

  19. Late Triassic (Carnian) lamproites from Noril'sk, polar Siberia: Evidence for melting of the recycled Archean crust and the question of lamproite source for some placer diamond deposits of the Siberian Craton

    NASA Astrophysics Data System (ADS)

    Ivanov, Alexei V.; Demonterova, Elena I.; Savatenkov, Valery M.; Perepelov, Alexander B.; Ryabov, Viktor V.; Shevko, Artem Y.

    2018-01-01

    Two typical lamproitic dykes were found in Noril'sk region of the north-western Siberian Craton, which according to mineralogical, geochemical and isotopic criteria belong to anorogenic, non-diamondiferous type of lamproites. According to the geologic relationships, they cut through the Noril'sk-1 intrusion of the Siberian flood basalt province and thus are younger than 251 Ma. 40Ar/39Ar dating of the two dykes yielded ages of 235.24 ± 0.19 Ma and 233.96 ± 0.19 Ma, showing that they were emplaced in Carnian of the Late Triassic, about 16 Ma after the flood basalt event. There are some indications that there were multiple lamproitic dyke emplacements, including probably emplacement of diamondiferous lamproites, which produced Carnian-age diamond-rich placer deposits in other parts of the Siberian Craton and in adjacent regions. Lead isotope modelling shows that the source of the studied lamproites was formed with participation of recycled crust, which underwent modification of its U/Pb ratio as early as 2.5 Ga. However, the exact mechanism of the recycling cannot be deciphered now. It could be either through delamination of the cratonic crust or subduction of a mix of ancient terrigenous sediments into the mantle transition zone.

  20. In vitro thrombolytic activity of purified streptokinase extracted from Streptococcus equinus VIT_VB2 isolated from bovine milk.

    PubMed

    Babu, Vaishnavi; Subathra Devi, C

    2015-01-01

    Streptokinase (SK) is an extracellular enzyme secreted by various strains of β-hemolytic Streptococci. The main focus of the current study is to evaluate the in vitro thrombolytic activity of purified SK extracted from Streptococcus equinus VIT_VB2 (Accession no. JX406835) isolated from milk sample. The growth rate of S. equinus VIT_VB2 strain was studied with pH and biomass content which has positive significant effect on enzyme yield. A temperature of 10 °C and pH of 6 was found to be optimum for maximum SK activity. The specific activity of the purified SK produced by VIT_VB2 strain was found to be 6,585 IU mg(-1). The molecular mass of the enzyme was determined as 47 kDa by SDS-PAGE. In vitro thrombolytic activity of purified SK was determined using synthetic chromogenic substrate S-2251, the activity of the purified enzyme was found to be 6,330 ± 2.2 IU. The purity of SK was compared with standard SK by HPLC. This is the first report which reveals the SK activity of S. equinus isolated from milk sample.

  1. Determination of perfluorinated alkylated substances in sediments and sediment core from the Gulf of Gdańsk, Baltic Sea.

    PubMed

    Falandysz, Jerzy; Rostkowski, Paweł; Jarzyńska, Grażyna; Falandysz, Jaromir J; Taniyasu, Sachi; Yamashita, Nobuyoshi

    2012-01-01

    Perfluorinated alkylated substances (PFAS) have been determined in surface sediments and sediment core from Gulf of Gdańsk, Baltic Sea. Perfluorooctanesulphonate (PFOS), perfluorohexanesulphonate (PFHxS), perfluorodecanoate (PFDA), perfluoronanoate (PFNA), perfluorooctanoate (PFOA), perfluoroheptanoate (PFHpA), perfluoroundecanote (PFUnDA), perfluorododecanoate (PFDoDA) and perfluorohexanoate (PFHxA) were quantified after isotopic dilution ((13)C(4) PFOS and (13)C(4) PFOA), liquid-liquid extractions by methanol and acetonitrile, cleanup by Envi-Carb, OasisWAX and Envi-Carb and final measurement by HPLC-MS/MS. PFOS, PFHxS, PFUnDA, PFDA, PFNA and PFOA were found in Baltic Sea sediments in concentrations exceeding the method limit of quantification (LOQ) of 2 pg/g. PFOS was detected in concentration up to 0.896 ng/g dry weight and PFHxS up to 0.326 ng/g dw, which shows on a weak pollution. PFOS (48-74%) or PFHxS (45-56%) dominated in PFAS composition of sediments surveyed. Supplemental materials are available for this article. Go to the publisher's online edition of the Journal of Environmental Science and Health, Part A to view the free supplemental file.

  2. Novel therapeutic mechanisms determine the effectiveness of lipid-core nanocapsules on melanoma models

    PubMed Central

    Drewes, Carine C; Fiel, Luana A; Bexiga, Celina G; Asbahr, Ana Carolina C; Uchiyama, Mayara K; Cogliati, Bruno; Araki, Koiti; Guterres, Sílvia S; Pohlmann, Adriana R; Farsky, Sandra P

    2016-01-01

    Melanoma is a severe metastatic skin cancer with poor prognosis and no effective treatment. Therefore, novel therapeutic approaches using nanotechnology have been proposed to improve therapeutic effectiveness. Lipid-core nanocapsules (LNCs), prepared with poly(ε-caprolactone), capric/caprylic triglyceride, and sorbitan monostearate and stabilized by polysorbate 80, are efficient as drug delivery systems. Here, we investigated the effects of acetyleugenol-loaded LNC (AcE-LNC) on human SK-Mel-28 melanoma cells and its therapeutic efficacies on melanoma induced by B16F10 in C57B6 mice. LNC and AcE-LNC had z-average diameters and zeta potential close to 210 nm and -10.0 mV, respectively. CytoViva® microscopy images showed that LNC and AcE-LNC penetrated into SK-Mel-28 cells, and remained in the cytoplasm. AcE-LNC in vitro treatment (18–90×109 particles/mL; 1 hour) induced late apoptosis and necrosis; LNC and AcE-LNC (3–18×109 particles/mL; 48 hours) treatments reduced cell proliferation and delayed the cell cycle. Elevated levels of nitric oxide were found in supernatant of LNC and AcE-LNC, which were not dependent on nitric oxide synthase expressions. Daily intraperitoneal or oral treatment (days 3–10 after tumor injection) with LNC or AcE-LNC (1×1012 particles/day), but not with AcE (50 mg/kg/day, same dose as AcE-LNC), reduced the volume of the tumor; nevertheless, intraperitoneal treatment caused toxicity. Oral LNC treatment was more efficient than AcE-LNC treatment. Moreover, oral treatment with nonencapsulated capric/caprylic triglyceride did not inhibit tumor development, implying that nanocapsule supramolecular structure is important to the therapeutic effects. Together, data herein presented highlight the relevance of the supramolecular structure of LNCs to toxicity on SK-Mel-28 cells and to the therapeutic efficacy on melanoma development in mice, conferring novel therapeutic mechanisms to LNC further than a drug delivery system. PMID:27099491

  3. [Sexual behaviors among students of the Medical University in Gdańsk].

    PubMed

    Grabowski, Karol; Wichowicz, Hubert M; Cubała, Wiesław Jerzy

    2006-01-01

    The aim of this study was to illustrate sexual behaviours, needs and expectations as well as attitudes towards contraception among students of the Faculty of Medicine, Medical University of Gdańsk. A group of 340 medical students aged 18 to 28 (195 females and 118 males) was examined with the questionnaire from October till December 2003. The examined group was representative for the investigated population (chi squared test: p> 0.005 for the entire group and both sexes). 313 questionnaires were qualified for further analysis. The questionnaire applied was composed of three groups of questions: questions concerning relationships and associated sexuality, questions concerning attitudes towards various aspects of sexuality and a set of questions dedicated to the role of sexology in the medical curriculum and general medical practice. The presented paper deals with the forms of sexual behaviours and attitudes towards contraception. The examined group is well educated in theoretical and practical aspects of sex-related phenomena. The efficient contraception methods are widely used in the studied population. A preferred form of contraception in the examined group is the use of condoms followed by oral contraceptives. The sexual initiation was mainly under the influence an emotional relationship. The examined individuals indicate love as the main factor positively influencing the quality of sexual life, followed by partner's loyalty and sexual satisfaction. The examined population has a positive attitude towards alternative forms of sexual practices. The students perceive sex as an important aspect of their functioning and are satisfied with their sexual partners.

  4. Neuroprotective effect of CNB-001, a novel pyrazole derivative of curcumin on biochemical and apoptotic markers against rotenone-induced SK-N-SH cellular model of Parkinson's disease.

    PubMed

    Jayaraj, Richard L; Tamilselvam, Kuppusamy; Manivasagam, Thamilarasan; Elangovan, Namasivayam

    2013-11-01

    Oxidative stress and mitochondrial dysfunction are underpinned for initiating a cascade of toxic events leading to dopaminergic neuronal death in Parkinson's disease (PD) and identified as vital target for therapeutic intervention. Curcumin, a potent antioxidant has been reported to display diverse neuroprotective properties against various neurodegenerative diseases including PD. In this present study, we investigated the protective effect of CNB-001, a pyrazole derivative of curcumin on rotenone-induced toxicity and its possible mechanisms in neuroblastoma SK-N-SH cells. Rotenone insult significantly reduced cell viability (MTT assay) and resulted in 78 % apoptosis (dual staining) by altering Bcl-2, Bax, caspase-3, and cytochrome C expression. Moreover, rotenone enhanced ROS production and disrupts mitochondrial membrane potential. These resultant phenotypes were distinctly alleviated by CNB-001. Pretreatment with CNB-001(2 μM) 2 h before rotenone exposure (100 nM) increased cell viability, decreased ROS formation, maintained normal physiological mitochondrial membrane potential, and reduced apoptosis. Furthermore, CNB-001 inhibited downstream apoptotic cascade by increasing the expression of vital antiapoptotic protein Bcl-2 and decreased the expression of Bax, caspase-3, and cytochrome C. Collectively, the results suggest that CNB-001 protects neuronal cell against toxicity through antioxidant and antiapoptotic properties through its action on mitochondria. Therefore, it may be concluded that CNB-001 can be further developed as a promising drug for treatment of PD.

  5. Induction of neutral endopeptidase (NEP) activity of SK-N-SH cells by natural compounds from green tea.

    PubMed

    Ayoub, Shereen; Melzig, Matthias F

    2006-04-01

    Deposition of amyloid beta-peptide as senile plaques in the brain is one of the neuropathological hallmarks of Alzheimer's disease, which is the most prevalent progressive neurodegenerative disease leading to dementia. Neutral endopeptidase is one of the major beta-amyloid-degrading enzymes in the brain. To examine the influence of different polyphenols and other natural products from green tea extract (from Camellia sinensis, Theaceae), we used the neuroblastoma cell line SK-N-SH and studied the changes in the specific cellular neutral endopeptidase activity after long-term treatment with these substances. We have shown that caffeine leads to an increase in specific cellular neutral endopeptidase activity more than theophylline, theobromine or theanine. We have also shown that the combination of epicatechin, epigallocatechin and epigallocatechingallate with caffeine, theobromine or theophylline induced cellular neutral endopeptidase activity. It is suggested that the enhancement of cellular neutral endopeptidase activity by green tea extract and its natural products might be correlated with an elevated level of intracellular cyclic adenosine monophosphate.

  6. Differential involvement of dopamine D-1 and D-2 receptors in the circling behaviour induced by apomorphine, SK & F 38393, pergolide and LY 171555 in 6-hydroxydopamine-lesioned rats.

    PubMed

    Arnt, J; Hyttel, J

    1985-01-01

    The antagonistic effect of dopamine (DA) D-1 and D-2 antagonists against circling behaviour induced by various DA agonists in 6-OHDA-lesioned rats has been investigated. DA D-1/D-2 selectivity of agonists in vitro was measured by the stimulatory effect on DA-sensitive adenylate cyclase in rat striatal homogenates (D-1), the inhibitory effect on electrically-induced release of 3H-DA in rabbit striatal slices (D-2) and the affinity to 3H-piflutixol (D-1) and 3H-spiroperidol (D-2) binding sites in rat striatal membranes. The contralateral circling behaviour induced by the DA D-1 agonist SK & F 38393 was blocked by the DA D-1 antagonist, SCH 23390, and by the mixed DA D-1/D-2 antagonist cis(Z)-flupentixol, but was not influenced by the DA D-2 antagonists spiroperidol and clebopride. In contrast, circling behaviour induced by the preferential DA D-2 agonists pergolide and LY 171555 was blocked by clebopride, spiroperidol, and cis(Z)-flupentixol, but weakly or not influenced by SCH 23390. Apomorphine-induced circling behaviour was blocked by cis(Z)-flupentixol, partially antagonized by SCH 23390 and clebopride but not inhibited by spiroperidol, although the time-course of circling was changed. Combinations of SCH 23390 with spiroperidol or clebopride in low doses completely blocked the effect of apomorphine. These results indicate that DA D-1 and D-2 receptors mediate circling behaviour through separate mechanisms which can be independently manipulated with respective agonists and antagonists.(ABSTRACT TRUNCATED AT 250 WORDS)

  7. Humoral anti-OV-TL 3 response after the intravenous administration of radiolabelled Fab' or F(ab')2 fragments in ovarian cancer patients.

    PubMed

    Tibben, J G; Thomas, C M; Massuger, L F; Segers, M F; Schijf, C P; Corstens, F H; Boerman, O C

    1995-10-01

    The human anti-mouse antibody (HAMA) response was determined in the serum of patients suspected of having ovarian cancer who underwent radioimmunoscintigraphy with either 99Tcm-OV-TL 3 Fab' (n = 20) or 111In-DTPA-OV-TL 3 F(ab')2 (n = 73). Blood samples were collected prior to and at several time points post-intravenous injection. The detection of HAMA was performed with an in-house OV-TL 3 F(ab')2-based sandwich-type immunoradiometric assay (IRMA). The homologous IRMA demonstrated that 8 of 20 (40%) patients had developed HAMA responses after injection of Fab' fragments and that 14 of 73 (19%) patients had developed HAMA responses after F(ab')2 administration. The subclass of the measured HAMA was analysed in a limited number of samples, showing IgG or IgM as well as mixed responses. The kinetics of the HAMA responses varied greatly. Our study showed the relevance of the sampling time and frequency: HAMA responses can be easily underestimated with a low sampling frequency. The homologous IRMA described in this study was able to quantify the OV-TL 3-specific HAMA responses. With additional assays, the subclass of the HAMA could be further analysed. Remarkably, the fraction of HAMA responders after injection of OV-TL 3 Fab' fragments was in the same range as the proportion of HAMA responders after F(ab')2 administration.

  8. Long non-coding RNA NNT-AS1 contributes to cell proliferation, metastasis and apoptosis in human ovarian cancer.

    PubMed

    Huang, Yaqing; Shi, Junyu; Xu, Yun

    2018-06-01

    Ovarian cancer is a markedly heterogeneous malignancy characterized by various histological subtypes. Molecular biomarkers have been indicated to serve significant functions in the early diagnosis and treatment of early-stage ovarian cancer. However, the detailed mechanism underlying the tumorigenesis of ovarian cancer remains unclear. The present study aimed to identify a novel long non-coding RNA in patients with ovarian cancer. Nicotinamide nucleotide transhydrogenase-antisense 1 (NNT-AS1) was markedly downregulated in patients with ovarian cancer and in cultured human ovarian cancer cells. Knockdown of NNT-AS1 in the human ovarian cancer cell lines HO-8910 and SK-OV-3 promoted colony formation and arrested the cell cycle at G 0 /G 1 phase. Furthermore, Transwell demonstrated that the downregulation of NNT-AS1 increased cell migration and invasion by ~60 and 70%, respectively, in HO-8910 and SK-OV-3 cells. Furthermore, cell apoptosis was inhibited by the transfection of siNNT-AS1 in the two cell lines, whereas the relative activities of caspase-3 and caspase-9 were decreased. These results indicated a protective function of NNT-AS1 in human ovarian cancer, providing novel insights into the diagnosis and treatment of ovarian cancer in clinical settings.

  9. In vivo targeting of OV-TL 3 immunoliposomes to ascitic ovarian carcinoma cells (OVCAR-3) in athymic nude mice.

    PubMed

    Nässander, U K; Steerenberg, P A; Poppe, H; Storm, G; Poels, L G; De Jong, W H; Crommelin, D J

    1992-02-01

    Specific binding of immunoliposomes to target tumor cells was investigated in a xenograft model (athymic nude mice) of i.p. growing human ovarian carcinoma (OVCAR-3). For the first time, quantitative evidence is presented that attachment of a tumor-specific antibody (OV-TL 3) dramatically enhances the association of liposomes with i.p. growing OVCAR-3 cells. The OV-TL 3-mediated binding of liposomes to the OVCAR-3 cells was rapid; 30 min after i.p. injection approximately 70% of the injected dose of OV-TL 3 immunoliposomes was associated with the OVCAR-3 cells while for unconjugated liposomes a value of only approximately 3% was obtained. At 2 h after injection, a maximal binding level of 84% was achieved in case of the OV-TL 3 immunoliposomes whereas the binding level of unconjugated liposomes was still about 3%. Twenty-four h after injection about 83% of the injected dose OV-TL 3 immunoliposomes still was associated with the OVCAR-3 cells, compared to about 10% of the injected dose of unconjugated liposomes. Accordingly, unconjugated liposomes disappeared from the peritoneal cavity much faster than the OV-TL 3 immunoliposomes. By comparison with immunoliposomes bearing irrelevant antibody, the specificity of the binding of the OV-TL 3 immunoliposomes to the OVCAR-3 cells was demonstrated. In addition, it was observed that the sustained high OV-TL 3 immunoliposome levels found in the peritoneal cavity are the result of both reduced particle clearance from the peritoneal cavity and the tenacious binding of the immunoliposomes to the tumor cells. Finally, data are presented showing that the degree of binding of OV-TL 3 immunoliposomes to OVCAR-3 cells in vitro and in vivo correlates positively with the antibody (Fab') density on the liposomes.

  10. Reduced Innate Immune Response to a Staphylococcus aureus Small Colony Variant Compared to Its Wild-Type Parent Strain

    PubMed Central

    Ou, Judy J. J.; Drilling, Amanda J.; Cooksley, Clare; Bassiouni, Ahmed; Kidd, Stephen P.; Psaltis, Alkis J.; Wormald, Peter J.; Vreugde, Sarah

    2016-01-01

    Background: Staphylococcus aureus (S. aureus) small colony variants (SCVs) can survive within the host intracellular milieu and are associated with chronic relapsing infections. However, it is unknown whether host invasion rates and immune responses differ between SCVs and their wild-type counterparts. This study used a stable S. aureus SCV (WCH-SK2SCV) developed from a clinical isolate (WCH-SK2WT) in inflammation-relevant conditions. Intracellular infection rates as well as host immune responses to WCH-SK2WT and WCH-SK2SCV infections were investigated. Method: NuLi-1 cells were infected with either WCH-SK2WT or WCH-SK2SCV, and the intracellular infection rate was determined over time. mRNA expression of cells infected with each strain intra- and extra-cellularly was analyzed using a microfluidic qPCR array to generate an expression profile of thirty-nine genes involved in the host immune response. Results: No difference was found in the intracellular infection rate between WCH-SK2WT and WCH-SK2SCV. Whereas, extracellular infection induced a robust pro-inflammatory response, intracellular infection elicited a modest response. Intracellular WCH-SK2WT infection induced mRNA expression of TLR2, pro-inflammatory cytokines (IL1B, IL6, and IL12) and tissue remodeling factors (MMP9). In contrast, intracellular WCH-SK2SCV infection induced up regulation of only TLR2. Conclusions: Whereas, host intracellular infection rates of WCH-SK2SCV and WCH-SK2WT were similar, WCH-SK2SCV intracellular infection induced a less widespread up regulation of pro-inflammatory and tissue remodeling factors in comparison to intracellular WCH-SK2WT infection. These findings support the current view that SCVs are able to evade host immune detection to allow their own survival. PMID:28083514

  11. Joint Fire Support in 2020: Development of a Future Joint Fires Systems Architecture for Immediate, Unplanned Targets

    DTIC Science & Technology

    2006-12-01

    Undersea Warfare Commander VHF Very High Frequency VoIP Voice over Internet Protocol WCS Weapons...USN surface and undersea warfare, USN and USA communications and networking operations, and USAF acquisition and aircraft maintenance projects. In...kin g 1 x x BN -R EG T- BD E Co or di na tio n 1 x x x x Pr ov id in g Ba tte ry Ta sk in g, E ng ag e, C oo rd in at io n 1 1 x x x Pr ov id in

  12. Enhancing effect of a plastic plasticizer, di-2-ethylhexyl phthalate on umu C gene expression in Salmonella typhimurium (TA 1535/pSK 1002).

    PubMed

    Okai, Y; Higashi-Okai, K

    2000-12-01

    Di-2-ethylhexyl phthalate (DEHP) is the most extensively used phthalate ester as a plasticizer for plastic products made of polyvinyl chloride (PVC), and previous mutagenic and genotoxic studies have shown positive and negative results of DEHP-induced genotoxicity. To elucidate this discrepancy, we reestimated the genotoxicity of DEHP in more detail using the umu C gene expression system in Salmonella typhimurium (TA 1535/pSK 1002) which reflects SOS response against genotoxin-induced DNA damage. Although DEHP itself did not have a significant effect on umu C gene expression in tester bacteria at 0.5 to 4 mM, higher concentrations of DEHP (2 and 4 mM) caused a weak induction of umu C gene expression in the presence of commercially available S-9 mixture. Rat liver S-9 fraction alone also showed a similar weak inducing activity in the absence of substrates for drug-metabolizing enzymes. When DEHP was preincubated with S-9 fraction of various rat organs and applied to the umu C gene expression assay, S-9 fraction of rat pancreas had the strongest inducing activity, and S-9 fractions of liver and intestine homogenates showed weak but significant activities. However, S-9 fractions of lung and kidney homogenates did not exhibit any significant activities. These S-9 fractions have proportional lipase activities comparable with umu C gene expression activities. Furthermore, when DEHP was treated with highly purified lipase from porcine pancreas, a significant umu C gene expression was observed and this expression was enhanced in the presence of 1 or 5 mM bile acids such as choric acid and deoxychoric acid. These results suggest that DEHP itself has no or very low genotoxicity, but enzymatic and non-enzymatic factors in specific tissues induce DEHP-dependent genotoxic activity.

  13. Osthole induces G2/M arrest and apoptosis in lung cancer A549 cells by modulating PI3K/Akt pathway

    PubMed Central

    2011-01-01

    Background To explore the effects of Osthole on the proliferation, cell cycle and apoptosis of human lung cancer A549 cells. Methods Human lung cancer A549 cells were treated with Osthole at different concentrations. Cell proliferation was measured using the MTT assay. Cell cycle was evaluated using DNA flow cytometry analysis. Induction of apoptosis was determined by flow cytometry and fluorescent microscopy. The expressions of Cyclin B1, p-Cdc2, Bcl-2, Bax, t-Akt and p-Akt were evaluated by Western blotting. Results Osthole inhibited the growth of human lung cancer A549 cells by inducing G2/M arrest and apoptosis. Western blotting demonstrated that Osthole down-regulated the expressions of Cyclin B1, p-Cdc2 and Bcl-2 and up-regulated the expressions of Bax in A549 cells. Inhibition of PI3K/Akt signaling pathway was also observed after treating A549 cells with Osthole. Conclusions Our findings suggest that Osthole may have a therapeutic application in the treatment of human lung cancer. PMID:21447176

  14. Immunotherapy in a human ovarian cancer xenograft model with two bispecific monoclonal antibodies: OV-TL 3/CD3 and OC/TR.

    PubMed

    van Ravenswaay Claasen, H H; Eggermont, A M; Nooyen, Y A; Warnaar, S O; Fieuren, G J

    1994-02-01

    The bispecific antibodies (bs-mAbs) OV-TL 3/CD3 and OC/TR (MOv18/CD3) efficiently mediate ovarian tumor cell lysis by cytotoxic T cells and activated peripheral blood lymphocytes (PBL) in vitro. OV-TL 3/CD3 and OC/TR are reactive with tumor-associated antigens on ovarian carcinoma cells (OA3 and CA-MOv18, respectively), and CD3 on activated PBL, bridging both cells and simultaneously inducing activation of the effector cells. In a comparative study we investigated the therapeutic efficacy of OV-TL 3/CD3 and OC/TR by targeting activated PBL with the bs-mAbs against intraperitoneally growing NIH:OVCAR-3 human ovarian carcinoma cells. As they have good tumor localization characteristics, HPLC-purified bispecific F(ab')2 fragments were used to target highly active PHA and IL-2-stimulated PBL effector cells. The efficacy of OV-TL 3/CD3 was compared to OC/TR with respect to tumor-associated antigen (TAA) binding on NIH:OVCAR-3 ascites cells and NIH:OVCAR-3 tumor cell lysis in vitro. In this report we show that ip ovarian cancer-bearing nude mice treated with IL-2 and activated PBL coated with bispecific F(ab')2 had a significantly longer survival than the untreated mice. No significant difference in survival was found between the OC/TR or OV-TL 3/CD3 bispecific antibody, although MOv18 expression was higher on NIH:OVCAR-3 ascites cells and PBL targeted with OC/TR induced slightly higher tumor cell lysis in vitro. Thus, the therapeutic efficacy of these bs-mAbs in vivo could not be predicted by TAA expression or bs-mAb-mediated tumor cell lysis in vitro.

  15. Diagnostic value of CD10 and Bcl2 expression in distinguishing cutaneous basal cell carcinoma from squamous cell carcinoma and seborrheic keratosis.

    PubMed

    Gaballah, Mohammad A; Ahmed, Rehab-Allah

    2015-12-01

    The distinction between cutaneous basal cell carcinoma (BCC), squamous cell carcinoma (SCC) and seborrheic keratosis (SK), which are common entities in clinical practice, can be difficult clinically and histologically. CD10 and Bcl2 antigens are important factors in tumor growth, survival and spread. The aim of the present study is to define the frequency of CD10 and Bcl2 expression in such cutaneous tumors and its relation to the clinicopathological characteristics as well as their possible diagnostic utility. CD10 and Bcl2 immunohistochemistry was performed on 30 BCC, 20 SCC and 15 SK. 93.3% of SK cases and 53.3% of BCC cases showed significant expression of CD10 in tumor cells when compared either with each other or with SCC cases (100% negative). Stromal CD10 expression was positive in 50% of BCC cases and 75% of SCC cases. Stromal CD10 expression was significantly higher in high risk BCC and BCC with infiltrating deep margins; furthermore, it showed a significant positive correlation with grade of SCC. A significant inverse correlation between CD10 expression in stromal and tumor cells of BCC was present. Bcl2 was significantly expressed in 93.3% of SK cases and 80% of BCC cases when compared with SCC cases (100% negative). It was found that for distinguishing BCC from SK, only CD10 expression in tumor cells provided a high diagnostic value with positive likelihood ratio (PLR) was 7.00. In addition, CD10 and Bcl2 expression in tumor cells could give convincing diagnostic value to distinguish SCC from SK (PLR=15.00 for each marker). Moreover, for differentiating BCC from SCC, only Bcl2 in the tumor cells could provide a high diagnostic value (PLR=5.5). In conclusion, CD10 and Bcl2 can help in differentiating cutaneous BCC from SK and SCC. The overexpression of CD10 in the stromal cells of SCC and some variants of BCC suggests the invasive properties of such tumors. Copyright © 2015 Elsevier GmbH. All rights reserved.

  16. Humic Acid Increases Amyloid β-Induced Cytotoxicity by Induction of ER Stress in Human SK-N-MC Neuronal Cells

    PubMed Central

    Li, Hsin-Hua; Lu, Fung-Jou; Hung, Hui-Chih; Liu, Guang-Yaw; Lai, Te-Jen; Lin, Chih-Li

    2015-01-01

    Humic acid (HA) is a possible etiological factor associated with for several vascular diseases. It is known that vascular risk factors can directly increase the susceptibility to Alzheimer’s disease (AD), which is a neurodegenerative disorder due to accumulation of amyloid β (Aβ) peptide in the brain. However, the role that HA contributes to Aβ-induced cytotoxicity has not been demonstrated. In the present study, we demonstrate that HA exhibits a synergistic effect enhancing Aβ-induced cytotoxicity in cultured human SK-N-MC neuronal cells. Furthermore, this deterioration was mediated through the activation of endoplasmic reticulum (ER) stress by stimulating PERK and eIF2α phosphorylation. We also observed HA and Aβ-induced cytotoxicity is associated with mitochondrial dysfunction caused by down-regulation of the Sirt1/PGC1α pathway, while in contrast, treating the cells with the ER stress inhibitor Salubrinal, or over-expression of Sirt1 significantly reduced loss of cell viability by HA and Aβ. Our findings suggest a new mechanism by which HA can deteriorate Aβ-induced cytotoxicity through modulation of ER stress, which may provide significant insights into the pathogenesis of AD co-occurring with vascular injury. PMID:25961951

  17. All-trans beta-carotene enhances mitogenic responses and ornithine decarboxylase activity of BALB/c 3T3 fibroblast cells induced by tumor promoter and fetal bovine serum but suppresses mutagen-dependent umu C gene expression in Salmonella typhimurium (TA 1535/pSK 1002).

    PubMed

    Okai, Y; Higashi-Okai, K; Yano, Y; Otani, S

    1996-01-19

    Although previous epidemiological studies have indicated that beta-carotene is an important agent for the chemical prevention against carcinogenesis, a recent prospective study has strikingly suggested that supplementation with beta-carotene significantly increased the incidence of some types of cancer (The alpha-Tocopherol and beta-Carotene Cancer Prevention Study Group, New Engl. J. Med., 330 (1994) 1031-1035). To analyze the discrepancy of this problem, the authors analyze the effects of beta-carotene on biochemical and biological events associated with carcinogenesis by in vitro experiments. (1) All-trans beta-carotene enhanced the proliferation and DNA synthesis of BALB/c 3T3 cells induced by a tumor promoter, 12-O-tetradecanoyl-phorbol-13-acetate (TPA) and fetal bovine serum, although beta-carotene itself did not show mitogenic activity. (2) All-trans beta-carotene caused a remarkable stimulation for the early induction of ornithine decarboxylase (ODC) activity after the stimulation of TPA and fetal bovine serum. (3) All-trans beta-carotene exhibited significant antimutagenic activity which suppresses umu C gene expression in Salmonella typhimurium (TA 1535/pSK 1002) induced by a typical mutagen, 2-aminoanthracene (2-AA). These experimental results suggest that all-trans beta-carotene might cause beneficial and harmful effects on different phases of carcinogenesis.

  18. Transfection Studies with Colloidal Systems Containing Highly Purified Bipolar Tetraether Lipids from Sulfolobus acidocaldarius.

    PubMed

    Engelhardt, Konrad H; Pinnapireddy, Shashank Reddy; Baghdan, Elias; Jedelská, Jarmila; Bakowsky, Udo

    2017-01-01

    Lipid vectors are commonly used to facilitate the transfer of nucleic acids into mammalian cells. In this study, two fractions of tetraether lipids from the archaea Sulfolobus acidocaldarius were extracted and purified using different methods. The purified lipid fractions polar lipid fraction E (PLFE) and hydrolysed glycerol-dialkyl-nonitol tetraether (hGDNT) differ in their structures, charge, size, and miscibility from conventional lipids. Liposomes were prepared by mixing tetraether lipids with cholesterol (CH) and 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP) resulting in stable vectors for gene delivery. Lipoplexes were prepared by complexation of liposomes with a luciferase expressing plasmid (pCMV-luc) at certain nitrogen-to-phosphorus (N/P) ratios and optimised for the transient transfection of ovarian adenocarcinoma cells (SK-OV-3). Complexation efficacy was investigated by gel-red fluorescence assay. Biophysical properties, like size, surface charge, and morphology, were investigated by differential light scattering (DLS), atomic force microscopy (AFM), and scanning electron microscopy (Cryo-SEM), respectively, revealing structural differences between liposomes and lipoplexes. A range of stable transfecting agents containing tetraether lipids were obtained by incorporating 5 mol% of tetraether lipids. Lipoplexes showed a decrease in free gel-red with increasing N/P ratios indicating efficient incorporation of plasmid DNA (pDNA) and remarkable stability. Transfection experiments of the lipoplexes revealed successful and superior transfection of SK-OV-3 cell line compared to the commercially available DOTAP and branched polyethyleneimine (25 kDa bPEI).

  19. Transfection Studies with Colloidal Systems Containing Highly Purified Bipolar Tetraether Lipids from Sulfolobus acidocaldarius

    PubMed Central

    Pinnapireddy, Shashank Reddy; Baghdan, Elias; Jedelská, Jarmila

    2017-01-01

    Lipid vectors are commonly used to facilitate the transfer of nucleic acids into mammalian cells. In this study, two fractions of tetraether lipids from the archaea Sulfolobus acidocaldarius were extracted and purified using different methods. The purified lipid fractions polar lipid fraction E (PLFE) and hydrolysed glycerol-dialkyl-nonitol tetraether (hGDNT) differ in their structures, charge, size, and miscibility from conventional lipids. Liposomes were prepared by mixing tetraether lipids with cholesterol (CH) and 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP) resulting in stable vectors for gene delivery. Lipoplexes were prepared by complexation of liposomes with a luciferase expressing plasmid (pCMV-luc) at certain nitrogen-to-phosphorus (N/P) ratios and optimised for the transient transfection of ovarian adenocarcinoma cells (SK-OV-3). Complexation efficacy was investigated by gel-red fluorescence assay. Biophysical properties, like size, surface charge, and morphology, were investigated by differential light scattering (DLS), atomic force microscopy (AFM), and scanning electron microscopy (Cryo-SEM), respectively, revealing structural differences between liposomes and lipoplexes. A range of stable transfecting agents containing tetraether lipids were obtained by incorporating 5 mol% of tetraether lipids. Lipoplexes showed a decrease in free gel-red with increasing N/P ratios indicating efficient incorporation of plasmid DNA (pDNA) and remarkable stability. Transfection experiments of the lipoplexes revealed successful and superior transfection of SK-OV-3 cell line compared to the commercially available DOTAP and branched polyethyleneimine (25 kDa bPEI). PMID:28239294

  20. 29 CFR 549.3 - Distinction between plan and trust.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 29 Labor 3 2010-07-01 2010-07-01 false Distinction between plan and trust. 549.3 Section 549.3... REQUIREMENTS OF A âBONA FIDE PROFIT-SHARING PLAN OR TRUSTâ § 549.3 Distinction between plan and trust. As used... profits; (b) Profit-sharing trust means any such program or arrangement as qualifies under this part which...

  1. Low-concentration vemurafenib induces the proliferation and invasion of human HaCaT keratinocytes through mitogen-activated protein kinase pathway activation.

    PubMed

    Roh, Mi Ryung; Kim, Jung Min; Lee, Sang Hee; Jang, Hong Sun; Park, Kyu Hyun; Chung, Kee Yang; Rha, Sun Young

    2015-09-01

    Cutaneous squamous cell carcinomas and keratoacanthomas commonly occur in patients treated with BRAF inhibitors. We investigated the effect of the BRAF inhibitor vemurafenib on normal immortalized human HaCaT keratinocytes to explore the mechanism of hyperproliferative cutaneous neoplasia associated with the use of BRAF inhibitors. Vemurafenib induced an increase in viable cell number in BRAF wild-type cell lines (SK-MEL-2 and HaCaT) but not in BRAF mutant cell lines (SK-MEL-24 and G361). In HaCaT keratinocytes, a low concentration (2 μmol/L) of vemurafenib increased cell proliferation and activated mitogen-activated protein kinase kinase/extracellular signal-regulated kinase in a CRAF-dependent manner. Invasiveness of HaCaT cells in a Matrigel assay significantly increased upon cultivation of cells with 2 μmol/L vemurafenib for 24 h. Gelatin zymography, reverse transcription polymerase chain reaction and western blot results revealed that 2 μmol/L vemurafenib treatment increased matrix metalloproteinase (MMP)-2 and MMP-9 expressions and activities in HaCaT cells. These results offer additional insight into the complex mechanism of paradoxical mitogen-activated protein kinase signaling involved in hyperproliferative cutaneous neoplasias that arise after BRAF inhibition and suggest a possible role for MMP in tumor progression and invasion. © 2015 Japanese Dermatological Association.

  2. Potent suppressive activity of nonpolyphenolic fraction of green tea (Camellia sinensis) against genotoxin-induced umu C gene expression in Salmonella typhimurium (TA 1535/pSK 1002), tumor promotor-dependent ornithine decarboxylase induction of BALB/c 3T3 fibroblast cells, and chemically induced mouse skin tumorigenesis.

    PubMed

    Okai, Y; Higashi-Okai, K

    Many experimental studies for anticarcinogenic activity of green tea (Camellia sinensis) and tea-derived polyphenols have been carried out. However, the anticarcinogenic activity of the nonpolyphenolic fraction of green tea has been poorly elucidated. To study this problem, the effect of the nonpolyphenolic fraction of green tea leaves was analyzed using in vitro and in vivo experiments associated with tumor initiation and promotion as follows: 1) The nonpolyphenolic fraction caused a strong suppressive effect on umu C gene expression in Salmonella typhimurium (TA 1535/pSK 1002) induced by genotoxic substances such as 2-amino-6-methyldipirido[1,2-a:3',2'-d]imidazole (Glu-P-1) and 2-aminoanthracene (2-AA) in the presence of a hepatic metabolizing enzyme mixture. 2) The same fraction showed a dose-dependent inhibition of ornithine decarboxylase (ODC) in BALB/c 3T3 fibroblasts induced by a tumor promotor, 12-O-tetradecanoylphorbol-13-acetate (TPA). 3) The same fraction also exhibited a significant suppression against mouse skin tumorigenesis induced by 7,12-dimethylbenz[a]anthracene (DMBA) (initiator) and TPA (promotor) through inhibition at both stages of tumor initiation and promotion. These results suggest that the nonpolyphenolic fraction of green tea has a potent suppressing activity against carcinogenesis associated with tumor initiation and promotion.

  3. Optimization of Antitumor Modulators of Pre-mRNA Splicing

    PubMed Central

    Lagisetti, Chandraiah; Palacios, Gustavo; Goronga, Tinopiwa; Freeman, Burgess; Caufield, William; Webb, Thomas R.

    2014-01-01

    The spliceosome regulates pre-mRNA splicing, which is a critical process in normal mammalian cells. Recently recurrent mutations in numerous spliceosomal proteins have been associated with a number of cancers. Previously natural product antitumor agents have been shown to interact with one of the proteins that is subject to recurrent mutations (SF3B1). We report the optimization of a class of tumor-selective spliceosome modulators, which demonstrate significant in vivo antitumor activity. This optimization culminated in the discovery of sudemycin D6, which shows potent cytotoxic activity in the melanoma line SK-MEL-2 (IC50= 39 nM) and other tumor lines, including: JeKo-1 (IC50= 26 nM), HeLa (IC50= 50 nM), and SK-N-AS (IC50= 81 nM). We also report improved processes for the synthesis of these compounds. Our work supports the idea that sudemycin D6 is worthy of further investigation as a novel preclinical anticancer agent with application in the treatment of numerous human cancers. PMID:24325474

  4. Metallothionein provides zinc-mediated protective effects against methamphetamine toxicity in SK-N-SH cells.

    PubMed

    Ajjimaporn, Amornpan; Swinscoe, John; Shavali, Shaik; Govitrapong, Piyarat; Ebadi, Manuchair

    2005-11-30

    Methamphetamine (METH) is a drug of abuse and neurotoxin that induces Parkinson's-like pathology after chronic usage by targeting dopaminergic neurons. Elucidation of the intracellular mechanisms that underlie METH-induced dopaminergic neuron toxicity may help in understanding the mechanism by which neurons die in Parkinson's disease. In the present study, we examined the role of reactive oxygen species (ROS) in the METH-induced death of human dopaminergic SK-N-SH cells and further assessed the neuroprotective effects of zinc and metallothionein (MT) against METH-induced toxicity in culture. METH significantly increased the production of reactive oxygen species, decreased intracellular ATP levels and reduced the cell viability. Pre-treatment with zinc markedly prevented the loss of cell viability caused by METH treatment. Zinc pre-treatment mainly increased the expression of metallothionein and prevented the generation of reactive oxygen species and ATP depletion caused by METH. Chelation of zinc by CaEDTA caused a significant decrease in MT expression and loss of protective effects of MT against METH toxicity. These results suggest that zinc-induced MT expression protects dopaminergic neurons via preventing the accumulation of toxic reactive oxygen species and halting the decrease in ATP levels. Furthermore, MT may prevent the loss of mitochondrial functions caused by neurotoxins. In conclusion, our study suggests that MT, a potent scavenger of free radicals is neuroprotective against dopaminergic toxicity in conditions such as drug of abuse and in Parkinson's disease.

  5. Multicomponent synthesis of some new (1S,4S)-2,5-diazabicyclo[2.2.1]heptane-dithiocarbamates and their in vitro anti-proliferative activity against CaSki, MDA-MB-231 and SK-Lu-1 tumour cells as apoptosis inducing agents without necrosis.

    PubMed

    Laskar, Sujay; Sánchez-Sánchez, Luis; López-Ortiz, Manuel; López-Muñoz, Hugo; Escobar-Sánchez, María L; Sánchez, Arturo T; Regla, Ignacio

    2017-12-01

    Identification of a new class of antitumor agent capable to induce apoptosis without triggering necrotic cell death event is challenging. The present communication describes the multicomponent synthesis of seven new (1S,4S)-2,5-diazabicyclo[2.2.1]heptane-dithiocarbamates and their in vitro antiproliferative activity on cervical cancer cell line (CaSki), breast cancer cell line (MDA-MB231), lung cancer cell line (SK-Lu-1) and human lymphocytes. Among the synthesized dithiocarbamates, compound 9e displayed significant antiproliferative activity without inducing any necrotic cell death (both on tumour cells and lymphocytes) and induced apoptosis in tumor cells by the caspase dependent apoptotic pathway. The compound 9e also exhibited greater tumor selectivity than human lymphocytes. In silico ADME predictions revealed that compound 9e has the potential to be developed as a drug candidate. Rapid chemical modifications of this lead are thus highly necessary for further investigation as a drug like safer antitumor candidate and also to achieve compounds with better activity profile.

  6. Human T-lymphotropic virus type 1 Tax protein complexes with P-TEFb and competes for Brd4 and 7SK snRNP/HEXIM1 binding.

    PubMed

    Cho, Won-Kyung; Jang, Moon Kyoo; Huang, Keven; Pise-Masison, Cynthia A; Brady, John N

    2010-12-01

    Positive transcription elongation factor b (P-TEFb) plays an important role in stimulating RNA polymerase II elongation for viral and cellular gene expression. P-TEFb is found in cells in either an active, low-molecular-weight (LMW) form or an inactive, high-molecular-weight (HMW) form. We report here that human T-lymphotropic virus type 1 (HTLV-1) Tax interacts with the cyclin T1 subunit of P-TEFb, forming a distinct Tax/P-TEFb LMW complex. We demonstrate that Tax can play a role in regulating the amount of HMW complex present in the cell by decreasing the binding of 7SK snRNP/HEXIM1 to P-TEFb. This is seen both in vitro using purified Tax protein and in vivo in cells transduced with Tax expression constructs. Further, we find that a peptide of cyclin T1 spanning the Tax binding domain inhibits the ability of Tax to disrupt HMW P-TEFb complexes. These results suggest that the direct interaction of Tax with cyclin T1 can dissociate P-TEFb from the P-TEFb/7SK snRNP/HEXIM1 complex for activation of the viral long terminal repeat (LTR). We also show that Tax competes with Brd4 for P-TEFb binding. Chromatin immunoprecipitation (ChIP) assays demonstrated that Brd4 and P-TEFb are associated with the basal HTLV-1 LTR, while Tax and P-TEFb are associated with the activated template. Furthermore, the knockdown of Brd4 by small interfering RNA (siRNA) activates the HTLV-1 LTR promoter, which results in an increase in viral expression and production. Our studies have identified Tax as a regulator of P-TEFb that is capable of affecting the balance between its association with the large inactive complex and the small active complex.

  7. Alterations in Skeletal Muscle Indicators of Mitochondrial Structure and Biogenesis in Patients with Type 2 Diabetes and Heart Failure: Effects of Epicatechin Rich Cocoa

    PubMed Central

    Taub, Pam R.; Ramirez‐Sanchez, Israel; Ciaraldi, Theodore P.; Perkins, Guy; Murphy, Anne N.; Naviaux, Robert; Hogan, Michael; Maisel, Alan S.; Henry, Robert R.; Ceballos, Guillermo

    2012-01-01

    Abstract (‐)‐Epicatechin (Epi), a flavanol in cacao stimulates mitochondrial volume and cristae density and protein markers of skeletal muscle (SkM) mitochondrial biogenesis in mice. Type 2 diabetes mellitus (DM2) and heart failure (HF) are diseases associated with defects in SkM mitochondrial structure/function. A study was implemented to assess perturbations and to determine the effects of Epi‐rich cocoa in SkM mitochondrial structure and mediators of biogenesis. Five patients with DM2 and stage II/III HF consumed dark chocolate and a beverage containing approximately 100 mg of Epi per day for 3 months. We assessed changes in protein and/or activity levels of oxidative phosphorylation proteins, porin, mitofilin, nNOS, nitric oxide, cGMP, SIRT1, PGC1α, Tfam, and mitochondria volume and cristae abundance by electron microscopy from SkM. Apparent major losses in normal mitochondria structure were observed before treatment. Epi‐rich cocoa increased protein and/or activity of mediators of biogenesis and cristae abundance while not changing mitochondrial volume density. Epi‐rich cocoa treatment improves SkM mitochondrial structure and in an orchestrated manner, increases molecular markers of mitochondrial biogenesis resulting in enhanced cristae density. Future controlled studies are warranted using Epi‐rich cocoa (or pure Epi) to translate improved mitochondrial structure into enhanced cardiac and/or SkM muscle function. Clin Trans Sci 2012; Volume 5: 43–47 PMID:22376256

  8. Proteasomal degradation of sphingosine kinase 1 and inhibition of dihydroceramide desaturase by the sphingosine kinase inhibitors, SKi or ABC294640, induces growth arrest in androgen-independent LNCaP-AI prostate cancer cells.

    PubMed

    McNaughton, Melissa; Pitman, Melissa; Pitson, Stuart M; Pyne, Nigel J; Pyne, Susan

    2016-03-29

    Sphingosine kinases (two isoforms termed SK1 and SK2) catalyse the formation of the bioactive lipid sphingosine 1-phosphate. We demonstrate here that the SK2 inhibitor, ABC294640 (3-(4-chlorophenyl)-adamantane-1-carboxylic acid (pyridin-4-ylmethyl)amide) or the SK1/SK2 inhibitor, SKi (2-(p-hydroxyanilino)-4-(p-chlorophenyl)thiazole)) induce the proteasomal degradation of SK1a (Mr = 42 kDa) and inhibit DNA synthesis in androgen-independent LNCaP-AI prostate cancer cells. These effects are recapitulated by the dihydroceramide desaturase (Des1) inhibitor, fenretinide. Moreover, SKi or ABC294640 reduce Des1 activity in Jurkat cells and ABC294640 induces the proteasomal degradation of Des1 (Mr = 38 kDa) in LNCaP-AI prostate cancer cells. Furthermore, SKi or ABC294640 or fenretinide increase the expression of the senescence markers, p53 and p21 in LNCaP-AI prostate cancer cells. The siRNA knockdown of SK1 or SK2 failed to increase p53 and p21 expression, but the former did reduce DNA synthesis in LNCaP-AI prostate cancer cells. Moreover, N-acetylcysteine (reactive oxygen species scavenger) blocked the SK inhibitor-induced increase in p21 and p53 expression but had no effect on the proteasomal degradation of SK1a. In addition, siRNA knockdown of Des1 increased p53 expression while a combination of Des1/SK1 siRNA increased the expression of p21. Therefore, Des1 and SK1 participate in regulating LNCaP-AI prostate cancer cell growth and this involves p53/p21-dependent and -independent pathways. Therefore, we propose targeting androgen-independent prostate cancer cells with compounds that affect Des1/SK1 to modulate both de novo and sphingolipid rheostat pathways in order to induce growth arrest.

  9. RNA Interference Directed against the Transglutaminase Gene Triggers Dysbiosis of Gut Microbiota in Drosophila*

    PubMed Central

    Sekihara, Sanae; Shibata, Toshio; Hyakkendani, Mai; Kawabata, Shun-ichiro

    2016-01-01

    We recently reported that transglutaminase (TG) suppresses immune deficiency pathway-controlled antimicrobial peptides (IMD-AMPs), thereby conferring immune tolerance to gut microbes, and that RNAi of the TG gene in flies decreases the lifespan compared with non-TG-RNAi flies. Here, analysis of the bacterial composition of the Drosophila gut by next-generation sequencing revealed that gut microbiota comprising one dominant genus of Acetobacter in non-TG-RNAi flies was shifted to that comprising two dominant genera of Acetobacter and Providencia in TG-RNAi flies. Four bacterial strains, including Acetobacter persici SK1 and Acetobacter indonesiensis SK2, Lactobacillus pentosus SK3, and Providencia rettgeri SK4, were isolated from the midgut of TG-RNAi flies. SK1 exhibited the highest resistance to the IMD-AMPs Cecropin A1 and Diptericin among the isolated bacteria. In contrast, SK4 exhibited considerably lower resistance against Cecropin A1, whereas SK4 exhibited high resistance to hypochlorous acid. The resistance of strains SK1–4 against IMD-AMPs in in vitro assays could not explain the shift of the microbiota in the gut of TG-RNAi flies. The lifespan was reduced in gnotobiotic flies that ingested both SK4 and SK1, concomitant with the production of reactive oxygen species and apoptosis in the midgut, whereas the survival rate was not altered in gnotobiotic flies that mono-ingested either SK4 or SK1. Interestingly, significant amounts of reactive oxygen species were detected in the midgut of gnotobiotic flies that ingested SK4 and SK2, concomitant with no significant apoptosis in the midgut. In gnotobiotic flies that co-ingested SK4 and SK1, an additional unknown factor(s) may be required to cause midgut apoptosis. PMID:27760824

  10. One new 19-nor cucurbitane-type triterpenoid from the stems of Momordica charantia.

    PubMed

    Li, Yong-chao; Xu, Xin-juan; Yang, Jing; Wu, Xing-gang; Fu, Qing-yun

    2016-01-01

    One new 19-nor cucurbitane-type triterpenoid (3β,9β,25-trihydroxy-7β-methoxy-19-nor-cucurbita-5,23(E)-diene) (1), together with other six known cucurbitane-type triterpenoids (2-7), were isolated from the stems of Momordica charantia L. The chemical structure of 1 was elucidated by extensive 1D NMR and 2D NMR (HSQC, HMBC, COSY and ROESY), MS experiments. Using MTT assay, compound 1 exhibited weak cytotoxicity against HL-60, A-549, and SK-BR-3 cell lines with the IC50 values at 27.3, 32.7 and 26.6 μM, respectively.

  11. Mutant human tumor suppressor p53 modulates the activation of mitogen-activated protein kinase and nuclear factor-kappaB, but not c-Jun N-terminal kinase and activated protein-1.

    PubMed

    Gulati, Anthony P; Yang, Yang-Ming; Harter, David; Mukhopadhyay, Asok; Aggarwal, Bharat B; Aggarwal, Bharat A; Benzil, Deborah L; Whysner, John; Albino, Anthony P; Murali, Raj; Jhanwar-Uniyal, Meena

    2006-01-01

    The roles of the mitogen-activated kinase protein (MAPK) pathway, nuclear factor-kappa B (NF-kappaB), and activator protein-1 (AP-1) in cellular responses to growth factors and mitogen are well established. However, the manner by which these proliferative pathways are affected by the tumor suppressor protein p53 is not fully understood. We report here the results of an investigation of the status of p53 on two human melanoma cell lines with wild-type p53 (SK-Mel-186) or mutant p53 (SK-Mel-110). The basal levels of the activated extracellular-signal regulated kinases 1 and 2 (ERK1/2) were high in cells with wild-type p53, but low in cells with mutant p53. The 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced activation of ERK1/2 through the phosphorylation of threonine and tyrosine at 202 and 204, respectively, was demonstrated in both cell lines, however, in a discrete manner. TPA-induced activation of ERK1/2 was sustained in wild-type p53 cells, while only a transient activation was seen in mutant p53 cells. Inhibition of MAPK kinase (MEK), an upstream kinase, by U0126, blocked TPA-induced activation of ERK1/2 in wild-type p53 cells and in mutant p53 cells. Treatment of wild-type p53 (SK-Mel 186) cells with small interfering RNA (siRNA) of p53 displayed a transient induction of activation of ERK1/2 following TPA treatment, indicating that p53 has a role in the regulation of the activation of ERK1/2. NF-kappaB activity decreased significantly in cells with wild-type p53, while enhanced NF-kappaB activity was evident in cells with mutant p53. The expression of either wild-type or mutant p53 had a similar effect on TPA-induced Jun N-terminal kinase (JNK) activation, indicating specificity for the ERK pathway. Similarly, AP-1 binding activity showed a transient variation in both cell lines after TPA treatment but with different kinetics. These observations suggest that both wild-type and mutant p53 can modulate the activation pathways for ERK1/2, and NF

  12. Multi-environment QTL analysis of grain morphology traits and fine mapping of a kernel-width QTL in Zheng58 × SK maize population.

    PubMed

    Raihan, Mohammad Sharif; Liu, Jie; Huang, Juan; Guo, Huan; Pan, Qingchun; Yan, Jianbing

    2016-08-01

    Sixteen major QTLs regulating maize kernel traits were mapped in multiple environments and one of them, qKW - 9.2 , was restricted to 630 Kb, harboring 28 putative gene models. To elucidate the genetic basis of kernel traits, a quantitative trait locus (QTL) analysis was conducted in a maize recombinant inbred line population derived from a cross between two diverse parents Zheng58 and SK, evaluated across eight environments. Construction of a high-density linkage map was based on 13,703 single-nucleotide polymorphism markers, covering 1860.9 cM of the whole genome. In total, 18, 26, 23, and 19 QTLs for kernel length, width, thickness, and 100-kernel weight, respectively, were detected on the basis of a single-environment analysis, and each QTL explained 3.2-23.7 % of the phenotypic variance. Sixteen major QTLs, which could explain greater than 10 % of the phenotypic variation, were mapped in multiple environments, implying that kernel traits might be controlled by many minor and multiple major QTLs. The major QTL qKW-9.2 with physical confidence interval of 1.68 Mbp, affecting kernel width, was then selected for fine mapping using heterogeneous inbred families. At final, the location of the underlying gene was narrowed down to 630 Kb, harboring 28 putative candidate-gene models. This information will enhance molecular breeding for kernel traits and simultaneously assist the gene cloning underlying this QTL, helping to reveal the genetic basis of kernel development in maize.

  13. Differential gene expression in porcine SK6 cells infected with wild-type and SAP domain-mutant foot-and-mouth disease virus.

    PubMed

    Ni, Zixin; Yang, Fan; Cao, Weijun; Zhang, Xiangle; Jin, Ye; Mao, Ruoqing; Du, Xiaoli; Li, Weiwei; Guo, Jianhong; Liu, Xiangtao; Zhu, Zixiang; Zheng, Haixue

    2016-06-01

    Foot-and-mouth disease virus (FMDV) is the causative agent of a highly contagious disease in livestock. The viral proteinase L(pro) of FMDV is involved in pathogenicity, and mutation of the L(pro) SAP domain reduces FMDV pathogenicity in pigs. To determine the gene expression profiles associated with decreased pathogenicity in porcine cells, we performed transcriptome analysis using next-generation sequencing technology and compared differentially expressed genes in SK6 cells infected with FMDV containing L(pro) with either a wild-type or mutated version of the SAP domain. This analysis yielded 1,853 genes that exhibited a2-fold change in expression and was validated by real-time quantitative PCR detection of several differentially expressed genes. Many of the differentially expressed genes correlated with antiviral responses corresponded to genes associated with transcription factors, immune regulation, cytokine production, inflammatory response, and apoptosis. Alterations in gene expression profiles may be responsible for the variations in pathogenicity observed between the two FMDV variants. Our results provided genes of interest for the further study of antiviral pathways and pathogenic mechanisms related to FMDV L(pro).

  14. [Astaxanthin inhibits proliferation and promotes apoptosis of A549 lung cancer cells via blocking JAK1/STAT3 pathway].

    PubMed

    Wu, Chuntao; Zhang, Jinji; Liu, Tienan; Jiao, Guimei; Li, Changzai; Hu, Baoshan

    2016-06-01

    Objective To investigate the anti-tumor effects of astaxanthin on A549 lung cancer cells and the related mechanisms. Methods A549 cells were cultured with various concentrations of astaxanthin (20, 40, 60, 80, 100 μmol/L), and DMSO at the same concentrations served as vehicle controls. The viability of A549 cells was detected by CCK-8 assay; cell cycle and apoptosis were observed by flow cytometry; and the expressions of B-cell lymphoma-2 (Bcl-2), Bcl-2 associated X protein (Bax), signal transducers and activators of transcription 3 (STAT3), and Janus kinase 1 (JAK1) were evaluated by Western blotting. Results CCK-8 assay showed that astaxanthin decreased the proliferation of A549 cells in a dose-dependent manner. Flow cytometry showed that astaxanthin increased the number of cells in the G0/G1 phase and induced apoptosis in A549 cells. Western blotting showed that astaxanthin up-regulated the expression of Bax and down-regulated the expressions of Bcl-2, STAT3 and JAK1. Conclusion Astaxanthin functions as a potent inhibitor of A549 lung cancer cell growth by targeting JAK1/STAT3 signaling pathway.

  15. Loss of the endothelial glycocalyx is associated with increased E-selectin mediated adhesion of lung tumour cells to the brain microvascular endothelium.

    PubMed

    Rai, Srijana; Nejadhamzeeigilani, Zaynab; Gutowski, Nicholas J; Whatmore, Jacqueline L

    2015-09-25

    Arrest of metastasising lung cancer cells to the brain microvasculature maybe mediated by interactions between ligands on circulating tumour cells and endothelial E-selectin adhesion molecules; a process likely to be regulated by the endothelial glycocalyx. Using human cerebral microvascular endothelial cells and non-small cell lung cancer (NSCLC) cell lines, we describe how factors secreted by NSCLC cells i.e. cystatin C, cathepsin L, insulin-like growth factor-binding protein 7 (IGFBP7), vascular endothelial growth factor (VEGF) and tumour necrosis factor-alpha (TNF-α), damage the glycocalyx and enhance initial contacts between lung tumour and cerebral endothelial cells. Endothelial cells were treated with tumour secreted-proteins or lung tumour conditioned medium (CM). Surface levels of E-selectin were quantified by ELISA. Adhesion of A549 and SK-MES-1 cells was examined under flow conditions (1 dyne/cm(2)). Alterations in the endothelial glycocalyx were quantified by binding of fluorescein isothiocyanate-linked wheat germ agglutinin (WGA-FITC). A549 and SK-MES-1 CM and secreted-proteins significantly enhanced endothelial surface E-selectin levels after 30 min and 4 h and tumour cell adhesion after 30 min, 4 and 24 h. Both coincided with significant glycocalyx degradation; A549 and SK-MES-1 CM removing 55 ± 12 % and 58 ± 18.7 % of WGA-FITC binding, respectively. Inhibition of E-selectin binding by monoclonal anti-E-selectin antibody completely attenuated tumour cell adhesion. These data suggest that metastasising lung cancer cells facilitate their own adhesion to the brain endothelium by secreting factors that damage the endothelial glycocalyx, resulting in exposure of the previously shielded adhesion molecules and engagement of the E-selectin-mediated adhesion axis.

  16. The Immunomodulatory Role of Adjuvants in Vaccines Formulated with the Recombinant Antigens Ov-103 and Ov-RAL-2 against Onchocerca volvulus in Mice.

    PubMed

    Hess, Jessica A; Zhan, Bin; Torigian, April R; Patton, John B; Petrovsky, Nikolai; Zhan, Tingting; Bottazzi, Maria Elena; Hotez, Peter J; Klei, Thomas R; Lustigman, Sara; Abraham, David

    2016-07-01

    In some regions in Africa, elimination of onchocerciasis may be possible with mass drug administration, although there is concern based on several factors that onchocerciasis cannot be eliminated solely through this approach. A vaccine against Onchocerca volvulus would provide a critical tool for the ultimate elimination of this infection. Previous studies have demonstrated that immunization of mice with Ov-103 and Ov-RAL-2, when formulated with alum, induced protective immunity. It was hypothesized that the levels of protective immunity induced with the two recombinant antigens formulated with alum would be improved by formulation with other adjuvants known to enhance different types of antigen-specific immune responses. Immunizing mice with Ov-103 and Ov-RAL-2 in conjunction with alum, Advax 2 and MF59 induced significant levels of larval killing and host protection. The immune response was biased towards Th2 with all three of the adjuvants, with IgG1 the dominant antibody. Improved larval killing and host protection was observed in mice immunized with co-administered Ov-103 and Ov-RAL-2 in conjunction with each of the three adjuvants as compared to single immunizations. Antigen-specific antibody titers were significantly increased in mice immunized concurrently with the two antigens. Based on chemokine levels, it appears that neutrophils and eosinophils participate in the protective immune response induced by Ov-103, and macrophages and neutrophils participate in immunity induced by Ov-RAL-2. The mechanism of protective immunity induced by Ov-103 and Ov-RAL-2, with the adjuvants alum, Advax 2 and MF59, appears to be multifactorial with roles for cytokines, chemokines, antibody and specific effector cells. The vaccines developed in this study have the potential of reducing the morbidity associated with onchocerciasis in humans.

  17. Recombinant streptokinase vs hydrocortisone suppositories in acute hemorrhoids: A randomized controlled trial.

    PubMed

    Hernández-Bernal, Francisco; Castellanos-Sierra, Georgina; Valenzuela-Silva, Carmen M; Catasús-Álvarez, Karem M; Martínez-Serrano, Osmany; Lazo-Diago, Odalys C; Bermúdez-Badell, Cimara H; Causa-García, José R; Domínguez-Suárez, Juan E

    2015-06-21

    To compare the efficacy and safety of recombinant streptokinase (rSK) vs hydrocortisone acetate-based suppositories in acute hemorrhoidal disease. A multicenter (11 sites), randomized (1:1:1), open, controlled trial with parallel groups was performed. All participating patients gave their written, informed consent. After inclusion, patients with acute symptoms of hemorrhoids were centrally randomized to receive, as outpatients, by the rectal route, suppositories of rSK 200000 IU of one unit every 8 h (first 3 units) and afterwards every 12 h until 8 administrations were completed (schedule A), one unit every 8 h until 6 units were completed (schedule B), or 25 mg hydrocortisone acetate once every 8 h up to a maximum of 24 administrations. Evaluations were performed at 3, 5, and 10 d post-inclusion. The main end-point was the 5(th)-day response (disappearance of pain and bleeding, and ≥ 70% reduction of the lesion size). Time to response and need for thrombectomy were secondary efficacy variables. Adverse events were also evaluated. Groups were homogeneous with regards to demographic and baseline characteristics. Fifth day complete response rates were 156/170 (91.8%; 95%CI: 87.3-96.2), 155/170 (91.2%; 95%CI: 86.6%-95.7%), and 46/170 (27.1%; 95%CI: 20.1%-34.0%) with rSK (schedule A and B) and hydrocortisone acetate suppositories, respectively. These 64.6% and 63.9% differences (95%CI: 56.7%-72.2% and 55.7%-72.0%) were highly significant (P < 0.001). This advantage was detected since the early 3(rd) day evaluation (68.8% and 64.1% vs 7.1% for the rSK and active control groups, respectively; P < 0.001) and was maintained even at the late 10(th) day assessment (97.1% and 93.5% vs 67.1% for rSK and hydrocortisone acetate, respectively; P < 0.001). Time to response was 3 d (95%CI: 2.9-3.1) for both rSK groups and 10 d (95%CI: 9.3-10.7) in the hydrocortisone acetate group. This difference was highly significant (P < 0.001). All subgroup stratified analyses (with or

  18. Comparison of the pharmacokinetics, biodistribution and dosimetry of monoclonal antibodies OC125, OV-TL 3, and 139H2 as IgG and F(ab')2 fragments in experimental ovarian cancer.

    PubMed

    Molthoff, C F; Pinedo, H M; Schlüper, H M; Nijman, H W; Boven, E

    1992-05-01

    Monoclonal antibody (MAb) 139H2 was previously shown to localise specifically into ovarian cancer xenografts in nude mice. MAb 139H2 was compared with MAbs OC125 and OV-TL 3, all reactive with ovarian carcinomas, for the binding characteristics as IgG and F(ab')2 fragments with the use of the OVCAR-3 cell line grown in vitro and as s.c. xenografts. Immunoperoxidase staining of OVCAR-3 tissue sections with MAbs OC125 and 139H2 was heterogeneous, whereas MAb OV-TL 3 showed homogeneity. No differences in binding were observed between IgG and F(ab')2. The avidity expressed as apparent affinity constants of MAbs OC125, OV-TL 3 and 139H2 for OVAR-3 cells were 1 x 10(9) M-1, 1 x 10(9) M-1, and 1 x 10(8) M-1, while the number of antigenic determinants were 5 x 10(6), 1 x 10(6) and 7 x 10(6), respectively. In OVCAR-3 bearing nude mice the blood half-lives of the MAbs as IgG and F(ab')2 were approximately 50 h and 6 h, respectively. Maximum tumour uptake for the whole MAbs OC125, OV-TL 3, 139H2 and a control MAb 2C7 was 8.5%, 17.7%, 11.1% and 2.5% of the injected dose g-1, reached at 72 h after injection. For the respective F(ab')2 fragments, the maximum values were 5.2%, 10.0%, 5.5% and 1.9% of the injected dose g-1, reached between 6 h and 15 h. Tumour to non-tumour ratios were more favourable for the F(ab')2 fragments as compared to those for MAbs as IgG. Biodistribution in mice bearing a control tumour confirmed the specificity of tumour localisation of MAbs OC125, OV-TL 3 and 139H2. After injection of a tracer dose of 10 microCi of radiolabelled MAbs OC125, OV-TL 3 and 139H2 as IgG, tumours received 38 cGy, and 9 cGy. In our OVCAR-3 model, a ranking in efficiency in tumour localisation would indicate MAb OV-TL 3 as most favourable MAb, but cross-reactivity with subpopulations of human white blood cells might hamper its clinical use. Dosimetric data indicate a 4-fold higher radiation absorbed dose to tumours for IgG compared with F(ab')2 fragments.

  19. Nitric oxide donors reduce the invasion ability of ovarian cancer cells in vitro.

    PubMed

    Kielbik, Michal; Szulc, Izabela; Brzezinska, Marta; Bednarska, Katarzyna; Przygodzka, Patrycja; Sulowska, Zofia; Nowak, Marek; Klink, Magdalena

    2014-11-01

    The most important factors involved in tumor metastasis and angiogenesis are metalloproteinases (MMPs), vascular endothelial growth factor, and multifunctional transforming growth factor β1. These factors are responsible for extracellular matrix degradation, induction of vascular permeability, and enhancement of tumor cells' invasion and metastasis. Elevated expression and secretion of the above-mentioned factors are correlated with the higher aggressiveness of tumors and low patient survival for example, patients with ovarian cancer. Therefore, regulation of the expression, secretion, and activity of these factors is still considered a potent target for therapeutic intervention in cancer patients. Nitric oxide (NO) donors belong to the class of agents with multivalent targeted activities in cancer cells and are considered potential anticancer therapeutics. Our studies have shown that NO donors such as spermine/NO and diethylenetriamine/NO decrease the secretion of vascular endothelial growth factor-A from the OVCAR-3 ovarian cancer cell line, but not from the SK-OV-3 ovarian cancer cell line. The release of MMP-2 from both cell lines was reduced in a soluble guanylate cyclase-dependent manner by spermine/NO and diethylenetriamine/NO. Nevertheless, MMP-2 activity was only affected in SK-OV-3 cells. Both NO donors reduced the transmigration of the ovarian cancer cell lines. We did not observe any significant effect of spermine/NO and diethylenetriamine/NO on mRNA expression of the tested aggressiveness factors. In conclusion, our data indicated that NO donors reduced the metastatic potential of ovarian cancer cells, but its impact is rather low and requires high concentrations of donors. Moreover, both the tested cell lines differed in the susceptibility to NO donors.

  20. A New In Vitro Co-Culture Model Using Magnetic Force-Based Nanotechnology.

    PubMed

    Takanari, Hiroki; Miwa, Keiko; Fu, XianMing; Nakai, Junichi; Ito, Akira; Ino, Kousuke; Honda, Hiroyuki; Tonomura, Wataru; Konishi, Satoshi; Opthof, Tobias; van der Heyden, Marcel Ag; Kodama, Itsuo; Lee, Jong-Kook

    2016-10-01

    Skeletal myoblast (SkMB) transplantation has been conducted as a therapeutic strategy for severe heart failure. However, arrhythmogenicity following transplantation remains unsolved. We developed an in vitro model of myoblast transplantation with "patterned" or "randomly-mixed" co-culture of SkMBs and cardiomyocytes enabling subsequent electrophysiological, and arrhythmogenic evaluation. SkMBs were magnetically labeled with magnetite nanoparticles and co-cultured with neonatal rat ventricular myocytes (NRVMs) on multi-electrode arrays. SkMBs were patterned by a magnet beneath the arrays. Excitation synchronicity was evaluated by Ca(2+) imaging using a gene-encoded Ca(2+) indicator, G-CaMP2. In the monoculture of NRVMs (control), conduction was well-organized. In the randomly-mixed co-culture of NRVMs and SkMBs (random group), there was inhomogeneous conduction from multiple origins. In the "patterned" co-culture where an en bloc SKMB-layer was inserted into the NRVM-layer, excitation homogenously propagated although conduction was distorted by the SkMB-area. The 4-mm distance conduction time (CT) in the random group was significantly longer (197 ± 126 ms) than in control (17 ± 3 ms). In the patterned group, CT through NRVM-area did not change (25 ± 3 ms), although CT through the SkMB-area was significantly longer (132 ± 77 ms). The intervals between spontaneous excitation varied beat-to-beat in the random group, while regular beating was recorded in the control and patterned groups. Synchronized Ca(2+) transients of NRVMs were observed in the patterned group, whereas those in the random group were asynchronous. Patterned alignment of SkMBs is feasible with magnetic nanoparticles. Using the novel in vitro model mimicking cell transplantation, it may become possible to predict arrhythmogenicity due to heterogenous cell transplantation. J. Cell. Physiol. 231: 2249-2256, 2016. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

  1. STS-65 Mission Specialist Chiao in front of IML-2 Rack 3 aboard OV-102

    NASA Technical Reports Server (NTRS)

    1994-01-01

    STS-65 Mission Specialist Leroy Chiao is seen in the International Microgravity Laboratory 2 (IML-2) spacelab science module in front of Rack 3 and above center aisle equipment. Chiao has just made an observation of the goldfish container (silver apparatus on left between his right hand and knee). The Rack 3 Aquatic Animal Experiment Unit (AAEU) also contains Medaka and newts. Chiao joined five other NASA astronauts and a Japanese payload specialist for two weeks of experimenting onboard the Space Shuttle Columbia, Orbiter Vehicle (OV) 102, in Earth orbit.

  2. Accumulation of 24S-hydroxycholesterol in neuronal SK-N-BE cells treated with hexacosanoic acid (C26:0): argument in favor of 24S-hydroxycholesterol as a potential biomarker of neurolipotoxicity.

    PubMed

    Zarrouk, A; Hammami, M; Moreau, T; Lizard, G

    2015-02-01

    Cholesterol oxide derivatives (oxysterols) are viewed as potential biomarkers of neurodegenerative diseases. 24S-hydroxycholesterol, an oxysterol produced only in brain neurons, is often found for unknown reasons in increased levels in the plasma in patients with neurodegenerative diseases. On human neuronal SK-N-BE cells treated with hexacosanoic acid (C26:0) identified at increased levels in the tissues and plasma of patients with peroxisomal leukodystrophies and Alzheimer's disease, we observed increased level of 24S-hydroxycholesterol associated with C26:0 induced lipotoxicity. This finding reinforces the hypothesis suggesting that 24S-hydroxycholesterol could constitute a biomarker of neurotoxicity. Copyright © 2014 Elsevier Masson SAS. All rights reserved.

  3. Quantitative comparison of the spreading and invasion of radial growth phase and metastatic melanoma cells in a three-dimensional human skin equivalent model.

    PubMed

    Haridas, Parvathi; McGovern, Jacqui A; McElwain, Sean D L; Simpson, Matthew J

    2017-01-01

    Standard two-dimensional (2D) cell migration assays do not provide information about vertical invasion processes, which are critical for melanoma progression. We provide information about three-dimensional (3D) melanoma cell migration, proliferation and invasion in a 3D melanoma skin equivalent (MSE) model. In particular, we pay careful attention to compare the structure of the tissues in the MSE with similarly-prepared 3D human skin equivalent (HSE) models. The HSE model is identically prepared to the MSE model except that melanoma cells are omitted. Using the MSE model, we examine melanoma migration, proliferation and invasion from two different human melanoma cell lines. One cell line, WM35, is associated with the early phase of the disease where spreading is thought to be confined to the epidermis. The other cell line, SK-MEL-28, is associated with the later phase of the disease where spreading into the dermis is expected. 3D MSE and HSE models are constructed using human de-epidermised dermis (DED) prepared from skin tissue. Primary fibroblasts and primary keratinocytes are used in the MSE and HSE models to ensure the formation of a stratified epidermis, with a well-defined basement membrane. Radial spreading of cells across the surface of the HSE and MSE models is observed. Vertical invasion of melanoma cells downward through the skin is observed and measured using immunohistochemistry. All measurements of invasion are made at day 0, 9, 15 and 20, providing detailed time course data. Both HSE and MSE models are similar to native skin in vivo , with a well-defined stratification of the epidermis that is separated from the dermis by a basement membrane. In the HSE and MSE we find fibroblast cells confined to the dermis, and differentiated keratinocytes in the epidermis. In the MSE, melanoma cells form colonies in the epidermis during the early part of the experiment. In the later stage of the experiment, the melanoma cells in the MSE invade deeper into the

  4. Quantitative comparison of the spreading and invasion of radial growth phase and metastatic melanoma cells in a three-dimensional human skin equivalent model

    PubMed Central

    Haridas, Parvathi; McGovern, Jacqui A.; McElwain, Sean D.L.

    2017-01-01

    Background Standard two-dimensional (2D) cell migration assays do not provide information about vertical invasion processes, which are critical for melanoma progression. We provide information about three-dimensional (3D) melanoma cell migration, proliferation and invasion in a 3D melanoma skin equivalent (MSE) model. In particular, we pay careful attention to compare the structure of the tissues in the MSE with similarly-prepared 3D human skin equivalent (HSE) models. The HSE model is identically prepared to the MSE model except that melanoma cells are omitted. Using the MSE model, we examine melanoma migration, proliferation and invasion from two different human melanoma cell lines. One cell line, WM35, is associated with the early phase of the disease where spreading is thought to be confined to the epidermis. The other cell line, SK-MEL-28, is associated with the later phase of the disease where spreading into the dermis is expected. Methods 3D MSE and HSE models are constructed using human de-epidermised dermis (DED) prepared from skin tissue. Primary fibroblasts and primary keratinocytes are used in the MSE and HSE models to ensure the formation of a stratified epidermis, with a well-defined basement membrane. Radial spreading of cells across the surface of the HSE and MSE models is observed. Vertical invasion of melanoma cells downward through the skin is observed and measured using immunohistochemistry. All measurements of invasion are made at day 0, 9, 15 and 20, providing detailed time course data. Results Both HSE and MSE models are similar to native skin in vivo, with a well-defined stratification of the epidermis that is separated from the dermis by a basement membrane. In the HSE and MSE we find fibroblast cells confined to the dermis, and differentiated keratinocytes in the epidermis. In the MSE, melanoma cells form colonies in the epidermis during the early part of the experiment. In the later stage of the experiment, the melanoma cells in the MSE

  5. Pyridostigmine but not 3,4-diaminopyridine exacerbates ACh receptor loss and myasthenia induced in mice by muscle-specific kinase autoantibody.

    PubMed

    Morsch, Marco; Reddel, Stephen W; Ghazanfari, Nazanin; Toyka, Klaus V; Phillips, William D

    2013-05-15

    In myasthenia gravis, the neuromuscular junction is impaired by the antibody-mediated loss of postsynaptic acetylcholine receptors (AChRs). Muscle weakness can be improved upon treatment with pyridostigmine, a cholinesterase inhibitor, or with 3,4-diaminopyridine, which increases the release of ACh quanta. The clinical efficacy of pyridostigmine is in doubt for certain forms of myasthenia. Here we formally examined the effects of these compounds in the antibody-induced mouse model of anti-muscle-specific kinase (MuSK) myasthenia gravis. Mice received 14 daily injections of IgG from patients with anti-MuSK myasthenia gravis. This caused reductions in postsynaptic AChR densities and in endplate potential amplitudes. Systemic delivery of pyridostigmine at therapeutically relevant levels from days 7 to 14 exacerbated the anti-MuSK-induced structural alterations and functional impairment at motor endplates in the diaphragm muscle. No such effect of pyridostigmine was found in mice receiving control human IgG. Mice receiving smaller amounts of MuSK autoantibodies did not display overt weakness, but 9 days of pyridostigmine treatment precipitated generalised muscle weakness. In contrast, one week of treatment with 3,4-diaminopyridine enhanced neuromuscular transmission in the diaphragm muscle. Both pyridostigmine and 3,4-diaminopyridine increase ACh in the synaptic cleft yet only pyridostigmine potentiated the anti-MuSK-induced decline in endplate ACh receptor density. These results thus suggest that ongoing pyridostigmine treatment potentiates anti-MuSK-induced AChR loss by prolonging the activity of ACh in the synaptic cleft.

  6. Synergistic effects of the sesquiterpene lactone, EPD, with cisplatin and paclitaxel in ovarian cancer cells.

    PubMed

    van Haaften, Caroline; Boot, Arnoud; Corver, Willem E; van Eendenburg, Jaap D H; Trimbos, Baptist J M Z; van Wezel, Tom

    2015-04-25

    Ovarian cancer remains still the leading cause of death of gynecological malignancy, in spite of first-line chemotherapy with cisplatin and paclitaxel. Although initial response is favorably, relapses are common and prognosis for women with advanced disease stays poor. Therefore efficacious approaches are needed. Previously, an anti-cancer agent, EPD exhibited potent cytotoxic effects towards ovarian cancer and not towards normal cells. Cell viability and cell cycle analysis studies were performed with EPD, in combination with cisplatin and/or paclitaxel, using the ovarian carcinoma cell lines: SK-OV-3, OVCAR-3, JC, JC-pl and normal fibroblasts. Cell viability was measured using Presto Blue and cell cycle analysis using a flow cytometer. Apoptosis was measured in JC and JC-pl , using the caspase 3 assay kit. In JC-pl, SK-OV-3 and JC, synergistic interactions between either EPD and cisplatin or EPD and paclitaxel were observed. For the first time the effects of EPD on the cell cycle of ovarian cancer cells and normal cells was studied. EPD and combinations of EPD with cisplatin and/ or paclitaxel showed cell cycle arrest in the G2/M phase. The combination of EPD and cisplatin showed a significant synergistic effect in cell line JC-pl, while EPD with paclitaxel showed synergistic interaction in JC. Additionally, synergistic drug combinations showed increased apoptosis. Our results showed a synergistic effect of EPD and cisplatin in an ovarian drug resistant cell line as well as a synergistic effect of EPD and paclitaxel in two other ovarian cell lines. These results might enhance clinical efficacy, compared to the existing regimen of paclitaxel and cisplatin.

  7. Effect of repeated ketoprofen administration during surgical castration of bulls on cortisol, immunological function, feed intake, growth, and behavior.

    PubMed

    Ting, S T L; Earley, B; Crowe, M A

    2003-05-01

    To determine the effect of repeated ketoprofen (K) administration to surgically castrated bulls on cortisol, acute-phase proteins, immune function, feed intake, growth and behavior, 50 Holstein x Friesian bulls (11 mo old; 300 +/- 3.3 kg) were assigned to one of five treatments: 1) untreated control (C); 2) surgical castration at 0 min (S); 3) S following an i.v. injection of 3 mg/kg of BW of K at -20 min (SK1); 4) S following 1.5 mg/kg of BW of K at -20 and 0 min (SK2); or 5) S following 1.5 mg/kg of BW of K at -20 and 0 min and 3 mg/kg of BW of K at 24 h (SK3). Castration acutely increased plasma cortisol concentrations in S- and K-treated animals compared with C, with no differences in peak and interval to peak cortisol responses among the castration groups. Overall, the integrated cortisol response was greater (P < 0.05) in the castrates than in C, whereas K treatments decreased (P < 0.05) this response compared with S alone, with no differences between K treatments. Plasma haptoglobin and fibrinogen concentrations were increased (P < 0.05) on d 3 in the castration groups compared with C as the result of tissue trauma induced by castration, whereas SK1 and SK2 had lower (P < 0.05) haptoglobin concentrations than S animals. On d 1, concanavalin A-induced interferon-gamma production was suppressed (P < 0.05) in S and SK3 compared with C, SK1, and SK2 animals. Overall from d 1 to 33, DMI were lower (P < 0.05) in S, SK1, and SK3 than in C animals. From d -1 to 35, ADG were lower (P < 0.05) in S, SK2, and SK3 compared with C animals. A higher (P < 0.05) incidence of standing postures and lower incidence of lying postures was observed in S compared with C during the first 6 h after treatment. However, the higher (P = 0.02) incidence of abnormal standing activities observed for S was reversed (P < 0.05) by the K treatments. In conclusion, surgical castration increased plasma cortisol and acute-phase proteins and decreased immune function, feed intake, and growth rate

  8. 29 CFR 549.2 - Disqualifying provisions.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... OF A âBONA FIDE PROFIT-SHARING PLAN OR TRUSTâ § 549.2 Disqualifying provisions. No plan or trust... fide profit-sharing plan or trust under section 7(e)(3)(b) of the Act: (a) If the share of any... terms of the plan or trust, is set at a predetermined fixed sum or is so limited as to provide in effect...

  9. The decays B → Ψ(2S)π(K),ηc(2S)π(K) in the pQCD approach beyond the leading-order

    NASA Astrophysics Data System (ADS)

    Zhang, Zhi-Qing

    2017-09-01

    Two body B meson decays involving the radially excited meson ψ (2 S) /ηc (2 S) in the final states are studied by using the perturbative QCD (pQCD) approach. We find that: (a) The branching ratios for the decays involving a K meson are predicted as Br (B+ → ψ (2 S)K+) = (5.37-2.22+1.85) ×10-4, Br (B0 → ψ (2 S)K0) = (4.98-2.06+1.71) ×10-4, Br (B+ →ηc (2 S)K+) = (3.54-3.09+3.18) ×10-4, which are consistent with the present data when the next-to-leading-order (NLO) effects are included. Here the NLO effects are from the vertex corrections and the NLO Wilson coefficients. The large errors in the decay B+ →ηc (2 S)K+ are mainly induced by using the decay constant f ηc (2 S) =0.243-0.111+0.079 GeV with large uncertainties. (b) While there seems to be some room left for other higher order corrections or the non-perturbative long distance contributions in the decays involving a π meson, Br (B+ → ψ (2 S)π+) = (1.17-0.50+0.42) ×10-5, Br (B0 → ψ (2 S)π0) =0.54-0.23+0.20 ×10-5, which are smaller than the present data. The results for other decays can be tested via running LHCb and forthcoming Super-B experiments. (c) There is no obvious evidence of the direct CP violation being seen in the decays B → ψ (2 S) π (K) ,ηc (2 S) π (K) in the present experiments, which is supported by our calculations. If a few percent value is confirmed in the future, this would definitely indicate the existence of new physics.

  10. Suppressive effects of the extracts of Japanese edible seaweeds on mutagen-induced umu C gene expression in Salmonella typhimurium (TA 1535/pSK 1002) and tumor promotor-dependent ornithine decarboxylase induction in BALB/c 3T3 fibroblast cells.

    PubMed

    Okai, Y; Higashi-Okai, K; Nakamura, S; Yano, Y; Otani, S

    1994-11-25

    Some of epidemiological data indicated that ubiquitous consumption of seaweeds in Japan may be a possible protective factor against some types of tumor. To analyse this problem, the authors studied the antimutagenic and antitumor promotion activities in methanol-soluble extracts of typical edible seaweeds which showed suppressive effects on 3-amino-1,4-dimethyl-5H-pyrido[4,3-b]indol (Trp-P-1)-induced umu C gene expression in SOS response of Salmonella typhimurium (TA 1535/pSK 1002) and 12-O-tetradecanoylphorbol-13-acetate (TPA)-dependent ornithine decarboxylase induction in BALB/c 3T3 fibroblast cells. Although eight varieties of edible seaweeds including chlorophyta, Phaenophyta and Rhodophyta showed significant antimutagenic and antipromotion activities, they expressed the activities different from each other. Among these seaweeds, Enteromorpha prolifera ('Sujiaonori' in Japanese) and Porphyra tenera ('Asakusanori') showed relatively strong suppressive activities in both antimutagenic and antipromotion assays compared with other seaweeds. These seaweeds contained considerable amounts of beta-carotene as a possible active principle with anticarcinogenic activity. This compound was partially associated with the antimutagenic activity in the seaweed extract, but did not contribute to the antipromotion activity of seaweed extract under our experimental conditions. These results strongly suggest that Japanese edible seaweeds have possible antimutagenic and antipromotion activities probably associated with antitumor activity.

  11. Dual-Labeled Near-Infrared/99mTc Imaging Probes Using PAMAM-Coated Silica Nanoparticles for the Imaging of HER2-Expressing Cancer Cells

    PubMed Central

    Yamaguchi, Haruka; Tsuchimochi, Makoto; Hayama, Kazuhide; Kawase, Tomoyuki; Tsubokawa, Norio

    2016-01-01

    We sought to develop dual-modality imaging probes using functionalized silica nanoparticles to target human epidermal growth factor receptor 2 (HER2)-overexpressing breast cancer cells and achieve efficient target imaging of HER2-expressing tumors. Polyamidoamine-based functionalized silica nanoparticles (PCSNs) for multimodal imaging were synthesized with near-infrared (NIR) fluorescence (indocyanine green (ICG)) and technetium-99m (99mTc) radioactivity. Anti-HER2 antibodies were bound to the labeled PCSNs. These dual-imaging probes were tested to image HER2-overexpressing breast carcinoma cells. In vivo imaging was also examined in breast tumor xenograft models in mice. SK-BR3 (HER2 positive) cells were imaged with stronger NIR fluorescent signals than that in MDA-MB231 (HER2 negative) cells. The increased radioactivity of the SK-BR3 cells was also confirmed by phosphor imaging. NIR images showed strong fluorescent signals in the SK-BR3 tumor model compared to muscle tissues and the MDA-MB231 tumor model. Automatic well counting results showed increased radioactivity in the SK-BR3 xenograft tumors. We developed functionalized silica nanoparticles loaded with 99mTc and ICG for the targeting and imaging of HER2-expressing cells. The dual-imaging probes efficiently imaged HER2-overexpressing cells. Although further studies are needed to produce efficient isotope labeling, the results suggest that the multifunctional silica nanoparticles are a promising vehicle for imaging specific components of the cell membrane in a dual-modality manner. PMID:27399687

  12. Dual-Labeled Near-Infrared/(99m)Tc Imaging Probes Using PAMAM-Coated Silica Nanoparticles for the Imaging of HER2-Expressing Cancer Cells.

    PubMed

    Yamaguchi, Haruka; Tsuchimochi, Makoto; Hayama, Kazuhide; Kawase, Tomoyuki; Tsubokawa, Norio

    2016-07-07

    We sought to develop dual-modality imaging probes using functionalized silica nanoparticles to target human epidermal growth factor receptor 2 (HER2)-overexpressing breast cancer cells and achieve efficient target imaging of HER2-expressing tumors. Polyamidoamine-based functionalized silica nanoparticles (PCSNs) for multimodal imaging were synthesized with near-infrared (NIR) fluorescence (indocyanine green (ICG)) and technetium-99m ((99m)Tc) radioactivity. Anti-HER2 antibodies were bound to the labeled PCSNs. These dual-imaging probes were tested to image HER2-overexpressing breast carcinoma cells. In vivo imaging was also examined in breast tumor xenograft models in mice. SK-BR3 (HER2 positive) cells were imaged with stronger NIR fluorescent signals than that in MDA-MB231 (HER2 negative) cells. The increased radioactivity of the SK-BR3 cells was also confirmed by phosphor imaging. NIR images showed strong fluorescent signals in the SK-BR3 tumor model compared to muscle tissues and the MDA-MB231 tumor model. Automatic well counting results showed increased radioactivity in the SK-BR3 xenograft tumors. We developed functionalized silica nanoparticles loaded with (99m)Tc and ICG for the targeting and imaging of HER2-expressing cells. The dual-imaging probes efficiently imaged HER2-overexpressing cells. Although further studies are needed to produce efficient isotope labeling, the results suggest that the multifunctional silica nanoparticles are a promising vehicle for imaging specific components of the cell membrane in a dual-modality manner.

  13. Apamin increases 5-HT cell firing in raphe dorsalis and extracellular 5-HT levels in amygdala: a concomitant in vivo study in anesthetized rats.

    PubMed

    Crespi, F

    2009-07-24

    The family of calcium-activated slow-potassium (SK) channels comprises 3 members, the SK1, SK2 and SK3 channels, all expressed in neurons, known to mediate the slow-afterhyperpolarization occurring after action potentials. In rats, the SK2 and SK3 channels are expressed in the ascending monoaminergic systems, in particular in the serotonin (5-HT) neurons of the raphe dorsalis nucleus (RDN). In mammals the amygdala, a limbic structure involved in the control of emotion and mood, receives 5-HT-containing projections originating in the RDN. The aim of the present study was to investigate the role of SK channels in mediating the release of 5-HT in the amygdala. Apamin, a polypeptiditic compound with SK2-SK3 channel selectivity, was used to block the channels. A dual probing methodology with Nafion coated carbon-fiber micro-electrode (Nafion-mCFE) was implemented to measure concomitantly the extracellular levels of 5-HT in the amygdala and the firing rate of 5-HT neurons in the RDN of anesthetized rats. Subcutaneous administration of apamin increased both the extracellular 5-HT levels in the amygdala and the firing rate of RDN neurons at doses as low as 12.5 microg. The recorded RDN neurons were of 5-HT phenotype, according to electrophysiologic signature and to the effects observed with peripheral administration of 8-hydroxy-2-(d-n-propyl-amino) tetralin (8-OH-DPAT) a 5-HT(1A) agonist known to selectively reduce the firing of 5-HT neurons in RDN. Increases of extracellular 5-HT levels in the amygdala were also seen when apamin was microinjected into the RDN, suggesting a role for 5-HT neurons of the RDN as target for subcutaneously administered apamin. The confirmation of the involvement of 5-HT neurons projecting from RDN to the amygdala in mediating the effects of apamin was obtained by micro-infusion of tetradotoxine into the bundle of 5-HT ascending fibers located in the region of the posterior amygdala. Attenuation of 5-HT release in the amygdala was observed in

  14. Distribution of OV-TL 3 and MOv18 in normal and malignant ovarian tissue.

    PubMed

    Buist, M R; Molthoff, C F; Kenemans, P; Meijer, C J

    1995-07-01

    To analyse the distribution of OV-TL 3 and MOv18 in normal ovarian tissue to determine which antibody is most suitable for (radio)immunotherapy of ovarian carcinoma. The distribution of OV-TL 3 and MOv18 was determined using immunohistochemistry and flow cytometry. Epithelial and other cells in many tissues, and leucocytes in peripheral blood, bone marrow and spleen stained positively with OV-TL 3. The staining pattern of MOv18 in normal tissues was more restricted and was confined to epithelial cells in the lung, kidney, pancreas, salivary gland, ovary, Fallopian tubes, and cervix. Reactivity was also observed with pneumocytes in the lung, tubuli in the kidney, acinar cells in the salivary gland and pancreas, in the placenta, and with Kupffer cells in the liver. The staining pattern of chimeric MOv18 was identical with the murine form. OV-TL 3 and MOv18 reacted with 100% and 98% (45/46) of the 46 tested epithelial ovarian cancers, respectively. In ovarian carcinoma tissue homogeneous staining of epithelial cells was observed with OV-TL 3 and more heterogeneous staining with MOv18. In 12 and nine patients, respectively, a difference in staining intensity for OV-TL 3 and MOv18 was observed between various tumour samples from the same patient. MOv18 has greater therapeutic potential because of its restricted reactivity with normal tissues and especially, in contrast to OV-TL 3, its lack of reactivity with haematopoietic cells.

  15. Recombinant streptokinase vs hydrocortisone suppositories in acute hemorrhoids: A randomized controlled trial

    PubMed Central

    Hernández-Bernal, Francisco; Castellanos-Sierra, Georgina; Valenzuela-Silva, Carmen M; Catasús-Álvarez, Karem M; Martínez-Serrano, Osmany; Lazo-Diago, Odalys C; Bermúdez-Badell, Cimara H; Causa-García, José R; Domínguez-Suárez, Juan E; Investigators, Pedro A López-Saura; THERESA-4 (Treatment of HEmorrhoids with REcombinant Streptokinase Application) Group of

    2015-01-01

    AIM: To compare the efficacy and safety of recombinant streptokinase (rSK) vs hydrocortisone acetate-based suppositories in acute hemorrhoidal disease. METHODS: A multicenter (11 sites), randomized (1:1:1), open, controlled trial with parallel groups was performed. All participating patients gave their written, informed consent. After inclusion, patients with acute symptoms of hemorrhoids were centrally randomized to receive, as outpatients, by the rectal route, suppositories of rSK 200000 IU of one unit every 8 h (first 3 units) and afterwards every 12 h until 8 administrations were completed (schedule A), one unit every 8 h until 6 units were completed (schedule B), or 25 mg hydrocortisone acetate once every 8 h up to a maximum of 24 administrations. Evaluations were performed at 3, 5, and 10 d post-inclusion. The main end-point was the 5th-day response (disappearance of pain and bleeding, and ≥ 70% reduction of the lesion size). Time to response and need for thrombectomy were secondary efficacy variables. Adverse events were also evaluated. RESULTS: Groups were homogeneous with regards to demographic and baseline characteristics. Fifth day complete response rates were 156/170 (91.8%; 95%CI: 87.3-96.2), 155/170 (91.2%; 95%CI: 86.6%-95.7%), and 46/170 (27.1%; 95%CI: 20.1%-34.0%) with rSK (schedule A and B) and hydrocortisone acetate suppositories, respectively. These 64.6% and 63.9% differences (95%CI: 56.7%-72.2% and 55.7%-72.0%) were highly significant (P < 0.001). This advantage was detected since the early 3rd day evaluation (68.8% and 64.1% vs 7.1% for the rSK and active control groups, respectively; P < 0.001) and was maintained even at the late 10th day assessment (97.1% and 93.5% vs 67.1% for rSK and hydrocortisone acetate, respectively; P < 0.001). Time to response was 3 d (95%CI: 2.9-3.1) for both rSK groups and 10 d (95%CI: 9.3-10.7) in the hydrocortisone acetate group. This difference was highly significant (P < 0.001). All subgroup stratified

  16. Solar neutrino measurements in Super-Kamiokande-IV

    NASA Astrophysics Data System (ADS)

    Abe, K.; Haga, Y.; Hayato, Y.; Ikeda, M.; Iyogi, K.; Kameda, J.; Kishimoto, Y.; Marti, Ll.; Miura, M.; Moriyama, S.; Nakahata, M.; Nakajima, T.; Nakayama, S.; Orii, A.; Sekiya, H.; Shiozawa, M.; Sonoda, Y.; Takeda, A.; Tanaka, H.; Takenaga, Y.; Tasaka, S.; Tomura, T.; Ueno, K.; Yokozawa, T.; Akutsu, R.; Irvine, T.; Kaji, H.; Kajita, T.; Kametani, I.; Kaneyuki, K.; Lee, K. P.; Nishimura, Y.; McLachlan, T.; Okumura, K.; Richard, E.; Labarga, L.; Fernandez, P.; Blaszczyk, F. d. M.; Gustafson, J.; Kachulis, C.; Kearns, E.; Raaf, J. L.; Stone, J. L.; Sulak, L. R.; Berkman, S.; Tobayama, S.; Goldhaber, M.; Bays, K.; Carminati, G.; Griskevich, N. J.; Kropp, W. R.; Mine, S.; Renshaw, A.; Smy, M. B.; Sobel, H. W.; Takhistov, V.; Weatherly, P.; Ganezer, K. S.; Hartfiel, B. L.; Hill, J.; Keig, W. E.; Hong, N.; Kim, J. Y.; Lim, I. T.; Park, R. G.; Akiri, T.; Albert, J. B.; Himmel, A.; Li, Z.; O'Sullivan, E.; Scholberg, K.; Walter, C. W.; Wongjirad, T.; Ishizuka, T.; Nakamura, T.; Jang, J. S.; Choi, K.; Learned, J. G.; Matsuno, S.; Smith, S. N.; Friend, M.; Hasegawa, T.; Ishida, T.; Ishii, T.; Kobayashi, T.; Nakadaira, T.; Nakamura, K.; Nishikawa, K.; Oyama, Y.; Sakashita, K.; Sekiguchi, T.; Tsukamoto, T.; Nakano, Y.; Suzuki, A. T.; Takeuchi, Y.; Yano, T.; Cao, S. V.; Hayashino, T.; Hiraki, T.; Hirota, S.; Huang, K.; Ieki, K.; Jiang, M.; Kikawa, T.; Minamino, A.; Murakami, A.; Nakaya, T.; Patel, N. D.; Suzuki, K.; Takahashi, S.; Wendell, R. A.; Fukuda, Y.; Itow, Y.; Mitsuka, G.; Muto, F.; Suzuki, T.; Mijakowski, P.; Frankiewicz, K.; Hignight, J.; Imber, J.; Jung, C. K.; Li, X.; Palomino, J. L.; Santucci, G.; Taylor, I.; Vilela, C.; Wilking, M. J.; Yanagisawa, C.; Fukuda, D.; Ishino, H.; Kayano, T.; Kibayashi, A.; Koshio, Y.; Mori, T.; Sakuda, M.; Takeuchi, J.; Yamaguchi, R.; Kuno, Y.; Tacik, R.; Kim, S. B.; Okazawa, H.; Choi, Y.; Ito, K.; Nishijima, K.; Koshiba, M.; Totsuka, Y.; Suda, Y.; Yokoyama, M.; Bronner, C.; Calland, R. G.; Hartz, M.; Martens, K.; Obayashi, Y.; Suzuki, Y.; Vagins, M. R.; Nantais, C. M.; Martin, J. F.; de Perio, P.; Tanaka, H. A.; Konaka, A.; Chen, S.; Sui, H.; Wan, L.; Yang, Z.; Zhang, H.; Zhang, Y.; Connolly, K.; Dziomba, M.; Wilkes, R. J.; Super-Kamiokande Collaboration

    2016-09-01

    Upgraded electronics, improved water system dynamics, better calibration and analysis techniques allowed Super-Kamiokande-IV to clearly observe very low-energy 8B solar neutrino interactions, with recoil electron kinetic energies as low as ˜3.5 MeV . Super-Kamiokande-IV data-taking began in September of 2008; this paper includes data until February 2014, a total livetime of 1664 days. The measured solar neutrino flux is (2.308 ±0.020 (stat)-0.040 +0.039(syst ))×1 06/(cm2 sec ) assuming no oscillations. The observed recoil electron energy spectrum is consistent with no distortions due to neutrino oscillations. An extended maximum likelihood fit to the amplitude of the expected solar zenith angle variation of the neutrino-electron elastic scattering rate in SK-IV results in a day/night asymmetry of (-3.6 ±1.6 (stat )±0.6 (syst ))% . The SK-IV solar neutrino data determine the solar mixing angle as sin2θ12=0.327-0.031+0.026 , all SK solar data (SK-I, SK-II, SK III and SK-IV) measures this angle to be sin2θ12=0.334-0.023+0.027 , the determined mass-squared splitting is Δ m212=4.8-0.8+1.5×10-5 eV2 .

  17. Proximal tubule sphingosine kinase-1 has a critical role in A1 adenosine receptor-mediated renal protection from ischemia

    PubMed Central

    Park, Sang Won; Kim, Mihwa; Kim, Joo Yun; Brown, Kevin M.; Haase, Volker H.; D’Agati, Vivette D.; Lee, H. Thomas

    2012-01-01

    Renal ischemia reperfusion injury is a major cause of acute kidney injury. We previously found that renal A1 adenosine receptor (A1AR) activation attenuated multiple cell death pathways including necrosis, apoptosis and inflammation. Here, we tested whether induction of cytoprotective sphingosine kinase (SK)-1 and sphingosine-1 phosphate (S1P) synthesis might be the mechanism of protection. A selective A1AR agonist (CCPA) increased the synthesis of S1P and selectively induced SK-1 in mouse kidney and HK-2 cells. This agonist failed to protect SK1-knockout but protected SK2-knockout mice against renal ischemia reperfusion injury indicating a critical role of SK1 in A1AR-mediated renal protection. Inhibition of SK prevented A1AR-mediated defense against necrosis and apoptosis in HK-2 cells. A selective S1P1R antagonist (W146) and global in vivo gene knockdown of S1P1Rs with small interfering RNA completely abolished the renal protection provided by CCPA. Mice selectively deficient in renal proximal tubule S1P1Rs (S1P1Rflox/flox PEPCKCre/−) were not protected against renal ischemia reperfusion injury by CCPA. Mechanistically, CCPA increased nuclear translocation of hypoxia inducible factor-1α in HK-2 cells and selective hypoxia inducible factor-1α inhibition blocked A1AR-mediated induction of SK1. Thus, proximal tubule SK-1 has a critical role in A1AR-mediated protection against renal ischemia reperfusion injury. PMID:22695326

  18. Evaluation of fluoren-NU as a novel antitumor agent.

    PubMed

    Mukherjee, Asama; Dutta, Sushanta; Chashoo, Gousia; Bhagat, Madhulika; Saxena, Ajit Kumar; Sanyal, Utpal

    2009-01-01

    A new nitrososourea derivative, namely fluoren-NU, 3-[2-(3-(2-chloroethyl)-3-nitrosouriedo}ethyl]-spiro[5,9'-fluorenyl]imidazolidine-2,4-dione (compound 2e), was synthesized from 3-(2-bromoethyl)-spiro [5,9'-fluorenyl]imidazolidine-2,4-dione via a four-step synthetic procedure. Its chemical alkylating activity was assessed by coupling with 4-(4-nitrobenzyl)pyridine. In vitro screening in six human tumor cell lines, namely SK-N-SH CNS, IMR-32 neuroblastoma, A549 lung, DU-145 prostate, HL-60 leukemia, and U-937 lymphoma, revealed its significant cytotoxicity in SK-N-SH. Its in vivo antitumoral potency was assessed in murine ascites tumors Ehrlich ascites carcinoma (EAC) and Sarcoma-180 (S-180) by measuring the increase in median survival times (MST) of drug-treated (T) over untreated control (C) mice. Results revealed significant tumor regression effects in both of these tumors. Life span of mice bearing advanced tumor for 5 days before the drug challenge was also considerably increased. In vivo toxicological assay at its optimum dose of 40 mg/kg for days 1-7 treatment schedule was conducted sequentially on day 9, 14, and 19 in normal and EAC-bearing mice. Results revealed that it did not adversely affect hematopoiesis or exhibit drug-induced hepatotoxicity and nephrotoxicity. It has shown minimal cytotoxic effect on human peripheral blood mononuclear cells (PBMC) having a high IC50 value of 792 microM. Compared to Mitonafide and CCNU used as standards it also significantly inhibited DNA and RNA synthesis in EAC tumor cells in vitro at 8 microM concentration.

  19. A Small Potassium Current in AgRP/NPY Neurons Regulates Feeding Behavior and Energy Metabolism.

    PubMed

    He, Yanlin; Shu, Gang; Yang, Yongjie; Xu, Pingwen; Xia, Yan; Wang, Chunmei; Saito, Kenji; Hinton, Antentor; Yan, Xiaofeng; Liu, Chen; Wu, Qi; Tong, Qingchun; Xu, Yong

    2016-11-08

    Neurons that co-express agouti-related peptide (AgRP) and neuropeptide Y (NPY) are indispensable for normal feeding behavior. Firing activities of AgRP/NPY neurons are dynamically regulated by energy status and coordinate appropriate feeding behavior to meet nutritional demands. However, intrinsic mechanisms that regulate AgRP/NPY neural activities during the fed-to-fasted transition are not fully understood. We found that AgRP/NPY neurons in satiated mice express high levels of the small-conductance calcium-activated potassium channel 3 (SK3) and are inhibited by SK3-mediated potassium currents; on the other hand, food deprivation suppresses SK3 expression in AgRP/NPY neurons, and the decreased SK3-mediated currents contribute to fasting-induced activation of these neurons. Genetic mutation of SK3 specifically in AgRP/NPY neurons leads to increased sensitivity to diet-induced obesity, associated with chronic hyperphagia and decreased energy expenditure. Our results identify SK3 as a key intrinsic mediator that coordinates nutritional status with AgRP/NPY neural activities and animals' feeding behavior and energy metabolism. Copyright © 2016 The Author(s). Published by Elsevier Inc. All rights reserved.

  20. Contamination of Runoff Water at Gdańsk Airport (Poland) by Polycyclic Aromatic Hydrocarbons (PAHs) and Polychlorinated Biphenyls (PCBs)

    PubMed Central

    Sulej, Anna Maria; Polkowska, Żaneta; Namieśnik, Jacek

    2011-01-01

    Airport runoff can contain high concentrations of various pollutants, in particular polycyclic aromatic hydrocarbons (PAHs) and polychlorinated biphenyls (PCBs), the environmental levels of which have to be monitored. Airport runoff water samples, collected at the Gdańsk-Rębiechowo Airport from 2008 to 2009, were analysed for PAHs and PCBs by gas chromatography. The aromatic fractions were separated by liquid-liquid extraction and analysed by GC/MS. Total PAH concentrations were 295–6,758 ng/L in 2008 and 180–1,924 ng/L in 2009, while total PCB levels in 2008 ranged from 0.14 to 0.44 μg/L and in 2009 from 0.06 to 0.23 μg/L. The PAH and PCB compositions in airport runoff waters were examined over a range of spatial and temporal scales to determine distributions, trends and possible sources. This pollution is mainly pyrolytic and related to anthropogenic activity. There were significant differences between the samples collected in the two seasons. An understanding of the magnitude of contamination due to airport runoff water is important for the effective management of airport infrastructure. PMID:22247699

  1. Pyridostigmine but not 3,4-diaminopyridine exacerbates ACh receptor loss and myasthenia induced in mice by muscle-specific kinase autoantibody

    PubMed Central

    Morsch, Marco; Reddel, Stephen W; Ghazanfari, Nazanin; Toyka, Klaus V; Phillips, William D

    2013-01-01

    In myasthenia gravis, the neuromuscular junction is impaired by the antibody-mediated loss of postsynaptic acetylcholine receptors (AChRs). Muscle weakness can be improved upon treatment with pyridostigmine, a cholinesterase inhibitor, or with 3,4-diaminopyridine, which increases the release of ACh quanta. The clinical efficacy of pyridostigmine is in doubt for certain forms of myasthenia. Here we formally examined the effects of these compounds in the antibody-induced mouse model of anti-muscle-specific kinase (MuSK) myasthenia gravis. Mice received 14 daily injections of IgG from patients with anti-MuSK myasthenia gravis. This caused reductions in postsynaptic AChR densities and in endplate potential amplitudes. Systemic delivery of pyridostigmine at therapeutically relevant levels from days 7 to 14 exacerbated the anti-MuSK-induced structural alterations and functional impairment at motor endplates in the diaphragm muscle. No such effect of pyridostigmine was found in mice receiving control human IgG. Mice receiving smaller amounts of MuSK autoantibodies did not display overt weakness, but 9 days of pyridostigmine treatment precipitated generalised muscle weakness. In contrast, one week of treatment with 3,4-diaminopyridine enhanced neuromuscular transmission in the diaphragm muscle. Both pyridostigmine and 3,4-diaminopyridine increase ACh in the synaptic cleft yet only pyridostigmine potentiated the anti-MuSK-induced decline in endplate ACh receptor density. These results thus suggest that ongoing pyridostigmine treatment potentiates anti-MuSK-induced AChR loss by prolonging the activity of ACh in the synaptic cleft. PMID:23440963

  2. [Maria Skłodowska-Curie and Piotr Curie an epoch-makingin year 1898].

    PubMed

    Wielogórski, Zbigniew

    2012-01-01

    For many reasons the year 1898 was unusual for Maria Skłodowska-Curie and her husband. After defining the subject of the doctoral thesis and choosing Henri Becqerel as thesis supervisor, Maria started intensive experimental work. In the allotted room called storeroom, in conditions that were far too inadequate, they managed to put up a unique measuring equipment composed of instruments whose originator was Pierre Curie. In the ionization chamber and in the piezoelectric quartz charges formed, whose mutual neutralization was shown by the quadrant electrometer. Ionization current, which was measured quantitatively, was proportional to the radiation of the sample. Studying many elements, their compounds and minerals enabled Maria to state that uranium is not the only element endowed with the power of radiation; the second one turned out to be thorium. Anomaly detected in the radiation of uranium minerals made it possible for Maria to draw an extremely important conclusion: radioactive uranium and thorium are not the only elements endowed with such an attribute. Pitchblende, which was studied by the Curie couple, had to contain also other radioactive substances. Gustave Bémont also participated in the chemical analysis of the uranium ore and it is worth reminding that he was involved in the discovery of polonium and uranium. The phenomenon of radioactivity couldn't have been explained if it was not for the sources of strong radioactivity. Those sources undoubtedly could have been the discovered elements but their scanty content in the uranium ore made their isolation very difficult and laborious. Access to industrial remains after procession of pitchblende from Jachymov (Sankt Joachimstahl), obtained owing to the mediation of Eduard Suess, provided the source of this raw material. From it, in a shack also called le hangar, the Curie couple isolated the first samples of the radium salt. This element, later extracted by discoverers on a grand scale and handed over in a

  3. Using clinical symptoms to predict adverse maternal and perinatal outcomes in women with preeclampsia: data from the PIERS (Pre-eclampsia Integrated Estimate of RiSk) study.

    PubMed

    Yen, Tin-Wing; Payne, Beth; Qu, Ziguang; Hutcheon, Jennifer A; Lee, Tang; Magee, Laura A; Walters, Barry N; von Dadelszen, Peter

    2011-08-01

    Preeclampsia is a leading cause of maternal morbidity. The clinical challenge lies in predicting which women with preeclampsia will suffer adverse outcomes and would benefit from treatment, while minimizing potentially harmful interventions. Our aim was to determine the ability of maternal symptoms (i.e., severe nausea or vomiting, headache, visual disturbance, right upper quadrant pain or epigastric pain, abdominal pain or vaginal bleeding, and chest pain or dyspnea) to predict adverse maternal or perinatal outcomes. We used data from the PIERS (Pre-eclampsia Integrated Estimate of RiSk) study, a multicentre, prospective cohort study designed to investigate the maternal risks associated with preeclampsia. Relative risks and receiver operating characteristic (ROC) curves were assessed for each preeclampsia symptom and outcome pair. Of 2023 women who underwent assessment, 52% experienced at least one preeclampsia symptom, with 5.2% and 5.3% respectively experiencing an adverse maternal or perinatal outcome. No symptom and outcome pair, in either of the maternal or perinatal groups, achieved an area under the ROC curve value > 0.7, which would be necessary to demonstrate a discriminatory predictive value. Maternal symptoms of preeclampsia are not independently valid predictors of maternal adverse outcome. Caution should be used when making clinical decisions on the basis of symptoms alone in the preeclamptic patient.

  4. Presence of gonadotropin-releasing hormone and its messenger ribonucleic acid in human ovarian epithelial carcinoma.

    PubMed

    Ohno, T; Imai, A; Furui, T; Takahashi, K; Tamaya, T

    1993-09-01

    The purpose of this study was to investigate the expression of gonadotropin-releasing hormone messenger ribonucleic acid and the presence of gonadotropin-releasing hormone in human ovarian carcinoma known to have gonadotropin-releasing hormone binding sites and to be affected by gonadotropin-releasing hormone analog. Human ovarian carcinomas surgically removed and human ovarian carcinoma cell lines were examined. Gonadotropin-releasing hormone was determined by a radioimmunoassay and a bioassay. Gonadotropin-releasing hormone messenger ribonucleic acid was determined by reverse transcription polymerase chain reaction using oligonucleotide primers synthesized according to the published human gonadotropin-releasing hormone sequence. Gonadotropin-releasing hormone was shown to be present in extracts of ovarian mucinous cystadenocarcinoma sample (0.8 +/- 0.12 pg/mg of protein) and ovarian adenocarcinoma cell line SK-OV3 (0.92 +/- 0.17 pg/mg of protein) but not in the normal ovary and placenta. Two of two extract samples from individual cases evoked dose-dependent phosphoinositide breakdown in rat granulosa cells similar to that caused by authentic gonadotropin-releasing hormone. Gonadotropin-releasing hormone messenger ribonucleic acid was detected in two of two mucinous cystadenocarcinoma specimens, one of one serous cystadenocarcinoma, and SK-OV3 cells but not in the dysgerminoma, mucinous cystadenoma, and normal ovary and placenta. The demonstration of gonadotropin-releasing hormone and its messenger ribonucleic acid raises the possibility that gonadotropin-releasing hormone may play an autocrine regulatory role in the growth of ovarian carcinoma.

  5. Protein Phosphatase 2A Signaling in Human Prostate Cancer

    DTIC Science & Technology

    2012-06-01

    analyses using specific antibodies . As expected, our sata showed a decreased phosphorylation of Akt and ERK in PPP2CA-overexpressing C4-2 and PC-3...Prostate 2006;66:1631-40. (7) Srivastava SK, Bhardwaj A, Singh S, Arora S, Wang B, Grizzle WE, et al. MicroRNA-150 directly targets MUC4 and

  6. Associations between cognitive abilities and life satisfaction in the oldest-old. Results from the longitudinal population study Good Aging in Skåne

    PubMed Central

    Enkvist, Åsa; Ekström, Henrik; Elmståhl, Sölve

    2013-01-01

    Introduction Studies on the associations between cognitive abilities and life satisfaction (LS) in the oldest-old are few. The aim of this study was to explore whether abilities in six different cognitive domains could predict LS in the oldest-old 3 years later. Methods The study population consisted of 681 individuals aged 78–98 years, drawn from the longitudinal population study “Good Aging in Skåne,” which is part of a national survey (The Swedish National Study on Aging and Care). Scores on 13 cognitive tests were related to scores on Neugartens’ LS index A (LSI-A) 3 years later. The cognitive tests were added into six different cognitive domains. A multiple regression analysis was constructed for each cognitive domain separately, with scores on the LSI-A as the dependent variable. The model was adjusted stepwise for sex, age, education, functional capacity, and depressive mood. Results Significant correlations were found between digit cancellation, word recall, verbal fluency (VF) A, VF animals, VF occupations, and mental rotations at baseline, as well as LSI-A at follow-up. The domains of spatial abilities (B = 0.453, P = 0.014) and processing speed (B = 0.118, P = 0.020) remained significantly associated with LSI-A 3 years later after adjustment. Conclusion The cognitive domains of spatial abilities and processing speed predicted LS 3 years later in the oldest-old. Clinical implications are discussed. PMID:23874091

  7. Effect of specific activity on neuroblastoma uptake of I-123-meta-iodobenzylguanidine in nude mice xenografted with SK-N-SH cells.

    PubMed

    Farahati, J; Coenen, H; Dutschka, K; Stuben, G; Knuhmann, K; Budach, W; Kremens, B; Reiners, C

    1997-01-01

    The effect of specific activity of meta[I-123]iodobenzylguanidine ([I-123]MIBG) on neuroblastoma uptake was studied in a nude mouse model (NMRI nu/nu) xenografted subcutaneously with SK-N-SH cells. Groups of eight animals received [I-123]MIBG intravenously with a specific activity of greater than or equal to 260 GBq/mu mol (no-carrier-added), 3.7 GBq/mu mol, 37 MBq/mu mol, and 0.37 MBq/mu mol, respectively. All animals in the group injected with 0.37 MBq/mu mol died immediately after the injection. Al 4 and 24 h, there was no significant effect of specific activity on tumor uptake of [I-123]MIBG in the different groups. The uptake of non-tumor tissue was in general lower with 37 MBq/mu mol compared to higher specific activities. The differences in blood, heart, liver, spleen and lungs were statistically significant at 24 h, whereas at 4 h significant differences were only present in the heart, liver and lungs. The results suggest that for the treatment of children with neuroblastoma a lower specific activity of radioiodinated MIBG may minimize the radiation exposure to non-tumor tissue but not to the tumor. Higher mass of MIBG >0.5 mu mol/g, however, is considered as lethal dose in our nude mice model and corresponding doses may cause toxic side effects in human.

  8. Inhibition of cell growth and induction of apoptosis in ovarian carcinoma cell lines CaOV3 and SKOV3 by natural withanolide Withaferin A.

    PubMed

    Zhang, Xuan; Samadi, Abbas K; Roby, Katherine F; Timmermann, Barbara; Cohen, Mark S

    2012-03-01

    Withaferin A, a natural withanolide, has shown anti-cancer properties in various cancers including breast cancer, but its effects in ovarian cancer remain unexplored. Notch 1 and Notch3 are critically involved in ovarian cancer progression. We decided to examine the effects of Withaferin A in ovarian carcinoma cell lines and its molecular mechanism of action including its regulation of Notch. The effects of Withaferin A were examined in CaOV3 and SKOV3 ovarian carcinoma cell lines using MTS assay, clonogenic assay, annexin V/propidium iodide flow cytometry, and cell cycle analysis. Western analysis was conducted to examine the molecular mechanisms of action. Withaferin A inhibited the growth and colony formation of CaOV3 and SKOV3 cells by inducing apoptosis and cell cycle arrest. These changes correlated with down-regulation of Notch1, Notch3, cdc25C, total and phosphorylated Akt, and bcl-2 proteins. Withaferin A inhibits CaOV3 and SKOV3 ovarian carcinoma cell growth, at least in part by targeting Notch1 and Notch3. Copyright © 2011 Elsevier Inc. All rights reserved.

  9. A Rare Early-type Star Revealed in the Wing of the Small Magellanic Cloud

    NASA Astrophysics Data System (ADS)

    Evans, C. J.; Hainich, R.; Oskinova, L. M.; Gallagher, J. S., III; Chu, Y.-H.; Gruendl, R. A.; Hamann, W.-R.; Hénault-Brunet, V.; Todt, H.

    2012-07-01

    Sk 183 is the visually brightest star in the N90 nebula, a young star-forming region in the Wing of the Small Magellanic Cloud (SMC). We present new optical spectroscopy from the Very Large Telescope which reveals Sk 183 to be one of the most massive O-type stars in the SMC. Classified as an O3-type dwarf on the basis of its nitrogen spectrum, the star also displays broadened He I absorption, which suggests a later type. We propose that Sk 183 has a composite spectrum and that it is similar to another star in the SMC, MPG 324. This brings the number of rare O2- and O3-type stars known in the whole of the SMC to a mere four. We estimate physical parameters for Sk 183 from analysis of its spectrum. For a single-star model, we estimate an effective temperature of 46 ± 2 kK, a low mass-loss rate of ~10-7 M ⊙ yr-1, and a spectroscopic mass of 46+9 -8 M ⊙ (for an adopted distance modulus of 18.7 mag to the young population in the SMC Wing). An illustrative binary model requires a slightly hotter temperature (~47.5 kK) for the primary component. In either scenario, Sk 183 is the earliest-type star known in N90 and will therefore be the dominant source of hydrogen-ionizing photons. This suggests Sk 183 is the primary influence on the star formation along the inner edge of the nebula.

  10. Twelve actin-encoding cDNAs from the American lobster, Homarus americanus: cloning and tissue expression of eight skeletal muscle, one heart, and three cytoplasmic isoforms.

    PubMed

    Kim, Bo Kwang; Kim, Kyoung Sun; Oh, Chul-Woong; Mykles, Donald L; Lee, Sung Gu; Kim, Hak Jun; Kim, Hyun-Woo

    2009-06-01

    Lobster muscles express a diverse array of myofibrillar protein isoforms. Three fiber types (fast, slow-twitch or S1, and slow-tonic or S2) differ qualitatively and quantitatively in myosin heavy and light chains, troponin-T, -I, and -C, paramyosin, and tropomyosin variants. However, little is known about the diversity of actin isoforms present in crustacean tissues. In this report we characterized cDNAs that encode twelve actin isoforms in the American lobster, Homarus americanus: eight from skeletal muscle (Ha-ActinSK1-8), one from heart (Ha-ActinHT1), and three cytoplasmic type actins from hepatopancreas (Ha-ActinCT1-3). All twelve cDNAs were products of distinct genes, as indicated by differences in the 3'-untranslated regions (UTRs). The open reading frames specified polypeptides 376 or 377 amino acids in length. Although key amino residues are conserved in the lobster actins, variations in nearby sequences may affect actin polymerization and/or interactions with other myofibrillar proteins. Quantitative reverse transcription-polymerase chain reaction showed muscle fiber type- and tissue-specific expression patterns. Ha-Actin-HT1 was expressed exclusively in heart (87% of the total; 12% of the total was Ha-ActinCT1). Ha-ActinCT1 was expressed in all tissues, while CT2 and CT3 were expressed only in hepatopancreas, with Ha-ActinCT2 as the major isoform (93% of the total). Ha-ActinSK1 and SK2 were the major isoforms (88% and 12% of the total, respectively) in the S1 fibers of crusher claw closer muscle. Fast fibers in the cutter claw closer and deep abdominal muscles differed in SK isoforms. Ha-ActinSK3, SK4, and SK5 were the major isoforms in cutter claw closer muscle (12%, 48%, and 37% of the total, respectively). Ha-ActinSK5 and SK8 were the major isoforms in deep abdominal flexor (31% and 65% of the total, respectively) and extensor (46% and 53% of the total, respectively) muscles, with SK6 and SK7 expressed at low levels. These data indicate that fast

  11. On the star partition dimension of comb product of cycle and path

    NASA Astrophysics Data System (ADS)

    Alfarisi, Ridho; Darmaji

    2017-08-01

    Let G = (V, E) be a connected graphs with vertex set V(G), edge set E(G) and S ⊆ V(G). Given an ordered partition Π = {S1, S2, S3, …, Sk} of the vertex set V of G, the representation of a vertex v ∈ V with respect to Π is the vector r(v|Π) = (d(v, S1), d(v, S2), …, d(v, Sk)), where d(v, Sk) represents the distance between the vertex v and the set Sk and d(v, Sk) = min{d(v, x)|x ∈ Sk }. A partition Π of V(G) is a resolving partition if different vertices of G have distinct representations, i.e., for every pair of vertices u, v ∈ V(G), r(u|Π) ≠ r(v|Π). The minimum k of Π resolving partition is a partition dimension of G, denoted by pd(G). The resolving partition Π = {S1, S2, S3, …, Sk } is called a star resolving partition for G if it is a resolving partition and each subgraph induced by Si, 1 ≤ i ≤ k, is a star. The minimum k for which there exists a star resolving partition of V(G) is the star partition dimension of G, denoted by spd(G). Finding the star partition dimension of G is classified to be a NP-Hard problem. In this paper, we will show that the partition dimension of comb product of cycle and path namely Cm⊳Pn and Pn⊳Cm for n ≥ 2 and m ≥ 3.

  12. Suppressive effects of retinoids, carotenoids and antioxidant vitamins on heterocyclic amine-induced umu C gene expression in Salmonella typhimurium (TA 1535/pSK 1002).

    PubMed

    Okai, Y; Higashi-Okai, K; Nakamura, S; Yano, Y; Otani, S

    1996-06-12

    Effects of retinoids, carotenoids and antioxidant vitamins were studied by mutagen-induced umu C gene expression system in Salmonella typhimurium (TA 1535/pSK 1002). Retinol (vitamin A), retinol acetate and retinoic acid showed remarkable inhibitory activities, whereas retinol palmitate exhibited significant but weak activity for umu C gene expression in tester bacteria induced by 3-amino-3,4-dimethyl-5H-pyrido[4.3-b]indol (Trp-P-1) in the presence of hepatic metabolizing enzymes (S9 mixture). Carotenoids having provitamin A activity (beta-carotene and canthaxanthin) exhibited moderate suppressive effects on the same experimental system. The ranks of suppressive activities were retinol > retinol acetate > retinoic acid > canthaxanthin > beta-carotene > retinol palmitate and their doses for inhibition by 50% (ID50) were estimated to be 1.2 x 10(-7), 3.0 x 10(-7), 5.4 x 10(-7), 1.5 x 10(-6), 4.0 x 10(-5) and 6.0 x 10(-5) M, respectively. However, they did not cause significant inhibition on umu C gene expression induced by direct-acting mutagen (adriamycin or mitomycin C) in the absence of S9 mixture. Inhibition of umu gene expression appears to be due to inhibition of P450-mediated metabolic activation of the heterocyclic amine Trp-P-1. Ascorbic acid (vitamin C) showed weak but significant suppressive activity at high-dose concentrations (3 x 10(-6) - 10(-4)M). However, alpha-tocopherol did not exhibit significant suppression at all dose concentrations. The significance of the experimental results is discussed from the viewpoint of the chemoprevention against genotoxicity associated with carcinogenesis.

  13. Improved outcome of 131I-mIBG treatment through combination with external beam radiotherapy in the SK-N-SH mouse model of neuroblastoma.

    PubMed

    Corroyer-Dulmont, Aurélien; Falzone, Nadia; Kersemans, Veerle; Thompson, James; Allen, Danny P; Able, Sarah; Kartsonaki, Christiana; Malcolm, Javian; Kinchesh, Paul; Hill, Mark A; Vojnovic, Boris; Smart, Sean C; Gaze, Mark N; Vallis, Katherine A

    2017-09-01

    To assess the efficacy of different schedules for combining external beam radiotherapy (EBRT) with molecular radiotherapy (MRT) using 131 I-mIBG in the management of neuroblastoma. BALB/c nu/nu mice bearing SK-N-SH neuroblastoma xenografts were assigned to five treatment groups: 131 I-mIBG 24h after EBRT, EBRT 6days after 131 I-mIBG, EBRT alone, 131 I-mIBG alone and control (untreated). A total of 56 mice were assigned to 3 studies. Study 1: Vessel permeability was evaluated using dynamic contrast-enhanced (DCE)-MRI (n=3). Study 2: Tumour uptake of 131 I-mIBG in excised lesions was evaluated by γ-counting and autoradiography (n=28). Study 3: Tumour volume was assessed by longitudinal MR imaging and survival was analysed (n=25). Tumour dosimetry was performed using Monte Carlo simulations of absorbed fractions with the radiation transport code PENELOPE. Given alone, both 131 I-mIBG and EBRT resulted in a seven-day delay in tumour regrowth. Following EBRT, vessel permeability was evaluated by DCE-MRI and showed an increase at 24h post irradiation that correlated with an increase in 131 I-mIBG tumour uptake, absorbed dose and overall survival in the case of combined treatment. Similarly, EBRT administered seven days after MRT to coincide with tumour regrowth, significantly decreased the tumour volume and increased overall survival. This study demonstrates that combining EBRT and MRT has an enhanced therapeutic effect and emphasizes the importance of treatment scheduling according to pathophysiological criteria such as tumour vessel permeability and tumour growth kinetics. Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.

  14. Photochemical activation of MH3-B1/rGel: a HER2-targeted treatment approach for ovarian cancer

    PubMed Central

    Bull-Hansen, Bente; Berstad, Maria B.; Berg, Kristian; Cao, Yu; Skarpen, Ellen; Fremstedal, Ane Sofie; Rosenblum, Michael G.; Peng, Qian; Weyergang, Anette

    2015-01-01

    HER2-targeted therapy has been shown to have limited efficacy in ovarian cancer despite frequent overexpression of this receptor. Photochemical internalization (PCI) is a modality for cytosolic drug delivery, currently undergoing clinical evaluation. In the present project we studied the application of PCI in combination with the HER2-targeted recombinant fusion toxin, MH3-B1/rGel, for the treatment of ovarian cancer. The SKOV-3 cell line, resistant to trastuzumab- and MH3-B1/rGel- monotherapy, was shown to respond strongly to PCI of MH3-B1/rGel to a similar extent as observed for the treatment-sensitive SK-BR-3 breast cancer cells. Extensive hydrolytic degradation of MH3-B1/rGel in acidic endocytic vesicles was indicated as the mechanism of MH3-B1/rGel resistance in SKOV-3 cells. This was shown by the positive Pearson's correlation coefficient between Alexa488-labeled MH3-B1/rGel and Lysotracker in SKOV-3 cells in contrast to the negative Pearson's correlation coefficient in SK-BR-3 cells. The application of PCI to induce the release of MH3-B1/rGel was also demonstrated to be effective on SKOV-3 xenografts. Application of PCI with MH3-B1/rGel was further found highly effective in the HER2 expressing HOC-7 and NuTu-19 ovarian cancer cell lines. The presented results warrant future development of PCI in combination with MH3-B1/rGel as a novel therapeutic approach in preclinical models of ovarian cancer. PMID:26002552

  15. On the partition dimension of comb product of path and complete graph

    NASA Astrophysics Data System (ADS)

    Darmaji, Alfarisi, Ridho

    2017-08-01

    For a vertex v of a connected graph G(V, E) with vertex set V(G), edge set E(G) and S ⊆ V(G). Given an ordered partition Π = {S1, S2, S3, …, Sk} of the vertex set V of G, the representation of a vertex v ∈ V with respect to Π is the vector r(v|Π) = (d(v, S1), d(v, S2), …, d(v, Sk)), where d(v, Sk) represents the distance between the vertex v and the set Sk and d(v, Sk) = min{d(v, x)|x ∈ Sk}. A partition Π of V(G) is a resolving partition if different vertices of G have distinct representations, i.e., for every pair of vertices u, v ∈ V(G), r(u|Π) ≠ r(v|Π). The minimum k of Π resolving partition is a partition dimension of G, denoted by pd(G). Finding the partition dimension of G is classified to be a NP-Hard problem. In this paper, we will show that the partition dimension of comb product of path and complete graph. The results show that comb product of complete grapph Km and path Pn namely p d (Km⊳Pn)=m where m ≥ 3 and n ≥ 2 and p d (Pn⊳Km)=m where m ≥ 3, n ≥ 2 and m ≥ n.

  16. Specific and nonspecific immunoassays to detect HAMA after administration of indium-111-labeled OV-TL 3 F(ab')2 monoclonal antibody to patients with ovarian cancer.

    PubMed

    Massuger, L F; Thomas, C M; Segers, M F; Corstens, F H; Verheijen, R H; Kenemans, P; Poels, L G

    1992-11-01

    The development of human anti-mouse antibodies (HAMA) may cause problems in radioimmunotargeting studies, but may also improve survival of patients. To identify the presence of HAMA in blood samples from patients intravenously injected with 1 mg of 111In-labeled OV-TL3-F(ab')2, we developed three specific OV-TL 3-based HAMA assays and tested these along with two commercially available nonspecific HAMA assays (Sorin and Immunomedics). The specific assays were positive for HAMA with 10 postinjection serum samples from 7 patients. Eight of the 10 samples were also HAMA positive with one or both nonspecific HAMA assays. Conflicting results were observed with half the number of samples. The two nonspecific assays also reacted positively with another 11 serum samples from 5 patients including their preinjection samples. Despite some contradictory results, the nonspecific HAMA assays identify both pre-existent and Mab-induced HAMA, whereas the specific OV-TL3-based HAMA assays identify specific immune-responses occurring after the OV-TL 3 injection.

  17. Oxoaporphine Metal Complexes (CoII, NiII, ZnII) with High Antitumor Activity by Inducing Mitochondria-Mediated Apoptosis and S-phase Arrest in HepG2

    PubMed Central

    Qin, Jiao-Lan; Shen, Wen-Ying; Chen, Zhen-Feng; Zhao, Li-Fang; Qin, Qi-Pin; Yu, Yan-Cheng; Liang, Hong

    2017-01-01

    Three new oxoaporphine Co(II), Ni(II) and Zn(II) complexes 1–3 have been synthesized and fully characterized. 1–3 have similar mononuclear structures with the metal and ligand ratio of 1:2. 1–3 exhibited higher cytotoxicity than the OD ligand and cisplatin against HepG2, T-24, BEL-7404, MGC80–3 and SK-OV-3/DDP cells, with IC50 value of 0.23−4.31 μM. Interestingly, 0.5 μM 1–3 significantly caused HepG2 arrest at S-phase, which was associated with the up-regulation of p53, p21, p27, Chk1 and Chk2 proteins, and decrease in cyclin A, CDK2, Cdc25A, PCNA proteins. In addition, 1–3 induced HepG2 apoptosis via a caspase-dependent mitochondrion pathway as evidenced by p53 activation, ROS production, Bax up-regulation and Bcl-2 down-regulation, mitochondrial dysfunction, cytochrome c release, caspase activation and PARP cleavage. Furthermore, 3 inhibited tumor growth in HepG2 xenograft model, and displayed more safety profile in vivo than cisplatin. PMID:28436418

  18. Oncogenic PIK3CA mutations occur in epidermal nevi and seborrheic keratoses with a characteristic mutation pattern

    PubMed Central

    Hafner, Christian; López-Knowles, Elena; Luis, Nuno M.; Toll, Agustí; Baselga, Eulàlia; Fernández-Casado, Alex; Hernández, Silvia; Ribé, Adriana; Mentzel, Thomas; Stoehr, Robert; Hofstaedter, Ferdinand; Landthaler, Michael; Vogt, Thomas; Pujol, Ramòn M.; Hartmann, Arndt; Real, Francisco X.

    2007-01-01

    Activating mutations of the p110 α subunit of PI3K (PIK3CA) oncogene have been identified in a broad spectrum of malignant tumors. However, their role in benign or preneoplastic conditions is unknown. Activating FGF receptor 3 (FGFR3) mutations are common in benign skin lesions, either as embryonic mutations in epidermal nevi (EN) or as somatic mutations in seborrheic keratoses (SK). FGFR3 mutations are also common in low-grade malignant bladder tumors, where they often occur in association with PIK3CA mutations. Therefore, we examined exons 9 and 20 of PIK3CA and FGFR3 hotspot mutations in EN (n = 33) and SK (n = 62), two proliferative skin lesions lacking malignant potential. Nine of 33 (27%) EN harbored PIK3CA mutations; all cases showed the E545G substitution, which is uncommon in cancers. In EN, R248C was the only FGFR3 mutation identified. By contrast, 10 of 62 (16%) SK revealed the typical cancer-associated PIK3CA mutations E542K, E545K, and H1047R. The same lesions displayed a wide range of FGFR3 mutations. Corresponding unaffected tissue was available for four EN and two mutant SK: all control samples displayed a WT sequence, confirming the somatic nature of the mutations found in lesional tissue. Forty of 95 (42%) lesions showed at least one mutation in either gene. PIK3CA and FGFR3 mutations displayed an independent distribution; 5/95 lesions harbored mutations in both genes. Our findings suggest that, in addition to their role in cancer, oncogenic PIK3CA mutations contribute to the pathogenesis of skin tumors lacking malignant potential. The remarkable genotype–phenotype correlation as observed in this study points to a distinct etiopathogenesis of the mutations in keratinocytes occuring either during fetal development or in adult life. PMID:17673550

  19. A wide reprogramming of histone H3 modifications during male meiosis I in rice is dependent on the Argonaute protein MEL1.

    PubMed

    Liu, Hua; Nonomura, Ken-Ichi

    2016-10-01

    The roles of epigenetic mechanisms, including small-RNA-mediated silencing, in plant meiosis largely remain unclear, despite their importance in plant reproduction. This study unveiled that rice chromosomes are reprogrammed during the premeiosis-to-meiosis transition in pollen mother cells (PMCs). This large-scale meiotic chromosome reprogramming (LMR) continued throughout meiosis I, during which time H3K9 dimethylation (H3K9me2) was increased, and H3K9 acetylation and H3S10 phosphorylation were broadly decreased, with an accompanying immunostaining pattern shift of RNA polymerase II. LMR was dependent on the rice Argonaute protein, MEIOSIS ARRESTED AT LEPTOTENE1 (MEL1), which is specifically expressed in germ cells prior to meiosis, because LMR was severely diminished in mel1 mutant anthers. Pivotal meiotic events, such as pre-synaptic centromere association, DNA double-strand break initiation and synapsis of homologous chromosomes, were also disrupted in this mutant. Interestingly, and as opposed to the LMR loss in most chromosomal regions, aberrant meiotic protein loading and hypermethylation of H3K9 emerged on the nucleolar organizing region in the mel1 PMCs. These results suggest that MEL1 plays important roles in epigenetic LMR to promote faithful homologous recombination and synapsis during rice meiosis. © 2016. Published by The Company of Biologists Ltd.

  20. Arsenic toxicity in the human nerve cell line SK-N-SH in the presence of chromium and copper

    PubMed Central

    HU, LIGANG; GREER, JUSTIN B.; SOLO-GABRIELE, HELENA; FIEBER, LYNNE A.; CAI, YONG

    2013-01-01

    As, Cr, and Cu represent one potential combination of multiple metals/metalloids exposures since these three elements are simultaneously leached from chromated copper arsenate (CCA)-treated wood, a common product used for building construction, at levels that can be potentially harmful. This study investigated the neurotoxicity of As associated with CCA-treated wood when accompanied by Cr and Cu. The toxicity was evaluated on basis of a cytotoxicity model using human neuroblastoma cell line SK-N-SH. The cells were cultured with CCA-treated wood leachates or with solutions containing arsenate [As(V)], divalent copper [Cu(II)], trivalent chromium [Cr(III)] alone or in different combinations of the three elements. The toxicity was evaluated using variations in cell replication compared to controls after 96 hrs exposure. Among the three elements present in wood leachates, As played the primary role in the observed toxic effects, which exerted through multiple pathways, including the generation of oxidative stress. DOM affected the absorption of metals/metalloids into the test cells, which however did not obviously appear to impact toxicity. As toxicity was enhanced by Cu(II) and inhibited by Cr(III) at concentrations below U.S. EPA’s allowable maximum contaminant levels in drinking waters. Thus assessing As toxicity in real environments is not sufficient if based solely on the result from As. PMID:23473430

  1. Biosynthesis of N,N-dimethyltryptamine (DMT) in a melanoma cell line and its metabolization by peroxidases.

    PubMed

    Gomes, Melissa M; Coimbra, Janine B; Clara, Renan O; Dörr, Felipe A; Moreno, Ana Carolina R; Chagas, Jair R; Tufik, Sérgio; Pinto, Ernani; Catalani, Luiz H; Campa, Ana

    2014-04-01

    Tryptophan (TRP) is essential for many physiological processes, and its metabolism changes in some diseases such as infection and cancer. The most studied aspects of TRP metabolism are the kynurenine and serotonin pathways. A minor metabolic route, tryptamine and N,N-dimethyltryptamine (DMT) biosynthesis, has received far less attention, probably because of the very low amounts of these compounds detected only in some tissues, which has led them to be collectively considered as trace amines. In a previous study, we showed a metabolic interrelationship for TRP in melanoma cell lines. Here, we identified DMT and N,N-dimethyl-N-formyl-kynuramine (DMFK) in the supernatant of cultured SK-Mel-147 cells. Furthermore, when we added DMT to the cell culture, we found hydroxy-DMT (OH-DMT) and indole acetic acid (IAA) in the cell supernatant at 24 h. We found that SK-Mel-147 cells expressed mRNA for myeloperoxidase (MPO) and also had peroxidase activity. We further found that DMT oxidation was catalyzed by peroxidases. DMT oxidation by horseradish peroxidase, H2O2 and MPO from PMA-activated neutrophils produced DMFK, N,N-dimethyl-kynuramine (DMK) and OH-DMT. Oxidation of DMT by peroxidases apparently uses the common peroxidase cycle involving the native enzyme, compound I and compound II. In conclusion, this study describes a possible alternative metabolic pathway for DMT involving peroxidases that has not previously been described in humans and identifies DMT and metabolites in a melanoma cell line. The extension of these findings to other cell types and the biological effects of DMT and its metabolites on cell proliferation and function are key questions for future studies. Copyright © 2014 Elsevier Inc. All rights reserved.

  2. Selective cytotoxicity of PAMAM G5 core–PAMAM G2.5 shell tecto-dendrimers on melanoma cells

    PubMed Central

    Schilrreff, Priscila; Mundiña-Weilenmann, Cecilia; Romero, Eder Lilia; Morilla, Maria Jose

    2012-01-01

    Background The controlled introduction of covalent linkages between dendrimer building blocks leads to polymers of higher architectural order known as tecto-dendrimers. Because of the few simple steps involved in their synthesis, tecto-dendrimers could expand the portfolio of structures beyond commercial dendrimers, due to the absence of synthetic drawbacks (large number of reaction steps, excessive monomer loading, and lengthy chromatographic separations) and structural constraints of high-generation dendrimers (reduction of good monodispersity and ideal dendritic construction due to de Gennes dense-packing phenomenon). However, the biomedical uses of tecto-dendrimers remain unexplored. In this work, after synthesizing saturated shell core–shell tecto-dendrimers using amine-terminated polyamidoamine (PAMAM) generation 5 (G5) as core and carboxyl-terminated PAMAM G2.5 as shell (G5G2.5 tecto-dendrimers), we surveyed for the first time the main features of their interaction with epithelial cells. Methods Structural characterization of G5G2.5 was performed by polyacrylamide gel electrophoresis, matrix-assisted laser desorption time-of-flight mass spectrometry, and microscopic techniques; their hydrodynamic size and Z-potential was also determined. Cellular uptake by human epidermal keratinocytes, colon adenocarcinoma, and epidermal melanoma (SK-Mel-28) cells was determined by flow cytometry. Cytotoxicity was determined by mitochondrial activity, lactate dehydrogenase release, glutathione depletion, and apoptosis/necrosis measurement. Results The resultant 60%–67% saturated shell, 87,000-dalton G5G2.5 (mean molecular weight) interacted with cells in a significantly different fashion in comparison to their building blocks and to its closest counterpart, PAMAM G6.5. After being actively taken up by epithelial cells, G5G2.5 caused cytotoxicity only on SK-Mel-28 cells, including depletion of intracellular glutathione and fast necrosis that was manifested above 5 μM G5

  3. The Mycobacterium marinum mel2 locus displays similarity to bacterial bioluminescence systems and plays a role in defense against reactive oxygen and nitrogen species

    PubMed Central

    Subbian, Selvakumar; Mehta, Parmod K; Cirillo, Suat LG; Cirillo, Jeffrey D

    2007-01-01

    Background Mycobacteria have developed a number of pathways that provide partial protection against both reactive oxygen species (ROS) and reactive nitrogen species (RNS). We recently identified a locus in Mycobacterium marinum, mel2, that plays a role during infection of macrophages. The molecular mechanism of mel2 action is not well understood. Results To better understand the role of the M. marinum mel2 locus, we examined these genes for conserved motifs in silico. Striking similarities were observed between the mel2 locus and loci that encode bioluminescence in other bacterial species. Since bioluminescence systems can play a role in resistance to oxidative stress, we postulated that the mel2 locus might be important for mycobacterial resistance to ROS and RNS. We found that an M. marinum mutant in the first gene in this putative operon, melF, confers increased susceptibility to both ROS and RNS. This mutant is more susceptible to ROS and RNS together than either reactive species alone. Conclusion These observations support a role for the M. marinum mel2 locus in resistance to oxidative stress and provide additional evidence that bioluminescence systems may have evolved from oxidative defense mechanisms. PMID:17239244

  4. Some recollections on acousto-optics research at the University of Gdańsk in the light of international cooperation and of the history of Spring Schools on Acousto-Optics and Applications.

    PubMed

    Kwiek, Piotr; Sliwiński, Antoni

    2009-03-01

    Some historical features of international cooperation that have been a background for originating the idea to organize since 1980 special international meetings named Spring Schools on Acousto-Optics and Applications and for continuing the events until now are presented. Thanks to the establishment of the Spring Schools, the group of acousto-opticians at the Gdańsk University have had the opportunity to exchange scientific experience and continue mutual cooperation in research with several acousto-optic centers and to publish a number of common papers. A few examples from these achievements and some historical facts illustrating research activity in the field of acousto-optics during past 30 years are recollected.

  5. Conformational and spectroscopic study of xanthogen ethyl formates, ROC(S)SC(O)OCH2CH3. Isolation of CH3CH2OC(O)SH

    NASA Astrophysics Data System (ADS)

    Juncal, Luciana C.; Cozzarín, Melina V.; Romano, Rosana M.

    2015-03-01

    ROC(S)SC(O)OCH2CH3, with R = CH3sbnd , (CH3)2CHsbnd and CH3(CH2)2sbnd , were obtained through the reaction between potassium xanthate salts, ROC(S)SK, and ethyl chloroformate, ClC(O)OCH2CH3. The liquid compounds were identified and characterized by 1H and 13C NMR and mass spectrometry. The conformations adopted by the molecules were studied by DFT methods. 6 conformers were theoretically predicted for R = CH3sbnd and (CH3)2CHsbnd , while the conformational flexibility of the n-propyl substituent increases the total number of feasible rotamers to 21. For the three molecules, the conformers can be associated in 3 groups, being the most stable the AS forms - the Cdbnd S double bond anti (A) with respect to the Csbnd S single bond and the Ssbnd C single bond syn (S) with respect to the Cdbnd O double bond - followed by AA and SS conformers. The vibrational spectra were interpreted in terms of the predicted conformational equilibrium, presenting the ν(Cdbnd O) spectral region signals corresponding to the three groups of conformers. A moderated pre-resonance Raman enhancement of the ν(Cdbnd S) vibrational mode of CH3(CH2)2OC(S)SC(O)OCH2CH3 was detected, when the excitation radiation approaches the energy of a n → π∗ electronic transition associated with the Cdbnd S chromophore. UV-visible spectra in different solvents were measured and interpreted in terms of TD-DFT calculations. The unknown molecule CH3CH2OC(O)SH was isolated by the UV-visible photolysis of CH3OC(S)SC(O)OCH2CH3 isolated in Ar matrix, and also obtained as a side-product of the reaction between potassium xanthate salts, ROC(S)SK, and ethyl chloroformate, ClC(O)OCH2CH3.

  6. Determination of trace level thorium and uranium in high purity gadolinium sulfate using ICP-MS with solid-phase chromatographic extraction resins

    NASA Astrophysics Data System (ADS)

    Ito, S.; Takaku, Y.; Ikeda, M.; Kishimoto, Y.

    2018-01-01

    The Super Kamiokand-Gadolinium (SK-Gd) project is the upgrade of the Super-Kamiokande (SK) detector in order to discover Supernova Relic Neutrinos (SRNs) by loading 0.2% of Gd2(SO4)3 into a 50 kton of the SK water tank. In order to continue solar neutrino measurement with low energy threshold at ˜3.5 MeV, main radioactive contamination, U and Th in Gd2(SO4)3, should be purified before loading. We developed solid-phase extraction technique to measure low concentration of U and Th in Gd2(SO4)3 by ICP-MS. The extraction technique and current status will be presented.

  7. Oviductal extracellular vesicles (oviductosomes, OVS) are conserved in humans: murine OVS play a pivotal role in sperm capacitation and fertility.

    PubMed

    Bathala, Pradeepthi; Fereshteh, Zeinab; Li, Kun; Al-Dossary, Amal A; Galileo, Deni S; Martin-DeLeon, Patricia A

    2018-03-01

    in sperm and their ability to interact with PMCA1, via co-immunoprecipitation. In vitro uptake of PMCA1 from OVS was analyzed in capacitated and uncapacitated sperm via quantitative western blot analysis, IF localization and flow cytometry. Caudal sperm were also assayed for uptake of tyrosine-phosphorylated proteins which were shown to be present in OVS. Finally, PMCA1 and PMCA4 in OVS and that delivered to sperm were assayed for enzymatic activity. Human fallopian tubes were flushed to recover luminal fluid which was processed for OVS via ultracentrifugation. Human OVS were negatively stained for transmission electron microscopy (TEM) and subjected to immunogold labeling, to detect PMCA4. Western analysis was used to detect HSC70 (an EV biomarker), PMCA1 and endothelial nitric oxide synthase (eNOS) which is a fertility-modulating protein delivered to human sperm by prostasomes. Oviducts of sexually mature female mice were sectioned after perfusion fixation for TEM tomography to obtain 3D information and to distinguish cross-sections of EVs from those of microvilli and cilia. Murine tissues, luminal fluids and EVs were assayed for PMCA1 (IF and western blot) or qRT-PCR. PMCA1 levels from western blots were quantified, using band densities and compared in WT and Pmca4-/- after induced estrus and in proestrus/estrus and metestrus/diestrus in cycling females. In vitro uptake of PMCA1 and tyrosine-phosphorylated proteins was quantified with flow cytometry and/or quantitative western blot. Ca2+-ATPase activity in OVS and sperm before and after PMCA1 and PMCA4 uptake was assayed, via the enzymatic hydrolysis rate of ATP. TEM revealed that human oviducts contain EVs (exosomal and microvesicular). These EVs contain PMCA4 (immunolabeling), eNOS and PMCA1 (western blot) in their cargo. TEM tomography showed the murine oviduct with EV-containing blebs which typify the apocrine pathway for EV biogenesis. Western blots revealed that during proestrus/estrus PMCA1 was

  8. EOVSA Implementation of a Spectral Kurtosis Correlator for Transient Detection and Classification

    NASA Astrophysics Data System (ADS)

    Nita, Gelu M.; Hickish, Jack; MacMahon, David; Gary, Dale E.

    2016-03-01

    We describe in general terms the practical use in astronomy of a higher-order statistical quantity called spectral kurtosis (SK), and describe the first implementation of SK-enabled firmware in the Fourier transform-engine (F-engine) of a digital FX correlator for the Expanded Owens Valley Solar Array (EOVSA). The development of the theory for SK is summarized, leading to an expression for generalized SK that is applicable to both SK spectrometers and those not specifically designed for SK. We also give the means for computing both the SK̂ estimator and thresholds for its application as a discriminator of RFI contamination. Tests of the performance of EOVSA as an SK spectrometer are shown to agree precisely with theoretical expectations, and the methods for configuring the correlator for correct SK operation are described.

  9. Inward rectifier potassium (Kir2.1) channels as end-stage boosters of endothelium-dependent vasodilators.

    PubMed

    Sonkusare, Swapnil K; Dalsgaard, Thomas; Bonev, Adrian D; Nelson, Mark T

    2016-06-15

    Increase in endothelial cell (EC) calcium activates calcium-sensitive intermediate and small conductance potassium (IK and SK) channels, thereby causing hyperpolarization and endothelium-dependent vasodilatation. Endothelial cells express inward rectifier potassium (Kir) channels, but their role in endothelium-dependent vasodilatation is not clear. In the mesenteric arteries, only ECs, but not smooth muscle cells, displayed Kir currents that were predominantly mediated by the Kir2.1 isoform. Endothelium-dependent vasodilatations in response to muscarinic receptor, TRPV4 (transient receptor potential vanilloid 4) channel and IK/SK channel agonists were highly attenuated by Kir channel inhibitors and by Kir2.1 channel knockdown. These results point to EC Kir channels as amplifiers of vasodilatation in response to increases in EC calcium and IK/SK channel activation and suggest that EC Kir channels could be targeted to treat endothelial dysfunction, which is a hallmark of vascular disorders. Endothelium-dependent vasodilators, such as acetylcholine, increase intracellular Ca(2+) through activation of transient receptor potential vanilloid 4 (TRPV4) channels in the plasma membrane and inositol trisphosphate receptors in the endoplasmic reticulum, leading to stimulation of Ca(2+) -sensitive intermediate and small conductance K(+) (IK and SK, respectively) channels. Although strong inward rectifier K(+) (Kir) channels have been reported in the native endothelial cells (ECs) their role in EC-dependent vasodilatation is not clear. Here, we test the idea that Kir channels boost the EC-dependent vasodilatation of resistance-sized arteries. We show that ECs, but not smooth muscle cells, of small mesenteric arteries have Kir currents, which are substantially reduced in EC-specific Kir2.1 knockdown (EC-Kir2.1(-/-) ) mice. Elevation of extracellular K(+) to 14 mm caused vasodilatation of pressurized arteries, which was prevented by endothelial denudation and Kir channel

  10. Inward rectifier potassium (Kir2.1) channels as end‐stage boosters of endothelium‐dependent vasodilators

    PubMed Central

    Dalsgaard, Thomas; Bonev, Adrian D.; Nelson, Mark T.

    2016-01-01

    Key points Increase in endothelial cell (EC) calcium activates calcium‐sensitive intermediate and small conductance potassium (IK and SK) channels, thereby causing hyperpolarization and endothelium‐dependent vasodilatation.Endothelial cells express inward rectifier potassium (Kir) channels, but their role in endothelium‐dependent vasodilatation is not clear.In the mesenteric arteries, only ECs, but not smooth muscle cells, displayed Kir currents that were predominantly mediated by the Kir2.1 isoform.Endothelium‐dependent vasodilatations in response to muscarinic receptor, TRPV4 (transient receptor potential vanilloid 4) channel and IK/SK channel agonists were highly attenuated by Kir channel inhibitors and by Kir2.1 channel knockdown.These results point to EC Kir channels as amplifiers of vasodilatation in response to increases in EC calcium and IK/SK channel activation and suggest that EC Kir channels could be targeted to treat endothelial dysfunction, which is a hallmark of vascular disorders. Abstract Endothelium‐dependent vasodilators, such as acetylcholine, increase intracellular Ca2+ through activation of transient receptor potential vanilloid 4 (TRPV4) channels in the plasma membrane and inositol trisphosphate receptors in the endoplasmic reticulum, leading to stimulation of Ca2+‐sensitive intermediate and small conductance K+ (IK and SK, respectively) channels. Although strong inward rectifier K+ (Kir) channels have been reported in the native endothelial cells (ECs) their role in EC‐dependent vasodilatation is not clear. Here, we test the idea that Kir channels boost the EC‐dependent vasodilatation of resistance‐sized arteries. We show that ECs, but not smooth muscle cells, of small mesenteric arteries have Kir currents, which are substantially reduced in EC‐specific Kir2.1 knockdown (EC‐Kir2.1 −/−) mice. Elevation of extracellular K+ to 14 mm caused vasodilatation of pressurized arteries, which was prevented by endothelial

  11. Plasma metabolomic profiling of dairy cows affected with ketosis using gas chromatography/mass spectrometry.

    PubMed

    Zhang, Hongyou; Wu, Ling; Xu, Chuang; Xia, Cheng; Sun, Lingwei; Shu, Shi

    2013-09-26

    Ketosis is an important problem for dairy cows` production performance. However, it is still little known about plasma metabolomics details of dairy ketosis. A gas chromatography/mass spectrometry (GC/MS) technique was used to investigate plasma metabolic differences in cows that had clinical ketosis (CK, n=22), subclinical ketosis (SK, n=32), or were clinically normal controls (NC, n=22). The endogenous plasma metabolome was measured by chemical derivatization followed by GC/MS, which led to the detection of 267 variables. A two-sample t-test of 30, 32, and 13 metabolites showed statistically significant differences between SK and NC, CK and NC, and CK and SK, respectively. Orthogonal signal correction-partial least-square discriminant analysis (OPLS-DA) revealed that the metabolic patterns of both CK and SK were mostly similar, with the exception of a few differences. The development of CK and SK involved disturbances in many metabolic pathways, mainly including fatty acid metabolism, amino acid metabolism, glycolysis, gluconeogenesis, and the pentose phosphate pathway. A diagnostic model arbitrary two groups was constructed using OPLS-DA and receiver-operator characteristic curves (ROC). Multivariate statistical diagnostics yielded the 19 potential biomarkers for SK and NC, 31 for CK and NC, and 8 for CK and SK with area under the curve (AUC) values. Our results showed the potential biomarkers from CK, SK, and NC, including carbohydrates, fatty acids, amino acids, even sitosterol and vitamin E isomers, etc. 2-piperidinecarboxylic acid and cis-9-hexadecenoic acid were closely associated with metabolic perturbations in ketosis as Glc, BHBA and NEFA for dealing with metabolic disturbances of ketosis in clinical practice. However, further research is needed to explain changes of 2,3,4-trihydroxybutyric acid, 3,4-dihydroxybutyric acid, α-aminobutyric acid, methylmalonic acid, sitosterol and α-tocopherol in CK and SK, and to reveal differences between CK and SK. Our

  12. Human liver microsomal cytochrome P-450 enzymes involved in the bioactivation of procarcinogens detected by umu gene response in Salmonella typhimurium TA 1535/pSK1002.

    PubMed

    Shimada, T; Iwasaki, M; Martin, M V; Guengerich, F P

    1989-06-15

    A total of 57 procarcinogens was examined for induction of umu gene response in the chimeric plasmid pSK1002, carried in Salmonella typhimurium TA 1535, after incubation with a series of human liver microsomal preparations which had been selected on the basis of characteristic levels of individual cytochrome P-450 (P-450) enzymes. The 18 most active compounds were selected and further analyzed using the umu gene response and correlative studies with a larger number of microsomal preparations, enzyme reconstitution studies involving purified enzymes, immunochemical inhibition, and patterns of stimulation and inhibition of catalytic activity by 7,8-benzoflavone. The results collectively indicate that 16 of these 18 most potent genotoxins examined are activated primarily either by P-450NF (the nifedipine oxidase) or P-450PA (the phenacetin O-deethylase). P-450NF appears to be the major enzyme involved in the bioactivation of aflatoxin B1, aflatoxin G1, sterigmatocystin, trans-7,8-dihydroxy-7,8-dihydrobenzo(a)pyrene, 6-aminochrysene, and tris-(2,3-dibromopropyl)phosphate in human liver. P-450PA appears to be the major enzyme involved in the bioactivation of 2-amino-3-methylimidazo[4,5-f]quinoline, 2-amino-3,5-dimethylimidazo[4, 5-f]quinoline, 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline, 2-aminoanthracene, 2-amino-6-methyldipyrido[1,2-a:3',2'-d]imidazole, 2-aminofluorene, 2-acetylaminofluorene, 4-aminobiphenyl, 3-amino-1-methyl-5H-pyrido[4,3-b] indole, and 2-aminodipyrido[1,2-a:3',2'-d]imidazole. More than one enzyme appears to contribute significantly to the bioactivation of the other two compounds examined, 3-amino-1,4-dimethyl-5H-pyrido[4,3-b] indole and 6-nitrochrysene. The literature suggests that the two human liver P-450s involved in activation of these 16 procarcinogens are highly inducible by barbiturates, macrolide antibodies, and certain steroids (P-450NF) and by smoking and ingestion of charcoal-containing food (P-450PA); noninvasive assays are available

  13. Downregulation of Endothelial Transient Receptor Potential Vanilloid Type 4 Channel and Small-Conductance of Ca2+-Activated K+ Channels Underpins Impaired Endothelium-Dependent Hyperpolarization in Hypertension.

    PubMed

    Seki, Takunori; Goto, Kenichi; Kiyohara, Kanako; Kansui, Yasuo; Murakami, Noboru; Haga, Yoshie; Ohtsubo, Toshio; Matsumura, Kiyoshi; Kitazono, Takanari

    2017-01-01

    Endothelium-dependent hyperpolarization (EDH)-mediated responses are impaired in hypertension, but the underlying mechanisms have not yet been determined. The activation of small- and intermediate-conductance of Ca 2+ -activated K + channels (SK Ca and IK Ca ) underpins EDH-mediated responses. It was recently reported that Ca 2+ influx through endothelial transient receptor potential vanilloid type 4 channel (TRPV4) is a prerequisite for the activation of SK Ca /IK Ca in endothelial cells in specific beds. Here, we attempted to determine whether the impairment of EDH in hypertension is attributable to the dysfunction of TRPV4 and S/IK Ca , using isolated superior mesenteric arteries of 20-week-old stroke-prone spontaneously hypertensive rats (SHRSP) and age-matched Wistar-Kyoto (WKY) rats. In the WKY arteries, EDH-mediated responses were reduced by a combination of SK Ca /IK Ca blockers (apamin plus TRAM-34; 1-[(2-chlorophenyl)diphenylmethl]-1H-pyrazole) and by the blockade of TRPV4 with the selective antagonist RN-1734 or HC-067047. In the SHRSP arteries, EDH-mediated hyperpolarization and relaxation were significantly impaired when compared with WKY. GSK1016790A, a selective TRPV4 activator, evoked robust hyperpolarization and relaxation in WKY arteries. In contrast, in SHRSP arteries, the GSK1016790A-evoked hyperpolarization was small and relaxation was absent. Hyperpolarization and relaxation to cyclohexyl-[2-(3,5-dimethyl-pyrazol-1-yl)-6-methyl-pyrimidin-4-yl]-amine, a selective SK Ca activator, were marginally decreased in SHRSP arteries compared with WKY arteries. The expression of endothelial TRPV4 and SK Ca protein was significantly decreased in the SHRSP mesenteric arteries compared with those of WKY, whereas function and expression of IK Ca were preserved in SHRSP arteries. These findings suggest that EDH-mediated responses are impaired in superior mesenteric arteries of SHRSP because of a reduction in both TRPV4 and SK Ca input to EDH. © 2016 American

  14. Growth, fatty acid profile in major lipid classes and lipid fluidity of Aurantiochytrium mangrovei SK-02 As a function of growth temperature.

    PubMed

    Chodchoey, Kanokwan; Verduyn, Cornelis

    2012-01-01

    Aurantiochytrium mangrovei Sk-02 was grown in a medium containing glucose (40 g/l), yeast extract (10 g/L) and sea salts (15 g/L) at temperatures ranging from 12 to 35°C. The fastest growth (µmax= 0.15 h(-1)) and highest fatty acid content of 415 mg/g-dry cell weight were found in the cells grown at 30°C. However, the cells grown at 12°C showed the highest percentage of polyunsaturated fatty acid (PUFA) (48.6% of total fatty acid). The percentage of docosahexaenoic acid (DHA) and pentadecanoic acid (C15:0) decreased with an increase in the growth temperature, whereas, palmitic acid (C16:0), stearic acid (C18:0) and DPA (C22:5n6) increased with an increase in the growth temperature. The composition of the major lipid class (%w/w) was slightly affected by the growth temperature. The fluidity of the organelle membrane or intracellular lipid (by DPH measurement) decreased with an increase in the growth temperatures, while the plasma membrane fluidity (by TMA-DPH measurement) could still maintain its fluidity in a wide range of temperatures (15 - 37°C). Furthermore, the distribution of DHA was found to be higher (36 - 54%) in phospholipid (PL) as compared to neutral lipid (NL) (20 - 41%).

  15. Delivery of tissue plasminogen activator and streptokinase magnetic nanoparticles to target vascular diseases.

    PubMed

    Tadayon, Ateke; Jamshidi, Reza; Esmaeili, Akbar

    2015-11-10

    Thrombolytic therapy for acute myocardial infarction standardly makes use of the medications streptokinase (SK) and tissue plasminogen activator (tPA). In this study, the potential of silica-coated magnetic nanoparticles (SiO2-MNPs) as nanocarriers clinical thrombolytic therapy was investigated. SiO2-MNPs for use in targeted therapeutic delivery of tPA and SK were prepared using a combined technique incorporating controlled precipitation and hydrothermal methods. Response surface methodology (RSM) was employed to evaluate the efficiency of the SiO2-MNPs. The production of SK secreted from Streptococcus equi was enhanced using random mutagenesis. The tPA and SK A were encapsulated by means of a silanizing agent with a surface rich in 3-aminopropyltrimethoxysilane layered around the SiO2-MNPs. Blood clot lysis assays and fibrin-containing agarose plates were used to carry out in vitro thrombolysis testing. The optimum conditions for producing MNPs were found to be at pH=13 and at a temperature of 75°C for 45 min. Culture conditions of 2.75% NaCl concentration at initial pH=7.5 for 90 s under UV resulted in maximum SK activity. The tPA/SK-conjugated SiO2-MNPs (SiO2-MNP-tPA-SK) increased operating stability in whole blood and storage stability in a buffer by 92%. More effective thrombolysis using magnetic targeting was indicated by a 38% reduction in blood clot lysis time achieved with SiO2-MNP-tPA-SK compared to administering the SiO2-MNPs without guidance. The silica-coated magnetic nanocarriers developed in this study show potential for improved clinical thrombolytic therapy. Copyright © 2015. Published by Elsevier B.V.

  16. Patient autoantibodies deplete postsynaptic muscle-specific kinase leading to disassembly of the ACh receptor scaffold and myasthenia gravis in mice

    PubMed Central

    Cole, R N; Ghazanfari, N; Ngo, S T; Gervásio, O L; Reddel, S W; Phillips, W D

    2010-01-01

    The postsynaptic muscle-specific kinase (MuSK) coordinates formation of the neuromuscular junction (NMJ) during embryonic development. Here we have studied the effects of MuSK autoantibodies upon the NMJ in adult mice. Daily injections of IgG from four MuSK autoantibody-positive myasthenia gravis patients (MuSK IgG; 45 mg day−1i.p. for 14 days) caused reductions in postsynaptic ACh receptor (AChR) packing as assessed by fluorescence resonance energy transfer (FRET). IgG from the patients with the highest titres of MuSK autoantibodies caused large (51–73%) reductions in postsynaptic MuSK staining (cf. control mice; P < 0.01) and muscle weakness. Among mice injected for 14 days with control and MuSK patient IgGs, the residual level of MuSK correlated with the degree of impairment of postsynaptic AChR packing. However, the loss of postsynaptic MuSK preceded this impairment of postsynaptic AChR. When added to cultured C2 muscle cells the MuSK autoantibodies caused tyrosine phosphorylation of MuSK and the AChR β-subunit, and internalization of MuSK from the plasma membrane. The results suggest a pathogenic mechanism in which MuSK autoantibodies rapidly deplete MuSK from the postsynaptic membrane leading to progressive dispersal of postsynaptic AChRs. Moreover, maintenance of postsynaptic AChR packing at the adult NMJ would appear to depend upon physical engagement of MuSK with the AChR scaffold, notwithstanding activation of the MuSK-rapsyn system of AChR clustering. PMID:20603331

  17. Genistein decreases A549 cell viability via inhibition of the PI3K/AKT/HIF‑1α/VEGF and NF‑κB/COX‑2 signaling pathways.

    PubMed

    Zhang, Juan; Su, Hongzheng; Li, Qingfeng; Li, Jing; Zhao, Qianfeng

    2017-04-01

    Genistein is an important chemopreventive agent against atherosclerosis and cancer. However, whether genistein is effective in the treatment of lung cancer, and its underlying mechanism, remains to be determined. The present study demonstrated that genistein treatment of A549 lung cancer cells decreased viability in a dose‑ and time‑dependent manner, and induced apoptosis. Additionally, A549 cells exhibited significantly increased reactive oxygen species formation and cytochrome‑c leakage, and activated caspase‑3, B‑cell lymphoma 2‑associated X protein and apoptosis inducing factor expression levels, which are involved in the mitochondrial apoptosis pathway. Furthermore, the phosphatidylinositol‑4,5‑biphosphate 3‑kinase (PI3K)/protein kinase B (AKT)/hypoxia‑inducible factor‑1α (HIF‑1α) and nuclear factor‑κB (NF‑κB)/cyclooxygenase‑2 (COX‑2) signaling pathways were significantly downregulated by genistein treatment. In conclusion, reduced proliferation and increased apoptosis in A549 lung cancer cells was associated with inhibition of the PI3K/AKT/HIF‑1α/ and NF‑κB/COX‑2 signaling pathways, which implicates genistein as a potential chemotherapeutic agent for the treatment of lung cancer.

  18. A de novo mutation in KCNN3 associated with autosomal dominant idiopathic non-cirrhotic portal hypertension.

    PubMed

    Koot, Bart G P; Alders, Marielle; Verheij, Joanne; Beuers, Ulrich; Cobben, Jan M

    2016-04-01

    Non-cirrhotic portal hypertension is characterized by histopathological abnormalities in the liver, mostly affecting small intrahepatic portal veins that cause portal hypertension in the absence of cirrhosis. It can be secondary to coagulation disorders or toxic agents. However, most cases are idiopathic non-cirrhotic portal hypertension (INCPH) and familial cases are rare. We report a family in which a father and three of his four children conceived with three different mothers are affected by INCPH. Whole exome and Sanger sequencing showed the father to have a de novo single nucleotide substitution c.1348G>C in the KCNN3 gene that was transmitted to all three of his affected offspring. The KCNN3 gene encodes small conductance calcium-activated potassium (SK) channel 3. SK channels are involved in the regulation of arterial and venous vascular tone by causing smooth muscle relaxation on activation. No data exist on the expression and function of SK channels in portal veins. The autosomal dominant inheritance in this unique pedigree and the single de novo mutation identified, strongly suggests that KCNN3 mutations have a pathogenetic role in INCPH. Copyright © 2015 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.

  19. Breast Cancer Therapy Using Antibody-Endostatin Fusion Proteins

    DTIC Science & Technology

    2008-04-01

    animal tumor and/or human xenograft models. Task 9. Anti-tumor activity in human tumor xenografts (Months 18 -24) Anti-tumor efficacy in human...breast cancer SK-BR-3 xenografts in SCID mice. SK-BR-3 was implanted on the flank of SCID mice.9 The treatment was repeated (shown below in Fig. 18 ...untreated group (p=0.0141) (Fig. 18 ). KEY RESEARCH ACCOMPLISHMENTS o Treatment of established SK-BR-3 xenografts in SCID mice with the αHER2

  20. 6-Methyl-1,2,4-benzenetriol, a new intermediate in penicillic acid biosynthesis in Penicillium cyclopium

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Sekiguchi, J.; Katayama, S.; Yamada, Y.

    1987-07-01

    Penicillic acid-negative mutants were obtained from a color mutant derived from Penicillium cyclopium NRRL 1888 through N-methyl-N'-nitro-N-nitrosoguanidine treatment. One mutant (SK2N6) accumulated 6-methyl-1,2,4-benzenetriol, which was not previously known to be a metabolite of P. cyclopium, in addition to orsellinic acid and orcinol. The radioactivity of (1-/sup 14/C)acetic acid was rapidly incorporated into 6-methyl-1,2,4-benzenetriol in a culture of P. cyclopium SK2N6. Moreover, the radioactivity of (/sup 14/C)6-methyl-1,2,4-benzenetriol was efficiently incorporated into penicillic acid in a culture of P. cyclopium NRRL 1888. These data indicate that 6-methyl-1,2,4-benzenetriol is a precursor for penicillic acid biosynthesis. The results on the addition of 1,4-dihydroxy-6-methoxy-2-methylbenzene, 6-methoxy-2-methylbenzoquinonemore » (1,4), and 1-O-methylorcinol to a culture of P. cyclopium SK2N6 indicated that only the former two compounds are converted to penicillic acid. Thus, a new portion of the penicillic acid biosynthetic pathway is proposed.« less

  1. Low energy neutrinos in Super-Kamiokande

    NASA Astrophysics Data System (ADS)

    Sekiya, Hiroyuki

    2016-05-01

    Super-Kamiokande (SK), a 50 kton water Cherenkov detector, observes 8B solar neutrinos via neutrino-electron elastic scattering. The analysis threshold was successfully lowered to 3.5 MeV (recoil electron kinetic energy) in SK-IV. To date SK has observed solar neutrinos for 18 years. An analysis regarding possible correlations between the solar neutrino flux and the 11 year solar activity cycle is shown. With large statistics, SK searches for distortions of the solar neutrino energy spectrum caused by the MSW resonance in the core of the sun. SK also searches for a day/night solar neutrino flux asymmetry induced by the matter in the Earth. The Super-Kamiokande Gd (SK-Gd) project is the upgrade of the SK detector via the addition of water-soluble gadolinium (Gd) salt. This modification will enable it to efficiently identify low energy anti-neutrinos. SK-Gd will pursue low energy physics currently inaccessible to SK due to backgrounds. The most important will be the world’s first observation of the diffuse supernova neutrino background. The main R&D program towards SK-Gd is EG ADS: a 200 ton, fully instrumented tank built in a new cavern in the Kamioka mine.

  2. DNA profiling analysis of endometrial and ovarian cell lines reveals misidentification, redundancy and contamination.

    PubMed

    Korch, Christopher; Spillman, Monique A; Jackson, Twila A; Jacobsen, Britta M; Murphy, Susan K; Lessey, Bruce A; Jordan, V Craig; Bradford, Andrew P

    2012-10-01

    Cell lines derived from human ovarian and endometrial cancers, and their immortalized non-malignant counterparts, are critical tools to investigate and characterize molecular mechanisms underlying gynecologic tumorigenesis, and facilitate development of novel therapeutics. To determine the extent of misidentification, contamination and redundancy, with evident consequences for the validity of research based upon these models, we undertook a systematic analysis and cataloging of endometrial and ovarian cell lines. Profiling of cell lines by analysis of DNA microsatellite short tandem repeats (STR), p53 nucleotide polymorphisms and microsatellite instability was performed. Fifty-one ovarian cancer lines were profiled with ten found to be redundant and five (A2008, OV2008, C13, SK-OV-4 and SK-OV-6) identified as cervical cancer cells. Ten endometrial cell lines were analyzed, with RL-92, HEC-1A, HEC-1B, HEC-50, KLE, and AN3CA all exhibiting unique, uncontaminated STR profiles. Multiple variants of Ishikawa and ECC-1 endometrial cancer cell lines were genotyped and analyzed by sequencing of mutations in the p53 gene. The profile of ECC-1 cells did not match the EnCa-101 tumor, from which it was reportedly derived, and all ECC-1 isolates were genotyped as Ishikawa cells, MCF-7 breast cancer cells, or a combination thereof. Two normal, immortalized endometrial epithelial cell lines, HES cells and the hTERT-EEC line, were identified as HeLa cervical carcinoma and MCF-7 breast cancer cells, respectively. Results demonstrate significant misidentification, duplication, and loss of integrity of endometrial and ovarian cancer cell lines. Authentication by STR DNA profiling is a simple and economical method to verify and validate studies undertaken with these models. Copyright © 2012 Elsevier Inc. All rights reserved.

  3. Antitumor evaluation and 3D-QSAR studies of a new series of the spiropyrroloquinoline isoindolinone/aza-isoindolinone derivatives by comparative molecular field analysis (CoMFA).

    PubMed

    Sadeghzadeh, Masoud; Salahinejad, Maryam; Zarezadeh, Nahid; Ghandi, Mehdi; Baghery, Maryam Keshavarz

    2017-11-01

    In current study, antitumor activity of two series of the newly synthesized spiropyrroloquinoline isoindolinone and spiropyrroloquinoline aza-isoindolinone scaffolds was evaluated against three human breast normal and cancer cell lines (MCF-10A, MCF-7 and SK-BR-3) and compared with cytotoxicity values of doxorubicin and colchicine as the standard drugs. It was found that several compounds were endowed with cytotoxicity in the low micromolar range. Among these two series, compounds 6i, 6j, 6k and 7l, 7m, 7n, 7o containing 3-ethyl-1H-indole moiety were found to be highly effective against both cancer cell lines ranging from [Formula: see text] to [Formula: see text] in comparison with the corresponding analogs. Compared with human cancer cells, the most potent compounds did not show high cytotoxicity against human breast normal MCF-10A cells. Generally, most of the evaluated compounds 6a-l and 7a-o series showed more antitumor activity against SK-BR-3 than MCF-7 cells. Moreover, comparative molecular field analysis (CoMFA) as a popular tools of three-dimensional quantitative structure-activity relationship (3D-QSAR) studies was carried out on 27 spiropyrroloquinolineisoindolinone and spiropyrroloquinolineaza-isoindolinone derivatives with antitumor activity against on SK-BR-3 cells. The obtained CoMFA models showed statistically excellent performance, which also possessed good predictive ability for an external test set. The results confirm the important effect of molecular steric and electrostatic interactions of these compounds on in vitro cytotoxicity against SK-BR-3.

  4. Alpha-Lipoic Acid Downregulates IL-1β and IL-6 by DNA Hypermethylation in SK-N-BE Neuroblastoma Cells.

    PubMed

    Dinicola, Simona; Proietti, Sara; Cucina, Alessandra; Bizzarri, Mariano; Fuso, Andrea

    2017-09-26

    Alpha-lipoic acid (ALA) is a pleiotropic molecule with antioxidant and anti-inflammatory properties, of which the effects are exerted through the modulation of NF-kB. This nuclear factor, in fact, modulates different inflammatory cytokines, including IL-1b and IL-6, in different tissues and cell types. We recently showed that IL-1b and IL-6 DNA methylation is modulated in the brain of Alzheimer's disease patients, and that IL-1b expression is associated to DNA methylation in the brain of patients with tuberous sclerosis complex. These results prompted us to ask whether ALA-induced repression of IL-1b and IL-6 was dependent on DNA methylation. Therefore, we profiled DNA methylation in the 5'-flanking region of the two aforementioned genes in SK-N-BE human neuroblastoma cells cultured in presence of ALA 0.5 mM. Our experimental data pointed out that the two promoters are hypermethylated in cells supplemented with ALA, both at CpG and non-CpG sites. Moreover, the observed hypermethylation is associated with decreased mRNA expression and decreased cytokine release. These results reinforce previous findings indicating that IL-1b and IL-6 undergo DNA methylation-dependent modulation in neural models and pave the road to study the epigenetic mechanisms triggered by ALA.

  5. Determination of trace levels of uranium and thorium in high purity gadolinium sulfate using the ICP-MS with solid-phase chromatographic extraction resin

    NASA Astrophysics Data System (ADS)

    Ito, S.; Takaku, Y.; Ikeda, M.; Kishimoto, Y.

    2017-11-01

    The new Super-Kamiokande-Gadolinium (SK-Gd) project is an upgrade of the Super-Kamiokande (SK) detector. In the SK-Gd project, 0.2% Gd_2(SO_4)_3 is loaded into the 50 kton water tank of the SK. One of the main purposes of the project is to discover supernova relic neutrinos. Neutrino measurements and proton decay searches will also be performed in the SK-Gd. In order to measure solar neutrinos with a low energy threshold of ˜3.5 MeV in the SK-Gd, the main radioactive contaminations, ^{238}U and ^{232}Th, in Gd_2(SO_4)_3{\\cdot}8H_2O, should be minimized before loading. Our maximum levels for U and Th are 5 mBq (U)/kg (Gd_2(SO_4)_3{\\cdot}8H_2O) and 0.05 mBq (Th)/kg (Gd_2(SO_4)_3{\\cdot}8H_2O). In order to measure such low concentrations of U and Th in Gd_2(SO_4)_3{\\cdot}8H_2O, we developed the solid-phase extraction technique. Using this method, about 90% or more U and Th could be efficiently extracted while Gd was reduced by a factor of about 104. This allowed these radioactivity contaminations to be measured precisely as 0.04 mBq/kg (Gd_2(SO_4)_3{\\cdot}8H_2O) for U and 0.01 mBq/kg (Gd_2(SO_4)_3{\\cdot}8H_2O) for Th. We measured three pure Gd_2(SO_4)_3{\\cdot}8H_2O samples using this method and estimated that the purest one contained <0.04 mBq (U)/kg (Gd_2(SO_4)_3{\\cdot}8H_2O) and 0.06 ± 0.01 mBq (Th)/kg (Gd_2(SO_4)_3{\\cdot}8H_2O) by the ICP-MS.

  6. [Serum potassium levels and long-term mortality in the elderly with hypertension].

    PubMed

    Heras Benito, M; Fernández-Reyes, M J; Guerrero Díaz, M T; Muñoz Pascual, A

    There is increasing evidence that small variations within the normal range (3.5-5mEq/L) of potassium are associated with mortality. To determine whether there is an association between serum potassium level (sK) and mortality in a cohort of elderly hypertensive patients. A retrospective, observational study was conducted on patients who had sK levels available in a period of clinical stability during their recruitment between January and April 2006 and followed-up for 10 years. The study obtained a total of 62 stable patients, with a mean age of 82.19±6 years (range 69-97), with 74.2% women, 33.9% diabetics, 20.3% with a history of heart failure, Ischaemic heart disease was observed in 19.4% and 44.3% received Angiotensin Converting Enzyme (ACE) inhibitors. An analysis was performed on the mortality rate during the 10 year period. The statistics were performed using the SPSS15.0 package. There were 49 deaths. The sK had a normal distribution. Baseline mean sK levels and median were 4.45±0.5mEq/L (range 3.1-5.5 mEq/L). Baseline sK levels were significantly higher in diabetic patients and patients on ACE inhibitors. The patients that died had higher sK levels (4.53±0.49mEq/L versus 4.14±0.40mEq/L, P=.011). Survival estimated using Kaplan Meier showed that patients with sK levels higher than the median and P75 had higher mortality. In our study, sK levels greater than 4.45mEq/L were associated with mortality. When selecting antihypertensive treatment in hypertensive elderly patients,, the use of ACE inhibitors should be assessed individually, with close monitoring at sK levels and try to keep them in the lower limit of the normal range (<4.45 mEq/L). Copyright © 2017 SEH-LELHA. Publicado por Elsevier España, S.L.U. All rights reserved.

  7. The relationship between solar keratoses and squamous cell carcinomas among Japanese.

    PubMed

    Takemiya, M; Ohtsuka, H; Miki, Y

    1990-06-01

    Between 1976 and 1988, 135 patients with solar keratosis (SK) and 53 patients with squamous cell carcinoma (SCC) on the sun-exposed skin, but without apparent preceding diseases such as burn scars, chronic radiodermatitis, chronic arsenic poisoning, or xeroderma pigmentosum, were encountered. Sixteen of the SCC patients also had SK on other areas of sun-exposed skin. There were 31 SCC patients also showing SK (SK-SCC) and 22 SCC not showing SK (DN-SCC) within the same histologic sections. The mean ages of the patients with SK-SCC and with DN-SCC were similar. Metastases to regional lymph nodes were observed in 5 SK-SCC patients, of whom 3 died of the disease, and in 5 DN-SCC patients, of whom 4 died of the disease. The five-year post-operative survival rates were 70% in SK-SCC and 74% in DN-SCC; the ten-year post-operative survival rates were 70% in SK-SCC and 44% in DN-SCC.

  8. Small-Conductance Ca2+-Activated Potassium Channels Negatively Regulate Aldosterone Secretion in Human Adrenocortical Cells.

    PubMed

    Yang, Tingting; Zhang, Hai-Liang; Liang, Qingnan; Shi, Yingtang; Mei, Yan-Ai; Barrett, Paula Q; Hu, Changlong

    2016-09-01

    Aldosterone, which plays a key role in maintaining water and electrolyte balance, is produced by zona glomerulosa cells of the adrenal cortex. Autonomous overproduction of aldosterone from zona glomerulosa cells causes primary hyperaldosteronism. Recent clinical studies have highlighted the pathological role of the KCNJ5 potassium channel in primary hyperaldosteronism. Our objective was to determine whether small-conductance Ca(2+)-activated potassium (SK) channels may also regulate aldosterone secretion in human adrenocortical cells. We found that apamin, the prototypic inhibitor of SK channels, decreased membrane voltage, raised intracellular Ca(2+) and dose dependently increased aldosterone secretion from human adrenocortical H295R cells. By contrast, 1-Ethyl-2-benzimidazolinone, an agonist of SK channels, antagonized apamin's action and decreased aldosterone secretion. Commensurate with an increase in aldosterone production, apamin increased mRNA expression of steroidogenic acute regulatory protein and aldosterone synthase that control the early and late rate-limiting steps in aldosterone biosynthesis, respectively. In addition, apamin increased angiotensin II-stimulated aldosterone secretion, whereas 1-Ethyl-2-benzimidazolinone suppressed both angiotensin II- and high K(+)-stimulated production of aldosterone in H295R cells. These findings were supported by apamin-modulation of basal and angiotensin II-stimulated aldosterone secretion from acutely prepared slices of human adrenals. We conclude that SK channel activity negatively regulates aldosterone secretion in human adrenocortical cells. Genetic association studies are necessary to determine whether mutations in SK channel subtype 2 genes may also drive aldosterone excess in primary hyperaldosteronism. © 2016 American Heart Association, Inc.

  9. TNF-alpha induction of GM2 expression on renal cell carcinomas promotes T cell dysfunction.

    PubMed

    Raval, Gira; Biswas, Soumika; Rayman, Patricia; Biswas, Kaushik; Sa, Gaurisankar; Ghosh, Sankar; Thornton, Mark; Hilston, Cynthia; Das, Tanya; Bukowski, Ronald; Finke, James; Tannenbaum, Charles S

    2007-05-15

    Previous studies from our laboratory demonstrated the role of tumor-derived gangliosides as important mediators of T cell apoptosis, and hence, as one mechanism by which tumors evade immune destruction. In this study, we report that TNF-alpha secreted by infiltrating inflammatory cells and/or genetically modified tumors augments tumor-associated GM2 levels, which leads to T cell death and immune dysfunction. The conversion of weakly apoptogenic renal cell carcinoma (RCC) clones to lines that can induce T cell death requires 3-5 days of TNF-alpha pretreatment, a time frame paralleling that needed for TNF-alpha to stimulate GM2 accumulation by SK-RC-45, SK-RC-54, and SK-RC-13. RCC tumor cell lines permanently transfected with the TNF-alpha transgene are similarly toxic for T lymphocytes, which correlates with their constitutively elevated levels of GM2. TNF-alpha increases GM2 ganglioside expression by enhancing the mRNA levels encoding its synthetic enzyme, GM2 synthase, as demonstrated by both RT-PCR and Southern analysis. The contribution of GM2 gangliosides to tumor-induced T cell death was supported by the finding that anti-GM2 Abs significantly blocked T cell apoptosis mediated by TNF-alpha-treated tumor cells, and by the observation that small interfering RNA directed against TNF-alpha abrogated GM2 synthase expression by TNF-transfected SK-RC-45, diminished its GM2 accumulation, and inhibited its apoptogenicity for T lymphocytes. Our results indicate that TNF-alpha signaling promotes RCC-induced killing of T cells by stimulating the acquisition of a distinct ganglioside assembly in RCC tumor cells.

  10. Cytotoxic triterpene diglycosides from the sea cucumber Stichopus horrens.

    PubMed

    Cuong, Nguyen Xuan; Vien, Le Thi; Hoang, Le; Hanh, Tran Thi Hong; Thao, Do Thi; Thanh, Nguyen Van; Nam, Nguyen Hoai; Thung, Do Cong; Kiem, Phan Van; Minh, Chau Van

    2017-07-01

    Using various chromatographic separation techniques, eight triterpene diglycosides (1-8), including four new compounds namely stichorrenosides A-D (1-4), were isolated from a methanol extract of the Vietnamese sea cucumber S. horrens. Their structures were elucidated based on spectroscopic analyses, including HR ESI MS, 1D and 2D NMR. Their in vitro cytotoxic activity against five human cancer cell lines, Hep-G2 (hepatoma cancer), KB (epidermoid carcinoma), LNCaP (prostate cancer), MCF7 (breast cancer), and SK-Mel2 (melanoma), was evaluated using SRB methods. Stichorrenoside D (4), stichoposide A (5), and 3β-O-[β-d-xylopyranosyl-(1→2)-β-d-xylopyranosyl]-23S-acetoxyholost-7-ene (7) showed strong cytotoxicity on all five tested cancer cell lines, whereas significant effect was observed for stichorrenoside C (3) and stichoposide B (6). Copyright © 2017 Elsevier Ltd. All rights reserved.

  11. Plasma metabolomic profiling of dairy cows affected with ketosis using gas chromatography/mass spectrometry

    PubMed Central

    2013-01-01

    Background Ketosis is an important problem for dairy cows` production performance. However, it is still little known about plasma metabolomics details of dairy ketosis. Results A gas chromatography/mass spectrometry (GC/MS) technique was used to investigate plasma metabolic differences in cows that had clinical ketosis (CK, n=22), subclinical ketosis (SK, n=32), or were clinically normal controls (NC, n=22). The endogenous plasma metabolome was measured by chemical derivatization followed by GC/MS, which led to the detection of 267 variables. A two-sample t-test of 30, 32, and 13 metabolites showed statistically significant differences between SK and NC, CK and NC, and CK and SK, respectively. Orthogonal signal correction-partial least-square discriminant analysis (OPLS-DA) revealed that the metabolic patterns of both CK and SK were mostly similar, with the exception of a few differences. The development of CK and SK involved disturbances in many metabolic pathways, mainly including fatty acid metabolism, amino acid metabolism, glycolysis, gluconeogenesis, and the pentose phosphate pathway. A diagnostic model arbitrary two groups was constructed using OPLS-DA and receiver–operator characteristic curves (ROC). Multivariate statistical diagnostics yielded the 19 potential biomarkers for SK and NC, 31 for CK and NC, and 8 for CK and SK with area under the curve (AUC) values. Our results showed the potential biomarkers from CK, SK, and NC, including carbohydrates, fatty acids, amino acids, even sitosterol and vitamin E isomers, etc. 2-piperidinecarboxylic acid and cis-9-hexadecenoic acid were closely associated with metabolic perturbations in ketosis as Glc, BHBA and NEFA for dealing with metabolic disturbances of ketosis in clinical practice. However, further research is needed to explain changes of 2,3,4-trihydroxybutyric acid, 3,4-dihydroxybutyric acid, α-aminobutyric acid, methylmalonic acid, sitosterol and α-tocopherol in CK and SK, and to reveal differences

  12. IgG4 autoantibodies against muscle-specific kinase undergo Fab-arm exchange in myasthenia gravis patients.

    PubMed

    Koneczny, Inga; Stevens, Jo A A; De Rosa, Anna; Huda, Saif; Huijbers, Maartje G; Saxena, Abhishek; Maestri, Michelangelo; Lazaridis, Konstantinos; Zisimopoulou, Paraskevi; Tzartos, Socrates; Verschuuren, Jan; van der Maarel, Silvère M; van Damme, Philip; De Baets, Marc H; Molenaar, Peter C; Vincent, Angela; Ricciardi, Roberta; Martinez-Martinez, Pilar; Losen, Mario

    2017-02-01

    Autoimmunity mediated by IgG4 subclass autoantibodies is an expanding field of research. Due to their structural characteristics a key feature of IgG4 antibodies is the ability to exchange Fab-arms with other, unrelated, IgG4 molecules, making the IgG4 molecule potentially monovalent for the specific antigen. However, whether those disease-associated antigen-specific IgG4 are mono- or divalent for their antigens is unknown. Myasthenia gravis (MG) with antibodies to muscle specific kinase (MuSK-MG) is a well-recognized disease in which the predominant pathogenic IgG4 antibody binds to extracellular epitopes on MuSK at the neuromuscular junction; this inhibits a pathway that clusters the acetylcholine (neurotransmitter) receptors and leads to failure of neuromuscular transmission. In vitro Fab-arm exchange-inducing conditions were applied to MuSK antibodies in sera, purified IgG4 and IgG1-3 sub-fractions. Solid-phase cross-linking assays were established to determine the extent of pre-existing and inducible Fab-arm exchange. Functional effects of the resulting populations of IgG4 antibodies were determined by measuring inhibition of agrin-induced AChR clustering in C2C12 cells. To confirm the results, κ/κ, λ/λ and hybrid κ/λ IgG4s were isolated and tested for MuSK antibodies. At least fifty percent of patients had IgG4, but not IgG1-3, MuSK antibodies that could undergo Fab-arm exchange in vitro under reducing conditions. Also MuSK antibodies were found in vivo that were divalent (monospecific for MuSK). Fab-arm exchange with normal human IgG4 did not prevent the inhibitory effect of serum derived MuSK antibodies on AChR clustering in C2C12 mouse myotubes. The results suggest that a considerable proportion of MuSK IgG4 could already be Fab-arm exchanged in vivo. This was confirmed by isolating endogenous IgG4 MuSK antibodies containing both κ and λ light chains, i.e. hybrid IgG4 molecules. These new findings demonstrate that Fab-arm exchanged antibodies

  13. Melatonin partially protects 661W cells from H2O2-induced death by inhibiting Fas/FasL-caspase-3.

    PubMed

    Sánchez-Bretaño, Aída; Baba, Kenkichi; Janjua, Uzair; Piano, Ilaria; Gargini, Claudia; Tosini, Gianluca

    2017-01-01

    Previous studies have shown that melatonin (MEL) signaling is involved in the modulation of photoreceptor viability during aging. Recent work by our laboratory suggested that MEL may protect cones by modulating the Fas/FasL-caspase-3 pathway. In this study, we first investigated the presence of MEL receptors (MT 1 and MT 2 ) in 661W cells, then whether MEL can prevent H 2 O 2 -induced cell death, and last, through which pathway MEL confers protection. The mRNA and proteins of the MEL receptors were detected with quantitative PCR (q-PCR) and immunocytochemistry, respectively. To test the protective effect of MEL, 661W cells were treated with H 2 O 2 for 2 h in the presence or absence of MEL, a MEL agonist, and an antagonist. To study the pathways involved in H 2 O 2 -mediated cell death, a Fas/FasL antagonist was used before the exposure to H 2 O 2 . Finally, Fas/FasL and caspase-3 mRNA was analyzed with q-PCR and immunocytochemistry in cells treated with H 2 O 2 and/or MEL. Cell viability was analyzed by using Trypan Blue. Both MEL receptors (MT 1 and MT 2 ) were detected at the mRNA and protein levels in 661W cells. MEL partially prevented H 2 O 2 -mediated cell death (20-25%). This effect was replicated with IIK7 (a melatonin receptor agonist) when used at a concentration of 1 µM. Preincubation with luzindole (a melatonin receptor antagonist) blocked MEL protection. Kp7-6, an antagonist of Fas/FasL, blocked cell death caused by H 2 O 2 similarly to what was observed for MEL. Fas, FasL, and caspase-3 expression was increased in cells treated with H 2 O 2 , and this effect was prevented by MEL. Finally, MEL treatment partially prevented the activation of caspase-3 caused by H 2 O 2 . The results demonstrate that MEL receptors are present and functional in 661W cells. MEL can prevent photoreceptor cell death induced by H 2 O 2 via the inhibition of the proapoptotic pathway Fas/FasL-caspase-3.

  14. Synthesis and Anticancer Activity of 2-(Alkyl-, Alkaryl-, Aryl-, Hetaryl-)-[1,2,4]triazolo[1,5-c]quinazolines

    PubMed Central

    Kovalenko, Sergiy I.; Antypenko, Lyudmyla M.; Bilyi, Andriy K.; Kholodnyak, Sergiy V.; Karpenko, Olexandr V.; Antypenko, Olexii M.; Mykhaylova, Natalya S.; Los, Tetyana I.; Kolomoets, Olexandra S.

    2013-01-01

    The combinatorial library of novel potential anticancer agents, namely, 2-(alkyl-, alkaryl-, aryl-, hetaryl-)[1,2,4]triazolo[1,5-c]quinazolines, was synthesized by the heterocyclization of the alkyl-, alkaryl-, aryl-, hetarylcarboxylic acid (3H-quinazoline-4-ylidene)hydrazides by oxidative heterocyclization of the 4-(arylidenehydrazino)quinazolines using bromine, and by the heterocyclization of N-(2-cyanophenyl)formimidic acid ethyl ester. The optimal method for synthesis of the s-triazolo[1,5-c]quinazolines appeared to be cyclocondensation of the corresponding carboxylic acid (3H-quinazoline-4-ylidene)hydrazides. The compounds’ structures were established by 1H, 13C NMR, LC- and EI-MS analysis. The in vitro screening of anticancer activity determined the most active compound to be 3,4,5-trimethoxy-N′-[quinazolin-4(3H)-ylidene]benzohydrazide (3.20) in micromolar concentrations with the GI50 level (MG_MID, GI50 is 2.29). Thus, the cancer cell lines whose growth is greatly inhibited by compound 3.20 are: non-small cell lung cancer (NCI-H522, GI50=0.34), CNS (SF-295, GI50=0.95), ovarian (OVCAR-3, GI50=0.33), prostate (PC-3, GI50=0.56), and breast cancer (MCF7, GI50=0.52), leukemia (K-562, GI50=0.41; SR, GI50=0.29), and melanoma (MDA-MB-435, GI50=0.31; SK-MEL-5, GI50=0.74; UACC-62, GI50=0.32). SAR-analysis is also discussed. PMID:23833709

  15. Triterpene tetraglycosides from the sea cucumber Stichopus horrens.

    PubMed

    Vien, Le Thi; Hoang, Le; Hanh, Tran Thi Hong; Thanh, Nguyen Van; Cuong, Nguyen Xuan; Nam, Nguyen Hoai; Thung, Do Cong; Kiem, Phan Van; Minh, Chau Van

    2018-05-01

    Using various chromatographic separations, three triterpene tetraglycosides (1-3), including one new compound, namely stichorrenoside E (1) along with thelenotoside B (2) and deacetyl thelenotoside B (3), were isolated from the MeOH extract of the Vietnamese sea cucumber Stichopus horrens. Their structures were confirmed by spectroscopic experiments, such as 1D and 2D NMR and HR-ESI-MS. Deacetylated thelenotoside B (3) is firstly isolated as a natural product. Among these compounds, thelenotoside B (2) showed strong cytotoxicities against five human cancer cell lines as HepG2, KB, LNCaP, MCF7 and SK-Mel2 with the IC 50 values from 0.95 ± 0.08 to 1.90 ± 0.13 μM, whereas stichorrenoside E (1) and deacetyl thelenotoside B (3) exhibited significant activities with the IC 50 values from 6.87 ± 0.25 to 11.62 ± 1.05 μM.

  16. Taspine derivative 12k suppressed A549 cell migration through the Wnt/β-catenin and EphrinB2 signaling pathway.

    PubMed

    Dai, Bingling; Ma, Yujiao; Yang, Tianfeng; Wang, Wenjie; Zhang, Yanmin

    2017-03-01

    12k, a taspine derivative, has been demonstrated to have the potent anti-tumor activity in lung cancer and colorectal cancer. The study aims to further explore the underlying mechanisms of 12k on A549 cell migration in vitro. Our data demonstrated that 12k negatively regulated Wnt signaling pathway by suppressing the phosphorylation of LRP5/6, and inhibiting the expression and nuclear translocation of β-catenin. 12k was shown to downregulate MMP3 and MMP7 expression which regulated by β-catenin interacts with TCF/LEF in the nucleus, and effectively impaired the related migration protein expression of MMP2 and MMP9 in A549 cells. In addition, 12k repressed the EphrinB2 and its PDZ protein, impairing the VEGFR2 and VEGFR3 expression in A549 cells, as well as inhibited the downstream of VEGFR2 included PI3K/AKT/mTOR and ERK/MAPK signaling pathways. Taken together, our findings revealed that 12k suppressed migration of A549 cells through the Wnt/β-catenin signaling pathway and EphrinB2 related signaling pathway. Copyright © 2016 Elsevier Masson SAS. All rights reserved.

  17. Magnolol and honokiol regulate the calcium-activated potassium channels signaling pathway in Enterotoxigenic Escherichia coli-induced diarrhea mice.

    PubMed

    Deng, Yanli; Han, Xuefeng; Tang, Shaoxun; Xiao, Wenjun; Tan, Zhiliang; Zhou, Chuanshe; Wang, Min; Kang, Jinghe

    2015-05-15

    To explore the regulatory mechanisms of magnolol and honokiol on calcium-activated potassium channels signaling pathway in Enterotoxigenic Escherichia coli (ETEC)-induced diarrhea mice, the concentrations of serum chloride ion (Cl(-)), sodium ion (Na(+)), potassium ion (K(+)) and calcium ion (Ca(2+)) were measured. Additionally, the mRNA expressions of calmodulin 1 (CaM), calcium/calmodulin-dependent protein kinase II alpha subunit (CaMKIIα) and beta subunit (CaMKIIβ), ryanodine receptor 1, inositol 1,4,5-trisphosphate receptors (IP3 receptors), protein kinases C (PKC), potassium intermediate/small conductance calcium-activated channels (SK) and potassium large conductance calcium-activated channels(BK)were determined. A diarrhea mouse model was established using ETEC suspensions (3.29×10(9)CFU/ml) at a dosage of 0.02ml/g live body weight (BW). Magnolol or honokiol was intragastrically administered at dosages of 100 (M100 or H100), 300 (M300 or H300) and 500 (M500 or H500) mg/kg BW according to a 3×3 factorial arrangement. Magnolol and honokiol increased the Cl(-) and K(+) concentrations, further, upregulated the CaM, BKα1 and BKβ3 mRNA levels but downregulated the IP3 receptors 1, PKC, SK1, SK2, SK3, SK4 and BKβ4 mRNA expressions. Magnolol and honokiol did not alter the CaMKIIα, CaMKIIβ, ryanodine receptor 1, IP3 receptor 2, IP3 receptor 3, BKβ1 and BKβ2 mRNA expressions. These results clarify that magnolol and honokiol, acting through Ca(2+) channel blockade, inhibit the activation of IP3 receptor 1 to regulate the IP3-Ca(2+) store release, activate CaM to inhibit SK channels, and effectively suppress PKC kinases to promote BKα1 and BKβ3 channels opening and BKβ4 channel closing, which modulates the intestinal ion secretion. Copyright © 2015 Elsevier B.V. All rights reserved.

  18. Synthesis and anti-lung cancer activity of a novel arsenomolybdate compound

    NASA Astrophysics Data System (ADS)

    Zhu, Tian-Tian; Wang, Juan; Chen, Song-Hu

    2017-12-01

    The new compound based on Wells-Dawson-type arsenomolybdate: [{Cu10(pz)11Cl4}{As2IIIAs2VMo6VMo12VIO62}]·H2O (1) has been hydrothermally synthesized and characterized by single-crystal X-ray diffraction analysis, X-ray powder diffraction (XRPD), XPS spectroscopy and thermogravimetric analysis (TG). Compound 1 is consisted of two As caps Wells-dawson-type arsenomolybdate and {Cu10(py)11} complexes by chloride bridge. In addition, the antitumor effects of the title compound 1 were studied on three human lung cancer cells (A549, SK-LU-1 and SW1573). The results showed that compared with the positive reference drug carboplatin, compound 1 displayed efficient antitumor activity.

  19. Calculating degree-based topological indices of dominating David derived networks

    NASA Astrophysics Data System (ADS)

    Ahmad, Muhammad Saeed; Nazeer, Waqas; Kang, Shin Min; Imran, Muhammad; Gao, Wei

    2017-12-01

    An important area of applied mathematics is the Chemical reaction network theory. The behavior of real world problems can be modeled by using this theory. Due to applications in theoretical chemistry and biochemistry, it has attracted researchers since its foundation. It also attracts pure mathematicians because it involves interesting mathematical structures. In this report, we compute newly defined topological indices, namely, Arithmetic-Geometric index (AG1 index), SK index, SK1 index, and SK2 index of the dominating David derived networks [1, 2, 3, 4, 5].

  20. Synthesis and characterization of a pH-sensitive conjugate of isoniazid with Fe3O4@SiO2 magnetic nanoparticles.

    PubMed

    Sedlák, Miloš; Bhosale, Dattatry Shivajirao; Beneš, Ludvík; Palarčík, Jiří; Kalendová, Andrea; Královec, Karel; Imramovský, Aleš

    2013-08-15

    The Letter describes the preparation and characterization of a conjugate of isoniazid (INH) with magnetic nanoparticles Fe3O4@SiO2 115±60 nm in size. The INH molecules were attached to the surface of nanoparticles by a covalent pH-sensitive amidine bond. The conjugate was characterized by X-ray diffraction, SEM, dynamic light scattering, IR spectroscopy and microanalysis. The conjugate released isoniazid under in vitro conditions (pH=4; 37 °C; t1/2≈115 s). In addition, the cytotoxicity of the Fe3O4@SiO2-INH conjugate was evaluated in SK-BR-3 cells using the xCELLigence system. Copyright © 2013 Elsevier Ltd. All rights reserved.

  1. TRIM25 is associated with cisplatin resistance in non-small-cell lung carcinoma A549 cell line via downregulation of 14-3-3σ.

    PubMed

    Qin, Xia; Qiu, Feng; Zou, Zhen

    2017-11-04

    Lung cancer, in particular, non-small cell lung cancer (NSCLC), is the leading cause of cancer-related mortality. Cis-Diamminedichloroplatinum (cisplatin, CDDP) as first-line chemotherapy for NSCLC, but resistance occurs frequently. We previously reported that Tripartite motif protein 25 (TRIM25) was highly expressed in cisplatin-resistant human lung adenocarcinoma A549 cells (A549/CDDP) in comparison with its parental A549 cells. Herein, we take a further step to demonstrate the association of TRIM25 and cisplatin resistance and also the underlying mechanisms. Knockdown of TRIM25 by RNA interference in A549/CDDP cells decreased half maximal inhibitory concentration (IC 50 ) values and promoted apoptosis in response to cisplatin, whereas overexpression of TRIM25 had opposite effects. More importantly, we found that concomitant knockdown of 14-3-3σ and TRIM25 absolutely reversed the decreased MDM2, increased p53, increased cleaved-Capsese3 and decreased IC 50 value induced by knockdown of TRIM25 individually, suggesting that TRIM25 mediated cisplatin resistance primarily through downregulation of 14-3-3σ. Our results indicate that TRIM25 is associated with cisplatin resistance and 14-3-3σ-MDM2-p53 signaling pathway is involved in this process, suggesting targeting TRIM25 may be a potential strategy for the reversal of cisplatin resistance. Copyright © 2017 Elsevier Inc. All rights reserved.

  2. Siderophile and chalcophile metals as tracers of the evolution of the Siberian Trap in the Noril'sk region, Russia

    NASA Astrophysics Data System (ADS)

    Brügmann, G. E.; Naldrett, A. J.; Asif, M.; Lightfoot, P. C.; Gorbachev, N. S.; Fedorenko, V. A.

    1993-05-01

    In this study Cu, Ni, and platinum-group elements (PGE) were determined in a sequence of basaltic and picritic lavas from the Siberian Trap in the Noril'sk area of Russia to constrain genetic relationships between the basalts and the petrogenesis of Ni-Cu-PGE sulfide deposits associated with the Talnakh and Noril'sk intrusions. In the most primitive basalts (8-19 wt% MgO) of the Tuklonsky (Tk) suite, Pt and Pd concentrations range from 4-13 ppb, increasing with decreasing MgO content; whereas Ir contents decrease with MgO from 0.8-0.05 ppb. The contrasting behavior of these elements, which all have very high sulfide-silicate partition coefficients, as well as the primitive mantle-like ratios of Cu/Y and Pd/Y, suggests that these magmas were not sulfide-saturated. The high PGE abundances imply that their parental magmas were also not sulfide saturated during partial melting in the mantle. Due to sulfide segregation, the overlying basalts of the Nadezhdinsky (Nd) series are low in Cu and Ni (52 and 38 ppm, respectively); highly depleted in all PGE; and have very low Cu/Y, Pd/Y, and Pd/Cu ratios. However, in stratigraphically higher levels, Cu, Ni, and PGE concentrations increase systematically through the Morongovsky (Mr) suite to reach a concentration plateau in the uppermost Mokulaevsky (Mk) suite (Pt 8 ppb; Pd: 9 ppb; Ir: 0.12 ppb; Rh: 0.4 ppb). At the same time, ratios such as Cu/Y increase and approach primitive mantle values. However, ratios involving PGE, such as Pd/Y, remain low, suggesting the removal of small amounts of sulfide (0.01-0.03%). The compositional variations in the basalts and the sulfide liquids can be quantitatively described by fractional segregation of a sulfide liquid in an open- or closed-system magma chamber. The latter model suggests that the basalts represent the eruption products of a zoned magma chamber in which light magma, with crustal components contaminated, overlies less contaminated, denser magma. Crustal contamination caused

  3. Na+-taurocholate cotransporting polypeptide (NTCP/SLC10A1) ortholog in the marine skate Leucoraja erinacea is not a physiological bile salt transporter

    PubMed Central

    Yu, Dongke; Zhang, Han; Lionarons, Daniel A.; Boyer, James L.

    2017-01-01

    The Na+-dependent taurocholate cotransporting polypeptide (NTCP/SLC10A1) is a hepatocyte-specific solute carrier, which plays an important role in maintaining bile salt homeostasis in mammals. The absence of a hepatic Na+-dependent bile salt transport system in marine skate and rainbow trout raises a question regarding the function of the Slc10a1 gene in these species. Here, we have characterized the Slc10a1 gene in the marine skate, Leucoraja erinacea. The transcript of skate Slc10a1 (skSlc10a1) encodes 319 amino acids and shares 46% identity to human NTCP (hNTCP) with similar topology to mammalian NTCP. SkSlc10a1 mRNA was mostly confined to the brain and testes with minimal expression in the liver. An FXR-bile salt reporter assay indicated that skSlc10a1 transported taurocholic acid (TCA) and scymnol sulfate, but not as effectively as hNTCP. An [3H]TCA uptake assay revealed that skSlc10a1 functioned as a Na+-dependent transporter, but with low affinity for TCA (Km = 92.4 µM) and scymnol sulfate (Ki = 31 µM), compared with hNTCP (TCA, Km = 5.4 µM; Scymnol sulfate, Ki = 3.5 µM). In contrast, the bile salt concentration in skate plasma was 2 µM, similar to levels seen in mammals. Interestingly, skSlc10a1 demonstrated transport activity for the neurosteroids dehydroepiandrosterone sulfate and estrone-3-sulfate at physiological concentration, similar to hNTCP. Together, our findings indicate that skSlc10a1 is not a physiological bile salt transporter, providing a molecular explanation for the absence of a hepatic Na+-dependent bile salt uptake system in skate. We speculate that Slc10a1 is a neurosteroid transporter in skate that gained its substrate specificity for bile salts later in vertebrate evolution. PMID:28077388

  4. Na+-taurocholate cotransporting polypeptide (NTCP/SLC10A1) ortholog in the marine skate Leucoraja erinacea is not a physiological bile salt transporter.

    PubMed

    Yu, Dongke; Zhang, Han; Lionarons, Daniel A; Boyer, James L; Cai, Shi-Ying

    2017-04-01

    The Na + -dependent taurocholate cotransporting polypeptide (NTCP/SLC10A1) is a hepatocyte-specific solute carrier, which plays an important role in maintaining bile salt homeostasis in mammals. The absence of a hepatic Na + -dependent bile salt transport system in marine skate and rainbow trout raises a question regarding the function of the Slc10a1 gene in these species. Here, we have characterized the Slc10a1 gene in the marine skate, Leucoraja erinacea The transcript of skate Slc10a1 (skSlc10a1) encodes 319 amino acids and shares 46% identity to human NTCP (hNTCP) with similar topology to mammalian NTCP. SkSlc10a1 mRNA was mostly confined to the brain and testes with minimal expression in the liver. An FXR-bile salt reporter assay indicated that skSlc10a1 transported taurocholic acid (TCA) and scymnol sulfate, but not as effectively as hNTCP. An [ 3 H]TCA uptake assay revealed that skSlc10a1 functioned as a Na + -dependent transporter, but with low affinity for TCA ( K m = 92.4 µM) and scymnol sulfate ( K i = 31 µM), compared with hNTCP (TCA, K m = 5.4 µM; Scymnol sulfate, K i = 3.5 µM). In contrast, the bile salt concentration in skate plasma was 2 µM, similar to levels seen in mammals. Interestingly, skSlc10a1 demonstrated transport activity for the neurosteroids dehydroepiandrosterone sulfate and estrone-3-sulfate at physiological concentration, similar to hNTCP. Together, our findings indicate that skSlc10a1 is not a physiological bile salt transporter, providing a molecular explanation for the absence of a hepatic Na + -dependent bile salt uptake system in skate. We speculate that Slc10a1 is a neurosteroid transporter in skate that gained its substrate specificity for bile salts later in vertebrate evolution. Copyright © 2017 the American Physiological Society.

  5. Properties of wine polymeric pigments formed from anthocyanin and tannins differing in size distribution and subunit composition.

    PubMed

    Bindon, Keren; Kassara, Stella; Hayasaka, Yoji; Schulkin, Alex; Smith, Paul

    2014-11-26

    To explore the effect of tannin composition on pigment formation, model ferments of purified 3-O-monoglucoside anthocyanins (ACN) were conducted either alone or in the presence of two different tannins. Tannins were isolated from grape seeds (Sd) or skins (Sk) following exhaustive extraction in 70% v/v acetone. The Sd and Sk tannin fractions had a mean degree of polymerization of 5.2 and 25.6, respectively. The Sd fraction was highly galloylated, at 22%, but galloylation was <2% in the Sk fraction. The Sk fraction was distinguished by a high proportion of prodelphinidin, at 58%. After a 6 month aging period, polymeric pigments were quantified and their color properties determined following isolation by solid-phase extraction. Wine color and polymeric pigment were highest in the treatment containing ACN+Sd and similar in the ACN+Sk and ACN treatments. The same trend between treatments was observed for total and polymeric nonbleachable pigments. Only minor changes in tannin subunit composition were found following ACN incorporation, but the size distribution of polymeric pigments determined by gel permeation chromatography decreased, in particular for the ACN+Sk treatment. Color incorporation in the higher molecular mass range was lower for ACN+Sk wines than for ACN+Sd wines. Compositional differences between the two tannin fractions may therefore limit the incorporation of ACNs in the colored form. The results suggest that in the ACN+Sk and ACN treatments, the formation of lower molecular mass oligomeric pigments was favored. In polymeric pigments derived from ACNs, the presence of ethyl- and vinyl-linked ACNs to the level of trimers was identified using mass spectrometry.

  6. Vertebrate rod photoreceptors express both BK and IK calcium-activated potassium channels, but only BK channels are involved in receptor potential regulation.

    PubMed

    Pelucchi, Bruna; Grimaldi, Annalisa; Moriondo, Andrea

    2008-01-01

    In salamander rods, Ca(2+)-activated K(+) current (I(KCa)) provides an effective "clamp" of the dark membrane potential to its normal resting level. By a combination of electrophysiological, pharmacological, and immunohistochemical approaches, we show that salamander rods functionally express large-conductance Ca(2+)- and voltage-dependent potassium (BK) channel and intermediate-conductance Ca(2+)-dependent potassium (IK) channel, but not small-conductance Ca(2+)-dependent potassium channel (SK) subtypes. Application of 100 nM iberiotoxin and 100 nM clotrimazole reduced net I(KCa) to 36% and 63%, respectively, whereas the current was unaffected by application of 1 microM apamin. Consistently, anti- SK1, -SK2, and -SK3 antibodies were unable to stain rod photoreceptors, whereas both anti-BK and -SK4/ IK1 antibodies heavily stained the ellipsoid region of the inner segments of the rods. Moreover, by using current-clamp experiments, it was clearly seen that the strong clamping effect of the total I(KCa) was lost when IbTx, but not CLTZ, was applied to the bath. This behavior strongly suggests that of BK and IK channels, only the former are responsible for the clamping effect on the photoreceptor membrane potential.

  7. PM2.5 induces Nrf2-mediated defense mechanisms against oxidative stress by activating PIK3/AKT signaling pathway in human lung alveolar epithelial A549 cells.

    PubMed

    Deng, Xiaobei; Rui, Wei; Zhang, Fang; Ding, Wenjun

    2013-06-01

    It has been well documented in in vitro studies that ambient airborne particulate matter (PM) with an aerodynamic diameter less than 2.5 μm (PM(2.5)) is capable of inducing oxidative stress, which plays a key role in PM(2.5)-mediated cytotoxicity. Although nuclear factor erythroid-2-related factor 2 (Nrf2) has been shown to regulate the intracellular defense mechanisms against oxidative stress, a potential of the Nrf2-mediated cellular defense against oxidative stress induced by PM(2.5) remains to be determined. This study was aimed to explore the potential signaling pathway of Nrf2-mediated defense mechanisms against PM(2.5)-induced oxidative stress in human type II alveolar epithelial A549 cells. We exposed A549 cells to PM(2.5) particles collected from Beijing at a concentration of 16 μg/cm(2). We observed that PM(2.5) triggered an increase of intracellular reactive oxygen species (ROS) in a time-dependent manner during a period of 2 h exposure. We also found that Nrf2 overexpression suppressed and Nrf2 knockdown increased PM(2.5)-induced ROS generation. Using Western blot and confocal microscopy, we found that PM(2.5) exposure triggered significant translocation of Nrf2 into nucleus, resulting in AKT phosphorylation and significant transcription of ARE-driven phases II enzyme genes, such as NAD(P)H:quinone oxidoreductase (NQO-1), heme oxygenase-1 (HO-1), and glutamate-cysteine ligase catalytic subunit (GCLC) in A549 cells. Evaluation of signaling pathways showed that a phosphatidylinositol 3-kinase (PI3K) inhibitor (LY294002), but not an ERK 1/2 inhibitor (PD98059) or a p38 MAPK (SB203580), significantly down-regulated PM(2.5)-induced Nrf2 nuclear translocation and HO-1 mRNA expression, indicating PI3K/AKT is involved in the signaling pathway leads to the PM(2.5)-induced nuclear translocation of Nrf2 and subsequent Nrf2-mediated HO-1 transcription. Taken together, our results suggest that PM(2.5)-induced ROS may function as signaling molecules to activate Nrf

  8. Study of energetic particle physics with advanced ECEI system on the HL-2A tokamak

    NASA Astrophysics Data System (ADS)

    Shi, Zhongbing; Jiang, Min; Yu, Liming; Chen, Wei; Shi, Peiwan; Zhong, Wulyu; Yang, Zengchen; Zhang, Boyu; Ji, Xiaoquan; Li, Yonggao; Zhou, Yan; Song, Shaodong; Huang, Mei; Song, Xianming; Li, Jiaxuan; Yuan, Baoshan; Fu, Bingzhong; Liu, Zetian; Ding, Xuantong; Xu, Yuhong; Yang, Qingwei; Duan, Xuru

    2017-07-01

    Understanding the physics of energetic particles (EP) is crucial for the burning plasmas in next generation fusion devices such as ITER. In this work, three types of internal kink modes (a saturated internal kink mode (SK), a resonant internal kink mode (RK), and a double e-fishbone) excited by energetic particles in the low density discharges during ECRH/ECCD heating have been studied by the newly developed 24(poloidal) × 16(radial) = 384 channel ECEI system on the HL-2A tokamak. The SK and RK rotate in the electron diamagnetic direction poloidally and are destabilized by the energetic trapped electrons. The SK is destabilized in the case of qmin > 1, while the RK is destabilized in the case of qmin < 1. The double e-fishbone, which has two m/n = 1/1 modes propagating in the opposite directions poloidally, has been observed during plasma current ramp-up with counter-ECCD. Strong thermal transfer and mode coupling between the two m/n = 1/1 modes have been studied.

  9. Synthesis of [closo-B12(OH)11NH3]-: a new heterobifunctional dodecaborane scaffold for drug delivery applications.

    PubMed

    Bondarev, Oleg; Khan, Aslam A; Tu, Xiaoyan; Sevryugina, Yulia V; Jalisatgi, Satish S; Hawthorne, M Frederick

    2013-09-04

    Effective utilization of [closo-B12H12](2-) derivatives in targeted drug delivery applications depends upon an efficient strategy to differentiate at least one of the 12 vertices on the B12(2-) core. Precursor molecules must also be able to withstand the initial harsh hydrogen peroxide treatment necessary for hydroxylation of the B-H vertices. We report here a method for preparation of the ammonio derivative [closo-B12(OH)11NH3](-) and also demonstrate its utility in construction of a targeted drug delivery scaffold. Treatment of the precursor [closo-B12H11NH3](-) with hydrogen peroxide gives the corresponding nitro derivative [closo-B12(OH)11NO2](2-) in good yield. The nitro group is easily reduced with hydrogen over a Raney nickel catalyst to produce [closo-B12(OH)11NH3](-). The 11 hydroxyl groups can then be readily converted to carbonates or carbamates. As a proof-of-principle of its utility as a drug delivery system, we used the resulting vertex-differentiated ammonio derivative to construct a platinated pro-drug possessing 11 copies of a carboplatin analogue conjugated to the B12(2-) core via carbamate linkage and a fluorescein molecule attached at the remaining vertex by an amide linkage. In vitro cytotoxicity assays demonstrated that activity of an untagged analog was similar to carboplatin against platinum-sensitive A459 cells and higher than carboplatin against platinum-resistant SK-OV-3 cells. Further fluorescence microscopy revealed that the fluorescein-tagged pro-drug localizes to the nuclei of A459 cells.

  10. Evaluation of multifrequency bioelectrical impedance analysis in assessing body composition of wrestlers.

    PubMed

    Utter, Alan C; Lambeth, Pamela G

    2010-02-01

    To evaluate the accuracy of multifrequency bioelectrical impedance analysis (MFBIA) in assessing fat-free mass (FFM) in comparison with hydrostatic weighing (HW) and skinfolds (SK) in high school wrestlers in a hydrated state. Body composition was determined by MFBIA, HW, and three-site SK in 72 high school wrestlers (mean +/- SD; age = 15.3 +/- 1.4 yr, height = 1.71 +/- 0.08 m, body mass = 67.3 +/- 13.4 kg). Hydration state was quantified by evaluating urine specific gravity. There were no significant differences for estimated FFM between MFBIA (57.2 +/- 9.5 kg) and HW (57.0 +/- 10.1 kg) or SK (56.4 +/- 8.8 kg). The SEE for FFM with HW as the reference method were 2.73 kg for MFBIA and 2.66 kg for SK. Correlations were found for FFM between HW and MFBIA (r = 0.96, P < 0.001) and between HW and SK (r = 0.97, P < 0.001). A systematic bias was found for MFBIA because the difference between MFBIA and HW correlated with the FFM average of the two methods (r = -0.22, P < 0.001). A bias was also seen between SK and HW and correlated with the FFM average (r = -0.47, P < 0.001). This study demonstrates that MFBIA provides similar estimates of FFM when compared with HW in a heterogeneous high school wrestling population during a hydrated state. MFBIA is an attractive assessment tool, easy to use, and may be considered as an alternative field-based method of estimating the FFM of high school wrestlers.

  11. Secondary metabolites of Centaurea calolepis and evaluation of cnicin for anti-inflammatory, antioxidant, and cytotoxic activities.

    PubMed

    Erel, Sura Baykan; Karaalp, Canan; Bedir, Erdal; Kaehlig, Hanspeter; Glasl, Sabine; Khan, Shabana; Krenn, Liselotte

    2011-08-01

    Centaurea L. (Astreaceae) species are used as herbal remedies in Turkey. Centaurea calolepis Boiss. is an endemic species of Anatolia that has not been subjected to phytochemical studies except essential oil analysis. Secondary metabolite determination, isolation and structure elucidation of pure compounds were performed on C. calolepis. Cnicin, which is the main component of several Centaurea species, was tested for its in vitro anti-inflammatory, antioxidant and cytotoxic activities. Chloroform and methanol extracts of the aerial parts of C. calolepis were subjected to isolation process using column chromatography. The structures of the compounds were characterized by 1D- and 2D-NMR experiments. Thin-layer chromatography and high performance liquid chromatography were used in determination of phenolics. Cnicin was subjected to a panel of cellular assays to test for inhibition of nuclear factor κB (NF-κB), inducible nitric oxide synthase (iNOS), reactive oxygen species and cytotoxicity. Cnicin, lucenin-2, schaftoside and 3-O-feruloylquinic acid were isolated from C. calolepis extracts. Vicenin-2, vitexin, isovitexin, homoorientin, rutin, orientin, luteolin-7-O-glycoside and chlorogenic acid were determined in fractions. Cnicin showed inhibition of NF-κB and inhibition of iNOS activity with IC₅₀ Values of 1.8 and 6.5 µM, respectively. Cytotoxic activity of cnicin was observed toward pig kidney epithelial (LLC-PK₁₁), human malignant melanoma (SK-MEL) and human ductal carcinoma (BT-549) cells with IC₅₀ values of 23.3, 14.0 and 18.3 µM, respectively. This is the first detailed report of secondary metabolites of C. calolepis. Evaluation of biological activity of cnicin establishes the potential of this compound as an anti-inflammatory and cytotoxic agent.

  12. Features of the supercritical CO2-assisted immobilization of fluorinated tetraphenylporphyrins into tetrafluoroethylene copolymers

    NASA Astrophysics Data System (ADS)

    Shershnev, I. V.; Cherkasova, A. V.; Kopylov, A. S.; Glagolev, N. N.; Bragina, N. A.; Solov'eva, A. B.

    2017-07-01

    The immobilization of fluorinated tetraphenylporphyrins (FTPPs) into tetrafluoroethylene copolymers (fluoroplast F-42 and MF-4SK, a perfluorinated sulfonic acid cation exchanger in H+-form) is conducted in supercritical CO2 (scCO2). The effects the conditions of immobilization (the temperature and pressure of scCO2, reaction time, and the addition of cosolvents) and the structure of the carrier polymer have on the content of porphyrin in these polymers is studied. The porphyrin-loaded polymer systems are shown to exhibit photosensitizing activity in anthracene and cholesterol oxidation in scCO2. Under conditions of photocatalysis, chemical and functional stability is a feature of only MF-4SK polymer systems; this is attributed to the formation of protonated forms of the porphyrins and their interaction with SO3 --groups of the polymer (an ion exchange process), which prevents leaching of the FTPP from the polymer matrix. The photocatalytic process actually occurs inside the matrix of the perfluorinated copolymer, with the protonated form of the porphyrin acting as a photosensitizer. The rate constant of anthracene photooxidation in the presence of FTPP-loaded MF-4SK films in scCO2 is found to pass through a maximum as a function of the porphyrin content and the polymer film thickness. The use of such catalytic systems for cholesterol photooxidation in scCO2 is shown to produce a virtual monoproduct (yield, 10%): 6-formyl-B-norcholestane-3,5-diol, a compound with high biological activity.

  13. Anesthetic Management of a Patient With Antimuscle-Specific Kinase Antibody-Positive Myasthenia Gravis Undergoing an Open Cholecystectomy: A Case Report.

    PubMed

    Akatsu, Masahiko; Ikegami, Yukihiro; Tase, Choichiro; Nishikawa, Koichi

    2017-03-15

    Myasthenia gravis (MG) is an autoimmune disease characterized by the production of antibodies against the acetylcholine receptor, muscle-specific kinase (MuSK), or other proteins at the neuromuscular junction. MG with antibodies against MuSK (MuSK-MG) has been described recently. Here, we report the first case of anesthetic management of a patient with MuSK-MG undergoing an open cholecystectomy. In our case, propofol and remifentanil-based anesthesia were used for successful management without using muscle relaxants. Patients with MuSK-MG have predominantly ocular, bulbar, and respiratory symptoms that may increase the risk of aspiration. Anesthesiologists need to pay attention to perioperative respiratory failure and respiratory crisis.

  14. Removal of Volatile Organics from Humidified Air Streams by Absorption.

    DTIC Science & Technology

    1987-12-01

    Type Comments Activated Carbons: SK-4 nut shell LCL coconut shell WV-B coal base CT coconut shell Specialty Carbons: Graphpac graphitized carbon area...Capacity, g/g Percentage Sorbent ( virgin ) (treated)D Change SK-4 0.114 0.117 +2.6 Carbosieve S-11 0.195 0.180 -7.7 Spherocarb 0.149 0.151 +1.3 CT...and WV-B with respect to their adsorption capacities. CT and SK-4 are both coconut -derived carbons produced by the same manufacturer. Differences

  15. Enhanced reactive oxygen species overexpression by CuO nanoparticles in poorly differentiated hepatocellular carcinoma cells

    NASA Astrophysics Data System (ADS)

    Kung, Mei-Lang; Hsieh, Shu-Ling; Wu, Chih-Chung; Chu, Tian-Huei; Lin, Yu-Chun; Yeh, Bi-Wen; Hsieh, Shuchen

    2015-01-01

    Copper oxide nanoparticles (CuO NPs) are known to exhibit toxic effects on a variety of cell types and organs. To determine the oxidative impact of CuO NPs on hepatocellular carcinoma (HCC) cells, well-differentiated (HepG2) and poorly differentiated (SK-Hep-1) cells were exposed to CuO NPs. Cell viability assay showed that the median inhibition concentration (IC50) for SK-Hep-1 and HepG2 cells was 25 μg ml-1 and 85 μg ml-1, respectively. Cellular fluorescence intensity using DCFH-DA staining analysis revealed significant intracellular reactive oxygen species (ROS) generation of up to 242% in SK-Hep-1 cells, compared with 86% in HepG2 cells. HPLC analysis demonstrated that a CuO NP treatment caused cellular GSH depletion of 58% and a GSH/GSSG ratio decrease to ~0.1 in SK-Hep-1 cells. The oxidative stress caused by enhanced superoxide anion production was observed in both HepG2 (146%) and SK-Hep-1 (192%) cells. The Griess assay verified that CuO NPs induced NO production (170%) in SK-Hep-1 cells. Comet assay and western blot further demonstrated that CuO NPs induced severe DNA strand breakage (70%) in SK-Hep-1 cells and caused DNA damage via increased γ-H2AX levels. These results suggest that well-differentiated HepG2 cells possess a robust antioxidant defense system against CuO NP-induced ROS stress and exhibit more tolerance to oxidative stress. Conversely, poorly differentiated SK-Hep-1 cells exhibited a deregulated antioxidant defense system that allowed accumulation of CuO NP-induced ROS and resulted in severe cytotoxicity.Copper oxide nanoparticles (CuO NPs) are known to exhibit toxic effects on a variety of cell types and organs. To determine the oxidative impact of CuO NPs on hepatocellular carcinoma (HCC) cells, well-differentiated (HepG2) and poorly differentiated (SK-Hep-1) cells were exposed to CuO NPs. Cell viability assay showed that the median inhibition concentration (IC50) for SK-Hep-1 and HepG2 cells was 25 μg ml-1 and 85 μg ml-1, respectively

  16. The effect of ethnicity on the vascular responses to cold exposure of the extremities.

    PubMed

    Maley, Matthew J; Eglin, Clare M; House, James R; Tipton, Michael J

    2014-11-01

    Cold injuries are more prevalent in individuals of African descent (AFD). Therefore, we investigated the effect of extremity cooling on skin blood flow (SkBF) and temperature (T sk) between ethnic groups. Thirty males [10 Caucasian (CAU), 10 Asian (ASN), 10 AFD] undertook three tests in 30 °C air whilst digit T sk and SkBF were measured: (i) vasomotor threshold (VT) test--arm immersed in 35 °C water progressively cooled to 10 °C and rewarmed to 35 °C to identify vasoconstriction and vasodilatation; (ii) cold-induced vasodilatation (CIVD) test--hand immersed in 8 °C water for 30 min followed by spontaneous warming; (iii) cold sensitivity (CS) test--foot immersed in 15 °C water for 2 min followed by spontaneous warming. Cold sensory thresholds of the forearm and finger were also assessed. In the VT test, vasoconstriction and vasodilatation occurred at a warmer finger T sk in AFD during cooling [21.2 (4.4) vs. 17.0 (3.1) °C, P = 0.034] and warming [22.0 (7.9) vs. 12.1 (4.1) °C, P = 0.002] compared with CAU. In the CIVD test, average SkBF during immersion was greater in CAU [42 (24) %] than ASN [25 (8) %, P = 0.036] and AFD [24 (13) %, P = 0.023]. Following immersion, SkBF was higher and rewarming faster in CAU [3.2 (0.4) °C min(-1)] compared with AFD [2.5 (0.7) °C min(-1), P = 0.037], but neither group differed from ASN [3.0 (0.6) °C min(-1)]. Responses to the CS test and cold sensory thresholds were similar between groups. AFD experienced a more intense protracted finger vasoconstriction than CAU during hand immersion, whilst ASN experienced an intermediate response. This greater sensitivity to cold may explain why AFD are more susceptible to cold injuries.

  17. COX-2 expression positively correlates with PD-L1 expression in human melanoma cells.

    PubMed

    Botti, Gerardo; Fratangelo, Federica; Cerrone, Margherita; Liguori, Giuseppina; Cantile, Monica; Anniciello, Anna Maria; Scala, Stefania; D'Alterio, Crescenzo; Trimarco, Chiara; Ianaro, Angela; Cirino, Giuseppe; Caracò, Corrado; Colombino, Maria; Palmieri, Giuseppe; Pepe, Stefano; Ascierto, Paolo Antonio; Sabbatino, Francesco; Scognamiglio, Giosuè

    2017-02-23

    The resistance to PD-1/PD-L1 inhibitors for the treatment of melanoma have prompted investigators to implement novel clinical trials which combine immunotherapy with different treatment modalities. Moreover is also important to investigate the mechanisms which regulate the dynamic expression of PD-L1 on tumor cells and PD-1 on T cells in order to identify predictive biomarkers of response. COX-2 is currently investigated as a major player of tumor progression in several type of malignancies including melanoma. In the present study we investigated the potential relationship between COX-2 and PD-L1 expression in melanoma. Tumor samples obtained from primary melanoma lesions and not matched lymph node metastases were analyzed for both PD-L1 and COX-2 expression by IHC analysis. Status of BRAF and NRAS mutations was analyzed by sequencing and PCR. Co-localization of PD-L1 and COX-2 expression was analyzed by double fluorescence staining. Lastly the BRAF V600E A375 and NRAS Q61R SK-MEL-2 melanoma cell lines were used to evaluate the effect of COX-2 inhibition by celecoxib on expression of PD-L1 in vitro. BRAF V600E/V600K and NRAS Q61R/Q61L were detected in 57.8 and 8.9% of the metastatic lesions, and in 65.9 and 6.8% of the primary tumors, respectively. PD-L1 and COX-2 expression were heterogeneously expressed in both primary melanoma lesions and not matched lymph node metastases. A significantly lower number of PD-L1 negative lesions was found in primary tumors as compared to not matched metastatic lesions (P = 0.002). COX-2 expression significantly correlated with PD-L1 expression in both primary (P = 0.001) and not matched metastatic (P = 0.048) lesions. Furthermore, in melanoma tumors, cancer cells expressing a higher levels of COX-2 also co-expressed a higher level of PD-L1. Lastly, inhibition of COX-2 activity by celecoxib down-regulated the expression of PD-L1 in both BRAF V600E A375 and NRAS Q61R SK-MEL-2 melanoma cell lines. COX-2 expression correlates

  18. Diversity and community composition of pico- and nanoplanktonic protists in the Vistula River estuary (Gulf of Gdańsk, Baltic Sea)

    NASA Astrophysics Data System (ADS)

    Piwosz, Kasia; Całkiewicz, Joanna; Gołębiewski, Marcin; Creer, Simon

    2018-07-01

    Pico- and nanoplanktonic protists (eukaryotic microorganisms with cell size of <3 μm and 3-20 μm, respectively) are the key component of plankton communities. However, their diversity and distribution patterns along environmental factors are still poorly recognized, largely due to their very large phylogenetic diversity that has been determined only via the application of molecular methods over the past two decades. Here, we compared diversity and composition of active communities of pico- and nanoplantonic protists from three zones of the Vistula River estuary (Gulf of Gdańsk): freshwater, mixing (salinity 3.5) and brackish (salinity 7), in four seasons, by pyrosequencing the V3-V4 fragment of 18S rRNA taxonomy marker gene libraries. Alpha diversity was the highest at the brackish site, but the OTU (Operational Taxonomic Units) richness was characteristic for specific protist groups at each site. The active protistan communities in the freshwater and mixing zones (salinity 0-3.5) were similar (sharing >72% of phylotypes) and included centric diatoms (Stephanodiscus minutulus), synurophytes from clades C, E and F, and cryptophytes. However, at salinity of 7 at the brackish site the communities were significantly different from those in freshwater/mixing zone, and showed higher contributions of Dinophyceae, Mamiellophyceae, Telonemia, and picobiliphytes. The high similarity between the freshwater and mixing site, as well as high dissimilarity of the brackish site was observed in all months, despite seasonal shifts in pico- and nanoplantonic protistan communities. Seventy five percent of the observed variability in the communities was explained by combinations of temperature, salinity, nutrients and geographical distance, indicating interplay between species sorting and mass effects in shaping the active protistan communities in the Vistula River estuary. Groups that were more active in freshwaters and the mixing zone seemed to be more affected by mass effects of

  19. Inhibition of Sphingosine Kinase 1 Ameliorates Angiotensin II-Induced Hypertension and Inhibits Transmembrane Calcium Entry via Store-Operated Calcium Channel

    PubMed Central

    Wilson, Parker C.; Fitzgibbon, Wayne R.; Garrett, Sara M.; Jaffa, Ayad A.; Luttrell, Louis M.; Brands, Michael W.

    2015-01-01

    Angiotensin II (AngII) plays a critical role in the regulation of vascular tone and blood pressure mainly via regulation of Ca2+ mobilization. Several reports have implicated sphingosine kinase 1 (SK1)/sphingosine 1-phosphate (S1P) in the mobilization of intracellular Ca2+ through a yet-undefined mechanism. Here we demonstrate that AngII-induces biphasic calcium entry in vascular smooth muscle cells, consisting of an immediate peak due to inositol tris-phosphate-dependent release of intracellular calcium, followed by a sustained transmembrane Ca2+ influx through store-operated calcium channels (SOCs). Inhibition of SK1 attenuates the second phase of transmembrane Ca2+ influx, suggesting a role for SK1 in AngII-dependent activation of SOC. Intracellular S1P triggers SOC-dependent Ca2+ influx independent of S1P receptors, whereas external application of S1P stimulated S1P receptor-dependent Ca2+ influx that is insensitive to inhibitors of SOCs, suggesting that the SK1/S1P axis regulates store-operated calcium entry via intracellular rather than extracellular actions. Genetic deletion of SK1 significantly inhibits both the acute hypertensive response to AngII in anaesthetized SK1 knockout mice and the sustained hypertensive response to continuous infusion of AngII in conscious animals. Collectively these data implicate SK1 as the missing link that connects the angiotensin AT1A receptor to transmembrane Ca2+ influx and identify SOCs as a potential intracellular target for SK1. PMID:25871850

  20. Study of pharmaceutical industrial problems

    NASA Technical Reports Server (NTRS)

    Pincus, J. H.

    1979-01-01

    The growth of a human colon carcinoma cell line (SK-CO-1) and its production of carcinoembryonic antigen (CEA) in monolayer culture and on single layers of glass beads in unit gravity were evaluated. The limitations of using a microsphere-cell growth system in unit gravity were identified and how these may be overcome in space was considered. The project had the following tasks: (1) growth of cultured human colon carcinoma cells on a monolayer and CEA production; (2) evaluation of CEA production and release by SK-CO-1 cells grown on glass beads; (3) evaluation of other microcarriers for growing SK-CO-1 cells and determination of the minimum amount of culture medium needed for cell growth; and (4) growth of SK-CO-1 cells on collagen monolayers and CEA production.

  1. Using Public Participation to Improve MELs Energy Data Collection

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Kloss, Margarita; Cheung, Iris; Brown, Richard

    Miscellaneous and electronic loads (MELs) comprise an increasing share of building energy consumption. Large-scale data collection is needed to inform meaningful energy reduction strategies because of the diversity of MELs and our lack of understanding about how people use them. Traditional methods of data collection, however, usually incur high labor and metering equipment expenses. As an alternative, this paper investigates the feasibility of crowdsourcing data collection to satisfy at least part of the data collection needs with acceptable accuracy. We assessed the reliability and accuracy of crowd-sourced data by recruiting 18 volunteers and testing our crowdsourcing protocol. The protocol askedmore » volunteers to perform measurement tasks for three MELs devices of increasing complexity 1) record power meter and MELs product characteristics, 2) identify and measure all power modes available, and 3) report the measured power. Volunteers performed reasonably well for devices with functionalities with which they were familiar, but many could not correctly identify all available power modes in complex devices. Accuracy may improve when participants measure the power used by familiar devices in their home, or by providing more specific instructions, e.g. videos. Furthermore, crowdsourcing data collection from individual homeowners has the potential to generate valuable information about MELs energy use in homes when integrated with existing programs such as Home Energy Saver and Building America.« less

  2. Physiological responses to changes in relative humidity under thermally neutral, warm and hot conditions.

    PubMed

    Kakitsuba, Naoshi

    2016-07-01

    Four hypothetical thermophysiological responses to changes in relative humidity (Rh) under thermally neutral, warm, and hot conditions were proposed for a person at rest. Under thermally neutral and warm conditions, the first hypothetical response to an increase in Rh was a decrease in mean skin temperature (T¯sk) due to increase in mean evaporation rate (E¯sk), and the second hypothetical response to a decrease in Rh was a decrease, an increase, or no change in T¯sk, depending on changes in the E¯sk. Under hot conditions, the third hypothetical response to an increase in the Rh was an increase in T¯sk or decrease in T¯sk upon decrease in the Rh due to changes in E¯sk, and the forth hypothetical response to an increase in Rh was an increase in T¯sk due to increase in the peripheral blood flow rate (SkBF). To test these hypotheses, the T¯sk and E¯sk of four young male volunteers were measured at 28°C, 30°C, or 32°C while the Rh was maintained at 40% or 80% Rh for 60min after 20min exposure at 60% Rh (control condition). In a second experiment, the T¯sk, E¯sk, and SkBF of five young male volunteers were measured at 34°C-40% Rh or 36°C-40% Rh, or 34°C-70% Rh or 36°C-70% Rh for 60min after 20min exposure at 28°C-60% Rh (control condition). The first hypothesis was partly supported by the findings that the T¯sk was lower than the control values at 28°C-80% Rh and the E¯sk was higher than the control values at 80% Rh at any tested temperature. The second hypothesis was partly supported by the findings that the T¯sk was lower than the control values at 28°C-40% Rh, and there were small changes in both T¯sk and E¯sk at 30°C-40% Rh. The third and fourth hypotheses were supported by the findings that the T¯sk at 36°C-70% Rh was significantly higher (p<0.01) than at 36°C-40% Rh, the E¯sk was significantly higher (p<0.01) at 70% Rh than at 40% Rh, and SkBF was positively correlated with T¯sk. Copyright © 2016 Elsevier Ltd. All rights reserved.

  3. Forearm Skin Blood Flow After Kinesiology Taping in Healthy Soccer Players: An Exploratory Investigation

    PubMed Central

    Woodward, Kirsty A.; Unnithan, Vish; Hopkins, Nicola D.

    2015-01-01

    Context Kinesiology tape (KT) has become popular among athletes for both injury prevention and rehabilitation due to its reported therapeutic effects, including facilitation of lymphatic flow and enhanced peripheral blood flow. However, evidence to support such claims is insufficient. Objective To determine whether KT improves skin blood flow (SkBF) responses in young, elite soccer players. Design Randomized crossover study. Setting Research laboratory. Patients or Other Participants Thirteen healthy, elite, adolescent male soccer players (age = 14.7 ± 0.6 years). Intervention(s) Participants completed 2 experimental trials; during trial 1, the volar aspect of the dominant forearm was taped. Forearm SkBF was measured within the taped area and 3 cm lateral to the taped area. During trial 2, no tape was applied to either site. Both trials were performed within 7 days. Main Outcome Measure(s) Baseline and maximal thermally (42°C) stimulated SkBF responses were assessed using laser Doppler flowmetry. Continuously measured SkBF and derived mean arterial pressure obtained at 5-minute intervals were used to calculate cutaneous vascular conductance (CVC), the primary outcome measure. Results No differences were observed for baseline SkBF or CVC between trials or measurement sites. After local heating, no differences were evident for SkBF or CVC between trials or measurement sites. Conclusions Our findings suggest that, in healthy, trained adolescent males, KT was not associated with increased forearm SkBF. PMID:26445024

  4. (−)-Epicatechin rich cocoa mediated modulation of oxidative stress regulators in skeletal muscle of heart failure and type 2 diabetes patients

    PubMed Central

    Ciaraldi, Theodore P.; Nogueira, Leonardo; Coe, Taylor; Perkins, Guy; Hogan, Michael; Maisel, Alan S.; Henry, Robert R.; Ceballos, Guillermo; Villarreal, Francisco

    2013-01-01

    Background Type 2 diabetes (T2D) and heart failure (HF) are associated with high levels of skeletal muscle (SkM) oxidative stress (OS). Health benefits attributed to flavonoids have been ascribed to antioxidation. However, for flavonoids with similar antioxidant potential, end-biological effects vary widely suggesting other mechanistic venues for reducing OS. Decreases in OS may follow the modulation of key regulatory pathways including antioxidant levels (e.g. glutathione) and enzymes such as mitochondrial superoxide dismutase (SOD2) and catalase. Methods We examined OS-related alterations in SkM in T2D/HF patients (as compared vs. healthy controls) and evaluated the effects of three-month treatment with (−)-epicatechin (Epi) rich cocoa (ERC). To evidence Epi as the mediator of the improved OS profile we examined the effects of pure Epi (vs. water) on SkM OS regulatory systems in a mouse model of insulin resistance and contrasted results vs. normal mice. Results There were severe alterations in OS regulatory systems in T2D/HF SkM as compared with healthy controls. Treatment with ERC induced recovery in glutathione levels and decreases in the nitrotyrosilation and carbonylation of proteins. With treatment, key transcriptional factors translocate into the nucleus leading to increases in SOD2 and catalase protein expression and activity levels. In insulin resistant mice, there were alterations in muscle OS and pure Epi replicated the beneficial effects of ERC found in humans. Conclusions Major perturbations in SkM OS can be reversed with ERC in T2D/HF patients. Epi likely mediates such effects and may provide an effective means to treat conditions associated with tissue OS. PMID:23870648

  5. Regulation of autophagy by sphingosine kinase 1 and its role in cell survival during nutrient starvation.

    PubMed

    Lavieu, Grégory; Scarlatti, Francesca; Sala, Giusy; Carpentier, Stéphane; Levade, Thierry; Ghidoni, Riccardo; Botti, Joëlle; Codogno, Patrice

    2006-03-31

    The sphingolipid ceramide induces macroautophagy (here called autophagy) and cell death with autophagic features in cancer cells. Here we show that overexpression of sphingosine kinase 1 (SK1), an enzyme responsible for the production of sphingosine 1-phosphate (S1P), in MCF-7 cells stimulates autophagy by increasing the formation of LC3-positive autophagosomes and the rate of proteolysis sensitive to the autophagy inhibitor 3-methyladenine. Autophagy was blocked in the presence of dimethylsphingosine, an inhibitor of SK activity, and in cells expressing a catalytically inactive form of SK1. In SK1(wt)-overexpressing cells, however, autophagy was not sensitive to fumonisin B1, an inhibitor of ceramide synthase. In contrast to ceramide-induced autophagy, SK1(S1P)-induced autophagy is characterized by (i) the inhibition of mammalian target of rapamycin signaling independently of the Akt/protein kinase B signaling arm and (ii) the lack of robust accumulation of the autophagy protein Beclin 1. In addition, nutrient starvation induced both the stimulation of autophagy and SK activity. Knocking down the expression of the autophagy protein Atg7 or that of SK1 by siRNA abolished starvation-induced autophagy and increased cell death with apoptotic hallmarks. In conclusion, these results show that SK1(S1P)-induced autophagy protects cells from death with apoptotic features during nutrient starvation.

  6. Effects of cholesterol oxides on cell death induction and calcium increase in human neuronal cells (SK-N-BE) and evaluation of the protective effects of docosahexaenoic acid (DHA; C22:6 n-3).

    PubMed

    Zarrouk, Amira; Nury, Thomas; Samadi, Mohammad; O'Callaghan, Yvonne; Hammami, Mohamed; O'Brien, Nora M; Lizard, Gérard; Mackrill, John J

    2015-07-01

    Some oxysterols are associated with neurodegenerative diseases. Their lipotoxicity is characterized by an oxidative stress and induction of apoptosis. To evaluate the capacity of these molecules to trigger cellular modifications involved in neurodegeneration, human neuronal cells SK-N-BE were treated with 7-ketocholesterol, 7α- and 7β-hydroxycholesterol, 6α- and 6β-hydroxycholesterol, 4α- and 4β-hydroxycholesterol, 24(S)-hydroxycholesterol and 27-hydroxycholesterol (50-100μM, 24h) without or with docosahexaenoic acid (50μM). The effects of these compounds on mitochondrial activity, cell growth, production of reactive oxygen species (ROS) and superoxide anions (O2(-)), catalase and superoxide dismutase activities were determined. The ability of the oxysterols to induce increases in Ca(2+) was measured after 10min and 24h of treatment using fura-2 videomicroscopy and Von Kossa staining, respectively. Cholesterol, 7-ketocholesterol, 7β-hydroxycholesterol, and 24(S)-hydroxycholesterol (100μM) induced mitochondrial dysfunction, cell growth inhibition, ROS overproduction and cell death. A slight increase in the percentage of cells with condensed and/or fragmented nuclei, characteristic of apoptotic cells, was detected. With 27-hydroxycholesterol, a marked increase of O2(-) was observed. Increases in intracellular Ca(2+) were only found with 7-ketocholesterol, 7β-hydroxycholesterol, 24(S)-hydroxycholesterol and 27-hydroxycholesterol. Pre-treatment with docosahexaenoic acid showed some protective effects depending on the oxysterol considered. According to the present data, 7-ketocholesterol, 7β-hydroxycholesterol, 24(S)-hydroxycholesterol and 27-hydroxycholesterol could favor neurodegeneration by their abilities to induce mitochondrial dysfunctions, oxidative stress and/or cell death associated or not with increases in cytosolic calcium levels. Copyright © 2015 Elsevier Ltd. All rights reserved.

  7. A conjugate of decyltriphenylphosphonium with plastoquinone can carry cyclic adenosine monophosphate, but not cyclic guanosine monophosphate, across artificial and natural membranes.

    PubMed

    Firsov, Alexander M; Rybalkina, Irina G; Kotova, Elena A; Rokitskaya, Tatyana I; Tashlitsky, Vadim N; Korshunova, Galina A; Rybalkin, Sergei D; Antonenko, Yuri N

    2018-02-01

    The present study demonstrated for the first time the interaction between adenosine 3',5'-cyclic monophosphate (cAMP), one of the most important signaling compounds in living organisms, and the mitochondria-targeted antioxidant plastoquinonyl-decyltriphenylphosphonium (SkQ1). The data obtained on model liquid membranes and human platelets revealed the ability of SkQ1 to selectively transport cAMP, but not guanosine 3',5'-cyclic monophosphate (cGMP), across both artificial and natural membranes. In particular, SkQ1 elicited translocation of cAMP from the source to the receiving phase of a Pressman-type cell, while showing low activity with cGMP. Importantly, only conjugate with plastoquinone, but not dodecyl-triphenylphosphonium, was effective in carrying cAMP. In human platelets, SkQ1 also appeared to serve as a carrier of cAMP, but not cGMP, from outside to inside the cell, as measured by phosphorylation of the vasodilator stimulated phosphoprotein. The SkQ1-induced transfer of cAMP across the plasma membrane found here can be tentatively suggested to interfere with cAMP signaling pathways in living cells. Copyright © 2017 Elsevier B.V. All rights reserved.

  8. Exosome Adherence and Internalization by Hepatic Stellate Cells Triggers Sphingosine 1-Phosphate-dependent Migration*

    PubMed Central

    Wang, Ruisi; Ding, Qian; Yaqoob, Usman; de Assuncao, Thiago M.; Verma, Vikas K.; Hirsova, Petra; Cao, Sheng; Mukhopadhyay, Debabrata; Huebert, Robert C.; Shah, Vijay H.

    2015-01-01

    Exosomes are cell-derived extracellular vesicles thought to promote intercellular communication by delivering specific content to target cells. The aim of this study was to determine whether endothelial cell (EC)-derived exosomes could regulate the phenotype of hepatic stellate cells (HSCs). Initial microarray studies showed that fibroblast growth factor 2 induced a 2.4-fold increase in mRNA levels of sphingosine kinase 1 (SK1). Exosomes derived from an SK1-overexpressing EC line increased HSC migration 3.2-fold. Migration was not conferred by the dominant negative SK1 exosome. Incubation of HSCs with exosomes was also associated with an 8.3-fold increase in phosphorylation of AKT and 2.5-fold increase in migration. Exosomes were found to express the matrix protein and integrin ligand fibronectin (FN) by Western blot analysis and transmission electron microscopy. Blockade of the FN-integrin interaction with a CD29 neutralizing antibody or the RGD peptide attenuated exosome-induced HSC AKT phosphorylation and migration. Inhibition of endocytosis with transfection of dynamin siRNA, the dominant negative dynamin GTPase construct Dyn2K44A, or the pharmacological inhibitor Dynasore significantly attenuated exosome-induced AKT phosphorylation. SK1 levels were increased in serum exosomes derived from mice with experimental liver fibrosis, and SK1 mRNA levels were up-regulated 2.5-fold in human liver cirrhosis patient samples. Finally, S1PR2 inhibition protected mice from CCl4-induced liver fibrosis. Therefore, EC-derived SK1-containing exosomes regulate HSC signaling and migration through FN-integrin-dependent exosome adherence and dynamin-dependent exosome internalization. These findings advance our understanding of EC/HSC cross-talk and identify exosomes as a potential target to attenuate pathobiology signals. PMID:26534962

  9. Exosome Adherence and Internalization by Hepatic Stellate Cells Triggers Sphingosine 1-Phosphate-dependent Migration.

    PubMed

    Wang, Ruisi; Ding, Qian; Yaqoob, Usman; de Assuncao, Thiago M; Verma, Vikas K; Hirsova, Petra; Cao, Sheng; Mukhopadhyay, Debabrata; Huebert, Robert C; Shah, Vijay H

    2015-12-25

    Exosomes are cell-derived extracellular vesicles thought to promote intercellular communication by delivering specific content to target cells. The aim of this study was to determine whether endothelial cell (EC)-derived exosomes could regulate the phenotype of hepatic stellate cells (HSCs). Initial microarray studies showed that fibroblast growth factor 2 induced a 2.4-fold increase in mRNA levels of sphingosine kinase 1 (SK1). Exosomes derived from an SK1-overexpressing EC line increased HSC migration 3.2-fold. Migration was not conferred by the dominant negative SK1 exosome. Incubation of HSCs with exosomes was also associated with an 8.3-fold increase in phosphorylation of AKT and 2.5-fold increase in migration. Exosomes were found to express the matrix protein and integrin ligand fibronectin (FN) by Western blot analysis and transmission electron microscopy. Blockade of the FN-integrin interaction with a CD29 neutralizing antibody or the RGD peptide attenuated exosome-induced HSC AKT phosphorylation and migration. Inhibition of endocytosis with transfection of dynamin siRNA, the dominant negative dynamin GTPase construct Dyn2K44A, or the pharmacological inhibitor Dynasore significantly attenuated exosome-induced AKT phosphorylation. SK1 levels were increased in serum exosomes derived from mice with experimental liver fibrosis, and SK1 mRNA levels were up-regulated 2.5-fold in human liver cirrhosis patient samples. Finally, S1PR2 inhibition protected mice from CCl4-induced liver fibrosis. Therefore, EC-derived SK1-containing exosomes regulate HSC signaling and migration through FN-integrin-dependent exosome adherence and dynamin-dependent exosome internalization. These findings advance our understanding of EC/HSC cross-talk and identify exosomes as a potential target to attenuate pathobiology signals. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

  10. Alterations in aerobic energy expenditure and neuromuscular function during a simulated cross-country skiathlon with the skating technique.

    PubMed

    Fabre, Nicolas; Mourot, Laurent; Zoppirolli, Chiara; Andersson, Erik; Willis, Sarah J; Holmberg, Hans-Christer

    2015-04-01

    Here, we tested the hypothesis that aerobic energy expenditure (AEE) is higher during a simulated 6-km (2 loops of 3-km each) "skiathlon" than during skating only on a treadmill and attempted to link any such increase to biomechanical and neuromuscular responses. Six elite male cross-country skiers performed two pre-testing time-trials (TT) to determine their best performances and to choose an appropriate submaximal speed for collection of physiological, biomechanical and neuromuscular data during two experimental sessions (exp). Each skier used, in randomized order, either the classical (CL) or skating technique (SK) for the first 3-km loop, followed by transition to the skating technique for the second 3-km loop. Respiratory parameters were recorded continuously. The EMG activity of the triceps brachii (TBr) and vastus lateralis (VLa) muscles during isometric contractions performed when the skiers were stationary (i.e., just before the first loop, during the transition, and after the second loop); their corresponding activity during dynamic contractions; and pole and plantar forces during the second loop were recorded. During the second 3-km of the TT, skating speed was significantly higher for the SK-SK than CL-SK. During this second loop, AEE was also higher (+1.5%) for CL-SKexp than SK-SKexp, in association with higher VLa EMG activity during both isometric and dynamic contractions, despite no differences in plantar or pole forces, poling times or cycle rates. Although the underlying mechanism remains unclear, during a skiathlon, the transition between the sections of classical skiing and skating alters skating performance (i.e., skiing speed), AEE and neuromuscular function. Copyright © 2015 Elsevier B.V. All rights reserved.

  11. Topical N-Acetylcysteine Accelerates Wound Healing in Vitro and in Vivo via the PKC/Stat3 Pathway

    PubMed Central

    Tsai, Min-Ling; Huang, Hui-Pei; Hsu, Jeng-Dong; Lai, Yung-Rung; Hsiao, Yu-Ping; Lu, Fung-Jou; Chang, Horng-Rong

    2014-01-01

    N-Acetylcysteine (Nac) is an antioxidant administered in both oral and injectable forms. In this study, we used Nac topically to treat burn wounds in vitro and in vivo to investigate mechanisms of action. In vitro, we monitored glutathione levels, cell proliferation, migration, scratch-wound healing activities and the epithelialization-related proteins, matrixmetalloproteinase-1 (MMP-1) and proteins involved in regulating the expression of MMP-1 in CCD-966SK cells treated with Nac. Various Nac concentrations (0.1, 0.5, and 1.0 mM) increased glutathione levels, cell viability, scratch-wound healing activities and migration abilities of CCD-966SK cells in a dose-dependent manner. The MMP-1 expression of CCD-966SK cells treated with 1.0 mM Nac for 24 h was significantly increased. Levels of phosphatidylinositol 3-kinase (PI3K), protein kinase C (PKC), janus kinase 1 (Jak1), signal transducer and activator of transcription 3 (Stat3), c-Fos and Jun, but not extracellular signal-regulated protein kinases 1 and 2 (Erk1/2), were also significantly increased in a dose-dependent manner compared to the controls. In addition, Nac induced collagenous expression of MMP-1 via the PKC/Stat3 signaling pathway. In vivo, a burn wound healing rat model was applied to assess the stimulation activity and histopathological effects of Nac, with 3.0% Nac-treated wounds being found to show better characteristics on re-epithelialization. Our results demonstrated that Nac can potentially promote wound healing activity, and may be a promising drug to accelerate burn wound healing. PMID:24798751

  12. HnRNP C, YB-1 and hnRNP L coordinately enhance skipping of human MUSK exon 10 to generate a Wnt-insensitive MuSK isoform

    PubMed Central

    Nasrin, Farhana; Rahman, Mohammad Alinoor; Masuda, Akio; Ohe, Kenji; Takeda, Jun-ichi; Ohno, Kinji

    2014-01-01

    Muscle specific receptor tyrosine kinase (MuSK) is an essential postsynaptic transmembrane molecule that mediates clustering of acetylcholine receptors (AChR). MUSK exon 10 is alternatively skipped in human, but not in mouse. Skipping of this exon disrupts a cysteine-rich region (Fz-CRD), which is essential for Wnt-mediated AChR clustering. To investigate the underlying mechanisms of alternative splicing, we exploited block-scanning mutagenesis with human minigene and identified a 20-nucleotide block that contained exonic splicing silencers. Using RNA-affinity purification, mass spectrometry, and Western blotting, we identified that hnRNP C, YB-1 and hnRNP L are bound to MUSK exon 10. siRNA-mediated knockdown and cDNA overexpression confirmed the additive, as well as the independent, splicing suppressing effects of hnRNP C, YB-1 and hnRNP L. Antibody-mediated in vitro protein depletion and scanning mutagenesis additionally revealed that binding of hnRNP C to RNA subsequently promotes binding of YB-1 and hnRNP L to the immediate downstream sites and enhances exon skipping. Simultaneous tethering of two splicing trans-factors to the target confirmed the cooperative effect of YB-1 and hnRNP L on hnRNP C-mediated exon skipping. Search for a similar motif in the human genome revealed nine alternative exons that were individually or coordinately regulated by hnRNP C and YB-1. PMID:25354590

  13. Dermoscopic Clues for Diagnosing Melanomas That Resemble Seborrheic Keratosis

    PubMed Central

    Segura, Sonia; Aguilera, Paula; Scalvenzi, Massimiliano; Longo, Caterina; Barreiro, Alicia; Broganelli, Paolo; Cavicchini, Stefano; Llambrich, Alex; Zaballos, Pedro; Thomas, Luc; Malvehy, Josep; Puig, Susana; Zalaudek, Iris

    2017-01-01

    Importance Melanomas that clinically mimic seborrheic keratosis (SK) can delay diagnosis and adequate treatment. However, little is known about the value of dermoscopy in recognizing these difficult-to-diagnose melanomas. Objective To describe the dermoscopic features of SK-like melanomas to understand their clinical morphology. Design, Setting, and Participants This observational retrospective study used 134 clinical and dermoscopic images of histopathologically proven melanomas in 134 patients treated in 9 skin cancer centers in Spain, France, Italy, and Austria. Without knowledge that the definite diagnosis for all the lesions was melanoma, 2 dermoscopy-trained observers evaluated the clinical descriptions and 48 dermoscopic features (including all melanocytic and nonmelanocytic criteria) of all 134 images and classified each dermoscopically as SK or not SK. The total dermoscopy score and the 7-point checklist score were assessed. Images of the lesions and patient data were collected from July 15, 2013, through July 31, 2014. Main Outcomes and Measures Frequencies of specific morphologic patterns of (clinically and dermoscopically) SK-like melanomas, patient demographics, and interobserver agreement of criteria were evaluated. Results Of the 134 cases collected from 72 men and 61 women, all of whom were white and who had a mean (SD) age of 55.6 (17.5) years, 110 (82.1%) revealed dermoscopic features suggestive of melanoma, including pigment network (74 [55.2%]), blue-white veil (72 [53.7%]), globules and dots (68 [50.7%]), pseudopods or streaks (47 [35.1%]), and blue-black sign (43 [32.3%]). The remaining 24 cases (17.9%) were considered likely SKs, even by dermoscopy. Overall, lesions showed a scaly and hyperkeratotic surface (45 [33.6%]), yellowish keratin (42 [31.3%]), comedo-like openings (41 [30.5%]), and milia-like cysts (30 [22.4%]). The entire sample achieved a mean (SD) total dermoscopy score of 4.7 (1.6) and a 7-point checklist score of 4.4 (2.3

  14. t-Darpp overexpression in HER2-positive breast cancer confers a survival advantage in lapatinib.

    PubMed

    Christenson, Jessica L; Denny, Erin C; Kane, Susan E

    2015-10-20

    Drug resistance is a major barrier to successful cancer treatment. For patients with HER2-positive breast cancer who initially respond to therapy, the majority develop resistance within one year of treatment. Patient outcomes could improve significantly if we can find and exploit common mechanisms of acquired resistance to different targeted therapies. Overexpression of t-Darpp, a truncated form of the dual kinase/phosphatase inhibitor Darpp-32, has been linked to acquired resistance to trastuzumab, a front-line therapy for HER2-positive breast cancer. Darpp-32 reverses t-Darpp's effect on trastuzumab resistance. In this study, we examined whether t-Darpp could be involved in resistance to lapatinib, another HER2-targeted therapeutic. Lapatinib-resistant SKBR3 cells (SK/LapR) showed a marked change in the Darpp-32:t-Darpp ratio toward a predominance of t-Darpp. Overexpression of t-Darpp alone was not sufficient to confer lapatinib resistance, but cells that overexpress t-Darpp partially mimicked the molecular resistance phenotype observed in SK/LapR cells exposed to lapatinib. SK/LapR cells failed to down-regulate Survivin and failed to induce BIM accumulation in response to lapatinib; cells overexpressing t-Darpp exhibited only the failed BIM accumulation. t-Darpp knock-down reversed this phenotype. Using a fluorescence-based co-culture system, we found that cells overexpressing t-Darpp formed colonies in lapatinib within 3-4 weeks, whereas parental cells in the same co-culture did not. Overall, t-Darpp appears to mediate a survival advantage in lapatinib, possibly linked to failed lapatinib-induced BIM accumulation. t-Darpp might also be relevant to acquired resistance to other cancer drugs that rely on BIM accumulation to induce apoptosis.

  15. A simple approach to design chitosan functionalized Fe3O4 nanoparticles for pH responsive delivery of doxorubicin for cancer therapy

    NASA Astrophysics Data System (ADS)

    Adimoolam, Mahesh G.; Amreddy, Narsireddy; Nalam, Madhusudana Rao; Sunkara, Manorama V.

    2018-02-01

    The use of magnetic nanoparticles (MNPs) in cancer therapy offer many advantages due to their unique size, physical and biocompatible properties. In this study we have developed a formulation, comprising of anti-cancer drug doxorubicin (Dox) conjugated to iron oxide nanoparticles via a pH sensitive imine linker. Different amounts of chitosan functionalized superparamagnetic iron oxide nanoparticles (Fe3O4-CHI) were synthesized in-situ by a simple hydrolysis method at room temperature. The synthesized nanoparticles were well characterized by TEM, Zeta Potential, TOC, XPS, TGA and VSM for their physicochemical properties. Dox was conjugated to the Fe3O4-CHI nanoparticles via a glutaraldehyde cross linker with the imine (sbnd Cdbnd Nsbnd) bond, which is sensitive to cleavage in the pH range of 4.4-6.4. The synthesized Fe3O4-Dox nanoparticles exhibited enhanced drug release in lower pH conditions which mimics the tumor microenvironment or intracellular organelles such as endosomes/lysosomes. The cell uptake and therapeutic efficacy of Fe3O4-Dox nanoparticles carried out in ovarian cancer cell (SK-OV-3) and breast cancer cell line (MCF7) showed improved therapeutic efficacy of Dox by nearly four-fold with Fe3O4-Dox nanoparticles.

  16. Purification and Characterization of Schwann Cells from Adult Human Skin and Nerve

    PubMed Central

    Stratton, Jo Anne; Shah, Prajay; Shapira, Yuval; Midha, Rajiv

    2017-01-01

    Abstract Despite its modest capacity for regeneration, peripheral nervous system injury often results in significant long-term disability. Supplementing peripheral nervous system injury with autologous Schwann cells (SCs) may serve to rejuvenate the postinjury environment to enhance regeneration and ultimately improve functional outcomes. However, human nerve-derived SC (hN-SC) collection procedures require invasive surgical resection. Here, we describe the characterization of SCs from adult human skin (hSk-SCs) of four male donors ranging between 27 and 46 years old. Within five weeks of isolating and culturing adherent mixed skin cells, we were able to obtain 3–5 million purified SCs. We found that hSk-SCs appeared transcriptionally indistinguishable from hN-SCs with both populations exhibiting expression of SC genes including: SOX10, SOX9, AP2A1, CDH19, EGR1, ETV5, PAX3, SOX2, CX32, DHH, NECL4, NFATC4, POU3F1, S100B, and YY1. Phenotypic analysis of hSk-SCs and hN-SCs cultures revealed highly enriched populations of SCs indicated by the high percentage of NES+ve, SOX10+ve, s100+ve and p75+ve cells, as well as the expression of a battery of other SC-associated proteins (PAX3, CDH19, ETV5, SOX2, POU3F1, S100B, EGR2, and YY1). We further show that both hSk-SCs and hN-SCs are capable of promoting axonal growth to similar degrees and that a subset of both associate with regenerating axons and form myelin following transplantation into the injured mouse sciatic nerve. Interestingly, although the majority of both hSk-SCs and hN-SCs maintained SOX10 immunoreactivity following transplant, only a subset of each activated the promyelinating factor, POU3F1, and were able to myelinate. Taken together, we demonstrate that adult hSk-SCs are genetically and phenotypically indistinguishable to hN-SCs. PMID:28512649

  17. DNA Hypomethylation in Intragenic and Intergenic Enhancer Chromatin of Muscle-Specific Genes Usually Correlates with their Expression

    PubMed Central

    Ehrlich, Kenneth C.; Paterson, Heather L.; Lacey, Michelle; Ehrlich, Melanie

    2016-01-01

    Tissue-specific enhancers are critical for gene regulation. In this study, we help elucidate the contribution of muscle-associated differential DNA methylation to the enhancer activity of highly muscle-specific genes. By bioinformatic analysis of 44 muscle-associated genes, we show that preferential gene expression in skeletal muscle (SkM) correlates with SkM-specific intragenic and intergenic enhancer chromatin and overlapping foci of DNA hypomethylation. Some genes, e.g., CASQ1 and FBXO32, displayed broad regions of both SkM- and heart-specific enhancer chromatin but exhibited focal SkM-specific DNA hypomethylation. Half of the genes had SkM-specific super-enhancers. In contrast to simple enhancer/gene-expression correlations, a super-enhancer was associated with the myogenic MYOD1 gene in both SkM and myoblasts even though SkM has < 1 percent as much MYOD1 expression. Local chromatin differences in this super-enhancer probably contribute to the SkM/myoblast differential expression. Transfection assays confirmed the tissue-specificity of the 0.3-kb core enhancer within MYOD1’s super-enhancer and demonstrated its repression by methylation of its three CG dinucleotides. Our study suggests that DNA hypomethylation increases enhancer tissue-specificity and that SkM super-enhancers sometimes are poised for physiologically important, rapid up-regulation. PMID:28018137

  18. Kinematic scaling relations of CALIFA galaxies: A dynamical mass proxy for galaxies across the Hubble sequence.

    NASA Astrophysics Data System (ADS)

    Aquino-Ortíz, E.; Valenzuela, O.; Sánchez, S. F.; Hernández-Toledo, H.; Ávila-Reese, V.; van de Ven, G.; Rodríguez-Puebla, A.; Zhu, L.; Mancillas, B.; Cano-Díaz, M.; García-Benito, R.

    2018-06-01

    We used ionized gas and stellar kinematics for 667 spatially resolved galaxies publicly available from the Calar Alto Legacy Integral Field Area survey (CALIFA) 3rd Data Release with the aim of studying kinematic scaling relations as the Tully & Fisher (TF) relation using rotation velocity, Vrot, the Faber & Jackson (FJ) relation using velocity dispersion, σ, and also a combination of Vrot and σ through the SK parameter defined as SK^2 = KV_{rot}^2 + σ ^2 with constant K. Late-type and early-type galaxies reproduce the TF and FJ relations. Some early-type galaxies also follow the TF relation and some late-type galaxies the FJ relation, but always with larger scatter. On the contrary, when we use the SK parameter, all galaxies, regardless of the morphological type, lie on the same scaling relation, showing a tight correlation with the total stellar mass, M⋆. Indeed, we find that the scatter in this relation is smaller or equal to that of the TF and FJ relations. We explore different values of the K parameter without significant differences (slope and scatter) in our final results with respect the case K = 0.5 besides than a small change in the zero point. We calibrate the kinematic SK^2 dynamical mass proxy in order to make it consistent with sophisticated published dynamical models within 0.15 dex. We show that the SK proxy is able to reproduce the relation between the dynamical mass and the stellar mass in the inner regions of galaxies. Our result may be useful in order to produce fast estimations of the central dynamical mass in galaxies and to study correlations in large galaxy surveys.

  19. Secretion of the Streptomyces tyrosinase is mediated through its trans-activator protein, MelC1.

    PubMed

    Leu, W M; Chen, L Y; Liaw, L L; Lee, Y H

    1992-10-05

    The tyrosinase of Streptomyces antibioticus is encoded by the second open reading frame, melC2 of the melanin operon (melC). The upstream open reading frame melC1 specifies a 146-amino acid protein with a typical NH2-terminal signal-peptide characteristic of a secretory protein. The MelC1 protein is involved in the transfer of copper ion to apotyrosinase MelC2 via binary complex formation (Lee, Y.-H. W., Chen, B.-F., Wu, S.-Y., Leu, W.-M., Lin, J.-J., Chen, C. W., and Lo, S. J. (1988) Gene (Amst.) 65, 71-81; Chen, L.-Y., Leu, W.-M., Wang, K.-T., and Lee, Y.-H.W. (1992) J. Biol. Chem. 267, 20100-20107). To investigate whether the export of tyrosinase is also dependent on MelC1, a mutational study of its signal-peptide sequence was performed. Four different mutants were obtained. Mutation at the positively charged region (mutant M-6LE, Arg6-Arg7----Leu6-Glu7) or the hydrophobic region (mutant M-16D, Val16----Asp16) led to Mel- phenotypes. These lesions caused a severe 7-10-fold reduction of the export of both the MelC1 and MelC2 proteins and a concomitant accumulation of the two proteins in the cytosolic fraction. The cell-associated tyrosinase activity in M-6LE but not in the M-16D mutant was dramatically reduced to 4% of the activity found in the wild type strain, suggesting that the basic NH2 terminus of MelC1 is also important for the trans-activation function of this protein. Nevertheless, the defects on the trans-activation and/or secretory functions of MelC1 in mutants M-6LE and M-16D are not due to the impairment of the formation of the MelC1.MelC2 complex. The translation of melanin operon genes in these two mutants also decreased. In contrast, the tyrosinase activity and the secretion of MelC2 were not affected if the mutations occurred at the putative cleavage site of the signal peptidase (e.g. mutant M-29SM, Arg29-Ala30----Ser29-Met30 or mutant 29-SMG, Arg29-Ala30-Asp31----Ser29-Med30-Gly31+ ++). Additionally, tyrosinase activity and its export were

  20. 31 CFR 549.303 - Entity.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 31 Money and Finance:Treasury 3 2011-07-01 2011-07-01 false Entity. 549.303 Section 549.303 Money... CONTROL, DEPARTMENT OF THE TREASURY LEBANON SANCTIONS REGULATIONS General Definitions § 549.303 Entity. The term entity means a partnership, association, trust, joint venture, corporation, group, subgroup...

  1. On the star partition dimension of comb product of cycle and complete graph

    NASA Astrophysics Data System (ADS)

    Alfarisi, Ridho; Darmaji; Dafik

    2017-06-01

    Let G = (V, E) be a connected graphs with vertex set V (G), edge set E(G) and S ⊆ V (G). For an ordered partition Π = {S 1, S 2, S 3, …, Sk } of V (G), the representation of a vertex v ∈ V (G) with respect to Π is the k-vectors r(v|Π) = (d(v, S 1), d(v, S 2), …, d(v, Sk )), where d(v, Sk ) represents the distance between the vertex v and the set Sk , defined by d(v, Sk ) = min{d(v, x)|x ∈ Sk}. The partition Π of V (G) is a resolving partition if the k-vektors r(v|Π), v ∈ V (G) are distinct. The minimum resolving partition Π is a partition dimension of G, denoted by pd(G). The resolving partition Π = {S 1, S 2, S 3, …, Sk} is called a star resolving partition for G if it is a resolving partition and each subgraph induced by Si , 1 ≤ i ≤ k, is a star. The minimum k for which there exists a star resolving partition of V (G) is the star partition dimension of G, denoted by spd(G). Finding a star partition dimension of G is classified to be a NP-Hard problem. Furthermore, the comb product between G and H, denoted by G ⊲ H, is a graph obtained by taking one copy of G and |V (G)| copies of H and grafting the i-th copy of H at the vertex o to the i-th vertex of G. By definition of comb product, we can say that V (G ⊲ H) = {(a, u)|a ∈ V (G), u ∈ V (H)} and (a, u)(b, v) ∈ E(G ⊲ H) whenever a = b and uv ∈ E(H), or ab ∈ E(G) and u = v = o. In this paper, we will study the star partition dimension of comb product of cycle and complete graph, namely Cn ⊲ Km and Km ⊲ Cn for n ≥ 3 and m ≥ 3.

  2. Structure and in vitro anticancer activity of sulfated O-polysaccharide from marine bacterium Poseidonocella pacifica KMM 9010T.

    PubMed

    Kokoulin, Maxim S; Kuzmich, Alexandra S; Kalinovsky, Anatoly I; Rubtsov, Eugene S; Romanenko, Lyudmila A; Mikhailov, Valery V; Komandrova, Nadezhda A

    2017-12-15

    We presented the structure of the sulfated polysaccharide moiety and anticancer activity in vitro of the О-deacylated lipopolysaccharide (DPS) isolated from the marine bacterium Poseidonocella pacifica KMM 9010 T . The structure of O-polysaccharide was investigated by chemical methods along with 1 H and 13 C NMR spectroscopy. The O-polysaccharide was built up of sulfated disaccharide repeating units consisted of d-rhamnose (d-Rhaр) and 3-deoxy-d-manno-oct-2-ulosonic acid (Kdop): →7)-β-Kdoр4Ac5S-(23)-β-d-Rhaр2S-(1→. We demonstrated that the DPS from P. pacifica KMM 9010 T non-toxic for normal mouse epidermal cells (JB6 Cl41 cell line) and inhibited colony formation of human colorectal carcinoma HT-29, breast adenocarcinoma MCF-7 and melanoma SK-MEL-5 cells in a dose-dependent manner. Copyright © 2017 Elsevier Ltd. All rights reserved.

  3. Potential application of immobilized streptokinase extracted from Streptococcus equinus VIT_VB2.

    PubMed

    Vaishnavi, B; Subathra Devi, C

    2017-11-26

    Streptokinase purified from Streptococcus equinus VIT_VB2 isolated from bovine milk sample was immobilized in various solid supports namely entrapment in agarose gel, calcium alginate beads and gelatin gel by cross-linking with formaldehyde. Immobilization of streptokinase in calcium alginate beads showed maximum efficiency (81.8 ± 1.06%) when compared with entrapment with agarose gel (55.6 ± 2.17%) and cross-linked gelatin formaldehyde gel (71.0 ± 1.54%). The purified SK activity was expressed maximum in calcium alginate (1%) and gelatin gel (0.25%) with 1292.68 ± 1.33 and 1121.9 ± 1.2 U mL -1 , respectively. Similarly, SK entrapped in gelatin gel and calcium alginate showed maximum in vitro blood clot lysis activity with 77.67 ± 2.64% and 76.16 ± 2.72%, respectively. The immobilized SK in gelatin gel showed complete clot lysis within 15 min; hence, this application of the study could be used in the treatment of superficial thrombophlebitis, phlebitis, and venous thrombosis. These beads were used for three repeated cycles to check the conversion of substrates into their products, and we concluded that SK can be immobilized in the suitable matrices. Therefore, this helps in the drug-delivery strategies in highly efficient way, moreover, economically competent process in the pharmaceutics.

  4. Comparative Computed Flow Dynamic Analysis of Different Optimization Techniques in Left Main Either Provisional or Culotte Stenting

    PubMed Central

    Dell’Avvocata, Fabio; Zuin, Marco; Giatti, Sara; Duong, Khanh; Pham, Trung; Tuan, Nguyen Si; Vassiliev, Dobrin; Daggubati, Ramesh; Nguyen, Thach

    2017-01-01

    Abstract Background and Objectives Provisional and culotte are the most commonly used techniques in left main (LM) stenting. The impact of different post-dilation techniques on fluid dynamic of LM bifurcation has not been yet investigated. The aim of this study is to evaluate, by means of computational fluid dynamic analysis (CFD), the impact of different post-dilation techniques including proximal optimization technique (POT), kissing balloon (KB), POT-Side-POT and POT–KB-POT, 2-steps Kissing (2SK) and Snuggle Kissing balloon (SKB) on flow dynamic profile after LM provisional or culotte stenting. Methods We considered an LM-LCA-LCX bifurcation reconstructed after reviewing 100 consecutive patients (mean age 71.4 ± 9.3 years, 49 males) with LM distal disease. The diameters of LAD and LCX were modelled according to the Finnet’s law as following: LM 4.5 mm, LAD 3.5 mm, LCX 2.75 mm, with bifurcation angle set up at 55°. Xience third-generation stent (Abbot Inc., USA) was reconstructed and virtually implanted in provisional/cross-over and culotte fashion. POT, KB, POT-side-POT, POT-KB-POT, 2SK and SKB were virtually applied and analyzed in terms of the wall shear stress (WSS). Results Analyzing the provisional stenting, the 2SK and KB techniques had a statistically significant lower impact on the WSS at the carina, while POT seemed to obtain a neutral effect. In the wall opposite to the carina, the more physiological profile has been obtained by KB and POT with higher WSS value and smaller surface area of the lower WSS. In culotte stenting, at the carina, POT-KB-POT and 2SK had a very physiological profile; while at the wall opposite to the carina, 2SK and POT–KB-POT decreased significantly the surface area of the lower WSS compared to the other techniques. Conclusion From the fluid dynamic point of view in LM provisional stenting, POT, 2SK and KB showed a similar beneficial impact on the bifurcation rheology, while in LM culotte stenting, POT-KB-POT and 2SK

  5. Comparative Computed Flow Dynamic Analysis of Different Optimization Techniques in Left Main Either Provisional or Culotte Stenting.

    PubMed

    Rigatelli, Gianluca; Dell'Avvocata, Fabio; Zuin, Marco; Giatti, Sara; Duong, Khanh; Pham, Trung; Tuan, Nguyen Si; Vassiliev, Dobrin; Daggubati, Ramesh; Nguyen, Thach

    2017-12-01

    Provisional and culotte are the most commonly used techniques in left main (LM) stenting. The impact of different post-dilation techniques on fluid dynamic of LM bifurcation has not been yet investigated. The aim of this study is to evaluate, by means of computational fluid dynamic analysis (CFD), the impact of different post-dilation techniques including proximal optimization technique (POT), kissing balloon (KB), POT-Side-POT and POT-KB-POT, 2-steps Kissing (2SK) and Snuggle Kissing balloon (SKB) on flow dynamic profile after LM provisional or culotte stenting. We considered an LM-LCA-LCX bifurcation reconstructed after reviewing 100 consecutive patients (mean age 71.4 ± 9.3 years, 49 males) with LM distal disease. The diameters of LAD and LCX were modelled according to the Finnet's law as following: LM 4.5 mm, LAD 3.5 mm, LCX 2.75 mm, with bifurcation angle set up at 55°. Xience third-generation stent (Abbot Inc., USA) was reconstructed and virtually implanted in provisional/cross-over and culotte fashion. POT, KB, POT-side-POT, POT-KB-POT, 2SK and SKB were virtually applied and analyzed in terms of the wall shear stress (WSS). Analyzing the provisional stenting, the 2SK and KB techniques had a statistically significant lower impact on the WSS at the carina, while POT seemed to obtain a neutral effect. In the wall opposite to the carina, the more physiological profile has been obtained by KB and POT with higher WSS value and smaller surface area of the lower WSS. In culotte stenting, at the carina, POT-KB-POT and 2SK had a very physiological profile; while at the wall opposite to the carina, 2SK and POT-KB-POT decreased significantly the surface area of the lower WSS compared to the other techniques. From the fluid dynamic point of view in LM provisional stenting, POT, 2SK and KB showed a similar beneficial impact on the bifurcation rheology, while in LM culotte stenting, POT-KB-POT and 2SK performed slightly better than the other techniques, probably

  6. Myosin phosphorylation improves contractile economy of mouse fast skeletal muscle during staircase potentiation.

    PubMed

    Bunda, Jordan; Gittings, William; Vandenboom, Rene

    2018-01-30

    Phosphorylation of the myosin regulatory light chain (RLC) by skeletal myosin light chain kinase (skMLCK) potentiates rodent fast twitch muscle but is an ATP-requiring process. Our objective was to investigate the effect of skMLCK-catalyzed RLC phosphorylation on the energetic cost of contraction and the contractile economy (ratio of mechanical output to metabolic input) of mouse fast twitch muscle in vitro (25°C). To this end, extensor digitorum longus (EDL) muscles from wild-type (WT) and from skMLCK-devoid (skMLCK -/- ) mice were subjected to repetitive low-frequency stimulation (10 Hz for 15 s) to produce staircase potentiation of isometric twitch force, after which muscles were quick frozen for determination of high-energy phosphate consumption (HEPC). During stimulation, WT muscles displayed significant potentiation of isometric twitch force while skMLCK -/- muscles did not (i.e. 23% versus 5% change, respectively). Consistent with this, RLC phosphorylation was increased ∼3.5-fold from the unstimulated control value in WT but not in skMLCK -/- muscles. Despite these differences, the HEPC of WT muscles was not greater than that of skMLCK -/- muscles. As a result of the increased contractile output relative to HEPC, the calculated contractile economy of WT muscles was greater than that of skMLCK -/- muscles. Thus, our results suggest that skMLCK-catalyzed phosphorylation of the myosin RLC increases the contractile economy of WT mouse EDL muscle compared with skMLCK -/- muscles without RLC phosphorylation. © 2018. Published by The Company of Biologists Ltd.

  7. New stable perfluoroallyl radical from perfluoro-2,4-dimethyl-3-ethylpentene-2 and its photoconversion reactions

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Shvedova, M.K.; Allayarov, S.R.; Barkalov, I.M.

    1988-05-01

    The thermally and chemically stable perfluoroally radical is formed in the radiolysis of perfluoro-2, 4-dimethyl-3-ethylpentene-2. A phototransformation of this radical into other radicals takes place under the action of UV light (/lambda/ < 300 nm) which are thermally unstable. Samples containing up to 90% FMP and 10% of its isomers were placed into ampules made of quartz or SK-4B glass, which gave to EPR signal after irradiation; the ampules were evacuated to a pressure of 0.13 Pa, frozen to 77 K, and irradiated with /sup 60/Co /lambda/-quanta at 77 K with doses up to 200 kGy. Irradiation with UV lightmore » was performed at 77 K with light from a DRSh-1000 lamp.« less

  8. Structure and anticancer activity of sulfated O-polysaccharide from marine bacterium Cobetia litoralis KMM 3880(T).

    PubMed

    Kokoulin, Maxim S; Kuzmich, Alexandra S; Kalinovsky, Anatoly I; Tomshich, Svetlana V; Romanenko, Lyudmila A; Mikhailov, Valery V; Komandrova, Nadezhda A

    2016-12-10

    We presented the structure of the polysaccharide moiety and anticancer activity in vitro of the sulfated lipopolysaccharide isolated from the marine bacterium Cobetia litoralis KMM 3880(T). The structure of O-polysaccharide was investigated by chemical methods along with (1)H and (13)C NMR spectroscopy. The O-polysaccharide was built up of branched trisaccharide repeating units consist of D-glucose (D-Glcр), D-mannose (D-Manр) and sulfated 3-deoxy-d-manno-oct-2-ulosonic acid (Kdo5S): →7-β-Kdoр4Ac5S-(2→4)-[β-d-Glcp-(1→2)-]-β-d-Manр6Ac-1→. We demonstrated that the lipopolysaccharide and О-deacetylated O-polysaccharide from Cobetia litoralis KMM 3880(T) inhibited a colony formation of human melanoma SK-MEL-28 and colorectal carcinoma HTC-116 cells. Copyright © 2016 Elsevier Ltd. All rights reserved.

  9. Depletion of hepatoma-derived growth factor-related protein-3 induces apoptotic sensitization of radioresistant A549 cells via reactive oxygen species-dependent p53 activation

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Yun, Hong Shik; Hong, Eun-Hee; Department of Chemistry, College of Natural Sciences, Hanyang University, Seoul 133-791

    2013-09-27

    Highlights: •HRP-3 is a radiation- and anticancer drug-responsive protein in A549 cells. •Depletion of HRP-3 induces apoptosis of radio- and chemoresistant A549 cells. •Depletion of HRP-3 promotes ROS generation via inhibition of the Nrf2/HO-1 pathway. •Depletion of HRP-3 enhances ROS-dependent p53 activation and PUMA expression. -- Abstract: Biomarkers based on functional signaling have the potential to provide greater insight into the pathogenesis of cancer and may offer additional targets for anticancer therapeutics. Here, we identified hepatoma-derived growth factor-related protein-3 (HRP-3) as a radioresistance-related gene and characterized the molecular mechanism by which its encoded protein regulates the radio- and chemoresistant phenotypemore » of lung cancer-derived A549 cells. Knockdown of HRP-3 promoted apoptosis of A549 cells and potentiated the apoptosis-inducing action of radio- and chemotherapy. This increase in apoptosis was associated with a substantial generation of reactive oxygen species (ROS) that was attributable to inhibition of the Nrf2/HO-1 antioxidant pathway and resulted in enhanced ROS-dependent p53 activation and p53-dependent expression of PUMA (p53 upregulated modulator of apoptosis). Therefore, the HRP-3/Nrf2/HO-1/ROS/p53/PUMA cascade is an essential feature of the A549 cell phenotype and a potential radiotherapy target, extending the range of targets in multimodal therapies against lung cancer.« less

  10. Design, synthesis, and biological evaluation of (2E)-(2-oxo-1, 2-dihydro-3H-indol-3-ylidene)acetate derivatives as anti-proliferative agents through ROS-induced cell apoptosis.

    PubMed

    Song, Zhuang; Chen, Cai-Ping; Liu, Jun; Wen, Xiaoan; Sun, Hongbin; Yuan, Haoliang

    2016-11-29

    A novel class of (2E)-(2-oxo-1, 2-dihydro-3H-indol-3-ylidene)acetate derivatives were designed and synthesized as potent anti-proliferative agents. Most of these compounds showed potent anti-proliferative activity against some tumor cell lines, including SK-BR-3, MDA-MB-231, HCT-116, SW480, Ovcar-3, HL-60, Saos-2 and HepG2. Compounds 8c and 11h were identified as the most potent ones, while HL-60, HCT116 and MDA-MB-231 were the most sensitive cell lines. Mechanistic study revealed that compound 8c enhanced reactive oxygen species level by inhibiting TrxR and then induced apoptosis by activating apoptosis proteins, bax and cleaved-caspase 3 in HCT116 cells. Preliminary SAR analysis indicated that modifications of the double bond and ester group made great effects on the anti-proliferative activity. Our findings suggested that it was worth further studies on the antitumor potency of (2E)-(2-oxo-1, 2-dihydro-3H-indol-3-ylidene)acetates. Copyright © 2016 Elsevier Masson SAS. All rights reserved.

  11. Type XVII collagen (BP180) can function as a cell-matrix adhesion molecule via binding to laminin 332

    PubMed Central

    Van den Bergh, F.; Eliason, S.L.; Giudice, G.J.

    2010-01-01

    Collagen XVII (COL17) is a transmembrane glycoprotein that is expressed on the basal surface of basal epidermal keratinocytes. Previous observations have led to the hypothesis that an interaction between COL17 and laminin 332, an extracellular matrix protein, contributes to the attachment of the basal keratinocyte to the basement membrane. In order to isolate and manipulate COL17 interactions with ECM components, we induced COL17 expression in two cells lines, SK-MEL1 and K562, that exhibit little or no capacity to attach to our test substrates, including laminin 332, types I and IV collagen, and fibronectin. Cells expressing high levels of COL17 preferentially adhered to a laminin 332 matrix, and, to a lesser extent, type IV collagen, while showing little or no binding to type I collagen or fibronectin. A quantitative analysis of cell adhesive forces revealed that, compared with COL17-negative cells, COL17-positive cells required over 7-fold greater force to achieve 50% detachment from a laminin 332 substrate. When a cell preparation (either K562 or SK-MEL1) with heterogeneous COL17 expression levels was allowed to attach to a laminin 332 matrix, the COL17-positive and COL17-negative cells differentially sorted to the bound and unbound cell fractions, respectively. COL17-dependent attachment to laminin 332 could be reduced or abolished by siRNA-mediated knockdown of COL17 expression or by adding to the assay wells specific antibodies against COL17 or laminin 332. These findings provide strong support for the hypothesis that cell surface COL17 can interact with laminin 332 and, together, participate in the adherence of a cell to the extracellular matrix. PMID:21034821

  12. Sesamol induced apoptotic effect in lung adenocarcinoma cells through both intrinsic and extrinsic pathways.

    PubMed

    Siriwarin, Boondaree; Weerapreeyakul, Natthida

    2016-07-25

    Sesamol is a phenolic lignan found in sesame seeds (Sesamum indicum L.) and sesame oil. The anticancer effects and molecular mechanisms underlying its apoptosis-inducing effect were investigated in human lung adenocarcinoma (SK-LU-1) cells. Sesamol inhibited SK-LU-1 cell growth with an IC50 value of 2.7 mM and exhibited less toxicity toward normal Vero cells after 48 h of treatment (Selective index = 3). Apoptotic bodies-the hallmark of apoptosis-were observed in sesamol-treated SK-LU-1 cells, stained with DAPI. Sesamol increased the activity of caspase 8, 9, and 3/7, indicating that apoptotic cell death occurred through both extrinsic and intrinsic pathways. Sesamol caused the loss of mitochondrial transmembrane potential signifying intrinsic apoptosis induction. Decreasing Bid expression revealed crosstalk between the intrinsic and extrinsic apoptotic pathways; demonstrating clearly that sesamol induces apoptosis through both pathways in human lung adenocarcinoma (SK-LU-1) cells. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

  13. Branching fraction measurement of J / ψ → K S K L and search for J / ψ → K S K S

    DOE PAGES

    Ablikim, M.; Achasov, M. N.; Ahmed, S.; ...

    2017-12-04

    Using a sample of 1.31×10 9J/ψ events collected with the BESIII detector at the BEPCII collider, we study the decays of J/ψ→ K SK L and K SK S. The branching fraction of J/ψ→ K SK L is determined to be B(J/ψ→ K SK L) = (1:93 0:01 (stat:) 0:05 (syst:))×10 -4, which signi cantly improves on previous measurements. No clear signal is observed for the J/ψ→ K SK S process, and the upper limit at the 95% con dence level for its branching fraction is determined to be B(J= ! K SK S) < 1:4×10 -8, which improves onmore » the previous searches by 2 orders in magnitude and reaches the order of the Einstein-Podolsky-Rosen expectation.« less

  14. Branching fraction measurement of J / ψ → K S K L and search for J / ψ → K S K S

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Ablikim, M.; Achasov, M. N.; Ahmed, S.

    Using a sample of 1.31×10 9J/ψ events collected with the BESIII detector at the BEPCII collider, we study the decays of J/ψ→ K SK L and K SK S. The branching fraction of J/ψ→ K SK L is determined to be B(J/ψ→ K SK L) = (1:93 0:01 (stat:) 0:05 (syst:))×10 -4, which signi cantly improves on previous measurements. No clear signal is observed for the J/ψ→ K SK S process, and the upper limit at the 95% con dence level for its branching fraction is determined to be B(J= ! K SK S) < 1:4×10 -8, which improves onmore » the previous searches by 2 orders in magnitude and reaches the order of the Einstein-Podolsky-Rosen expectation.« less

  15. Tumour cells down-regulate CCN2 gene expression in co-cultured fibroblasts in a Smad7- and ERK-dependent manner.

    PubMed

    van Rooyen, Beverley A; Schäfer, Georgia; Leaner, Virna D; Parker, M Iqbal

    2013-10-03

    Recent studies have revealed that interactions between tumour cells and the surrounding stroma play an important role in facilitating tumour growth and invasion. Stromal fibroblasts produce most of the extracellular matrix components found in the stroma. The aim of this study was to investigate mechanisms involved in tumour cell-mediated regulation of extracellular matrix and adhesion molecules in co-cultured fibroblasts. To this end, microarray analysis was performed on CCD-1068SK human fibroblast cells after direct co-culture with MDA-MB-231 human breast tumour cells. We found that the expression of both connective tissue growth factor (CTGF/CCN2) and type I collagen was negatively regulated in CCD-1068SK fibroblast cells under direct co-culture conditions. Further analysis revealed that Smad7, a known negative regulator of the Smad signalling pathway involved in CCN2 promoter regulation, was increased in directly co-cultured fibroblasts. Inhibition of Smad7 expression in CCD-1068SK fibroblasts resulted in increased CCN2 expression, while Smad7 overexpression had the opposite effect. Silencing CCN2 gene expression in fibroblasts led, in turn, to a decrease in type I collagen mRNA and protein levels. ERK signalling was also shown to be impaired in CCD-1068SK fibroblasts after direct co-culture with MDA-MB-231 tumour cells, with Smad7 overexpression in fibroblasts leading to a similar decrease in ERK activity. These effects were not, however, seen in fibroblasts that were indirectly co-cultured with tumour cells. We therefore conclude that breast cancer cells require close contact with fibroblasts in order to upregulate Smad7 which, in turn, leads to decreased ERK signalling resulting in diminished expression of the stromal proteins CCN2 and type I collagen.

  16. Validating tyrosinase homologue melA as a photoacoustic reporter gene for imaging Escherichia coli

    NASA Astrophysics Data System (ADS)

    Paproski, Robert J.; Li, Yan; Barber, Quinn; Lewis, John D.; Campbell, Robert E.; Zemp, Roger

    2015-10-01

    To understand the pathogenic processes for infectious bacteria, appropriate research tools are required for replicating and characterizing infections. Fluorescence and bioluminescence imaging have primarily been used to image infections in animal models, but optical scattering in tissue significantly limits imaging depth and resolution. Photoacoustic imaging, which has improved depth-to-resolution ratio compared to conventional optical imaging, could be useful for visualizing melA-expressing bacteria since melA is a bacterial tyrosinase homologue which produces melanin. Escherichia coli-expressing melA was visibly dark in liquid culture. When melA-expressing bacteria in tubes were imaged with a VisualSonics Vevo LAZR system, the signal-to-noise ratio of a 9× dilution sample was 55, suggesting that ˜20 bacteria cells could be detected with our system. Multispectral (680, 700, 750, 800, 850, and 900 nm) analysis of the photoacoustic signal allowed unmixing of melA-expressing bacteria from blood. To compare photoacoustic reporter gene melA (using Vevo system) with luminescent and fluorescent reporter gene Nano-lantern (using Bruker Xtreme In-Vivo system), tubes of bacteria expressing melA or Nano-lantern were submerged 10 mm in 1% Intralipid, spaced between <1 and 20 mm apart from each other, and imaged with the appropriate imaging modality. Photoacoustic imaging could resolve the two tubes of melA-expressing bacteria even when the tubes were less than 1 mm from each other, while bioluminescence and fluorescence imaging could not resolve the two tubes of Nano-lantern-expressing bacteria even when the tubes were spaced 10 mm from each other. After injecting 100-μL of melA-expressing bacteria in the back flank of a chicken embryo, photoacoustic imaging allowed visualization of melA-expressing bacteria up to 10-mm deep into the embryo. Photoacoustic signal from melA could also be separated from deoxy- and oxy-hemoglobin signal observed within the embryo and

  17. Maintenance plasma exchange treatment for muscle specific kinase antibody positive myasthenia gravis patients.

    PubMed

    Yamada, Chisa; Teener, James W; Davenport, Robertson D; Cooling, Laura

    2015-10-01

    Anti-muscle specific kinase antibody positive myasthenia gravis (MuSK MG) is often characterized by a relatively severe and progressive course, refractoriness to standard myasthenia gravis (MG) medications, and an increased risk of myasthenic crisis. We report here successful management of three MuSK MG patients using maintenance therapeutic plasma exchange (TPE) treatment for up to 4.5 years. The study was a 5-year retrospective review of all MG patients treated with TPE between 2008 and 2013 at University of Michigan. Inclusion criteria of MuSK MG were positive for anti-MuSK antibodies and a diagnosis of MuSK MG by staff neurologists. Patient data included age, gender, diagnostic testing results, medications, and the dates and response to TPE treatments. A total of 153 MG patients underwent at least one course of TPE between 2008 and 2013. A total of 12 patients (7.8%) were positive for anti-MuSK antibodies. Patients were predominantly female (83.3%) and a median age of onset was 46-years old. Three MuSK MG patients were successfully managed with maintenance TPE. Maintenance TPE may be an effective option for MuSK MG patients. The key of successful maintenance treatment at our institution has been to tailor the TPE frequency for each individual, and to modify the treatment interval in conjunction with medical management. © 2014 Wiley Periodicals, Inc.

  18. The role of immunosuppression in squamous cell carcinomas arising in seborrheic keratosis.

    PubMed

    Conic, Ruzica Z; Napekoski, Karl; Schuetz, Heidi; Piliang, Melissa; Bergfeld, Wilma; Atanaskova Mesinkovska, Natasha

    2017-06-01

    Seborrheic keratoses (SK) are common skin neoplasms considered to be benign. Reports of associated squamous cell carcinoma arising within seborrheic keratosis (SCC-SK) have been described. To describe the histopathologic characteristics of SCC-SK and identify predisposing factors in formation of these rare lesions. There were 162 cases of SCC-SK in a span of a decade (2003-2014). All of the histopathologic specimens and medical records were reviewed. Data from these patients were compared to a control group with seborrheic keratosis who were matched by age, sex, and location of lesion from the same time period (n = 162). SCC-SK has the classic histopathologic features of SK, such as hyperkeratosis, parakeratosis, papillomatosis, and pseudohorn cysts. The areas of squamous cell carcinoma were characterized by areas of squamous dysplasia (100%), hypogranulosis (79.6%), squamous eddies (79.6%), solar elastosis (80.9%), and brown pigmentation (59.9%). Patients with a history of immunosuppression had an increased risk for developing SCC-SK (19% vs 3%; P < .01), particularly when inhibition was transplant-associated (10% vs 0%; P < .01). This was a single center, retrospective study. SCC-SK occurs more often in elderly men with a history of immunosuppression associated with organ transplants. Copyright © 2016 American Academy of Dermatology, Inc. Published by Elsevier Inc. All rights reserved.

  19. DOE Office of Scientific and Technical Information (OSTI.GOV)

    Evans, C. J.; Hainich, R.; Oskinova, L. M.

    Sk 183 is the visually brightest star in the N90 nebula, a young star-forming region in the Wing of the Small Magellanic Cloud (SMC). We present new optical spectroscopy from the Very Large Telescope which reveals Sk 183 to be one of the most massive O-type stars in the SMC. Classified as an O3-type dwarf on the basis of its nitrogen spectrum, the star also displays broadened He I absorption, which suggests a later type. We propose that Sk 183 has a composite spectrum and that it is similar to another star in the SMC, MPG 324. This brings themore » number of rare O2- and O3-type stars known in the whole of the SMC to a mere four. We estimate physical parameters for Sk 183 from analysis of its spectrum. For a single-star model, we estimate an effective temperature of 46 {+-} 2 kK, a low mass-loss rate of {approx}10{sup -7} M{sub Sun} yr{sup -1}, and a spectroscopic mass of 46{sup +9}{sub -8} M{sub Sun} (for an adopted distance modulus of 18.7 mag to the young population in the SMC Wing). An illustrative binary model requires a slightly hotter temperature ({approx}47.5 kK) for the primary component. In either scenario, Sk 183 is the earliest-type star known in N90 and will therefore be the dominant source of hydrogen-ionizing photons. This suggests Sk 183 is the primary influence on the star formation along the inner edge of the nebula.« less

  20. Probing the role of helix α1 in the acid-tolerance and thermal stability of the Streptomyces sp. SK Glucose Isomerase by site-directed mutagenesis.

    PubMed

    Hajer, Ben Hlima; Dorra, Zouari Ayadi; Monia, Mezghani; Samir, Bejar; Nushin, Aghajari

    2014-03-10

    In order to investigate the role of helix α1 in the different biochemical properties between class I and class II Glucose Isomerases, a histidine and a phenylalanine residue were inserted at position 17 and 19 of Streptomyces sp. SK Glucose Isomerase (SKGI). In addition, W16 was substituted by a histidine. The H17/F19 insertion displaced the optimal pH of SKGI from 6.5 to 7-8 and slightly decreased the thermostability. As for the W16H mutant, a shift in optimal pH of SKGI from 6.5 to 6 was observed along with a decrease in the enzyme thermostability at 85°C with a half-life time reduced twice compared to the wild-type enzyme. Three-dimensional structure analysis suggested that the insertion of a histidine at position 17 results in the formation of new hydrogen bond with D287, thereby preventing it from deprotonating the O2 hydroxyl of the sugar at low pH, while the substitution W16H induced opposite effect by preventing hydrogen bond formation between D287 and W16 and thereby probably facilitating the hydrogen transfer during the isomerization reaction. The findings highlight the essential role of helix α1, which bears the three introduced mutations, in the acid-tolerance and the thermostability of SKGI and of glucose isomerases in general. Copyright © 2014 Elsevier B.V. All rights reserved.

  1. Beneficial effects of dark chocolate on exercise capacity in sedentary subjects: underlying mechanisms. A double blind, randomized, placebo controlled trial.

    PubMed

    Taub, Pam R; Ramirez-Sanchez, Israel; Patel, Minal; Higginbotham, Erin; Moreno-Ulloa, Aldo; Román-Pintos, Luis Miguel; Phillips, Paul; Perkins, Guy; Ceballos, Guillermo; Villarreal, Francisco

    2016-09-14

    In heart failure patients the consumption of (-)-epicatechin ((-)-Epi)-rich cocoa can restore skeletal muscle (SkM) mitochondrial structure and decrease biomarkers of oxidative stress. However, nothing is known about its effects on exercise capacity and underlying mechanisms in normal, sedentary subjects. Twenty normal, sedentary subjects (∼50 years old) were randomized to placebo or dark chocolate (DC) groups and consumed 20 g of the products for 3 months. Subjects underwent before and after treatment, bicycle ergometry to assess VO2 max and work, SkM biopsy to assess changes in mitochondrial density, function and oxidative stress and blood sampling to assess metabolic endpoints. Seventeen subjects completed the trial. In the DC group (n = 9), VO2 max increased (17% increase, p = 0.056) as well as maximum work (watts) achieved (p = 0.026) with no changes with placebo (n = 8). The DC group evidenced increases in HDL levels (p = 0.005) and decreased triglycerides (p = 0.07). With DC, SkM evidenced significant increases in protein levels for LKB1, AMPK and PGC1α and in their active forms (phosphorylated AMPK and LKB1) as well as in citrate synthase activity while no changes were observed in mitochondrial density. With DC, significant increases in SkM reduced glutathione levels and decreases in protein carbonylation were observed. Improvements in maximum work achieved and VO2 max may be due to DC activation of upstream control systems and enhancement of SkM mitochondria efficiency. Larger clinical studies are warranted to confirm these observations.

  2. Preparation and biologic evaluation of a novel radioiodinated benzylpiperazine, 123I-MEL037, for malignant melanoma.

    PubMed

    Pham, Tien Q; Berghofer, Paula; Liu, Xiang; Greguric, Ivan; Dikic, Branko; Ballantyne, Patrice; Mattner, Filomena; Nguyen, Vu; Loc'h, Christian; Katsifis, Andrew

    2007-08-01

    Radiopharmaceuticals that can target the random metastatic dissemination of melanoma tumors may present opportunities for imaging and staging the disease as well as potential radiotherapeutic applications. A novel molecule, 2-(2-(4-(4-(123)I-iodobenzyl)piperazin-1-yl)-2-oxoethyl)isoindoline-1,3-dione (MEL037), was synthesized, labeled with 123I, and evaluated for application in melanoma tumor scintigraphy and radiotherapy. The tumor imaging potential of 123I-MEL037 was studied in vivo in C57BL/6J female mice bearing the B16F0 murine melanoma tumor and in BALB/c nude mice bearing the A375 human amelanotic melanoma tumor by biodistribution, competition studies, and SPECT. 123I-MEL037 exhibited high and rapid uptake in the B16F0 melanoma tumor at 1 h (13 %ID/g [percentage injected dose per gram]), increasing with time to reach 25 %ID/g at 6 h. A significant uptake was also observed in the eyes (2 %ID, at 3-6 h after injection) of black mice. No uptake was observed in the tumor or in the eyes of nude mice bearing the A375 tumor. Because of high uptake and long retention in the tumor and rapid body clearance, the mean contrast ratios (MCR) of 123I-MEL037 were 30 and 60, at 24 and 48 h after injection, respectively. At 24 h after injection of mice bearing the B16 melanoma, SPECT indicated that the radioactivity was located predominately in the tumor followed by the eyes, whereas no specific localization of the radioactivity was noted in mice bearing the A375 human amelanotic tumor. In competition experiments, uptake of 123I-MEL037 in brain, lung, heart, and kidney--organs known to contain sigma-receptors--was not significantly different in haloperidol-treated animals compared with control animals. Therefore, reduction of uptake in tumor and eyes of the pigmented mice bearing the B16F0 tumor suggested that the mechanism of tumor uptake was likely due to an interaction with melanin. These findings suggested that 123I-MEL037, which displays a rapid and very high tumor

  3. Stable isotopes from Torneträsk, northern Sweden provide a millennial length reconstruction of summer sunshine and its relationship to Arctic circulation

    NASA Astrophysics Data System (ADS)

    Griffiths, R.; Loader, N.; Grudd, H.; Young, G.; McCarroll, D.

    2012-12-01

    Results are presented from the first 1,100 years of a long stable carbon isotope chronology currently in development from Scots Pine (Pinus sylvestris L.) trees growing in the Torneträsk region of northern Sweden. The isotope record currently comprises a total of 74 trees with a mean annual replication of >12, thereby enabling it to be directly compared with other tree-ring based palæoclimate reconstructions from this region. The resulting carbon isotope series is calibrated against instrumental data from the closest meteorological station at Abisko (AD1913-2008) to provide a record of June-August sunshine for northern Fennoscandia. This parameter is closely linked to the direct control of assimilation rate; Photosynthetically Active Radiation (PAR) and the indirect measures; mean July-August temperature and percent cloud cover. The coupled response of summer sunshine and temperature in this region permits a multi-parameter comparison with a local reconstruction of past temperature variability based upon tree growth proxies to explore the stability of this coupling through time. Several periods are identified where the temperature (X-ray density) and sunshine (stable carbon isotope ratio) records diverge. The most significant and sustained of these occur between c. AD1200-1380 and c. AD1550-1780, providing evidence for a cool, sunny two-phase "Little Ice Age". Whilst summer sunshine reconstructed for the 20th century is statistically different from the mean of the last 1,100 (P<0.01) years, conditions during the early Mediæval period are similar to those experienced in northern Fennoscandia during the 20th century (P>0.01), and as such it is the 17th-18th, and to a lesser extent, the 13th Centuries rather than the early Mediæval period that appear anomalous when viewed within the context of the last 1,100 years.

  4. Inhibition of Staphylococcus epidermidis Biofilm Formation by Traditional Thai Herbal Recipes Used for Wound Treatment.

    PubMed

    Chusri, S; Sompetch, K; Mukdee, S; Jansrisewangwong, S; Srichai, T; Maneenoon, K; Limsuwan, S; Voravuthikunchai, S P

    2012-01-01

    Development of biofilm is a key mechanism involved in Staphylococcus epidermidis virulence during device-associated infections. We aimed to investigate antibiofilm formation and mature biofilm eradication ability of ethanol and water extracts of Thai traditional herbal recipes including THR-SK004, THR-SK010, and THR-SK011 against S. epidermidis. A biofilm forming reference strain, S. epidermidis ATCC 35984 was employed as a model for searching anti-biofilm agents by MTT reduction assay. The results revealed that the ethanol extract of THR-SK004 (THR-SK004E) could inhibit the formation of S. epidermidis biofilm on polystyrene surfaces. Furthermore, treatments with the extract efficiently inhibit the biofilm formation of the pathogen on glass surfaces determined by scanning electron microscopy and crystal violet staining. In addition, THR-SK010 ethanol extract (THR-SK010E; 0.63-5 μg/mL) could decrease 30 to 40% of the biofilm development. Almost 90% of a 7-day-old staphylococcal biofilm was destroyed after treatment with THR-SK004E (250 and 500 μg/mL) and THR-SK010E (10 and 20 μg/mL) for 24 h. Therefore, our results clearly demonstrated THR-SK004E could prevent the staphylococcal biofilm development, whereas both THR-SK004E and THR-SK010E possessed remarkable eradication ability on the mature staphylococcal biofilm.

  5. Multilayer Spheroids To Quantify Drug Uptake and Diffusion in 3D

    PubMed Central

    2015-01-01

    There is a need for new quantitative in vitro models of drug uptake and diffusion to help assess drug toxicity/efficacy as well as new more predictive models for drug discovery. We report a three-dimensional (3D) multilayer spheroid model and a new algorithm to quantitatively study uptake and inward diffusion of fluorescent calcein via gap junction intercellular communication (GJIC). When incubated with calcein-AM, a substrate of the efflux transporter P-glycoprotein (Pgp), spheroids from a variety of cell types accumulated calcein over time. Accumulation decreased in spheroids overexpressing Pgp (HEK-MDR) and was increased in the presence of Pgp inhibitors (verapamil, loperamide, cyclosporin A). Inward diffusion of calcein was negligible in spheroids that lacked GJIC (OVCAR-3, SK-OV-3) and was reduced in the presence of an inhibitor of GJIC (carbenoxolone). In addition to inhibiting Pgp, verapamil and loperamide, but not cyclosporin A, inhibited inward diffusion of calcein, suggesting that they also inhibit GJIC. The dose response curves of verapamil’s inhibition of Pgp and GJIC were similar (IC50: 8 μM). The method is amenable to many different cell types and may serve as a quantitative 3D model that more accurately replicates in vivo barriers to drug uptake and diffusion. PMID:24641346

  6. Neuronal Uptake and Neuroprotective Properties of Curcumin-Loaded Nanoparticles on SK-N-SH Cell Line: Role of Poly(lactide-co-glycolide) Polymeric Matrix Composition.

    PubMed

    Djiokeng Paka, Ghislain; Doggui, Sihem; Zaghmi, Ahlem; Safar, Ramia; Dao, Lé; Reisch, Andreas; Klymchenko, Andrey; Roullin, V Gaëlle; Joubert, Olivier; Ramassamy, Charles

    2016-02-01

    Curcumin, a neuroprotective agent with promising therapeutic approach has poor brain bioavailability. Herein, we demonstrate that curcumin-encapsulated poly(lactide-co-glycolide) (PLGA) 50:50 nanoparticles (NPs-Cur 50:50) are able to prevent the phosphorylation of Akt and Tau proteins in SK-N-SH cells induced by H2O2 and display higher anti-inflammatory and antioxidant activities than free curcumin. PLGA can display various physicochemical and degradation characteristics for controlled drug release applications according to the matrix used. We demonstrate that the release of curcumin entrapped into a PLGA 50:50 matrix (NPs-Cur 50:50) is faster than into PLGA 65:35. We have studied the effects of the PLGA matrix on the expression of some key antioxidant- and neuroprotective-related genes such as APOE, APOJ, TRX, GLRX, and REST. NPs-Cur induced the elevation of GLRX and TRX while decreasing APOJ mRNA levels and had no effect on APOE and REST expressions. In the presence of H2O2, both NPs-Cur matrices are more efficient than free curcumin to prevent the induction of these genes. Higher uptake was found with NPs-Cur 50:50 than NPs-Cur 65:35 or free curcumin. By using PLGA nanoparticles loaded with the fluorescent dye Lumogen Red, we demonstrated that PLGA nanoparticles are indeed taken up by neuronal cells. These data highlight the importance of polymer composition in the therapeutic properties of the nanodrug delivery systems. Our study demonstrated that NPs-Cur enhance the action of curcumin on several pathways implicated in the pathophysiology of Alzheimer's disease (AD). Overall, these results suggest that PLGA nanoparticles are a promising strategy for the brain delivery of drugs for the treatment of AD.

  7. Complete stable remission and autoantibody specificity in myasthenia gravis.

    PubMed

    Baggi, Fulvio; Andreetta, Francesca; Maggi, Lorenzo; Confalonieri, Paolo; Morandi, Lucia; Salerno, Franco; Bernasconi, Pia; Montomoli, Cristina; Barberis, Massimo; Mantegazza, Renato; Antozzi, Carlo

    2013-01-08

    Patients with myasthenia gravis (MG) are subgrouped as acetylcholine receptor (AChR)-positive, muscle-specific kinase (MuSK)-positive, and AChR/MuSK-negative MG (or double negative [DN]) on the basis of autoantibody assay. We investigated the relationships between autoantibody specificity, main clinical features, and outcome of the disease, in particular the occurrence of complete stable remission (CSR), by means of a retrospective study on a cohort of 677 Italian patients with MG. A total of 517 (76%) patients with AChR-positive MG, 55 (8%) patients with MuSK-positive MG, and 105 (16%) patients with DN MG were included in the study. Kaplan-Meier and Cox proportional hazard regression analyses were used to evaluate associations between baseline characteristics, antibody specificity, and CSR. Clinical stage at onset and at maximal worsening was more severe for MuSK-positive patients: bulbar impairment at maximal worsening was found in 83.6% of MuSK-positive patients compared with 58.6% of AChR-positive patients and 43.8% of DN patients (p < 0.001). Baseline characteristics of AChR-positive and DN patients were similar. CSR was observed in 3.6% of MuSK-positive patients compared with 22.2% of AChR-positive and 21.9% of DN patients. In the whole MG cohort, onset before age 40 (hazard ratio [HR] = 1.96, 95% confidence interval [CI] 1.27-3.02, p = 0.002) and ocular and generalized clinical stages at maximal worsening were associated with CSR (ocular, HR = 8.05, 95% CI 1.88-34.53, p = 0.005; generalized, HR = 3.71, 95% CI 1.16-11.90, p = 0.023; bulbar, HR = 3.16, 95% CI 1.00-10.05, p = 0.051). MuSK antibodies identify a clinically distinguishable, more severe form of MG since the disease onset, with a lower occurrence of CSR. These features should be considered by the clinician in the management of this particular form of MG.

  8. Orthostatic intolerance predicts mild cognitive impairment: incidence of mild cognitive impairment and dementia from the Swedish general population cohort Good Aging in Skåne

    PubMed Central

    Elmståhl, Sölve; Widerström, Elisabet

    2014-01-01

    Introduction Contradictory results have been reported on the relationship between orthostatic hypotension (OH) and mild cognitive impairment (MCI). Objective To study the incidence of MCI and dementia and their relationship to OH and subclinical OH with orthostatic symptoms (orthostatic intolerance). Study design and setting This study used a prospective general population cohort design and was based on data from the Swedish Good Aging in Skåne study (GÅS-SNAC), they were studied 6 years after baseline of the present study, with the same study protocol at baseline and at follow-up. The study sample comprised 1,480 randomly invited subjects aged 60 to 93 years, and had a participation rate of 82% at follow-up. OH test included assessment of blood pressure and symptoms of OH. Results The 6-year incidence of MCI was 8%, increasing from 12.1 to 40.5 per 1,000 person-years for men and 6.9 to 16.9 per 1,000 person-years for women aged 60 to >80 years. The corresponding 6-year incidence of dementia was 8%. Orthostatic intolerance during uprising was related to risk for MCI at follow-up (odds ratio [OR] =1.84 [1.20–2.80][95% CI]), adjusted for age and education independently of blood pressure during testing. After stratification for hypertension (HT), the corresponding age-adjusted OR for MCI in the non-HT group was 1.71 (1.10–2.31) and 1.76 (1.11–2.13) in the HT group. Among controls, the proportion of those with OH was 16%; those with MCI 24%; and those with dementia 31% (age-adjusted OR 1.93 [1.19–3.14]). Conclusion Not only OH, but also symptoms of OH, seem to be a risk factor for cognitive decline and should be considered in the management of blood pressure among the elderly population. PMID:25429211

  9. Potent suppressing activity of the non-polyphenolic fraction of green tea (Camellia sinensis) against genotoxin-induced umu C gene expression in Salmonella typhimurium (TA 1535/pSK 1002)--association with pheophytins a and b.

    PubMed

    Okai, Y; Higashi-Okai, K

    1997-11-25

    Antigenotoxic and antimutagenic activities of green tea extract and tea-derived polyphenols have been studied using in vitro and in vivo experiments. However, antigenotoxic substances in the non-polyphenolic fraction of green tea have been poorly elucidated. In the present study, the effect of the non-polyphenolic fraction of green tea on genotoxin-induced umu C gene expression was analyzed using a tester bacteria, and potent antigenotoxic substances in the non-polyphenolic fraction were identified. The non-polyphenolic fraction of green tea showed strong suppressive activities against umu C gene expression in Salmonella typhimurium (TA 1535/pSK 1002) induced by 3-amino-1,4-dimethyl-5H-pyrido[4,3-b]indol (Trp-P-1) or mitomycin C (MMC) in the presence or absence of S9 metabolizing enzyme mixture. The non-polyphenolic fraction of green tea exhibited major two-color bands in a silica gel TLC and they were identified as chlorophyll-related compounds, pheophytins a and b, judged by their specific colors, Rf values in silica gel TLC and absorption spectra. These pigments showed significant suppressive activities against umu C gene expression in tester bacteria induced by Trp-P- and MMC in a dose-dependent manner. These results suggest that the non-polyphenolic fraction of green tea contains pheophytins a and b as potent antigenotoxic substances.

  10. Pore water phosphate and ammonia below the permanent halocline in the south-eastern Baltic Sea and their benthic fluxes under anoxic conditions

    NASA Astrophysics Data System (ADS)

    Graca, Bożena; Witek, Zbigniew; Burska, Dorota; Białkowska, Izabela; Łukawska-Matuszewska, Katarzyna; Bolałek, Jerzy

    2006-12-01

    In this paper the results of a study on the distribution of pore water phosphates and ammonia, and their fluxes under anoxic condition in a deep (> 70 m) accumulation-type bottom of the south-eastern Baltic Sea, namely in the Gdańsk Deep and the adjacent areas, are presented. All measurements were taken during the growth period, i.e. in September 2000, April 2001 and June 2002. Benthic phosphate and ammonia fluxes were estimated using Fick's First Law. Phosphate and ammonia concentrations ranged from 7.5 to 266.3 μmol dm - 3 and from 53.6 to 1248.3 μmol dm - 3 , respectively. The values recorded in the central part of the Gdańsk Deep were lower than those found both on its slopes and on the SW slope of the Gotland Deep. The lowest phosphate contents were typical of the Oblique Sill which separates the Gdańsk and Gotland Deeps. In 1993-2002, as a result of anoxia the sediments in the Gdańsk Deep released about 5.1 × 10 3 t P and 22.8 × 10 3 t N. These loads supplied on average 1.5% and 0.9% of phytoplankton's demand for P and N, respectively. In comparison to the total external load of nutrients discharged to the Gulf of Gdańsk (i.e. 8.79 × 10 3 t year - 1 P tot and 130.79 × 10 3 t year - 1 N tot; [Witek, Z., Humborg, Ch., Savchuk, O., Grelowski, A. and Łysiak-Pastuszak, E., 2003. Nitrogen and phosphorus budgets of the Gulf of Gdańsk (Baltic Sea). Est. Coast. Shelf Sci., 57:239-248.]), the return flux of P and N from the anoxic sediments to the water column in the Gdańsk Deep was a minor source of these elements.

  11. Seborrheic inclusion cyst of the skin positive for cytoplasmic inclusion bodies and HPV antigen.

    PubMed

    Terada, Tadashi

    2012-01-01

    Seborrheic inclusion cyst (SIC) is a very rare variant of epidermal cyst of the skin. SIC shows seborrheic keratosis (SK)-like lesion in epidermal cyst. SIC is extremely rare; only 6 case reports have been published in the English literature. However, no immunohistochemical study of SIC has been reported. A 41-year-old Japanese man noticed a subcutaneous tumor in the neck. Physical examination showed slightly mobile tumor in the subcutaneous tissue, and total excision was performed. Grossly, the tumor (1 x 1 x 0.8 cm) was cyst containing atheromatous keratin. Microscopically, the lesion is a cyst containing keratins. About one half of the cyst showed features of epidermal cyst consisting of mature squamous epithelium with granular layers. The other one half showed SK-like epidermal proliferation. The SK-like area showed basaloid cell proliferation with pseudohorn cysts. No significant atypia was noted. Many eosinophilic cytoplasmic inclusion bodies were noted in the SK-like area. Immunohistochemically, the SK-like area was positive for pancytokeratin AE1/3, pancytokeratin CAM5.2, p63, and Ki-67 (labeling=8%) and HPV, but negative for p53. The pathological diagnosis was SIC.

  12. Near-infrared quantum dots for HER2 localization and imaging of cancer cells.

    PubMed

    Rizvi, Sarwat B; Rouhi, Sepideh; Taniguchi, Shohei; Yang, Shi Yu; Green, Mark; Keshtgar, Mo; Seifalian, Alexander M

    2014-01-01

    Quantum dots are fluorescent nanoparticles with unique photophysical properties that allow them to be used as diagnostic, therapeutic, and theranostic agents, particularly in medical and surgical oncology. Near-infrared-emitting quantum dots can be visualized in deep tissues because the biological window is transparent to these wavelengths. Their small sizes and free surface reactive groups that can be conjugated to biomolecules make them ideal probes for in vivo cancer localization, targeted chemotherapy, and image-guided cancer surgery. The human epidermal growth factor receptor 2 gene (HER2/neu) is overexpressed in 25%-30% of breast cancers. The current methods of detection for HER2 status, including immunohistochemistry and fluorescence in situ hybridization, are used ex vivo and cannot be used in vivo. In this paper, we demonstrate the application of near-infrared-emitting quantum dots for HER2 localization in fixed and live cancer cells as a first step prior to their in vivo application. Near-infrared-emitting quantum dots were characterized and their in vitro toxicity was established using three cancer cell lines, ie, HepG2, SK-BR-3 (HER2-overexpressing), and MCF7 (HER2-underexpressing). Mouse antihuman anti-HER2 monoclonal antibody was conjugated to the near-infrared-emitting quantum dots. In vitro toxicity studies showed biocompatibility of SK-BR-3 and MCF7 cell lines with near-infrared-emitting quantum dots at a concentration of 60 μg/mL after one hour and 24 hours of exposure. Near-infrared-emitting quantum dot antiHER2-antibody bioconjugates successfully localized HER2 receptors on SK-BR-3 cells. Near-infrared-emitting quantum dot bioconjugates can be used for rapid localization of HER2 receptors and can potentially be used for targeted therapy as well as image-guided surgery.

  13. Novel copper(II) complexes as new promising antitumour agents. A crystal structure of [Cu(1,10-phenanthroline-5,6-dione)2(OH2)(OClO3)](ClO4).

    PubMed

    Pivetta, Tiziana; Trudu, Federica; Valletta, Elisa; Isaia, Francesco; Castellano, Carlo; Demartin, Francesco; Tuveri, Rossana; Vascellari, Sarah; Pani, Alessandra

    2014-12-01

    The cytotoxic properties of copper(II) complexes with 1,10-phenanthroline (phen) can be modified by substitution in the phen backbone. For this purpose, Cu(II) complexes with phen, 1,10-phenanthrolin-5,6-dione (phendione) and 1,10-phenanthrolin-5,6-diol (phendiol) have been synthesised and characterised. The crystal structure of [Cu(phendione)2(OH2)(OClO3)](ClO4) is discussed. The complex formation equilibria between Cu(II) and phen or phendione were studied by potentiometric measurements at 25 and 37°C in 0.1 M ionic strength (NaCl). The antitumour activity of the compounds has been tested in vitro against a panel of tumour (DU-145, HEP-G2, SK-MES-1, CCRF-CEM, CCRF-SB) and normal (CRL-7065) human cell lines. The studied compounds generally present an antiproliferative effect greater than that of cisplatin. The phen and phendione ligands present a similar antiproliferative effect against all the tested cells. Phendiol presents an antiproliferative effect 1.3 to 18 times greater than that of phen or phendione for leukemic, lung, prostatic and fibroblast cells, while it presents less activity towards hepatic cells. Complexes with two ligands are more cytotoxic towards all the tested cell lines than complexes with one ligand and are generally more cytotoxic than the ligand alone. Complexes [Cu(phendiol)2(OH2)](ClO4)2 and [Cu(phendione)2(OH2)(OClO3)](ClO4) appear to be the most active compounds for the treatment of SK-MES-1 and HEP-G2 cells, respectively, being at least 18 times more cytotoxic than cisplatin. The studied Cu(II) complexes are characterised by a strong DNA affinity and were found to interact with DNA mainly by groove binding or electrostatic interactions. The complexes appear to act on cells with a mechanism different from that of cisplatin. Copyright © 2014 Elsevier Inc. All rights reserved.

  14. SK channel blocker apamin attenuates the effect of SSRI fluoxetine upon cell firing in dorsal raphe nucleus: a concomitant electrophysiological and electrochemical in vivo study reveals implications for modulating extracellular 5-HT.

    PubMed

    Crespi, Francesco

    2010-06-02

    A dual probing methodology was implemented so that combined in vivo voltammetric (electrochemical) and in vivo electrophysiological analysis could be carried out concomitantly in two distinct brain regions of the same anaesthetized animal, i.e., cell body such as the dorsal raphe nucleus (DRN) and related terminal region such as the hippocampus, the frontal cortex, and the amygdala. In particular, this methodology allowed: In addition, the dual probing methodology has been applied to verify the original proposal that a combined treatment with a potassium (SK) channel blocker such as apamin and an SSRI (i.e., fluoxetine) could overcome the slow onset of the SSRI upon central 5-HT activity that could be related to the slow onset of its therapeutic action. Briefly, the effect of apamin either alone or followed by fluoxetine upon cell firing in the DRN (in vivo electrophysiology) and concomitantly upon 5-HT levels (in vivo voltammetry) in the amygdala (forebrain structure involved in mood regulation and innervated by ascending 5-HT projections from the DRN) was studied. Copyright 2010 Elsevier B.V. All rights reserved.

  15. Phloretin induces apoptosis of non-small cell lung carcinoma A549 cells via JNK1/2 and p38 MAPK pathways.

    PubMed

    Min, Jie; Huang, Kenan; Tang, Hua; Ding, Xinyu; Qi, Chen; Qin, Xiong; Xu, Zhifei

    2015-12-01

    Phloretin (Ph) existing in apples, pears and various vegetables is known to have antitumor activities in several cancer cell lines. However, little is known about its effect on human lung cancer cells. The aim of the present study was to see whether Ph could induce apoptosis of non-small cell lung cancer (NSCLC) cells, and explore the possible underlying mechanism of action. We found that Ph markedly induced cell apoptosis of NSCLC cell line A549, and inhibited the migration of A549 cells in a dose-dependent manner. The expression level of BAX, cleaved caspase-3 and -9, and degraded form of PARP was increased and Bcl-2 was decreased after Ph treatment. In addition, the phosphorylation of P38 MAPK, ERK1/2 and JNK1/2 was increased in a dose‑dependent manner in parallel with Ph treatment. Inhibition of P38 MAPK and JNK1/2 by specific inhibitors significantly abolished the Ph-induced activation of the caspase-3 and -9. In vivo tumor-suppression assay further indicated that Ph (20 mg/kg) displayed a more significant inhibitory effect on A549 xenografts in tumor growth. All these findings indicate that Ph is able to inhibit NSCLC A549 cell growth by inducing apoptosis through P38 MAPK and JNK1/2 pathways, and therefore may prove to be an adjuvant to the treatment of NSCLC.

  16. Stationkeeping Monte Carlo Simulation for the James Webb Space Telescope

    NASA Technical Reports Server (NTRS)

    Dichmann, Donald J.; Alberding, Cassandra M.; Yu, Wayne H.

    2014-01-01

    The James Webb Space Telescope (JWST) is scheduled to launch in 2018 into a Libration Point Orbit (LPO) around the Sun-Earth/Moon (SEM) L2 point, with a planned mission lifetime of 10.5 years after a six-month transfer to the mission orbit. This paper discusses our approach to Stationkeeping (SK) maneuver planning to determine an adequate SK delta-V budget. The SK maneuver planning for JWST is made challenging by two factors: JWST has a large Sunshield, and JWST will be repointed regularly producing significant changes in Solar Radiation Pressure (SRP). To accurately model SRP we employ the Solar Pressure and Drag (SPAD) tool, which uses ray tracing to accurately compute SRP force as a function of attitude. As an additional challenge, the future JWST observation schedule will not be known at the time of SK maneuver planning. Thus there will be significant variation in SRP between SK maneuvers, and the future variation in SRP is unknown. We have enhanced an earlier SK simulation to create a Monte Carlo simulation that incorporates random draws for uncertainties that affect the budget, including random draws of the observation schedule. Each SK maneuver is planned to optimize delta-V magnitude, subject to constraints on spacecraft pointing. We report the results of the Monte Carlo simulations and discuss possible improvements during flight operations to reduce the SK delta-V budget.

  17. Inhibition of Staphylococcus epidermidis Biofilm Formation by Traditional Thai Herbal Recipes Used for Wound Treatment

    PubMed Central

    Chusri, S.; Sompetch, K.; Mukdee, S.; Jansrisewangwong, S.; Srichai, T.; Maneenoon, K.; Limsuwan, S.; Voravuthikunchai, S. P.

    2012-01-01

    Development of biofilm is a key mechanism involved in Staphylococcus epidermidis virulence during device-associated infections. We aimed to investigate antibiofilm formation and mature biofilm eradication ability of ethanol and water extracts of Thai traditional herbal recipes including THR-SK004, THR-SK010, and THR-SK011 against S. epidermidis. A biofilm forming reference strain, S. epidermidis ATCC 35984 was employed as a model for searching anti-biofilm agents by MTT reduction assay. The results revealed that the ethanol extract of THR-SK004 (THR-SK004E) could inhibit the formation of S. epidermidis biofilm on polystyrene surfaces. Furthermore, treatments with the extract efficiently inhibit the biofilm formation of the pathogen on glass surfaces determined by scanning electron microscopy and crystal violet staining. In addition, THR-SK010 ethanol extract (THR-SK010E; 0.63–5 μg/mL) could decrease 30 to 40% of the biofilm development. Almost 90% of a 7-day-old staphylococcal biofilm was destroyed after treatment with THR-SK004E (250 and 500 μg/mL) and THR-SK010E (10 and 20 μg/mL) for 24 h. Therefore, our results clearly demonstrated THR-SK004E could prevent the staphylococcal biofilm development, whereas both THR-SK004E and THR-SK010E possessed remarkable eradication ability on the mature staphylococcal biofilm. PMID:22919409

  18. Air-kerma strength determination of a new directional 103Pd source

    PubMed Central

    Reed, Joshua L.; DeWerd, Larry A.; Culberson, Wesley S.

    2015-01-01

    Purpose: A new directional 103Pd planar source array called a CivaSheet™ has been developed by CivaTech Oncology, Inc., for potential use in low-dose-rate (LDR) brachytherapy treatments. The array consists of multiple individual polymer capsules called CivaDots, containing 103Pd and a gold shield that attenuates the radiation on one side, thus defining a hot and cold side. This novel source requires new methods to establish a source strength metric. The presence of gold material in such close proximity to the active 103Pd region causes the source spectrum to be significantly different than the energy spectra of seeds normally used in LDR brachytherapy treatments. In this investigation, the authors perform air-kerma strength (SK) measurements, develop new correction factors for these measurements based on an experimentally verified energy spectrum, and test the robustness of transferring SK to a well-type ionization chamber. Methods: SK measurements were performed with the variable-aperture free-air chamber (VAFAC) at the University of Wisconsin Medical Radiation Research Center. Subsequent measurements were then performed in a well-type ionization chamber. To realize the quantity SK from a directional source with gold material present, new methods and correction factors were considered. Updated correction factors were calculated using the mcnp 6 Monte Carlo code in order to determine SK with the presence of gold fluorescent energy lines. In addition to SK measurements, a low-energy high-purity germanium (HPGe) detector was used to experimentally verify the calculated spectrum, a sodium iodide (NaI) scintillating counter was used to verify the azimuthal and polar anisotropy, and a well-type ionization chamber was used to test the feasibility of disseminating SK values for a directional source within a cylindrically symmetric measurement volume. Results: The UW VAFAC was successfully used to measure the SK of four CivaDots with reproducibilities within 0.3%. Monte

  19. DCEBIO facilitates myogenic differentiation via intermediate conductance Ca2+ activated K+ channel activation in C2C12 myoblasts.

    PubMed

    Tanaka, Shoko; Ono, Yuko; Sakamoto, Kazuho

    2017-04-01

    Membrane hyperpolarization is suggested to be a trigger for skeletal muscle differentiation. We investigated whether DCEBIO, an opener of the small/intermediate conductance Ca 2+ activated K + (SK Ca /IK Ca ) channels, increase myogenic differentiation in C2C12 skeletal myoblasts. DCEBIO significantly increased myotube formation, protein expression level of myosin heavy chain II, and mRNA expression level of myogenin in C2C12 myoblasts cultured in differentiation medium. DCEBIO induced myotube formation and hyperpolarization were reduced by the IK Ca channel blocker TRAM-34, but not by the SK Ca channel blocker apamin. These findings show that DCEBIO increases myogenic differentiation by activating IK Ca channels. Copyright © 2017 The Authors. Production and hosting by Elsevier B.V. All rights reserved.

  20. DOE Office of Scientific and Technical Information (OSTI.GOV)

    Fowler, C.J.; Ahlgren, P.C.; O'Neill, C.

    IMR-32 and SK-N-MC cells were found to contain ({sup 3}H)quinuclidinyl benzilate specific binding sites inhibited by pirenzepine in a manner suggesting the presence of both M1-type and M2-type muscarinic receptor recognition sites. Neither cell had detectable ({sup 3}H)8-OH-DPAT binding sites. Carbachol stimulated the rate of inositol phospholipid breakdown in IMR-32 and SK-N-MC human neuroblastoma cells with an EC{sub 50} value of about 50 {mu}M in both cases. Pirenzepine inhibited the carbachol stimulated inositol phospholipid breakdown in both cells with Hill slopes of unity and IC{sub 50} values of 15 nM (IMR-32) and 12 nM (SK-N-MC). The 5-HT{sub 1A} receptor agonistmore » 8-OH-DPAT competitively inhibited carbachol-stimulated inositol phospholipid breakdown with pA{sub 2} values of 5.78 (IMR-32) and 5.61 (SK-N-MC). The 5-HT agonists 5-MeODMT and buspirone at micromolar concentrations inhibited carbachol-stimulated breakdown in IMR-32 cells. The inhibition by 8-OH-DPAT and 5-MeODMT was not affected by preincubation with (-)alprenolol. 5-HT was without effect on either basal or carbachol-stimulated breakdown. It is concluded that IMR-32 and SK-N-MC neuroblastoma cells express muscarinic M1-type but not serotoninergic receptors coupled to phosphoinositide-specific phospholipase C. 8-OH-DPAT acts as a weak antagonist at these muscarinic receptors.« less

  1. Anemia and Micronutrient Status of Women of Childbearing Age and Children 6-59 Months in the Democratic Republic of the Congo.

    PubMed

    Harvey-Leeson, Sarah; Karakochuk, Crystal D; Hawes, Meaghan; Tugirimana, Pierrot L; Bahizire, Esto; Akilimali, Pierre Z; Michaux, Kristina D; Lynd, Larry D; Whitfield, Kyly C; Moursi, Mourad; Boy, Erick; Foley, Jennifer; McLean, Judy; Houghton, Lisa A; Gibson, Rosalind S; Green, Tim J

    2016-02-17

    Little is known about the micronutrient status of women and children in the Democratic Republic of the Congo, which is critical for the design of effective nutrition interventions. We recruited 744 mother-child pairs from South Kivu (SK) and Kongo Central (KC). We determined hemoglobin (Hb), serum zinc, vitamin B12, folate, ferritin, soluble transferrin receptor (sTfR), retinol binding protein (RBP), C-reactive protein, and α-1 acid glycoprotein concentrations. Anemia prevalence was determined using Hb adjusted for altitude alone and Hb adjusted for both altitude and ethnicity. Anemia prevalence was lower after Hb adjustment for altitude and ethnicity, compared to only altitude, among women (6% vs. 17% in SK; 10% vs. 32% in KC), children 6-23 months (26% vs. 59% in SK; 25% vs. 42% in KC), and children 24-59 months (14% vs. 35% in SK; 23% vs. 44% in KC), respectively. Iron deficiency was seemingly higher with sTfR as compared to inflammation-adjusted ferritin among women (18% vs. 4% in SK; 21% vs. 5% in KC), children 6-23 months (51% vs. 14% in SK; 74% vs. 10% in KC), and children 24-59 months (23% vs. 4% in SK; 58% vs. 1% in KC). Regardless of indicator, iron deficiency anemia (IDA) never exceeded 3% in women. In children, IDA reached almost 20% when sTfR was used but was only 10% with ferritin. Folate, B12, and vitamin A (RBP) deficiencies were all very low (<5%); RBP was 10% in children. The prevalence of anemia was unexpectedly low. Inflammation-adjusted zinc deficiency was high among women (52% in SK; 58% in KC), children 6-23 months (23% in SK; 20% in KC), and children 24-59 months (25% in SK; 27% in KC). The rate of biochemical zinc deficiency among Congolese women and children requires attention.

  2. Preparation and quality test of superparamagnetic iron oxide labeled antisense oligodeoxynucleotide probe: a preliminary study.

    PubMed

    Wen, Ming; Li, Bibo; Ouyang, Yu; Luo, Yi; Li, Shaolin

    2009-06-01

    transfected SK-Br-3 oncocytes by antisense probe had the lowest signal of all. The SPIO-labeled ASODN probe shows unique features including well-distributed spherical morphology, high conjugating rate and loading efficiency, and the signal intensity of SPIO-labeled ASODN-transfected SK-Br-3 oncocytes is reduced in MR imaging. These results indicate that the SPIO-labeled ASODN probe is potentially useful as a MR targeting contrast enhancing agent to specifically diagnose tumors which had over-expression of the c-erbB2 oncogene at an early stage.

  3. Body temperatures of hibernating little brown bats reveal pronounced behavioural activity during deep torpor and suggest a fever response during white-nose syndrome.

    PubMed

    Mayberry, Heather W; McGuire, Liam P; Willis, Craig K R

    2018-03-01

    Hibernating animals use torpor [reduced body temperature (T b ) and metabolic rate] to reduce energy expenditure during winter. Periodic arousals to normal T b are energetically expensive, so hibernators trade off arousal benefits against energetic costs. This is especially important for bats with white-nose syndrome (WNS), a fungal disease causing increased arousal frequency. Little brown bats (Myotis lucifugus) with WNS show upregulation of endogenous pyrogens and sickness behaviour. Therefore, we hypothesized that WNS should cause a fever response characterized by elevated T b . Hibernators could also accrue some benefits of arousals with minimal T b increase, thus avoiding full arousal costs. We compared skin temperature (T sk ) of captive Myotis lucifugus inoculated with the WNS-causing fungus to T sk of sham-inoculated controls. Infected bats re-warmed to higher T sk during arousals which is consistent with a fever response. Torpid T sk did not differ. During what we term "cold arousals", bats exhibited movement following T sk increases of only 2.2 ± 0.3 °C, compared to >20 °C increases during normal arousals. Cold arousals occurred in both infected and control bats, suggesting they are not a pathophysiological consequence of WNS. Fever responses are energetically costly and could exacerbate energy limitation and premature fat depletion for bats with WNS. Cold arousals could represent an energy-saving mechanism for both healthy and WNS-affected bats when complete arousals are unnecessary or too costly. A few cold arousals were observed mid-hibernation, typically in response to disturbances. Cold arousals may, therefore, represent a voluntary restriction of arousal temperature instead of loss of thermoregulatory control.

  4. Clinical and Histopathological Investigation of Seborrheic Keratosis

    PubMed Central

    Roh, Nam Kyung; Hahn, Hyung Jin; Lee, Yang Won; Choe, Yong Beom

    2016-01-01

    Background Seborrheic keratosis (SK) is one of the most common epidermal tumors of the skin. However, only a few large-scale clinicohistopathological investigations have been conducted on SK or on the possible correlation between histopathological SK subtype and location. Objective The aim of this study was to analyze the clinical and histopathological features of a relatively large number of cases of diagnosed SK. Methods Two hundred and seventy-one pathology slides of skin tissue from patients with clinically diagnosed SK and 206 cases of biopsy-proven SK were analyzed. The biopsy-proven cases of SK were assessed for histopathological subclassification. The demographic, clinical, and histopathological data of the patients were collected for analysis of associated factors. Results The most frequent histopathological subtype was the acanthotic type, followed by mixed, hyperkeratotic, melanoacanthoma, clonal, irritated, and adenoid types; an unexpectedly high percentage (9.2%) of the melanoacanthoma variant was observed. The adenoid type was more common in sun-exposed sites than in sun-protected sites (p=0.028). Premalignant and malignant entities together represented almost one-quarter (24.2%) of the clinicopathological mismatch cases (i.e., mismatch between the clinical and histopathological diagnoses). Regarding the location of SK development, the frequency of mismatch for the sun-exposed areas was significantly higher than that for sun-protected areas (p=0.043). Conclusion The adenoid type was more common in sun-exposed sites. Biopsy sampling should be performed for lesions situated in sun-exposed areas to exclude other premalignant or malignant diseases. PMID:27081260

  5. ROS Production Is Essential for the Apoptotic Function of E2F1 in Pheochromocytoma and Neuroblastoma Cell Lines

    PubMed Central

    Espada, Lilia; Meo-Evoli, Nathalie; Sancho, Patricia; Real, Sebastian; Fabregat, Isabel; Ambrosio, Santiago; Tauler, Albert

    2012-01-01

    In this study we demonstrate that accumulation of reactive oxygen species (ROS) is essential for E2F1 mediated apoptosis in ER-E2F1 PC12 pheochromocytoma, and SH-SY5Y and SK-N-JD neuroblastoma stable cell lines. In these cells, the ER-E2F1 fusion protein is expressed in the cytosol; the addition of 4-hydroxytamoxifen (OHT) induces its translocation to the nucleus and activation of E2F1target genes. Previously we demonstrated that, in ER-E2F1 PC12 cells, OHT treatment induced apoptosis through activation of caspase-3. Here we show that caspase-8 activity did not change upon treatment with OHT. Moreover, over-expression of Bcl-xL arrested OHT-induced apoptosis; by contrast, over-expression of c-FLIP, did not have any effect on OHT-induced apoptosis. OHT addition induces BimL expression, its translocation to mitochondria and activation of Bax, which is paralleled by diminished mitochondrial enrichment of Bcl-xL. Treatment with a Bax-inhibitory peptide reduced OHT-induced apoptosis. These results point out the essential role of mitochondria on the apoptotic process driven by E2F1. ROS accumulation followed E2F1 induction and treatment with the antioxidant N-acetylcysteine, inhibited E2F1-induced Bax translocation to mitochondria and subsequent apoptosis. The role of ROS in mediating OHT-induced apoptosis was also studied in two neuroblastoma cell lines, SH-SY5Y and SK-N-JD. In SH-SY5Y cells, activation of E2F1 by the addition of OHT induced ROS production and apoptosis, whereas over-expression of E2F1 in SK-N-JD cells failed to induce either response. Transcriptional profiling revealed that many of the genes responsible for scavenging ROS were down-regulated following E2F1-induction in SH-SY5Y, but not in SK-N-JD cells. Finally, inhibition of GSK3β blocked ROS production, Bax activation and the down regulation of ROS scavenging genes. These findings provide an explanation for the apparent contradictory role of E2F1 as an apoptotic agent versus a cell cycle activator

  6. Apoptosis in the areas of squamous differentiation of irritated seborrheic keratosis.

    PubMed

    Pesce, C; Scalora, S

    2000-03-01

    Seborrheic keratosis (SK) consists of a localized proliferation of basaloid keratinocytes, often accompanied by hyperkeratosis and hyperpigmentation. In irritated SK, these features are associated with areas of squamous differentiation with larger keratinocytes and squamous cell eddies. This work is concerned with the evaluation of apoptosis, as demonstrated by the TUNEL method, in the different varieties of SK. Apoptosis was highly expressed in the areas of squamous differentiation of irritated SK, but only mildly increased in the other varieties of SK. These data support the hypothesis that apoptosis has a role in the squamous differentiation of irritated SK. In consideration also of previous data showing that irritated SK is associated with downregulation of EGF-R expression and 125I-EGF binding, we postulate that the morphologic features of irritated SK could correspond to an involution phase of the disease, characterized by altered cell balance with inadequate cell renewal and increased cell loss.

  7. Polynomial interpolation and sums of powers of integers

    NASA Astrophysics Data System (ADS)

    Cereceda, José Luis

    2017-02-01

    In this note, we revisit the problem of polynomial interpolation and explicitly construct two polynomials in n of degree k + 1, Pk(n) and Qk(n), such that Pk(n) = Qk(n) = fk(n) for n = 1, 2,… , k, where fk(1), fk(2),… , fk(k) are k arbitrarily chosen (real or complex) values. Then, we focus on the case that fk(n) is given by the sum of powers of the first n positive integers Sk(n) = 1k + 2k + ṡṡṡ + nk, and show that Sk(n) admits the polynomial representations Sk(n) = Pk(n) and Sk(n) = Qk(n) for all n = 1, 2,… , and k ≥ 1, where the first representation involves the Eulerian numbers, and the second one the Stirling numbers of the second kind. Finally, we consider yet another polynomial formula for Sk(n) alternative to the well-known formula of Bernoulli.

  8. Thermal conductivity of pure silica MEL and MFI zeolite thin films

    NASA Astrophysics Data System (ADS)

    Coquil, Thomas; Lew, Christopher M.; Yan, Yushan; Pilon, Laurent

    2010-08-01

    This paper reports the room temperature cross-plane thermal conductivity of pure silica zeolite (PSZ) MEL and MFI thin films. PSZ MEL thin films were prepared by spin coating a suspension of MEL nanoparticles in 1-butanol solution onto silicon substrates followed by calcination and vapor-phase silylation with trimethylchlorosilane. The mass fraction of nanoparticles within the suspension varied from 16% to 55%. This was achieved by varying the crystallization time of the suspension. The thin films consisted of crystalline MEL nanoparticles embedded in a nonuniform and highly porous silica matrix. They featured porosity, relative crystallinity, and MEL nanoparticles size ranging from 40% to 59%, 23% to 47% and 55 nm to 80 nm, respectively. PSZ MFI thin films were made by in situ crystallization, were b-oriented, fully crystalline, and had a 33% porosity. Thermal conductivity of these PSZ thin films was measured at room temperature using the 3ω method. The cross-plane thermal conductivity of the MEL thin films remained nearly unchanged around 1.02±0.10 W m-1 K-1 despite increases in (i) relative crystallinity, (ii) MEL nanoparticle size, and (iii) yield caused by longer nanoparticle crystallization time. Indeed, the effects of these parameters on the thermal conductivity were compensated by the simultaneous increase in porosity. PSZ MFI thin films were found to have similar thermal conductivity as MEL thin films even though they had smaller porosity. Finally, the average thermal conductivity of the PSZ films was three to five times larger than that reported for amorphous sol-gel mesoporous silica thin films with similar porosity and dielectric constant.

  9. Ghrelin ameliorates the human alveolar epithelial A549 cell apoptosis induced by lipopolysaccharide

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Huang, Chunrong; Zheng, Haichong; He, Wanmei

    Ghrelin is a gastric acyl-peptide that plays an inhibitory role in cell apoptosis. Herein we investigate the protective effects of ghrelin in LPS-induced apoptosis of human alveolar epithelial A549 cells, along with the possible molecular mechanisms. LPS exposure impaired cell viability and increased apoptosis of A549 cells significantly in concentration- and time-dependent manners embodied in increased Bax and cleaved caspase-3 production, coupled with decreased Bcl-2 levels. Simultaneously, LPS remarkably decreased the expression of phosphatidylinositol 3 kinase/protein kinase B (PI3K/Akt) and extracellular signal-regulated kinas (ERK) in A549 cells. However, ghrelin'pretreatment ameliorated LPS-caused alterations in the ratio of Bax/Bcl-2 and cleaved caspase-3 expression, whereas activatedmore » the PI3K/Akt and ERK signaling. These results demonstrate that ghrelin lightens LPS-induced apoptosis of human alveolar epithelial cells partly through activating the PI3K/Akt and ERK pathway and thereby might benefit alleviating septic ALI. -- Graphical abstract: Ghrelin ameliorates the human alveolar epithelial A549 cells apoptosis induced by lipopolysaccharide partly through activating the PI3K/Akt and ERK pathway. Display Omitted -- Highlights: •It has been observed that LPS insult significantly increased apoptosis in A549 cells. •Both Akt and ERK signaling are critical adapter molecules to mediate the ghrelin-mediated proliferative effect. •Ghrelin may have a therapeutic effect in the prevention of LPS-induced apoptosis.« less

  10. Logistics Study -- Shelter Stockage.

    DTIC Science & Technology

    1979-11-01

    Supplies such as paper cups, toilet tissue, sanitary napkins , commode 4 chemicals, etc., must be provided. 4 WATER REQUIREMENTS Potable water and its...PROPOSED SANITATION KITS ITEM UI SK-III SK-IV SK-V Container ea 1 1 1 * Toilet Tissue ro 5 10 10 Can Opener ea 1 1 - Sanitary Napkins ea 36 60 60 Water...34 3.3.5 Unit No. 5. -- One set of assembly instructions as delineated in Figure 4. -he instructions shall be printed in black or blue ink on a sheet

  11. Melphalan plus total body irradiation (MEL-TBI) or cyclophosphamide (MEL-CY) as a conditioning regimen with second autotransplant in responding patients with myeloma is inferior compared to historical controls receiving tandem transplants with melphalan alone.

    PubMed

    Desikan, K R; Tricot, G; Dhodapkar, M; Fassas, A; Siegel, D; Vesole, D H; Jagannath, S; Singhal, S; Mehta, J; Spoon, D; Anaissie, E; Barlogie, B; Munshi, N

    2000-03-01

    The role of more intense conditioning for second transplant was evaluated in myeloma patients achieving at least partial remission (PR) after first transplant with melphalan at 200 mg/m2. Forty-three patients received more intensive conditioning for the second transplant. Nineteen patients received cyclophosphamide 120 mg/kg along with melphalan 200 g/m2 (MEL-CY; group 1) while 24 patients received total body irradiation (1125 cGy) in conjunction with melphalan 140 mg/m2 (MEL-TBI; group 2). Forty-three matched control patients were identified from 450 patients receiving melphalan alone for second transplant (MEL200; group 3). Engraftment and toxicities were comparable among the groups with the exception of increased treatment-related mortality of 8% in group 2 compared to none in groups 1 and 3 (P = 0.07). Despite identical CR rates of 74, 71 and 70%, respectively, in groups 1, 2 and 3 (P = 1.0), event-free survival (median: 27, 15 and 61; P < 0.0001) and overall survival (median: 39, 25 and 76 months; P = 0.003) were significantly decreased in patients receiving more intensive conditioning (groups 1 and 2). Lymphocyte recovery, evaluated as a surrogate for immune recovery, was inferior in more intensively treated patients (groups 1 and 2 compared to group 3). Our findings suggest that more intense conditioning appears to have no benefit in patients responding to their first cycle of high-dose therapy and may even be detrimental in this setting. Bone Marrow Transplantation (2000) 25, 483-487.

  12. Phloretin induces apoptosis of non-small cell lung carcinoma A549 cells via JNK1/2 and p38 MAPK pathways

    PubMed Central

    MIN, JIE; LI, XU; HUANG, KENAN; TANG, HUA; DING, XINYU; QI, CHEN; QIN, XIONG; XU, ZHIFEI

    2015-01-01

    Phloretin (Ph) existing in apples, pears and various vegetables is known to have antitumor activities in several cancer cell lines. However, little is known about its effect on human lung cancer cells. The aim of the present study was to see whether Ph could induce apoptosis of non-small cell lung cancer (NSCLC) cells, and explore the possible underlying mechanism of action. We found that Ph markedly induced cell apoptosis of NSCLC cell line A549, and inhibited the migration of A549 cells in a dose-dependent manner. The expression level of BAX, cleaved caspase-3 and -9, and degraded form of PARP was increased and Bcl-2 was decreased after Ph treatment. In addition, the phosphorylation of P38 MAPK, ERK1/2 and JNK1/2 was increased in a dose-dependent manner in parallel with Ph treatment. Inhibition of P38 MAPK and JNK1/2 by specific inhibitors significantly abolished the Ph-induced activation of the caspase-3 and -9. In vivo tumor-suppression assay further indicated that Ph (20 mg/kg) displayed a more significant inhibitory effect on A549 xenografts in tumor growth. All these findings indicate that Ph is able to inhibit NSCLC A549 cell growth by inducing apoptosis through P38 MAPK and JNK1/2 pathways, and therefore may prove to be an adjuvant to the treatment of NSCLC. PMID:26503828

  13. Validating tyrosinase homologue MelA as a photoacoustic reporter gene for imaging Escherichia coli

    NASA Astrophysics Data System (ADS)

    Paproski, Robert J.; Li, Yan; Barber, Quinn; Lewis, John D.; Campbell, Robert; Zemp, Roger

    2015-03-01

    Antibiotic drug resistance is a major worldwide issue. Development of new therapies against pathogenic bacteria requires appropriate research tools for replicating and characterizing infections. Previously fluorescence and bioluminescence modalities have been used to image infectious burden in animal models but scattering significantly limits imaging depth and resolution. We hypothesize that photoacoustic imaging, which has improved depth-toresolution ratio, could be useful for visualizing MelA-expressing bacteria since MelA is a bacterial tyrosinase homologue involved in melanin production. Using an inducible expression system, E. coli expressing MelA were visibly black in liquid culture. Phosphate buffered saline (PBS), MelA-expressing bacteria (at different dilutions in PBS), and chicken embryo blood were injected in plastic tubes which were imaged using a VisualSonics Vevo LAZR system. Photoacoustic imaging at 6 different wavelengths (680, 700, 750, 800, 850 and 900nm) enabled spectral de-mixing to distinguish melanin signals from blood. The signal to noise ratio of 9x diluted MelA bacteria was 55, suggesting that ~20 bacteria cells could be detected with our system. When MelA bacteria were injected as a 100 μL bolus into a chicken embryo, photoacoustic signals from deoxy- and oxy- hemoglobin as well as MelA-expressing bacteria could be separated and overlaid on an ultrasound image, allowing visualization of the bacterial location. Photoacoustic imaging may be a useful tool for visualizing bacterial infections and further work incorporating photoacoustic reporters into infectious bacterial strains is warranted.

  14. Secretion pathway of liver IGF-1 via JAK2/STAT3 in chick embryo under the monochromatic light.

    PubMed

    Wang, Tuanjie; Dong, Yulan; Wang, Zixu; Cao, Jing; Chen, Yaoxing

    2016-02-01

    This study reveals mechanism of monochromatic light on the IGF-1 secretion of chick embryo liver. The chick embryos were incubated and exposed to continuous red, green, blue light or a dark environment. Compared to other light-treated groups, green light increased IGF-1 and melatonin concentrations both in plasma and liver, and Mel1a, Mel1b and Mel1c receptors expressions in liver but decreased p-JAK2, p-STAT3 and ROS in liver. IGF-1 had a positive correlation with melatonin, but a negative relevance with p-JAK2 and p-STAT3. In vitro, the IGF-1 level in the hepatocyte supernatant was enhanced by melatonin with lower p-JAK2/p-STAT3 and ROS levels, which was suppressed by Mel1c antagonist but not Mel1a/Mel1b or Mel1b antagonists. AG490 (JAK/STAT inhibitor) promoted role of melatonin-Mel1c modulated IGF-1 secretion. These results suggest the antioxidant effect of melatonin mediated the green light-enhanced IGF-1 secretion of chick embryo liver through Mel1c receptor to inhibit the JAK2/STAT3 pathway.

  15. Tumour detection and localization using 99Tcm-labelled OV-TL 3 Fab' in patients suspected of ovarian cancer.

    PubMed

    Tibben, J G; Massuger, L F; Claessens, R A; Schijf, C P; Pak, K Y; Strijk, S P; Kenemans, P; Corstens, F H

    1992-12-01

    Fab' fragments of the monoclonal antibody OV-TL 3, that recognizes an ovarian carcinoma-associated antigen (OA3), were labelled with 99Tcm using D-glucarate as a ligand. Twenty patients suspected of having primary or recurrent ovarian cancer received intravenously 1 mg of the Fab' labelled with 740 MBq 99Tcm. Both planar and single photon emission computed tomographic (SPECT) scintigraphy were performed up to 30 h after intravenous infusion. In 19 out of 20 patients surgical and histopathological evaluation was performed between 2 and 6 days postinfusion. Imaging results were compared with X-ray computed tomography (CT), ultrasonography (US) and CA 125 serum level. Blood clearance was fast with median t1/2 beta of 9.5 h. Thirty-seven per cent of the injected dose (% ID) was excreted in the urine within the first 24 h, whereas 7% ID was excreted in the 24 h faeces. In one patient with an OA3 negative ovarian carcinoma, radioimmunoscintigraphy (RIS) did not visualize the tumour. In two other patients a benign ovarian cyst was found, also showing no elevated uptake. In 13 out of 17 patients ovarian cancer lesions were detected with RIS, whereas CT and US detected lesions in, respectively, 15 and 12 patients. Of 36 surgically defined tumour deposits larger than 1 cm in diameter, 53% were detected and localized with RIS, whereas CT and US detected 61 and 40%, respectively. Radioimmunoscintigraphy with 99Tcm-OV-TL 3 Fab' is less distressing for the patients but the overall imaging performance is not improved when compared with 111In-OV-TL 3 F(ab')2.

  16. Antiurolithiatic Effect of Sirupeelai Samoola Kudineer: A Polyherbal Siddha Decoction on Ethylene Glycol-induced Renal Calculus in Experimental Rats

    PubMed Central

    Vasanthi, A. Hannah Rachel; Muthulakshmi, V.; Gayathri, V.; Manikandan, R.; Ananthi, S.; Kuruvilla, Sarah

    2017-01-01

    Background: Sirupeelai Samoola Kudineer (SK), a polyherbal decoction containing four medicinal plants has been used in Siddha system of medicine, practiced in Southern parts of India for the management of urolithiasis. Objective: The present study is carried out to scientifically validate the traditional claim and to study the mechanism of action of the drug. Materials and Methods: In the present study, anti-urolithiatic effect of SK was evaluated in Sprague-Dawley rats using ethylene glycol through drinking water and intraperitoneal injection of sodium oxalate. Renal damage was confirmed by the increased production of thiobarbituric acid reactive substance (TBARS). Results: Co-treatment with SK to urolithiatic rats for 21 days significantly prevented the elevation of renal and urinary stone biomarkers in plasma and renal tissue thereby preventing renal damage and the formation of renal calculi. Administration of SK at all doses and cystone restored the antioxidant (glutathione) levels by preventing the elevation of TBARS in the kidney tissue, which was further confirmed by histological sections. Conclusions: SK treatment promotes diuresis which leads to flushing of the renal stones and maintains the alkaline environment in the urinary system which probably mediates the antilithiatic activity. SK provides structural and functional protection to the kidneys by enhancing its physiological function against stone formation and validates its clinical use. SUMMARY SK exhibited antilithiatic and diuretic potential in ethylene glycol and sodium oxalate induced urolithiasis in ratsElevated urinary stone markers (Calcium, oxalate, uric acid, magnesium and phosphates) in plasma and renal tubular enzymes (LDH, GGT, ALP, AST ALT) in urolithiatic rats were reversed by SK treatmentSK administration significantly reduced the level of renal stress markers like Urea, Creatinine, LPO and elevated SOD, GPx, GSH levels aiding in nephroprotectionSK also provides structural and

  17. Alstonine as a potential fluorescent marker for tiny tumor detection and imaging

    NASA Astrophysics Data System (ADS)

    Viallet, Pierre M.; Vo-Dinh, Tuan; Salmon, Jean-Marie; Watts, Wendi; Rocchi, Emmanuelle; Isola, Narayana R.; Rebillard, Xavier

    1997-06-01

    3,4,5,6,16,17-Hexadehydro-16-(methoxycarbolyl)-19(alpha) - methyl-20(alpha) -oxyohimbanium (alstonine) is a fluorescent alcaloid which is known to stain tumor cells more efficiently than normal. The interactions between alstonine and biological macromolecules were first investigated to provide the rationale for preferential labelling. Molecular filtration and spectrosfluorometric techniques with different macromolecules and isopolynucleotides have demonstrated that binding occurs only in the presence of uridyl rings. For the binding affect only the fluorescence intensity of alstonine it can be assumed that it involves only the side chain of the fluorescent compound. The capability for preferential staining was verified in culture using SK-OV-3 cells and rat hepatocarcinoma cells as tumor cells and Mouse fibroblasts or rat liver cells as controls. Techniques of image analysis have demonstrated the efficiency of cellular labelling even in aggregates of rat hepatocarcinoma. These experiments lead the way to the detection of tiny tumors developed on thin visceral walls, using a fiber optic device.

  18. Towards a Learning Society: A Framework for Research.

    ERIC Educational Resources Information Center

    Schuller, Tom

    1992-01-01

    This research framework has three levels: (1) societal (individual, household/family, work organization, regional/national/international); (2) disciplinary approach (social anthropology, sociology, industrial relations, economics, politics); and (3) time (historical/diachronic, life course, routines). (SK)

  19. MEL-18 loss mediates estrogen receptor–α downregulation and hormone independence

    PubMed Central

    Lee, Jeong-Yeon; Won, Hee-Young; Park, Ji-Hye; Kim, Hye-Yeon; Choi, Hee-Joo; Shin, Dong-Hui; Kang, Ju-Hee; Woo, Jong-Kyu; Oh, Seung-Hyun; Son, Taekwon; Choi, Jin-Woo; Kim, Sehwan; Kim, Hyung-Yong; Yi, Kijong; Jang, Ki-Seok; Oh, Young-Ha; Kong, Gu

    2015-01-01

    The polycomb protein MEL-18 has been proposed as a tumor suppressor in breast cancer; however, its functional relevance to the hormonal regulation of breast cancer remains unknown. Here, we demonstrated that MEL-18 loss contributes to the hormone-independent phenotype of breast cancer by modulating hormone receptor expression. In multiple breast cancer cohorts, MEL-18 was markedly downregulated in triple-negative breast cancer (TNBC). MEL-18 expression positively correlated with the expression of luminal markers, including estrogen receptor–α (ER-α, encoded by ESR1). MEL-18 loss was also associated with poor response to antihormonal therapy in ER-α–positive breast cancer. Furthermore, whereas MEL-18 loss in luminal breast cancer cells resulted in the downregulation of expression and activity of ER-α and the progesterone receptor (PR), MEL-18 overexpression restored ER-α expression in TNBC. Consistently, in vivo xenograft experiments demonstrated that MEL-18 loss induces estrogen-independent growth and tamoxifen resistance in luminal breast cancer, and that MEL-18 overexpression confers tamoxifen sensitivity in TNBC. MEL-18 suppressed SUMOylation of the ESR1 transactivators p53 and SP1, thereby driving ESR1 transcription. MEL-18 facilitated the deSUMOylation process by inhibiting BMI-1/RING1B-mediated ubiquitin-proteasomal degradation of SUMO1/sentrin-specific protease 1 (SENP1). These findings demonstrate that MEL-18 is a SUMO-dependent regulator of hormone receptors and suggest MEL-18 expression as a marker for determining the antihormonal therapy response in patients with breast cancer. PMID:25822021

  20. β-adrenergic blockade does not impair the skin blood flow sensitivity to local heating in burned and non-burned skin under neutral and hot environments in children

    PubMed Central

    Rivas, Eric; McEntire, Serina J.; Herndon, David N.; Mlcak, Ronald P.; Suman, Oscar E.

    2017-01-01

    Objective Tested the hypothesis that propranolol, a drug given to burn patients to reduce hypermetabolism/cardiac stress, may inhibit heat dissipation by changing the sensitivity of skin blood flow (SkBF) to local heating under neutral and hot conditions. Methods In a randomized double-blind study, a placebo was given to 8 burned children while propranolol was given to 13 burned children with similar characteristics (mean ± SD: 11.9±3y, 147±20cm, 45±23kg, 56±12% TBSA). Non-burned children (n=13, 11.4±3y, 152±15cm, 52±13kg) served as healthy controls. A progressive local heating protocol characterized SkBF responses in burned and unburned skin and non-burned control skin under the two environmental conditions (23°C and 34°C) via laser-Doppler flowmetry. Results Resting SkBF was greater in burned and unburned skin compared to the non-burned control (main effect: skin, P<0.0001; 57±32 burned; 38±36 unburned vs 9±8 control %SkBFmax). No difference was found for maximal SkBF capacity to local heating between groups. Additionally, dose response curves for the sensitivity of SkBF to local heating were not different among burned or unburned skin, and non-burned control skin (EC50, P>0.05) under either condition. Conclusion Therapeutic propranolol does not negatively affect SkBF under neutral or hot environmental conditions and further compromise temperature regulation in burned children. PMID:28071840