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Sample records for extracellular domain ecd

  1. Immunization with a novel chimeric peptide representing B and T cell epitopes from HER2 extracellular domain (HER2 ECD) for breast cancer.

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    Mahdavi, Manijeh; Keyhanfar, Mehrnaz; Jafarian, Abbas; Mohabatkar, Hassan; Rabbani, Mohammad

    2014-12-01

    Because of direct stimulating immune system against disease, vaccination or active immunotherapy is preferable compared to passive immunotherapy. For this purpose, a newly designed chimeric peptide containing epitopes for both B and T cells from HER2 ECD subdomain III was proposed. To evaluate the effects of the active immunization, a discontinuous B cell epitope peptide was selected based on average antigenicity by bioinformatics analysis. The selected peptide was collinearly synthesized as a chimera with a T helper epitope from the protein sequence of measles virus fusion (208-302) using the GPSL linker. Three mice were immunized with the chimeric peptide. Reactive antibodies with HER2 protein in ELISA and immunofluorescence assays with no cross-reactivity were generated. The 3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide (MTT) assay indicated that the anti-peptide sera had inhibitory effects on proliferation of SK-BR-3 cells. Hence, the newly designed, discontinuous chimeric peptide representing B and T cell epitopes from subdomain III of HER2-ECD can form the basis for future vaccines design, where these data can be applied for monoclonal antibody production targeting the distinct epitope of HER2 receptor compared to the two broadly used anti-HER2 monoclonal antibodies, Herceptin and pertuzumab.

  2. Soluble FGFR4 extracellular domain inhibits FGF19-induced activation of FGFR4 signaling and prevents nonalcoholic fatty liver disease

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    Chen, Qiang [State Key Laboratory of Stress Cell Biology, School of Life Sciences, Xiamen University, Xiamen (China); The First Affiliated Hospital of Xiamen University, Xiamen (China); Jiang, Yuan; An, Yuan; Zhao, Na; Zhao, Yang [State Key Laboratory of Stress Cell Biology, School of Life Sciences, Xiamen University, Xiamen (China); Yu, Chundong, E-mail: cdyu@xmu.edu.cn [State Key Laboratory of Stress Cell Biology, School of Life Sciences, Xiamen University, Xiamen (China)

    2011-06-17

    Highlights: {yields} Soluble FGFR4 extracellular domain (FGFR4-ECD) was effectively expressed. {yields} FGFR4-ECD inhibited FGF19-induced activation of FGFR4 signaling. {yields} FGFR4-ECD reduced palmitic acid-induced steatosis of HepG2 cells. {yields} FGFR4-ECD reduced tetracycline-induced fatty liver in mice. {yields} FGFR4-ECD partially restored tetracycline-repressed PPAR{alpha} expression. -- Abstract: Fibroblast growth factor receptor 4 (FGFR4) is a transmembrane tyrosine kinase receptor that plays a crucial role in the regulation of hepatic bile acid and lipid metabolism. FGFR4 underlies high-fat diet-induced hepatic steatosis, suggesting that inhibition of FGFR4 activation may be an effective way to prevent or treat nonalcoholic fatty liver disease (NAFLD). To determine whether neutralization of FGFR4 ligands by soluble FGFR4 extracellular domain (FGFR4-ECD) can inhibit the activation of FGFR4, we constructed FGFR4-ECD expression vector and showed that FGFR4-ECD was effectively expressed in cells and secreted into culture medium. FGFR4-ECD inhibited FGF19-induced activation of FGFR4 signaling and reduced steatosis of HepG2 induced by palmitic acid in vitro. Furthermore, in a tetracycline-induced fatty liver model, expression of FGFR4-ECD in mouse liver reduced the accumulation of hepatic lipids and partially restored the expression of peroxisome proliferator-activated receptor {alpha} (PPAR{alpha}), which promotes the mitochondrial fatty acid beta-oxidation but is repressed by tetracycline. Taken together, these results demonstrate that FGFR4-ECD can block FGFR4 signaling and prevent hepatic steatosis, highlighting the potential value of inhibition of FGFR4 signaling as a method for therapeutic intervention against NAFLD.

  3. Soluble FGFR4 extracellular domain inhibits FGF19-induced activation of FGFR4 signaling and prevents nonalcoholic fatty liver disease

    International Nuclear Information System (INIS)

    Chen, Qiang; Jiang, Yuan; An, Yuan; Zhao, Na; Zhao, Yang; Yu, Chundong

    2011-01-01

    Highlights: → Soluble FGFR4 extracellular domain (FGFR4-ECD) was effectively expressed. → FGFR4-ECD inhibited FGF19-induced activation of FGFR4 signaling. → FGFR4-ECD reduced palmitic acid-induced steatosis of HepG2 cells. → FGFR4-ECD reduced tetracycline-induced fatty liver in mice. → FGFR4-ECD partially restored tetracycline-repressed PPARα expression. -- Abstract: Fibroblast growth factor receptor 4 (FGFR4) is a transmembrane tyrosine kinase receptor that plays a crucial role in the regulation of hepatic bile acid and lipid metabolism. FGFR4 underlies high-fat diet-induced hepatic steatosis, suggesting that inhibition of FGFR4 activation may be an effective way to prevent or treat nonalcoholic fatty liver disease (NAFLD). To determine whether neutralization of FGFR4 ligands by soluble FGFR4 extracellular domain (FGFR4-ECD) can inhibit the activation of FGFR4, we constructed FGFR4-ECD expression vector and showed that FGFR4-ECD was effectively expressed in cells and secreted into culture medium. FGFR4-ECD inhibited FGF19-induced activation of FGFR4 signaling and reduced steatosis of HepG2 induced by palmitic acid in vitro. Furthermore, in a tetracycline-induced fatty liver model, expression of FGFR4-ECD in mouse liver reduced the accumulation of hepatic lipids and partially restored the expression of peroxisome proliferator-activated receptor α (PPARα), which promotes the mitochondrial fatty acid beta-oxidation but is repressed by tetracycline. Taken together, these results demonstrate that FGFR4-ECD can block FGFR4 signaling and prevent hepatic steatosis, highlighting the potential value of inhibition of FGFR4 signaling as a method for therapeutic intervention against NAFLD.

  4. The mannosylated extracellular domain of Her2/neu produced in P. pastoris induces protective antitumor immunity

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    Dimitriadis, Alexios; Gontinou, Chrysanthi; Baxevanis, Constantin N; Mamalaki, Avgi

    2009-01-01

    Her2/neu is overexpressed in various human cancers of epithelial origin and is associated with increased metastatic potential and poor prognosis. Several attempts have been made using the extracellular domain of Her2/neu (ECD/Her2) as a prophylactic vaccine in mice with no success in tumor prevention. The extracellular domain of Her2/neu (ECD/Her2) was expressed in yeast P. pastoris, in a soluble highly mannosylated form. The immune response of the immunization with this recombinant ECD/Her2 was analyzed using immunoprecipitation and western blot analysis, proliferation and cytotoxicity assays as well as specific tumor growth assays. Mannosylated ECD/Her2 elicited a humoral response with HER2/neu specific antibodies in vaccinated mice, which were able to reduce the proliferation rate of cancer cells in vitro. Moreover, it elicited a cellular response with Her2/neu-specific CTL capable of lysing tumor cells, in vitro. When immunized Balb/c and HHD mice were challenged with Her2/neu-overexpressing cells, tumor growth was inhibited. Here we report on the efficacy of the extracellular domain of human Her2/neu produced in yeast P. pastoris, which confers mannosylation of the protein, to act as a potent anti-tumor vaccine against Her2/neu overexpressing tumors. Specific cellular and humoral responses were observed as well as efficacy

  5. Differential Requirement of the Extracellular Domain in Activation of Class B G Protein-coupled Receptors.

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    Zhao, Li-Hua; Yin, Yanting; Yang, Dehua; Liu, Bo; Hou, Li; Wang, Xiaoxi; Pal, Kuntal; Jiang, Yi; Feng, Yang; Cai, Xiaoqing; Dai, Antao; Liu, Mingyao; Wang, Ming-Wei; Melcher, Karsten; Xu, H Eric

    2016-07-15

    G protein-coupled receptors (GPCRs) from the secretin-like (class B) family are key players in hormonal homeostasis and are important drug targets for the treatment of metabolic disorders and neuronal diseases. They consist of a large N-terminal extracellular domain (ECD) and a transmembrane domain (TMD) with the GPCR signature of seven transmembrane helices. Class B GPCRs are activated by peptide hormones with their C termini bound to the receptor ECD and their N termini bound to the TMD. It is thought that the ECD functions as an affinity trap to bind and localize the hormone to the receptor. This in turn would allow the hormone N terminus to insert into the TMD and induce conformational changes of the TMD to activate downstream signaling. In contrast to this prevailing model, we demonstrate that human class B GPCRs vary widely in their requirement of the ECD for activation. In one group, represented by corticotrophin-releasing factor receptor 1 (CRF1R), parathyroid hormone receptor (PTH1R), and pituitary adenylate cyclase activating polypeptide type 1 receptor (PAC1R), the ECD requirement for high affinity hormone binding can be bypassed by induced proximity and mass action effects, whereas in the other group, represented by glucagon receptor (GCGR) and glucagon-like peptide-1 receptor (GLP-1R), the ECD is required for signaling even when the hormone is covalently linked to the TMD. Furthermore, the activation of GLP-1R by small molecules that interact with the intracellular side of the receptor is dependent on the presence of its ECD, suggesting a direct role of the ECD in GLP-1R activation. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

  6. The extracellular domain of Her2 in serum as a biomarker of breast cancer.

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    Perrier, Alexandre; Gligorov, Joseph; Lefèvre, Guillaume; Boissan, Mathieu

    2018-02-28

    Breast cancer is a major health problem worldwide. In ~15% of breast cancers, the epidermal growth factor receptor HER2, a transmembrane protein, is overexpressed. This HER2 overexpression is associated with an aggressive form of the disease and a poor clinical prognosis. The extracellular domain (ECD) of HER2 is released into the blood by a proteolytic mechanism known as "ECD shedding". This proteolytic shedding leaves a constitutively active truncated receptor in the membrane that is 10-100-fold more oncogenic than the full-length receptor and promotes the growth and survival of cancer cells. Shedding of the HER2 ECD is increased during metastasis: whereas 15% of primary breast cancer patients have elevated levels of serum HER2 ECD (sHER2 ECD), the levels reach 45% in patients with metastatic disease. Thus, sHER2 ECD has been proposed as a promising biomarker for cancer recurrence and for monitoring the disease status of patients overexpressing HER2. Nevertheless, in 2016, the American Society of Clinical Oncology advises clinicians not to use soluble HER2 levels to guide their choice of adjuvant therapy for patients with HER2-positive breast cancer, because the evidence was considered not strong enough. Currently, biomarkers such as carcinoembryonic antigen and cancer antigen 15-3 are widely used to monitor metastatic breast cancer disease even if the level of evidence of clinical impact of this monitoring is poor. In this article, we review the evidence that sHER2 ECD might be used in some situations as a biomarker for breast cancer. Although this serum biomarker will not replace the direct measurement of tumor HER2 status for diagnosis of early-stage tumors; it might be especially useful in metastatic disease for prognosis, as an indicator of cancer progression and of therapy response, particularly to anti-HER2 therapies. Owing to these data, sHER2 ECD should be considered as a promising biomarker to detect cancer recurrence and metastasis.

  7. Serum HER 2 extracellular domain level is correlated with tissue HER 2 status in metastatic gastric or gastro-oesophageal junction adenocarcinoma.

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    Shu-Qin Dai

    Full Text Available BACKGROUND: To explore the association between serum human epidermal growth factor receptor 2 (HER 2 extracellular domain (ECD levels and tissue HER 2 status in metastatic gastric cancer. PATIENTS AND METHODS: HER 2 status was retrospectively analyzed in 219 advanced gastric or gastroesophageal junction (GEJ patients. Serum HER 2 ECD was measured by chemiluminescent assay and tissue HER 2 was assessed by fluorescent in situ hybridisation (FISH and immunohistochemistry (IHC assay. RESULTS: Significant associations were found between serum HER 2 ECD levels and tissue HER 2 status. Twenty-four patients had HER 2 ECD levels >16.35 ng/mL, which has a sensitivity of 51.4% and a specificity of 97.3% to predict tissue HER 2 status. When the cut-off value was increased to 22 ng/mL, then all 12 patients with serum HER 2 ECD levels>22 ng/mL were tissue HER 2 positive, corresponding to a specificity of 100% and a sensitivity of 32.4%. High serum HER 2 ECD levels were strongly associated with the intestinal histological type (Lauren's classification, liver metastasis, multiple metastasis (>2 and increased LDH levels, but not with overall survival. CONCLUSIONS: The high specificity of the serum HER 2 ECD assay in predicting tissue HER 2 status suggests its potential as a surrogate marker of the HER 2 status in gastric cancer.

  8. The glycosylated IgII extracellular domain of EMMPRIN is implicated in the induction of MMP-2.

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    Papadimitropoulou, Adriana; Mamalaki, Avgi

    2013-07-01

    EMMPRIN is a widely expressed transmembrane glycoprotein that plays important roles in many physiological and pathological processes, such as tumor invasion and metastasis. It stimulates the production of matrix metalloproteinase (MMPs) by tumor-associated fibroblasts. In the present study, our aim was to (a) to investigate if the IgII loop domain of the extracellular domain (ECD) of EMMPRIN contributes to the MMP production by fibroblasts and (b) to evaluate the significance of glycosylation in this process. For this purpose, we expressed the ECD, IgI, or IgII domains of EMMPRIN, in their glycosylated and non-glycosylated forms, in the heterologous expression systems of P. pastoris and E. coli, respectively. Dermal fibroblasts were treated with purified recombinant domains and proteins from cell extracts and supernatants were analyzed by Western blot and zymography assays. Fibroblasts treated with ECD-, IgI-, and IgII-glycosylated domains of EMMPRIN significantly stimulated the gelatinolytic activity of MMP-2, compared to untreated fibroblasts, whereas no significant effect was observed after treatment with the non-glycosylated ECD, IgI, and IgII domains. Western blot analysis from cell extracts and supernatants revealed that only the glycosylated forms were able to stimulate MMP-2 production and secretion, respectively. Quantitative PCR revealed that this effect was not attributed to transcriptional alterations. This study showed that N-glycosylation was a prerequisite for efficient MMP-2 production, with the IgII loop domain contributing significantly to this process. Perturbation of the function of IgII-EMMPRIN loop could have potential therapeutic value in the inhibition of MMP-2-dependent cancer cell invasion and metastasis.

  9. The extracellular domain of neurotrophin receptor p75 as a candidate biomarker for amyotrophic lateral sclerosis.

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    Stephanie R Shepheard

    Full Text Available Objective biomarkers for amyotrophic lateral sclerosis would facilitate the discovery of new treatments. The common neurotrophin receptor p75 is up regulated and the extracellular domain cleaved from injured neurons and peripheral glia in amyotrophic lateral sclerosis. We have tested the hypothesis that urinary levels of extracellular neurotrophin receptor p75 serve as a biomarker for both human motor amyotrophic lateral sclerosis and the SOD1(G93A mouse model of the disease. The extracellular domain of neurotrophin receptor p75 was identified in the urine of amyotrophic lateral sclerosis patients by an immuno-precipitation/western blot procedure and confirmed by mass spectrometry. An ELISA was established to measure urinary extracellular neurotrophin receptor p75. The mean value for urinary extracellular neurotrophin receptor p75 from 28 amyotrophic lateral sclerosis patients measured by ELISA was 7.9±0.5 ng/mg creatinine and this was significantly higher (p<0.001 than 12 controls (2.6±0.2 ng/mg creatinine and 19 patients with other neurological disease (Parkinson's disease and Multiple Sclerosis; 4.1±0.2 ng/mg creatinine. Pilot data of disease progression rates in 14 MND patients indicates that p75NTR(ECD levels were significantly higher (p = 0.0041 in 7 rapidly progressing patients as compared to 7 with slowly progressing disease. Extracellular neurotrophin receptor p75 was also readily detected in SOD1(G93A mice by immuno-precipitation/western blot before the onset of clinical symptoms. These findings indicate a significant relation between urinary extracellular neurotrophin receptor p75 levels and disease progression and suggests that it may be a useful marker of disease activity and progression in amyotrophic lateral sclerosis.

  10. X-ray crystallographic studies of the extracellular domain of the first plant ATP receptor, DORN1, and the orthologous protein from Camelina sativa

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    Li, Zhijie; Chakraborty, Sayan; Xu, Guozhou (NCSU)

    2016-10-26

    Does not respond to nucleotides 1 (DORN1) has recently been identified as the first membrane-integral plant ATP receptor, which is required for ATP-induced calcium response, mitogen-activated protein kinase activation and defense responses inArabidopsis thaliana. In order to understand DORN1-mediated ATP sensing and signal transduction, crystallization and preliminary X-ray studies were conducted on the extracellular domain of DORN1 (atDORN1-ECD) and that of an orthologous protein,Camelina sativalectin receptor kinase I.9 (csLecRK-I.9-ECD or csI.9-ECD). A variety of deglycosylation strategies were employed to optimize the glycosylated recombinant atDORN1-ECD for crystallization. In addition, the glycosylated csI.9-ECD protein was crystallized at 291 K. X-ray diffraction data were collected at 4.6 Å resolution from a single crystal. The crystal belonged to space groupC222 orC2221, with unit-cell parametersa= 94.7,b= 191.5,c= 302.8 Å. These preliminary studies have laid the foundation for structural determination of the DORN1 and I.9 receptor proteins, which will lead to a better understanding of the perception and function of extracellular ATP in plants.

  11. Crystal structure of the PAC1R extracellular domain unifies a consensus fold for hormone recognition by class B G-protein coupled receptors.

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    Shiva Kumar

    Full Text Available Pituitary adenylate cyclase activating polypeptide (PACAP is a member of the PACAP/glucagon family of peptide hormones, which controls many physiological functions in the immune, nervous, endocrine, and muscular systems. It activates adenylate cyclase by binding to its receptor, PAC1R, a member of class B G-protein coupled receptors (GPCR. Crystal structures of a number of Class B GPCR extracellular domains (ECD bound to their respective peptide hormones have revealed a consensus mechanism of hormone binding. However, the mechanism of how PACAP binds to its receptor remains controversial as an NMR structure of the PAC1R ECD/PACAP complex reveals a different topology of the ECD and a distinct mode of ligand recognition. Here we report a 1.9 Å crystal structure of the PAC1R ECD, which adopts the same fold as commonly observed for other members of Class B GPCR. Binding studies and cell-based assays with alanine-scanned peptides and mutated receptor support a model that PAC1R uses the same conserved fold of Class B GPCR ECD for PACAP binding, thus unifying the consensus mechanism of hormone binding for this family of receptors.

  12. Insertion of tetracysteine motifs into dopamine transporter extracellular domains.

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    Deanna M Navaroli

    Full Text Available The neuronal dopamine transporter (DAT is a major determinant of extracellular dopamine (DA levels and is the primary target for a variety of addictive and therapeutic psychoactive drugs. DAT is acutely regulated by protein kinase C (PKC activation and amphetamine exposure, both of which modulate DAT surface expression by endocytic trafficking. In order to use live imaging approaches to study DAT endocytosis, methods are needed to exclusively label the DAT surface pool. The use of membrane impermeant, sulfonated biarsenic dyes holds potential as one such approach, and requires introduction of an extracellular tetracysteine motif (tetraCys; CCPGCC to facilitate dye binding. In the current study, we took advantage of intrinsic proline-glycine (Pro-Gly dipeptides encoded in predicted DAT extracellular domains to introduce tetraCys motifs into DAT extracellular loops 2, 3, and 4. [(3H]DA uptake studies, surface biotinylation and fluorescence microscopy in PC12 cells indicate that tetraCys insertion into the DAT second extracellular loop results in a functional transporter that maintains PKC-mediated downregulation. Introduction of tetraCys into extracellular loops 3 and 4 yielded DATs with severely compromised function that failed to mature and traffic to the cell surface. This is the first demonstration of successful introduction of a tetracysteine motif into a DAT extracellular domain, and may hold promise for use of biarsenic dyes in live DAT imaging studies.

  13. The YARHG domain: an extracellular domain in search of a function.

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    Penny Coggill

    Full Text Available We have identified a new bacterial protein domain that we hypothesise binds to peptidoglycan. This domain is called the YARHG domain after the most highly conserved sequence-segment. The domain is found in the extracellular space and is likely to be composed of four alpha-helices. The domain is found associated with protein kinase domains, suggesting it is associated with signalling in some bacteria. The domain is also found associated with three different families of peptidases. The large number of different domains that are found associated with YARHG suggests that it is a useful functional module that nature has recombined multiple times.

  14. The non-small cell lung cancer EGFR extracellular domain mutation, M277E, is oncogenic and drug-sensitive

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    Yu S

    2017-09-01

    Full Text Available Su Yu,1,2 Yang Zhang,1 Yunjian Pan,1 Chao Cheng,1,3 Yihua Sun,1,3 Haiquan Chen1–4 1Department of Thoracic Surgery, Fudan University Shanghai Cancer Center, Shanghai, China; 2Cancer Research Center, Fudan University Shanghai Cancer Center, Shanghai, China; 3Department of Oncology, Shanghai Medical College, Fudan University, Shanghai, China; 4Institutes of Biomedical Sciences, Fudan University, Shanghai, China Purpose: To identify novel oncogenic mutations in non-small cell lung cancer patient specimens that lack mutations in known targetable genes (“pan-negative” patients.Methods: Comprehensive mutational analyses were performed on 1,356 lung adenocarcinoma specimens. In this cohort of patients, common lung cancer oncogenic driver mutations were detected in the epidermal growth factor receptor (EGFR kinase domain, the human epidermal growth factor receptor 2 kinase domain, as well as the KRAS, BRAF, ALK, ROS1 and RET genes. A sub-cohort of pan-negative patient specimens was assayed for mutations in the EGFR extracellular domain (ECD. Additionally, EGFR mutant NIH-3T3 stable cell lines were constructed and assessed for protein content, anchorage-independent growth, and tumor formation in xenograft models to identify oncogenic mutations. BaF3 lymphocytes were also used to test sensitivities of the mutations to tyrosine kinase inhibitors.Results: In pan-negative lung adenocarcinoma cases, a novel oncogenic EGFR ECD mutation was identified (M277E. EGFR M277E mutations encoded oncoproteins that transformed NIH-3T3 cells to grow in the absence of exogenous epidermal growth factor. Transformation was further evidenced by anchorage-independent growth and tumor formation in immunocompromised xenograft mouse models. Finally, as seen in the canonical EGFR L858R mutation, the M277E mutation conferred sensitivity to both erlotinib and cetuximab in BaF3 cell lines and to erlotinib in xenograft models.Conclusion: Here, a new EGFR driver mutation, M277E

  15. A chimeric prokaryotic-eukaryotic pentameric ligand gated ion channel reveals interactions between the extracellular and transmembrane domains shape neurosteroid modulation.

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    Ghosh, Borna; Tsao, Tzu-Wei; Czajkowski, Cynthia

    2017-10-01

    Pentameric ligand-gated ion channels (pLGICs) are the targets of several clinical and endogenous allosteric modulators including anesthetics and neurosteroids. Molecular mechanisms underlying allosteric drug modulation are poorly understood. Here, we constructed a chimeric pLGIC by fusing the extracellular domain (ECD) of the proton-activated, cation-selective bacterial channel GLIC to the transmembrane domain (TMD) of the human ρ1 chloride-selective GABA A R, and tested the hypothesis that drug actions are regulated locally in the domain that houses its binding site. The chimeric channels were proton-gated and chloride-selective demonstrating the GLIC ECD was functionally coupled to the GABAρ TMD. Channels were blocked by picrotoxin and inhibited by pentobarbital, etomidate and propofol. The point mutation, ρ TMD W328M, conferred positive modulation and direct gating by pentobarbital. The data suggest that the structural machinery mediating general anesthetic modulation resides in the TMD. Proton-activation and neurosteroid modulation of the GLIC-ρ chimeric channels, however, did not simply mimic their respective actions on GLIC and GABAρ revealing that across domain interactions between the ECD and TMD play important roles in determining their actions. Proton-induced current responses were biphasic suggesting that the chimeric channels contain an additional proton sensor. Neurosteroid modulation of the GLIC-ρ chimeric channels by the stereoisomers, 5α-THDOC and 5β-THDOC, were swapped compared to their actions on GABAρ indicating that positive versus negative neurosteroid modulation is not encoded solely in the TMD nor by neurosteroid isomer structure but is dependent on specific interdomain connections between the ECD and TMD. Our data reveal a new mechanism for shaping neurosteroid modulation. Copyright © 2017 Elsevier Ltd. All rights reserved.

  16. Measurement of cetuximab and panitumumab-unbound serum EGFR extracellular domain using an assay based on slow off-rate modified aptamer (SOMAmer reagents.

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    Noh Jin Park

    Full Text Available Response to cetuximab (Erbitux® and panitumumab (Vectibix® varies among individuals, and even those who show response ultimately gain drug resistance. One possible etiologic factor is differential interaction between the drug and target. We describe the development of an assay based on Slow Off-rate Modified Aptamer (SOMAmer(™ reagents that can distinguish drug-bound from unbound epidermal growth factor receptor (EGFR.This quantitative assay uses a SOMAmer reagent specific for EGFR extracellular domain (ECD as a capturing reagent. Captured SOMAmer is quantitated using PCR. Linearity and accuracy (recovery of the assay were assessed using normal sera and purified EGFR ECD.This EGFR ECD assay showed linearity between 2.5 and 600 ng/mL. Average recovery was 101%. The assay detected EGFR but showed little cross-reactivity to other ErbB proteins: 0.4% for ErbB2, 6.9% for ErbB3, and 1.3% for ErbB4. Preincubation of normal serum with either cetuximab or panitumumab resulted in a dose-dependent decrease in EGFR ECD levels measured using the SOMAmer assay; preincubation did not affect measurement with an ELISA.This SOMAmer-based serum EGFR ECD assay accurately and specifically measures EGFR in serum. Detection of significant amounts of drug-unbound EGFR in patients undergoing cetuximab or panitumumab treatment could be an indicator of poor drug response. Further studies are needed to evaluate the utility of the assay as an indicator of drug efficacy or as a guide to dosing.

  17. On the function of chitin synthase extracellular domains in biomineralization.

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    Weiss, Ingrid M; Lüke, Florian; Eichner, Norbert; Guth, Christina; Clausen-Schaumann, Hauke

    2013-08-01

    Molluscs with various shell architectures evolved around 542-525 million years ago, as part of a larger phenomenon related to the diversification of metazoan phyla. Molluscs deposit minerals in a chitin matrix. The mollusc chitin is synthesized by transmembrane enzymes that contain several unique extracellular domains. Here we investigate the assembly mechanism of the chitin synthase Ar-CS1 via its extracellular domain ArCS1_E22. The corresponding transmembrane protein ArCS1_E22TM accumulates in membrane fractions of the expression host Dictyostelium discoideum. Soluble recombinant ArCS1_E22 proteins can be purified as monomers only at basic pH. According to confocal fluorescence microscopy experiments, immunolabeled ArCS1_E22 proteins adsorb preferably to aragonitic nacre platelets at pH 7.75. At pH 8.2 or pH 9.0 the fluorescence signal is less intense, indicating that protein-mineral interaction is reduced with increasing pH. Furthermore, ArCS1_E22 forms regular nanostructures on cationic substrates as revealed by atomic force microscopy (AFM) experiments on modified mica cleavage planes. These experiments suggest that the extracellular domain ArCS1_E22 is involved in regulating the multiple enzyme activities of Ar-CS1 such as chitin synthesis and myosin movements by interaction with mineral surfaces and eventually by protein assembly. The protein complexes could locally probe the status of mineralization according to pH unless ions and pCO2 are balanced with suitable buffer substances. Taking into account that the intact enzyme could act as a force sensor, the results presented here provide further evidence that shell formation is coordinated physiologically with precise adjustment of cellular activities to the structure, topography and stiffness at the mineralizing interface. Copyright © 2013 Elsevier Inc. All rights reserved.

  18. Domain organizations of modular extracellular matrix proteins and their evolution.

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    Engel, J

    1996-11-01

    Multidomain proteins which are composed of modular units are a rather recent invention of evolution. Domains are defined as autonomously folding regions of a protein, and many of them are similar in sequence and structure, indicating common ancestry. Their modular nature is emphasized by frequent repetitions in identical or in different proteins and by a large number of different combinations with other domains. The extracellular matrix is perhaps the largest biological system composed of modular mosaic proteins, and its astonishing complexity and diversity are based on them. A cluster of minireviews on modular proteins is being published in Matrix Biology. These deal with the evolution of modular proteins, the three-dimensional structure of domains and the ways in which these interact in a multidomain protein. They discuss structure-function relationships in calcium binding domains, collagen helices, alpha-helical coiled-coil domains and C-lectins. The present minireview is focused on some general aspects and serves as an introduction to the cluster.

  19. Structural elucidation of transmembrane domain zero (TMD0) of EcdL: A multidrug resistance-associated protein (MRP) family of ATP-binding cassette transporter protein revealed by atomistic simulation.

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    Bera, Krishnendu; Rani, Priyanka; Kishor, Gaurav; Agarwal, Shikha; Kumar, Antresh; Singh, Durg Vijay

    2017-09-20

    ATP-Binding cassette (ABC) transporters play an extensive role in the translocation of diverse sets of biologically important molecules across membrane. EchnocandinB (antifungal) and EcdL protein of Aspergillus rugulosus are encoded by the same cluster of genes. Co-expression of EcdL and echinocandinB reflects tightly linked biological functions. EcdL belongs to Multidrug Resistance associated Protein (MRP) subfamily of ABC transporters with an extra transmembrane domain zero (TMD0). Complete structure of MRP subfamily comprising of TMD0 domain, at atomic resolution is not known. We hypothesized that the transportation of echonocandinB is mediated via EcdL protein. Henceforth, it is pertinent to know the topological arrangement of TMD0, with other domains of protein and its possible role in transportation of echinocandinB. Absence of effective template for TMD0 domain lead us to model by I-TASSER, further structure has been refined by multiple template modelling using homologous templates of remaining domains (TMD1, NBD1, TMD2, NBD2). The modelled structure has been validated for packing, folding and stereochemical properties. MD simulation for 0.1 μs has been carried out in the biphasic environment for refinement of modelled protein. Non-redundant structures have been excavated by clustering of MD trajectory. The structural alignment of modelled structure has shown Z-score -37.9; 31.6, 31.5 with RMSD; 2.4, 4.2, 4.8 with ABC transporters; PDB ID 4F4C, 4M1 M, 4M2T, respectively, reflecting the correctness of structure. EchinocandinB has been docked to the modelled as well as to the clustered structures, which reveals interaction of echinocandinB with TMD0 and other TM helices in the translocation path build of TMDs.

  20. The minimal essential unit for cadherin-mediated intercellular adhesion comprises extracellular domains 1 and 2

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    Shan, Weisong; Yagita, Yoshiki; Wang, Zhaohui

    2004-01-01

    of the extracellular domains of N-cadherin and produced various cell lines to examine adhesion properties. We show that the first domain of N-cadherin alone on the cell surface fails to generate adhesive activity and that the first two domains of N-cadherin form the "minimal essential unit" to mediate cell adhesion...... domains of N-cadherin have distinct roles in cell adhesion, i.e. the first two domains are responsible for homophilic adhesion activity, and the other domains promote adhesion efficiency most likely by positioning essential domains relatively far out from the cell surface....

  1. Recombinant expression in E. coli of human FGFR2 with its transmembrane and extracellular domains

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    Adam Bajinting

    2017-06-01

    Full Text Available Fibroblast growth factor receptors (FGFRs are a family of receptor tyrosine kinases containing three domains: an extracellular receptor domain, a single transmembrane helix, and an intracellular tyrosine kinase domain. FGFRs are activated by fibroblast growth factors (FGFs as part of complex signal transduction cascades regulating angiogenesis, skeletal formation, cell differentiation, proliferation, cell survival, and cancer. We have developed the first recombinant expression system in E. coli to produce a construct of human FGFR2 containing its transmembrane and extracellular receptor domains. We demonstrate that the expressed construct is functional in binding heparin and dimerizing. Size exclusion chromatography demonstrates that the purified FGFR2 does not form a complex with FGF1 or adopts an inactive dimer conformation. Progress towards the successful recombinant production of intact FGFRs will facilitate further biochemical experiments and structure determination that will provide insight into how extracellular FGF binding activates intracellular kinase activity.

  2. Sequestration of latent TGF-β binding protein 1 into CADASIL-related Notch3-ECD deposits.

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    Kast, Jessica; Hanecker, Patrizia; Beaufort, Nathalie; Giese, Armin; Joutel, Anne; Dichgans, Martin; Opherk, Christian; Haffner, Christof

    2014-08-13

    Cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL) represents the most common hereditary form of cerebral small vessel disease characterized by early-onset stroke and premature dementia. It is caused by mutations in the transmembrane receptor Notch3, which promote the aggregation and accumulation of the Notch3 extracellular domain (Notch3-ECD) within blood vessel walls. This process is believed to mediate the abnormal recruitment and dysregulation of additional factors including extracellular matrix (ECM) proteins resulting in brain vessel dysfunction. Based on recent evidence indicating a role for the transforming growth factor-β (TGF-β) pathway in sporadic and familial small vessel disease we studied fibronectin, fibrillin-1 and latent TGF-β binding protein 1 (LTBP-1), three ECM constituents involved in the regulation of TGF-β bioavailability, in post-mortem brain tissue from CADASIL patients and control subjects. Fibronectin and fibrillin-1 were found to be enriched in CADASIL vessels without co-localizing with Notch3-ECD deposits, likely as a result of fibrotic processes secondary to aggregate formation. In contrast, LTBP-1 showed both an accumulation and a striking co-localization with Notch3-ECD deposits suggesting specific recruitment into aggregates. We also detected increased levels of the TGF-β prodomain (also known as latency-associated peptide, LAP) indicating dysregulation of the TGF-β pathway in CADASIL development. In vitro analyses revealed a direct interaction between LTBP-1 and Notch3-ECD and demonstrated a specific co-aggregation of LTBP-1 with mutant Notch3. We propose LTBP-1 as a novel component of Notch3-ECD deposits and suggest its involvement in pathological processes triggered by Notch3-ECD aggregation.

  3. DC-SIGN neck domain is a pH-sensor controlling oligomerization: SAXS and hydrodynamic studies of extracellular domain.

    Science.gov (United States)

    Tabarani, Georges; Thépaut, Michel; Stroebel, David; Ebel, Christine; Vivès, Corinne; Vachette, Patrice; Durand, Dominique; Fieschi, Franck

    2009-08-07

    DC-SIGN is a C-type lectin receptor of dendritic cells and is involved in the early stages of numerous infectious diseases. DC-SIGN is organized into a tetramer enabling multivalent interaction with pathogens. Once formed, the DC-SIGN-pathogen complex can be internalized into compartments of increasing acidity. We have studied the pH dependence of the oligomerization state and conformation of the entire extracellular domain and neck region. We present evidence for equilibrium between the monomeric and tetrameric states of the extracellular domain, which exhibits a marked dependence with respect to both pH and ionic strength. Using solution x-ray scattering we have obtained a molecular envelope of the extracellular domain in which a model has been built. Our results highlight the central role of the neck domain in the pH-sensitive control of the oligomerization state, in the extended conformation of the protein, and in carbohydrate recognition domain organization and presentation. This work opens new insight into the molecular mechanism of ligand release and points to new avenues to block the first step of this important infection pathway.

  4. The extracellular domain of neurotrophin receptor p75 as a candidate biomarker for amyotrophic lateral sclerosis.

    Science.gov (United States)

    Shepheard, Stephanie R; Chataway, Tim; Schultz, David W; Rush, Robert A; Rogers, Mary-Louise

    2014-01-01

    Objective biomarkers for amyotrophic lateral sclerosis would facilitate the discovery of new treatments. The common neurotrophin receptor p75 is up regulated and the extracellular domain cleaved from injured neurons and peripheral glia in amyotrophic lateral sclerosis. We have tested the hypothesis that urinary levels of extracellular neurotrophin receptor p75 serve as a biomarker for both human motor amyotrophic lateral sclerosis and the SOD1(G93A) mouse model of the disease. The extracellular domain of neurotrophin receptor p75 was identified in the urine of amyotrophic lateral sclerosis patients by an immuno-precipitation/western blot procedure and confirmed by mass spectrometry. An ELISA was established to measure urinary extracellular neurotrophin receptor p75. The mean value for urinary extracellular neurotrophin receptor p75 from 28 amyotrophic lateral sclerosis patients measured by ELISA was 7.9±0.5 ng/mg creatinine and this was significantly higher (pneurotrophin receptor p75 was also readily detected in SOD1(G93A) mice by immuno-precipitation/western blot before the onset of clinical symptoms. These findings indicate a significant relation between urinary extracellular neurotrophin receptor p75 levels and disease progression and suggests that it may be a useful marker of disease activity and progression in amyotrophic lateral sclerosis.

  5. Characterization of the protease activity that cleaves the extracellular domain of β-dystroglycan

    International Nuclear Information System (INIS)

    Zhong Di; Saito, Fumiaki; Saito, Yuko; Nakamura, Ayami; Shimizu, Teruo; Matsumura, Kiichiro

    2006-01-01

    Dystroglycan (DG) complex, composed of αDG and βDG, provides a link between the extracellular matrix (ECM) and cortical cytoskeleton. Although the proteolytic processing of βDG was reported in various physiological and pathological conditions, its exact mechanism remains unknown. In this study, we addressed this issue using the cell culture system of rat schwannoma cell line RT4. We found that the culture medium of RT4 cells was enriched with the protease activity that degrades the fusion protein construct of the extracellular domain of βDG specifically. This activity was suppressed by the inhibitor of matrix metalloproteinase-2 (MMP-2) and MMP-9, but not by the inhibitors of MMP-1, MMP-3, MMP-8, and MMP-13. Zymography and RT-PCR analysis showed that RT4 cells secreted MMP-2 and MMP-9 into the culture medium. Finally, active MMP-2 and MMP-9 enzymes degraded the fusion protein construct of the extracellular domain of βDG. These results indicate (1) that RT4 cells secrete the protease activity that degrades the extracellular domain of βDG specifically and (2) that MMP-2 and MMP-9 may be involved in this process

  6. A dual role of the extracellular domain of Drosophila Crumbs for morphogenesis of the embryonic neuroectoderm

    Directory of Open Access Journals (Sweden)

    Shradha Das

    2018-01-01

    Full Text Available Epithelia are highly polarised tissues and several highly conserved polarity protein complexes serve to establish and maintain polarity. The transmembrane protein Crumbs (Crb, the central component of the Crb protein complex, is required, among others, for the maintenance of polarity in most epithelia in the Drosophila embryo. However, different epithelia exhibit different phenotypic severity upon loss of crb. Using a transgenomic approach allowed us to more accurately define the role of crb in different epithelia. In particular, we provide evidence that the loss of epithelial tissue integrity in the ventral epidermis of crb mutant embryos is due to impaired actomyosin activity and an excess number of neuroblasts. We demonstrate that the intracellular domain of Crb could only partially rescue this phenotype, while it is able to completely restore tissue integrity in other epithelia. Based on these results we suggest a dual role of the extracellular domain of Crb in the ventral neuroectoderm. First, it is required for apical enrichment of the Crb protein, which in turn regulates actomyosin activity and thereby ensures tissue integrity; and second, the extracellular domain of Crb stabilises the Notch receptor and thereby ensures proper Notch signalling and specification of the correct number of neuroblasts.

  7. Thyrotropin-luteinizing hormone/chorionic gonadotropin receptor extracellular domain chimeras as probes for thyrotropin receptor function

    International Nuclear Information System (INIS)

    Nagayama, Yuji; Wadsworth, H.L.; Chazenbalk, G.D.; Russo, D.; Seto, Pui; Rapoport, B.

    1991-01-01

    To define the sites in the extracellular domain of the human thyrotropin (TSH) receptor that are involved in TSH binding and signal transduction the authors constructed chimeric thyrotropin-luteinizing hormone/chorionic gonadotropin (TSH-LH/CG) receptors. The extracellular domain of the human TSH receptor was divided into five regions that were replaced, either singly or in various combinations, with homologous regions of the rat LH/CG receptor. The chimeric receptors were stably expressed in Chinese hamster ovary cells. The data obtained suggest that the carboxyl region of the extracellular domain (amino acid residues 261-418) and particularly the middle region (residues 171-260) play a role in signal transduction. The possibility is also raised of an interaction between the amino and carboxyl regions of the extracellular domain in the process of signal transduction. In summary, these studies suggest that the middle region and carboxyl half of the extracellular domain of the TSH receptor are involved in signal transduction and that the TSH-binding region is likely to span the entire extracellular domain, with multiple discontinuous contact sites

  8. Epidermal growth factor receptor activation in glioblastoma through novel missense mutations in the extracellular domain.

    Directory of Open Access Journals (Sweden)

    Jeffrey C Lee

    2006-12-01

    Full Text Available Protein tyrosine kinases are important regulators of cellular homeostasis with tightly controlled catalytic activity. Mutations in kinase-encoding genes can relieve the autoinhibitory constraints on kinase activity, can promote malignant transformation, and appear to be a major determinant of response to kinase inhibitor therapy. Missense mutations in the EGFR kinase domain, for example, have recently been identified in patients who showed clinical responses to EGFR kinase inhibitor therapy.Encouraged by the promising clinical activity of epidermal growth factor receptor (EGFR kinase inhibitors in treating glioblastoma in humans, we have sequenced the complete EGFR coding sequence in glioma tumor samples and cell lines. We identified novel missense mutations in the extracellular domain of EGFR in 13.6% (18/132 of glioblastomas and 12.5% (1/8 of glioblastoma cell lines. These EGFR mutations were associated with increased EGFR gene dosage and conferred anchorage-independent growth and tumorigenicity to NIH-3T3 cells. Cells transformed by expression of these EGFR mutants were sensitive to small-molecule EGFR kinase inhibitors.Our results suggest extracellular missense mutations as a novel mechanism for oncogenic EGFR activation and may help identify patients who can benefit from EGFR kinase inhibitors for treatment of glioblastoma.

  9. ECDS - a Swedish Research Infrastructure for the Open Sharing of Environment and Climate Data

    Directory of Open Access Journals (Sweden)

    T Klein

    2013-02-01

    Full Text Available Environment Climate Data Sweden (ECDS is a new Swedish research infrastructure, furthering the reuse of scientific data in the domains of environment and climate. ECDS consists of a technical infrastructure and a service organization, supporting the management, exchange, and re-use of scientific data. The technical components of ECDS include a portal and an underlying data catalogue with information on datasets. The datasets are described using a metadata profile compliant with international standards. The datasets accessible through ECDS can be hosted by universities, institutes, or research groups or at the new Swedish federated data storage facility Swestore of the Swedish National Infrastructure for Computing (SNIC.

  10. The soluble extracellular domain of E-cadherin interferes with EPEC adherence via interaction with the Tir:intimin complex.

    Science.gov (United States)

    Login, Frédéric H; Jensen, Helene H; Pedersen, Gitte A; Amieva, Manuel R; Nejsum, Lene N

    2018-06-19

    Enteropathogenic Escherichia coli (EPEC) causes watery diarrhea when colonizing the surface of enterocytes. The translocated intimin receptor (Tir):intimin receptor complex facilitates tight adherence to epithelial cells and formation of actin pedestals beneath EPEC. We found that the host cell adherens junction protein E-cadherin (Ecad) was recruited to EPEC microcolonies. Live-cell and confocal imaging revealed that Ecad recruitment depends on, and occurs after, formation of the Tir:intimin complex. Combinatorial binding experiments using wild-type EPEC, isogenic mutants lacking Tir or intimin, and E. coli expressing intimin showed that the extracellular domain of Ecad binds the bacterial surface in a Tir:intimin-dependent manner. Finally, addition of the soluble extracellular domain of Ecad to the infection medium or depletion of Ecad extracellular domain from the cell surface reduced EPEC adhesion to host cells. Thus, the soluble extracellular domain of Ecad may be used in the design of intervention strategies targeting EPEC adherence to host cells.-Login, F. H., Jensen, H. H., Pedersen, G. A., Amieva, M. R., Nejsum, L. N. The soluble extracellular domain of E-cadherin interferes with EPEC adherence via interaction with the Tir:intimin complex.

  11. Crystal structure of a prolactin receptor antagonist bound to the extracellular domain of the prolactin receptor

    DEFF Research Database (Denmark)

    Svensson, L Anders; Bondensgaard, Kent; Nørskov-Lauritsen, Leif

    2008-01-01

    The crystal structure of the complex between an N-terminally truncated G129R human prolactin (PRL) variant and the extracellular domain of the human prolactin receptor (PRLR) was determined at 2.5A resolution by x-ray crystallography. This structure represents the first experimental structure...... studies, the structural data imply that the definition of PRL binding site 1 should be extended to include residues situated in the N-terminal part of loop 1 and in the C terminus. Comparison of the structure of the receptor-bound PRL variant with the structure reported for the unbound form of a similar...... scale rearrangements and structuring occur in the flexible N-terminal part of loop 1. Hydrogen exchange mass spectrometry data imply that the dynamics of the four-helix bundle in solution generally become stabilized upon receptor interaction at binding site 1....

  12. SAXS analysis of a soluble cytosolic NgBR construct including extracellular and transmembrane domains.

    Directory of Open Access Journals (Sweden)

    Joshua Holcomb

    Full Text Available The Nogo-B receptor (NgBR is involved in oncogenic Ras signaling through directly binding to farnesylated Ras. It recruits farnesylated Ras to the non-lipid-raft membrane for interaction with downstream effectors. However, the cytosolic domain of NgBR itself is only partially folded. The lack of several conserved secondary structural elements makes this domain unlikely to form a complete farnesyl binding pocket. We find that inclusion of the extracellular and transmembrane domains that contain additional conserved residues to the cytosolic region results in a well folded protein with a similar size and shape to the E.coli cis-isoprenyl transferase (UPPs. Small Angle X-ray Scattering (SAXS analysis reveals the radius of gyration (Rg of our NgBR construct to be 18.2 Å with a maximum particle dimension (Dmax of 61.0 Å. Ab initio shape modeling returns a globular molecular envelope with an estimated molecular weight of 23.0 kD closely correlated with the calculated molecular weight. Both Kratky plot and pair distribution function of NgBR scattering reveal a bell shaped peak which is characteristic of a single globularly folded protein. In addition, circular dichroism (CD analysis reveals that our construct has the secondary structure contents similar to the UPPs. However, this result does not agree with the currently accepted topological orientation of NgBR which might partition this construct into three separate domains. This discrepancy suggests another possible NgBR topology and lends insight into a potential molecular basis of how NgBR facilitates farnesylated Ras recruitment.

  13. Crystal structure of the extracellular domain of human myelin protein zero

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Zhigang; Wang, Yong; Yedidi, Ravikiran S.; Brunzelle, Joseph S.; Kovari, Iulia A.; Sohi, Jasloveleen; Kamholz, John; Kovari, Ladislau C. (WSU-MED); (NWU)

    2012-03-27

    Charcot-Marie-Tooth disease (CMT), a hereditary motor and sensory neuropathy, is the most common genetic neuropathy with an incidence of 1 in 2600. Several forms of CMT have been identified arising from different genomic abnormalities such as CMT1 including CMT1A, CMT1B, and CMTX. CMT1 with associated peripheral nervous system (PNS) demyelination, the most frequent diagnosis, demonstrates slowed nerve conduction velocities and segmental demyelination upon nerve biopsy. One of its subtypes, CMT1A, presents a 1.5-Mb duplication in the p11-p12 region of the human chromosome 17 which encodes peripheral myelin protein 22 (PMP22). CMT1B, a less common form, arises from the mutations in the myelin protein zero (MPZ) gene on chromosome 1, region q22-q23, which encodes the major structural component of the peripheral myelin. A rare type of CMT1 has been found recently and is caused by point mutations in early growth response gene 2 (EGR2), encoding a zinc finger transcription factor in Schwann cells. In addition, CMTX, an X-linked form of CMT, arises from a mutation in the connexin-32 gene. Myelin protein zero, associated with CMT1B, is a transmembrane protein of 219 amino acid residues. Human MPZ consists of three domains: 125 residues constitute the glycosylated immunoglobulin-like extracellular domain; 27 residues span the membrane; and 67 residues comprise the highly basic intracellular domain. MPZ makes up approximately 50% of the protein content of myelin, and is expressed predominantly in Schwann cells, the myelinating cell of the PNS. Myelin protein zero, a homophilic adhesion molecule, is a member of the immunoglobulin super-family and is essential for normal myelin structure and function. In addition, MPZ knockout mice displayed abnormal myelin that severely affects the myelination pathway, and overexpression of MPZ causes congenital hypomyelination of peripheral nerves. Myelin protein zero mutations account for {approx}5% of patients with CMT. To date, over 125

  14. Crystal Structure of the Extracellular Cholinesterase-Like Domain from Neuroligin-2

    Energy Technology Data Exchange (ETDEWEB)

    Koehnke,J.; Jin, X.; Budreck, E.; Posy, S.; Scheiffele, P.; Hnoig, B.; Shapiro, L.

    2008-01-01

    Neuroligins (NLs) are catalytically inactive members of a family of cholinesterase-like transmembrane proteins that mediate cell adhesion at neuronal synapses. Postsynaptic neuroligins engage in Ca2+-dependent transsynaptic interactions via their extracellular cholinesterase domain with presynaptic neurexins (NRXs). These interactions may be regulated by two short splice insertions (termed A and B) in the NL cholinesterase domain. Here, we present the 3.3- Angstroms crystal structure of the ectodomain from NL2 containing splice insertion A (NL2A). The overall structure of NL2A resembles that of cholinesterases, but several structural features are unique to the NL proteins. First, structural elements surrounding the esterase active-site region differ significantly between active esterases and NL2A. On the opposite surface of the NL2A molecule, the positions of the A and B splice insertions identify a candidate NRX interaction site of the NL protein. Finally, sequence comparisons of NL isoforms allow for mapping the location of residues of previously identified mutations in NL3 and NL4 found in patients with autism spectrum disorders. Overall, the NL2 structure promises to provide a valuable model for dissecting NL isoform- and synapse-specific functions.

  15. The study on mutations of the gene of extracellular domain of human thyrotropin receptor in the patients with thyroid diseases

    International Nuclear Information System (INIS)

    Zhang Zuncheng; Fang Peihua; Tan Jian; Lu Mei

    2002-01-01

    Objective: To define the sequence of the gene of extracellular domain of normal human thyrotropin receptor (hTSHR) and to investigate the mutations of the gene in the patients with thyroid diseases. Methods: Total RNAs were extracted from the thyroid tissue of four normal controls, twelve Graves' disease, four Hashimoto's thyroiditis and eleven nodular goiter patients. The extracellular domain of hTSHR genes were amplified by reverse transcription-polymerase chain reaction (RT-PCR) and sequenced with CEQ 2000 Genetic Analyzer. Results: The normal controls and the patients with thyroid disease had the same gene sequences of the extracellular domain of hTSHR. No mutation was found, except a silent base exchange in exon 7 (Asn187) at 661 base, in which 20 samples were 'T', 11 samples were 'C', without changes of amino acid of the TSHR. Conclusions: This study has not revealed mutations in the gene of extracellular domain of hTSHR. Other molecular pathogenetic mechanisms may be involved and more research is demanded

  16. Glucagon-like peptide-1 receptor ligand interactions: structural cross talk between ligands and the extracellular domain.

    Directory of Open Access Journals (Sweden)

    Graham M West

    Full Text Available Activation of the glucagon-like peptide-1 receptor (GLP-1R in pancreatic β-cells potentiates insulin production and is a current therapeutic target for the treatment of type 2 diabetes mellitus (T2DM. Like other class B G protein-coupled receptors (GPCRs, the GLP-1R contains an N-terminal extracellular ligand binding domain. N-terminal truncations on the peptide agonist generate antagonists capable of binding to the extracellular domain, but not capable of activating full length receptor. The main objective of this study was to use Hydrogen/deuterium exchange (HDX to identify how the amide hydrogen bonding network of peptide ligands and the extracellular domain of GLP-1R (nGLP-1R were altered by binding interactions and to then use this platform to validate direct binding events for putative GLP-1R small molecule ligands. The HDX studies presented here for two glucagon-like peptide-1 receptor (GLP-1R peptide ligands indicates that the antagonist exendin-4[9-39] is significantly destabilized in the presence of nonionic detergents as compared to the agonist exendin-4. Furthermore, HDX can detect stabilization of exendin-4 and exendin-4[9-39] hydrogen bonding networks at the N-terminal helix [Val19 to Lys27] upon binding to the N-terminal extracellular domain of GLP-1R (nGLP-1R. In addition we show hydrogen bonding network stabilization on nGLP-1R in response to ligand binding, and validate direct binding events with the extracellular domain of the receptor for putative GLP-1R small molecule ligands.

  17. Solution structure and dynamics of C-terminal regulatory domain of Vibrio vulnificus extracellular metalloprotease

    Energy Technology Data Exchange (ETDEWEB)

    Yun, Ji-Hye; Kim, Heeyoun [Department of Biochemistry, College of Life Science and Biotechnology, Yonsei University, Seoul 120-749 (Korea, Republic of); Park, Jung Eun [Department of Biotechnology, College of Natural Sciences, Chosun University, Gwangju 501-759 (Korea, Republic of); Lee, Jung Sup, E-mail: jsplee@mail.chosun.ac.kr [Department of Biotechnology, College of Natural Sciences, Chosun University, Gwangju 501-759 (Korea, Republic of); Lee, Weontae, E-mail: wlee@spin.yonsei.ac.kr [Department of Biochemistry, College of Life Science and Biotechnology, Yonsei University, Seoul 120-749 (Korea, Republic of)

    2013-01-11

    Highlights: Black-Right-Pointing-Pointer We have determined solution structures of vEP C-terminal regulatory domain. Black-Right-Pointing-Pointer vEP C-ter100 has a compact {beta}-barrel structure with eight anti-parallel {beta}-strands. Black-Right-Pointing-Pointer Solution structure of vEP C-ter100 shares its molecular topology with that of the collagen-binding domain of collagenase. Black-Right-Pointing-Pointer Residues in the {beta}3 region of vEP C-ter100 might be important in putative ligand/receptor binding. Black-Right-Pointing-Pointer vEP C-ter100 interacts strongly with iron ion. -- Abstract: An extracellular metalloprotease (vEP) secreted by Vibrio vulnificus ATCC29307 is a 45-kDa proteolytic enzyme that has prothrombin activation and fibrinolytic activities during bacterial infection. The action of vEP could result in clotting that could serve to protect the bacteria from the host defense machinery. Very recently, we showed that the C-terminal propeptide (C-ter100), which is unique to vEP, is involved in regulation of vEP activity. To understand the structural basis of this function of vEP C-ter100, we have determined the solution structure and backbone dynamics using multidimensional nuclear magnetic resonance spectroscopy. The solution structure shows that vEP C-ter100 is composed of eight anti-parallel {beta}-strands with a unique fold that has a compact {beta}-barrel formation which stabilized by hydrophobic and hydrogen bonding networks. Protein dynamics shows that the overall structure, including loops, is very rigid and stabilized. By structural database analysis, we found that vEP C-ter100 shares its topology with that of the collagen-binding domain of collagenase, despite low sequence homology between the two domains. Fluorescence assay reveals that vEP C-ter100 interacts strongly with iron (Fe{sup 3+}). These findings suggest that vEP protease might recruit substrate molecules, such as collagen, by binding at C-ter100 and that vEP participates

  18. Crystal structure of the ligand-bound glucagon-like peptide-1 receptor extracellular domain.

    Science.gov (United States)

    Runge, Steffen; Thøgersen, Henning; Madsen, Kjeld; Lau, Jesper; Rudolph, Rainer

    2008-04-25

    The glucagon-like peptide-1 receptor (GLP-1R) belongs to Family B1 of the seven-transmembrane G protein-coupled receptors, and its natural agonist ligand is the peptide hormone glucagon-like peptide-1 (GLP-1). GLP-1 is involved in glucose homeostasis, and activation of GLP-1R in the plasma membrane of pancreatic beta-cells potentiates glucose-dependent insulin secretion. The N-terminal extracellular domain (nGLP-1R) is an important ligand binding domain that binds GLP-1 and the homologous peptide Exendin-4 with differential affinity. Exendin-4 has a C-terminal extension of nine amino acid residues known as the "Trp cage", which is absent in GLP-1. The Trp cage was believed to interact with nGLP-1R and thereby explain the superior affinity of Exendin-4. However, the molecular details that govern ligand binding and specificity of nGLP-1R remain undefined. Here we report the crystal structure of human nGLP-1R in complex with the antagonist Exendin-4(9-39) solved by the multiwavelength anomalous dispersion method to 2.2A resolution. The structure reveals that Exendin-4(9-39) is an amphipathic alpha-helix forming both hydrophobic and hydrophilic interactions with nGLP-1R. The Trp cage of Exendin-4 is not involved in binding to nGLP-1R. The hydrophobic binding site of nGLP-1R is defined by discontinuous segments including primarily a well defined alpha-helix in the N terminus of nGLP-1R and a loop between two antiparallel beta-strands. The structure provides for the first time detailed molecular insight into ligand binding of the human GLP-1 receptor, an established target for treatment of type 2 diabetes.

  19. Peptide ligands for targeting the extracellular domain of EGFR: Comparison between linear and cyclic peptides.

    Science.gov (United States)

    Williams, Tyrslai M; Sable, Rushikesh; Singh, Sitanshu; Vicente, Maria Graca H; Jois, Seetharama D

    2018-02-01

    Colorectal cancer (CRC) is the third most common solid internal malignancy among cancers. Early detection of cancer is key to increasing the survival rate of colorectal cancer patients. Overexpression of the EGFR protein is associated with CRC. We have designed a series of peptides that are highly specific for the extracellular domain of EGFR, based on our earlier studies on linear peptides. The previously reported linear peptide LARLLT, known to bind to EGFR, was modified with the goals of increasing its stability and its specificity toward EGFR. Peptide modifications, including D-amino acid substitution, cyclization, and chain reversal, were investigated. In addition, to facilitate labeling of the peptide with a fluorescent dye, an additional lysine residue was introduced onto the linear (KLARLLT) and cyclic peptides cyclo(KLARLLT) (Cyclo.L1). The lysine residue was also converted into an azide group in both a linear and reversed cyclic peptide sequences cyclo(K(N3)larllt) (Cyclo.L1.1) to allow for subsequent "click" conjugation. The cyclic peptides showed enhanced binding to EGFR by SPR. NMR and molecular modeling studies suggest that the peptides acquire a β-turn structure in solution. In vitro stability studies in human serum show that the cyclic peptide is more stable than the linear peptide. © 2017 John Wiley & Sons A/S.

  20. The MUC1 extracellular domain subunit is found in nuclear speckles and associates with spliceosomes.

    Directory of Open Access Journals (Sweden)

    Priyadarsini Kumar

    Full Text Available MUC1 is a large transmembrane glycoprotein and oncogene expressed by epithelial cells and overexpressed and underglycosylated in cancer cells. The MUC1 cytoplasmic subunit (MUC1-C can translocate to the nucleus and regulate gene expression. It is frequently assumed that the MUC1 extracellular subunit (MUC1-N does not enter the nucleus. Based on an unexpected observation that MUC1 extracellular domain antibody produced an apparently nucleus-associated staining pattern in trophoblasts, we have tested the hypothesis that MUC1-N is expressed inside the nucleus. Three different antibodies were used to identify MUC1-N in normal epithelial cells and tissues as well as in several cancer cell lines. The results of immunofluorescence and confocal microscopy analyses as well as subcellular fractionation, Western blotting, and siRNA/shRNA studies, confirm that MUC1-N is found within nuclei of all cell types examined. More detailed examination of its intranuclear distribution using a proximity ligation assay, subcellular fractionation, and immunoprecipitation suggests that MUC1-N is located in nuclear speckles (interchromatin granule clusters and closely associates with the spliceosome protein U2AF65. Nuclear localization of MUC1-N was abolished when cells were treated with RNase A and nuclear localization was altered when cells were incubated with the transcription inhibitor 5,6-dichloro-1-b-d-ribofuranosylbenzimidazole (DRB. While MUC1-N predominantly associated with speckles, MUC1-C was present in the nuclear matrix, nucleoli, and the nuclear periphery. In some nuclei, confocal microscopic analysis suggest that MUC1-C staining is located close to, but only partially overlaps, MUC1-N in speckles. However, only MUC1-N was found in isolated speckles by Western blotting. Also, MUC1-C and MUC1-N distributed differently during mitosis. These results suggest that MUC1-N translocates to the nucleus where it is expressed in nuclear speckles and that MUC1-N and MUC

  1. Structure of the LDL receptor extracellular domain at endosomalpH

    Energy Technology Data Exchange (ETDEWEB)

    Rudenko, Gabby; Henry, Lisa; Henderson, Keith; Ichtchenko,Konstantin; Brown, Michael S.; Goldstein, Joseph L.; Deisenhofer, Johann

    2002-09-05

    The structure of the low-density lipoprotein receptor extracellular portion has been determined. The document proposes a mechanism for the release of lipoprotein in the endosome. Without this release, the mechanism of receptor recycling cannot function.

  2. Role of fibronectin in collagen deposition: Fab' to the gelatin-binding domain of fibronectin inhibits both fibronectin and collagen organization in fibroblast extracellular matrix

    OpenAIRE

    1982-01-01

    We report the effect of Fab' (anti-60k) to a 60,000 mol wt gelatin binding domain of fibronectin (1981, J. Biol. Chem. 256:5583) on diploid fibroblast (IMR-90) extracellular fibronectin and collagen organization. Anti-60k Fab' did not inhibit IMR-90 attachment or proliferation in fibronectin-depleted medium. Fibroblasts cultured with preimmune Fab' deposited a dense extracellular network of fibronectin and collagen detectable by immunofluorescence, while anti-60k Fab' prevented extracellular ...

  3. Evidence of positive selection at codon sites localized in extracellular domains of mammalian CC motif chemokine receptor proteins

    Directory of Open Access Journals (Sweden)

    Metzger Kelsey J

    2010-05-01

    Full Text Available Abstract Background CC chemokine receptor proteins (CCR1 through CCR10 are seven-transmembrane G-protein coupled receptors whose signaling pathways are known for their important roles coordinating immune system responses through targeted trafficking of white blood cells. In addition, some of these receptors have been identified as fusion proteins for viral pathogens: for example, HIV-1 strains utilize CCR5, CCR2 and CCR3 proteins to obtain cellular entry in humans. The extracellular domains of these receptor proteins are involved in ligand-binding specificity as well as pathogen recognition interactions. In mammals, the majority of chemokine receptor genes are clustered together; in humans, seven of the ten genes are clustered in the 3p21-24 chromosome region. Gene conversion events, or exchange of DNA sequence between genes, have been reported in chemokine receptor paralogs in various mammalian lineages, especially between the cytogenetically closely located pairs CCR2/5 and CCR1/3. Datasets of mammalian orthologs for each gene were analyzed separately to minimize the potential confounding impact of analyzing highly similar sequences resulting from gene conversion events. Molecular evolution approaches and the software package Phylogenetic Analyses by Maximum Likelihood (PAML were utilized to investigate the signature of selection that has acted on the mammalian CC chemokine receptor (CCR gene family. The results of neutral vs. adaptive evolution (positive selection hypothesis testing using Site Models are reported. In general, positive selection is defined by a ratio of nonsynonymous/synonymous nucleotide changes (dN/dS, or ω >1. Results Of the ten mammalian CC motif chemokine receptor sequence datasets analyzed, only CCR2 and CCR3 contain amino acid codon sites that exhibit evidence of positive selection using site based hypothesis testing in PAML. Nineteen of the twenty codon sites putatively indentified as likely to be under positive

  4. Molecular mechanism of Zn2+ agonism in the extracellular domain of GPR39

    DEFF Research Database (Denmark)

    Storjohann, Laura; Holst, Birgitte; Schwartz, Thue W

    2008-01-01

    -terminal segment. Surprisingly, substitution of Asp313 located in extracellular loop 3 greatly increased ligand-independent signaling and apparently eliminated Zn2+-induced activation. It is proposed that Zn2+ acts as an agonist for GPR39, not in the classical manner by directly stabilizing an active conformation...

  5. Differential recognition of terminal extracellular Plasmodium falciparum VAR2CSA domains by sera from multigravid, malaria-exposed Malian women.

    Science.gov (United States)

    Travassos, Mark A; Coulibaly, Drissa; Bailey, Jason A; Niangaly, Amadou; Adams, Matthew; Nyunt, Myaing M; Ouattara, Amed; Lyke, Kirsten E; Laurens, Matthew B; Pablo, Jozelyn; Jasinskas, Algis; Nakajima, Rie; Berry, Andrea A; Takala-Harrison, Shannon; Kone, Abdoulaye K; Kouriba, Bourema; Rowe, J Alexandra; Doumbo, Ogobara K; Thera, Mahamadou A; Laufer, Miriam K; Felgner, Philip L; Plowe, Christopher V

    2015-06-01

    The Plasmodium falciparum erythrocyte membrane protein 1 (PfEMP1) family mediates parasite sequestration in small capillaries through tissue-specific cytoadherence. The best characterized of these proteins is VAR2CSA, which is expressed on the surface of infected erythrocytes that bind to chondroitin sulfate in the placental matrix. Antibodies to VAR2CSA prevent placental cytoadherence and protect against placental malaria. The size and complexity of the VAR2CSA protein pose challenges for vaccine development, but smaller constitutive domains may be suitable for subunit vaccine development. A protein microarray was printed to include five overlapping fragments of the 3D7 VAR2CSA extracellular region. Malian women with a history of at least one pregnancy had antibody recognition of four of these fragments and had stronger reactivity against the two distal fragments than did nulliparous women, children, and men from Mali, suggesting that the C-terminal extracellular VAR2CSA domains are a potential focus of protective immunity. With carefully chosen sera from longitudinal studies of pregnant women, this approach has the potential to identify seroreactive VAR2CSA domains associated with protective immunity against pregnancy-associated malaria. © The American Society of Tropical Medicine and Hygiene.

  6. A genomic point mutation in the extracellular domain of the thyrotropin receptor in patients with Graves` ophthalmopathy

    Energy Technology Data Exchange (ETDEWEB)

    Bahn, R.S.; Dutton, C.M.; Heufelder, A.E.; Sarkar, G. [Mayo Clinic/Foundation, Rochester, MN (United States)]|[Ludwig-Maximilians-Universitat, Munich (Germany)

    1994-02-01

    Orbital and pretibial fibroblasts are targets of autoimmune attack in Graves` ophthalmopathy (GO) and pretibial dermopathy (PTD). The fibroblast autoantigen involved in these peripheral manifestations of Graves` disease and the reason for the association of GO and PTD with hyperthyroidism are unknown. RNA encoding the full-length extracellular domain of the TSH receptor has been demonstrated in orbital and dermal fibroblasts from patients with GO and normal subjects, suggesting a possible antigenic link between fibroblasts and thyrocytes. RNA was isolated from cultured orbital, pretibial, and abdominal fibroblasts obtained from patients with severe GO (n = 22) and normal subjects (n = 5). RNA was reverse transcribed, and the resulting cDNA was amplified by the polymerase chain reaction, using primers spanning overlapping regions of the entire extracellular domain of the TSH receptor. Nucleotide sequence analysis showed an A for C substitution in the first position of codon 52 in 2 of the patients, both of whom had GO, PTD, and acropachy. Genomic DNA isolated from the 2 affected patients, and not from an additional 12 normal subjects, revealed the codon 52 mutation by direct sequencing and AciI restriction enzyme digestions. In conclusion, the authors have demonstrated the presence of a genomic point mutation, leading to a threonine for proline amino acid shift in the predicted peptide, in the extracellular domain of the TSH receptor in two patients with severe GO, PTD, acropachy, and high thyroid-stimulating immunoglobulin levels. RNA encoding this mutant product was demonstrated in the fibroblasts of these patients. They suggest that the TSH receptor may be an important fibroblast autoantigen in GO and PTD, and that this mutant form of the receptor may have unique immunogenic properties. 28 refs., 3 figs., 2 tabs.

  7. Crystal Structure of Glucagon-like Peptide-1 in Complex with the Extracellular Domain of the Glucagon-like Peptide-1 Receptor*

    Science.gov (United States)

    Underwood, Christina Rye; Garibay, Patrick; Knudsen, Lotte Bjerre; Hastrup, Sven; Peters, Günther H.; Rudolph, Rainer; Reedtz-Runge, Steffen

    2010-01-01

    GLP-1 (glucagon-like peptide-1) is an incretin released from intestinal L-cells in response to food intake. Activation of the GLP-1 receptor potentiates the synthesis and release of insulin from pancreatic β-cells in a glucose-dependent manner. The GLP-1 receptor belongs to class B of the G-protein-coupled receptors, a subfamily characterized by a large N-terminal extracellular ligand binding domain. Exendin-4 and GLP-1 are 50% identical, and exendin-4 is a full agonist with similar affinity and potency for the GLP-1 receptor. We recently solved the crystal structure of the GLP-1 receptor extracellular domain in complex with the competitive antagonist exendin-4(9–39). Interestingly, the isolated extracellular domain binds exendin-4 with much higher affinity than the endogenous agonist GLP-1. Here, we have solved the crystal structure of the extracellular domain in complex with GLP-1 to 2.1 Åresolution. The structure shows that important hydrophobic ligand-receptor interactions are conserved in agonist- and antagonist-bound forms of the extracellular domain, but certain residues in the ligand-binding site adopt a GLP-1-specific conformation. GLP-1 is a kinked but continuous α-helix from Thr13 to Val33 when bound to the extracellular domain. We supplemented the crystal structure with site-directed mutagenesis to link the structural information of the isolated extracellular domain with the binding properties of the full-length receptor. The data support the existence of differences in the binding modes of GLP-1 and exendin-4 on the full-length GLP-1 receptor. PMID:19861722

  8. Crystal structure of glucagon-like peptide-1 in complex with the extracellular domain of the glucagon-like peptide-1 receptor.

    Science.gov (United States)

    Underwood, Christina Rye; Garibay, Patrick; Knudsen, Lotte Bjerre; Hastrup, Sven; Peters, Günther H; Rudolph, Rainer; Reedtz-Runge, Steffen

    2010-01-01

    GLP-1 (glucagon-like peptide-1) is an incretin released from intestinal L-cells in response to food intake. Activation of the GLP-1 receptor potentiates the synthesis and release of insulin from pancreatic beta-cells in a glucose-dependent manner. The GLP-1 receptor belongs to class B of the G-protein-coupled receptors, a subfamily characterized by a large N-terminal extracellular ligand binding domain. Exendin-4 and GLP-1 are 50% identical, and exendin-4 is a full agonist with similar affinity and potency for the GLP-1 receptor. We recently solved the crystal structure of the GLP-1 receptor extracellular domain in complex with the competitive antagonist exendin-4(9-39). Interestingly, the isolated extracellular domain binds exendin-4 with much higher affinity than the endogenous agonist GLP-1. Here, we have solved the crystal structure of the extracellular domain in complex with GLP-1 to 2.1 Aresolution. The structure shows that important hydrophobic ligand-receptor interactions are conserved in agonist- and antagonist-bound forms of the extracellular domain, but certain residues in the ligand-binding site adopt a GLP-1-specific conformation. GLP-1 is a kinked but continuous alpha-helix from Thr(13) to Val(33) when bound to the extracellular domain. We supplemented the crystal structure with site-directed mutagenesis to link the structural information of the isolated extracellular domain with the binding properties of the full-length receptor. The data support the existence of differences in the binding modes of GLP-1 and exendin-4 on the full-length GLP-1 receptor.

  9. eCD4-Ig variants that more potently neutralize HIV-1.

    Science.gov (United States)

    Fetzer, Ina; Gardner, Matthew R; Davis-Gardner, Meredith E; Prasad, Neha R; Alfant, Barnett; Weber, Jesse A; Farzan, Michael

    2018-03-28

    The HIV-1 entry inhibitor eCD4-Ig is a fusion of CD4-Ig and a coreceptor-mimetic peptide. eCD4-Ig is markedly more potent than CD4-Ig, with neutralization efficiencies approaching those of HIV-1 broadly neutralizing antibodies (bNAbs). However, unlike bNAbs, eCD4-Ig neutralizes all HIV-1, HIV-2 and SIV isolates that it has been tested against, suggesting that it may be useful in clinical settings where antibody escape is a concern. Here we characterize three new eCD4-Ig variants, each with different architectures and each utilizing D1.22, a stabilized form of CD4 domain 1. These variants were 10- to 20-fold more potent than our original eCD4-Ig variant, with a construct bearing four D1.22 domains (eD1.22-HL-Ig) exhibiting the greatest potency. However, this variant mediated less efficient antibody-dependent cell-mediated cytotoxicity (ADCC) activity than eCD4-Ig itself or several other eCD4-Ig variants, including the smallest variant (eD1.22-Ig). A variant with the same architecture as original eCD4-Ig (eD1.22-D2-Ig) showed modestly higher thermal stability and best prevented promotion of infection of CCR5-positive, CD4-negative cells. All three variants, and eCD4-Ig itself, mediated more efficient shedding of the HIV-1 envelope glycoprotein gp120 than did CD4-Ig. Finally, we show that only three D1.22 mutations contributed to the potency of eD1.22-D2-Ig, and that introduction of these changes into eCD4-Ig resulted in a variant 9-fold more potent than eCD4-Ig and 2-fold more potent than eD1.22-D2-Ig. These studies will assist in developing eCD4-Ig variants with properties optimized for prophylaxis, therapy, and cure applications. IMPORTANCE HIV-1 bNAbs have properties different from antiretroviral compounds. Specifically, antibodies can enlist immune effector cells to eliminate infected cells, whereas antiretroviral compounds simply interfere with various steps in the viral lifecycle. Unfortunately, HIV-1 is adept at evading antibody recognition, limiting the

  10. Comparative molecular dynamics study of neuromyelitis optica-immunoglobulin G binding to aquaporin-4 extracellular domains.

    Science.gov (United States)

    Alberga, Domenico; Trisciuzzi, Daniela; Lattanzi, Gianluca; Bennett, Jeffrey L; Verkman, Alan S; Mangiatordi, Giuseppe Felice; Nicolotti, Orazio

    2017-08-01

    Neuromyelitis optica (NMO) is an inflammatory demyelinating disease of the central nervous system in which most patients have serum autoantibodies (called NMO-IgG) that bind to astrocyte water channel aquaporin-4 (AQP4). A potential therapeutic strategy in NMO is to block the interaction of NMO-IgG with AQP4. Building on recent observation that some single-point and compound mutations of the AQP4 extracellular loop C prevent NMO-IgG binding, we carried out comparative Molecular Dynamics (MD) investigations on three AQP4 mutants, TP 137-138 AA, N 153 Q and V 150 G, whose 295-ns long trajectories were compared to that of wild type human AQP4. A robust conclusion of our modeling is that loop C mutations affect the conformation of neighboring extracellular loop A, thereby interfering with NMO-IgG binding. Analysis of individual mutations suggested specific hydrogen bonding and other molecular interactions involved in AQP4-IgG binding to AQP4. Copyright © 2017 Elsevier B.V. All rights reserved.

  11. Site-specific chemical conjugation of human Fas ligand extracellular domain using trans-cyclooctene - methyltetrazine reactions.

    Science.gov (United States)

    Muraki, Michiro; Hirota, Kiyonori

    2017-07-03

    Fas ligand plays a key role in the human immune system as a major cell death inducing protein. The extracellular domain of human Fas ligand (hFasLECD) triggers apoptosis of malignant cells, and therefore is expected to have substantial potentials in medical biotechnology. However, the current application of this protein to clinical medicine is hampered by a shortage of the benefits relative to the drawbacks including the side-effects in systemic administration. Effective procedures for the engineering of the protein by attaching useful additional functions are required to overcome the problem. A procedure for the site-specific chemical conjugation of hFasLECD with a fluorochrome and functional proteins was devised using an inverse-electron-demand Diels-Alder reaction between trans-cyclooctene group and methyltetrazine group. The conjugations in the present study were attained by using much less molar excess amounts of the compounds to be attached as compared with the conventional chemical modification reactions using maleimide derivatives in the previous study. The isolated conjugates of hFasLECD with sulfo-Cy3, avidin and rabbit IgG Fab' domain presented the functional and the structural integrities of the attached molecules without impairing the specific binding activity toward human Fas receptor extracellular domain. The present study provided a new fundamental strategy for the production of the engineered hFasLECDs with additional beneficial functions, which will lead to the developments of the improved diagnostic systems and the effective treatment methods of serious diseases by using this protein as a component of novel molecular tools.

  12. Novel receptor-like kinases in cacao contain PR-1 extracellular domains.

    Science.gov (United States)

    Teixeira, Paulo José Pereira Lima; Costa, Gustavo Gilson Lacerda; Fiorin, Gabriel Lorencini; Pereira, Gonçalo Amarante Guimarães; Mondego, Jorge Maurício Costa

    2013-08-01

    Members of the pathogenesis-related protein 1 (PR-1) family are well-known markers of plant defence responses, forming part of the arsenal of the secreted proteins produced on pathogen recognition. Here, we report the identification of two cacao (Theobroma cacao L.) PR-1s that are fused to transmembrane regions and serine/threonine kinase domains, in a manner characteristic of receptor-like kinases (RLKs). These proteins (TcPR-1f and TcPR-1g) were named PR-1 receptor kinases (PR-1RKs). Phylogenetic analysis of RLKs and PR-1 proteins from cacao indicated that PR-1RKs originated from a fusion between sequences encoding PR-1 and the kinase domain of a LecRLK (Lectin Receptor-Like Kinase). Retrotransposition marks surround TcPR-1f, suggesting that retrotransposition was involved in the origin of PR-1RKs. Genes with a similar domain architecture to cacao PR-1RKs were found in rice (Oryza sativa), barrel medic (Medicago truncatula) and a nonphototrophic bacterium (Herpetosiphon aurantiacus). However, their kinase domains differed from those found in LecRLKs, indicating the occurrence of convergent evolution. TcPR-1g expression was up-regulated in the biotrophic stage of witches' broom disease, suggesting a role for PR-1RKs during cacao defence responses. We hypothesize that PR-1RKs transduce a defence signal by interacting with a PR-1 ligand. © 2013 BSPP AND JOHN WILEY & SONS LTD.

  13. A chitinase with two catalytic domains is required for organization of the cuticular extracellular matrix of a beetle.

    Directory of Open Access Journals (Sweden)

    Mi Young Noh

    2018-03-01

    Full Text Available Insect cuticle or exoskeleton is an extracellular matrix formed primarily from two different structural biopolymers, chitin and protein. During each molt cycle, a new cuticle is deposited simultaneously with degradation of the inner part of the chitinous procuticle of the overlying old exoskeleton by molting fluid enzymes including epidermal chitinases. In this study we report a novel role for an epidermal endochitinase containing two catalytic domains, TcCHT7, from the red flour beetle, Tribolium castaneum, in organizing chitin in the newly forming cuticle rather than in degrading chitin present in the prior one. Recombinant TcCHT7 expressed in insect cells is membrane-bound and capable of hydrolyzing an extracellular chitin substrate, whereas in vivo, this enzyme is also released from the plasma membrane and co-localizes with chitin in the entire procuticle. RNAi of TcCHT7 reveals that this enzyme is nonessential for any type of molt or degradation of the chitinous matrix in the old cuticle. In contrast, TcCHT7 is required for maintaining the integrity of the cuticle as a compact structure of alternating electron-dense and electron-lucent laminae. There is a reduction in thickness of elytral and leg cuticles after RNAi for TcCHT7. TcCHT7 is also required for formation of properly oriented long chitin fibers inside pore canals that are vertically oriented columnar structures, which contribute to the mechanical strength of a light-weight, yet rigid, adult cuticle. The conservation of CHT7-like proteins harboring such a unique domain configuration among many insect and other arthropod species indicates a critical role for the group III class of chitinases in the higher ordered organization of chitin fibers for development of the structural integrity of many invertebrate exoskeletons.

  14. Biophysical characterization of the olfactomedin domain of myocilin, an extracellular matrix protein implicated in inherited forms of glaucoma.

    Directory of Open Access Journals (Sweden)

    Susan D Orwig

    Full Text Available Myocilin is an eye protein found in the trabecular extracellular matrix (TEM, within the anatomic region that controls fluid flow. Variants of myocilin, localized to its olfactomedin (OLF domain, have been linked to inherited forms of glaucoma, a disease associated with elevated intraocular pressure. OLF domains have also been implicated in psychiatric diseases and cancers by their involvement in signaling, neuronal growth, and development. However, molecular characterization of OLFs has been hampered by challenges in recombinant expression, a hurdle we have recently overcome for the myocilin OLF domain (myoc-OLF. Here, we report the first detailed solution biophysical characterization of myoc-OLF to gain insight into its structure and function. Myoc-OLF is stable in the presence of glycosaminoglycans, as well as in a wide pH range in buffers with functional groups reminiscent of such glycosaminoglycans. Circular dichroism (CD reveals significant β-sheet and β-turn secondary structure. Unexpectedly, the CD signature is reminiscent of α-chymotrypsin as well as another ocular protein family, the βγ-crystallins. At neutral pH, intrinsic tryptophan fluorescence and CD melts indicate a highly cooperative transition with a melting temperature of ∼55 °C. Limited proteolysis combined with mass spectrometry reveals that the compact core structural domain of OLF consists of approximately residues 238-461, which retains the single disulfide bond and is as stable as the full myoc-OLF construct. The data presented here inform new testable hypotheses for interactions with specific TEM components, and will assist in design of therapeutic agents for myocilin glaucoma.

  15. eCD4-Ig promotes ADCC activity of sera from HIV-1-infected patients.

    Directory of Open Access Journals (Sweden)

    Meredith E Davis-Gardner

    2017-12-01

    Full Text Available Antibody-dependent cell-mediated cytotoxity (ADCC can eliminate HIV-1 infected cells, and may help reduce the reservoir of latent virus in infected patients. Sera of HIV-1 positive individuals include a number of antibodies that recognize epitopes usually occluded on HIV-1 envelope glycoprotein (Env trimers. We have recently described eCD4-Ig, a potent and exceptionally broad inhibitor of HIV-1 entry that can be used to protect rhesus macaques from multiple high-dose challenges with simian-human immunodeficiency virus AD8 (SHIV-AD8. Here we show that eCD4-Ig bearing an IgG1 Fc domain (eCD4-IgG1 can mediate efficient ADCC activity against HIV-1 isolates with differing tropisms, and that it does so at least 10-fold more efficiently than CD4-Ig, even when more CD4-Ig molecules bound cell surface-expressed Env. An ADCC-inactive IgG2 form of eCD4-Ig (eCD4-IgG2 exposes V3-loop and CD4-induced epitopes on cell-expressed trimers, and renders HIV-1-infected cells susceptible to ADCC mediated by antibodies of these classes. Moreover, eCD4-IgG2, but not IgG2 forms of the broadly neutralizing antibodies VRC01 and 10-1074, enhances the ADCC activities of serum antibodies from patients by 100-fold, and significantly enhanced killing of two latently infected T-cell lines reactivated by vorinostat or TNFα. Thus eCD4-Ig is qualitatively different from CD4-Ig or neutralizing antibodies in its ability to mediate ADCC, and it may be uniquely useful in treating HIV-1 infection or reducing the reservoir of latently infected cells.

  16. The extracellular domain of myelin oligodendrocyte glycoprotein elicits atypical experimental autoimmune encephalomyelitis in rat and Macaque species.

    Directory of Open Access Journals (Sweden)

    Alan D Curtis

    Full Text Available Atypical models of experimental autoimmune encephalomyelitis (EAE are advantageous in that the heterogeneity of clinical signs appears more reflective of those in multiple sclerosis (MS. Conversely, models of classical EAE feature stereotypic progression of an ascending flaccid paralysis that is not a characteristic of MS. The study of atypical EAE however has been limited due to the relative lack of suitable models that feature reliable disease incidence and severity, excepting mice deficient in gamma-interferon signaling pathways. In this study, atypical EAE was induced in Lewis rats, and a related approach was effective for induction of an unusual neurologic syndrome in a cynomolgus macaque. Lewis rats were immunized with the rat immunoglobulin variable (IgV-related extracellular domain of myelin oligodendrocyte glycoprotein (IgV-MOG in complete Freund's adjuvant (CFA followed by one or more injections of rat IgV-MOG in incomplete Freund's adjuvant (IFA. The resulting disease was marked by torticollis, unilateral rigid paralysis, forelimb weakness, and high titers of anti-MOG antibody against conformational epitopes of MOG, as well as other signs of atypical EAE. A similar strategy elicited a distinct atypical form of EAE in a cynomolgus macaque. By day 36 in the monkey, titers of IgG against conformational epitopes of extracellular MOG were evident, and on day 201, the macaque had an abrupt onset of an unusual form of EAE that included a pronounced arousal-dependent, transient myotonia. The disease persisted for 6-7 weeks and was marked by a gradual, consistent improvement and an eventual full recovery without recurrence. These data indicate that one or more boosters of IgV-MOG in IFA represent a key variable for induction of atypical or unusual forms of EAE in rat and Macaca species. These studies also reveal a close correlation between humoral immunity against conformational epitopes of MOG, extended confluent demyelinating plaques in

  17. The extracellular domain of myelin oligodendrocyte glycoprotein elicits atypical experimental autoimmune encephalomyelitis in rat and Macaque species.

    Science.gov (United States)

    Curtis, Alan D; Taslim, Najla; Reece, Shaun P; Grebenciucova, Elena; Ray, Richard H; Rosenbaum, Matthew D; Wardle, Robert L; Van Scott, Michael R; Mannie, Mark D

    2014-01-01

    Atypical models of experimental autoimmune encephalomyelitis (EAE) are advantageous in that the heterogeneity of clinical signs appears more reflective of those in multiple sclerosis (MS). Conversely, models of classical EAE feature stereotypic progression of an ascending flaccid paralysis that is not a characteristic of MS. The study of atypical EAE however has been limited due to the relative lack of suitable models that feature reliable disease incidence and severity, excepting mice deficient in gamma-interferon signaling pathways. In this study, atypical EAE was induced in Lewis rats, and a related approach was effective for induction of an unusual neurologic syndrome in a cynomolgus macaque. Lewis rats were immunized with the rat immunoglobulin variable (IgV)-related extracellular domain of myelin oligodendrocyte glycoprotein (IgV-MOG) in complete Freund's adjuvant (CFA) followed by one or more injections of rat IgV-MOG in incomplete Freund's adjuvant (IFA). The resulting disease was marked by torticollis, unilateral rigid paralysis, forelimb weakness, and high titers of anti-MOG antibody against conformational epitopes of MOG, as well as other signs of atypical EAE. A similar strategy elicited a distinct atypical form of EAE in a cynomolgus macaque. By day 36 in the monkey, titers of IgG against conformational epitopes of extracellular MOG were evident, and on day 201, the macaque had an abrupt onset of an unusual form of EAE that included a pronounced arousal-dependent, transient myotonia. The disease persisted for 6-7 weeks and was marked by a gradual, consistent improvement and an eventual full recovery without recurrence. These data indicate that one or more boosters of IgV-MOG in IFA represent a key variable for induction of atypical or unusual forms of EAE in rat and Macaca species. These studies also reveal a close correlation between humoral immunity against conformational epitopes of MOG, extended confluent demyelinating plaques in spinal cord and

  18. Plumbing the depths: extracellular water storage in specialized leaf structures and its functional expression in a three-domain pressure -volume relationship.

    Science.gov (United States)

    Nguyen, Hoa T; Meir, Patrick; Wolfe, Joe; Mencuccini, Maurizio; Ball, Marilyn C

    2017-07-01

    A three-domain pressure-volume relationship (PV curve) was studied in relation to leaf anatomical structure during dehydration in the grey mangrove, Avicennia marina. In domain 1, relative water content (RWC) declined 13% with 0.85 MPa decrease in leaf water potential, reflecting a decrease in extracellular water stored primarily in trichomes and petiolar cisternae. In domain 2, RWC decreased by another 12% with a further reduction in leaf water potential to -5.1 MPa, the turgor loss point. Given the osmotic potential at full turgor (-4.2 MPa) and the effective modulus of elasticity (~40 MPa), domain 2 emphasized the role of cell wall elasticity in conserving cellular hydration during leaf water loss. Domain 3 was dominated by osmotic effects and characterized by plasmolysis in most tissues and cell types without cell wall collapse. Extracellular and cellular water storage could support an evaporation rate of 1 mmol m -2 s -1 for up to 54 and 50 min, respectively, before turgor loss was reached. This study emphasized the importance of leaf anatomy for the interpretation of PV curves, and identified extracellular water storage sites that enable transient water use without substantive turgor loss when other factors, such as high soil salinity, constrain rates of water transport. © 2016 John Wiley & Sons Ltd.

  19. Structure of the extracellular domain of matrix protein 2 of influenza A virus in complex with a protective monoclonal antibody.

    Science.gov (United States)

    Cho, Ki Joon; Schepens, Bert; Seok, Jong Hyeon; Kim, Sella; Roose, Kenny; Lee, Ji-Hye; Gallardo, Rodrigo; Van Hamme, Evelien; Schymkowitz, Joost; Rousseau, Frederic; Fiers, Walter; Saelens, Xavier; Kim, Kyung Hyun

    2015-04-01

    The extracellular domain of influenza A virus matrix protein 2 (M2e) is conserved and is being evaluated as a quasiuniversal influenza A vaccine candidate. We describe the crystal structure at 1.6 Å resolution of M2e in complex with the Fab fragment of an M2e-specific monoclonal antibody that protects against influenza A virus challenge. This antibody binds M2 expressed on the surfaces of cells infected with influenza A virus. Five out of six complementary determining regions interact with M2e, and three highly conserved M2e residues are critical for this interaction. In this complex, M2e adopts a compact U-shaped conformation stabilized in the center by the highly conserved tryptophan residue in M2e. This is the first description of the three-dimensional structure of M2e. M2e of influenza A is under investigation as a universal influenza A vaccine, but its three-dimensional structure is unknown. We describe the structure of M2e stabilized with an M2e-specific monoclonal antibody that recognizes natural M2. We found that the conserved tryptophan is positioned in the center of the U-shaped structure of M2e and stabilizes its conformation. The structure also explains why previously reported in vivo escape viruses, selected with a similar monoclonal antibody, carried proline residue substitutions at position 10 in M2. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  20. Deletion of the calmodulin-binding domain of Grb7 impairs cell attachment to the extracellular matrix and migration

    Energy Technology Data Exchange (ETDEWEB)

    García-Palmero, Irene; Villalobo, Antonio, E-mail: antonio.villalobo@iib.uam.es

    2013-06-28

    Highlights: •Grb7 is a calmodulin (CaM)-binding protein. •Deleting the CaM-binding site impairs cell attachment and migration. •CaM antagonists inhibit Grb7-mediated cell migration. •We conclude that CaM controls Grb7-mediated cell migration. -- Abstract: The adaptor Grb7 is a calmodulin (CaM)-binding protein that participates in signaling pathways involved in cell migration, proliferation and the control of angiogenesis, and plays a significant role in tumor growth, its metastatic spread and tumor-associated neo-vasculature formation. In this report we show that deletion of the CaM-binding site of Grb7, located in the proximal region of its pleckstrin homology (PH) domain, impairs cell migration, cell attachment to the extracellular matrix, and the reorganization of the actin cytoskeleton occurring during this process. Moreover, we show that the cell-permeable CaM antagonists N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide (W-7) and N-(4-aminobutyl)-5-chloro-2-naphthalenesulfonamide (W-13) both retard the migration of cells expressing wild type Grb7, but not the migration of cells expressing the mutant protein lacking the CaM-binding site (Grb7Δ), underscoring the proactive role of CaM binding to Grb7 during this process.

  1. Preparation and quality control of ECD kit

    International Nuclear Information System (INIS)

    Yassin, T.; Dadokh, M.; Almalki, R.

    2005-05-01

    ECD Kit for brain imaging after labeling with technetium 99m was prepared according to an optimum conditions, Each vial contains 1 mg of ECD and 0.155 mg of stannous chloride anhydrous SnCl 2 , 24 mg of Mannitol and 0.5 mg of Na 2 Ca EDTA, The prepared kit showed high quality satisfying the requirements of international pharmacopoeias from the points of physical, chemical, radiochemical and biological purities, and its validity for human injection. And the labeling yield exceeded 95 % with average value of about 97.63 ± 0.89% for 2mCi /2ml radioactivity , This study also showed that each vial content can be labeled with maximum activity of 99m Tc of about 50 mCi (Authors)

  2. Inactivation of Mechanically Activated Piezo1 Ion Channels Is Determined by the C-Terminal Extracellular Domain and the Inner Pore Helix

    Directory of Open Access Journals (Sweden)

    Jason Wu

    2017-11-01

    Full Text Available Piezo proteins form mechanically activated ion channels that are responsible for our sense of light touch, proprioception, and vascular blood flow. Upon activation by mechanical stimuli, Piezo channels rapidly inactivate in a voltage-dependent manner through an unknown mechanism. Inactivation of Piezo channels is physiologically important, as it modulates overall mechanical sensitivity, gives rise to frequency filtering of repetitive mechanical stimuli, and is itself the target of numerous human disease-related channelopathies that are not well understood mechanistically. Here, we identify the globular C-terminal extracellular domain as a structure that is sufficient to confer the time course of inactivation and a single positively charged lysine residue at the adjacent inner pore helix as being required for its voltage dependence. Our results are consistent with a mechanism for inactivation that is mediated through voltage-dependent conformations of the inner pore helix and allosteric coupling with the C-terminal extracellular domain.

  3. SagE induces highly effective protective immunity against Streptococcus iniae mainly through an immunogenic domain in the extracellular region.

    Science.gov (United States)

    Sun, Yun; Sun, Li; Xing, Ming-qing; Liu, Chun-sheng; Hu, Yong-hua

    2013-11-12

    Streptococcus iniae is a Gram-positive bacterium and a severe pathogen of a wide range of farmed fish. S. iniae possesses a virulence-associated streptolysin S cluster composed of several components, one of which is SagE. SagE a transmembrane protein with one major extracellular region named ECR. This study aimed to develop a SagE-based DNA candidate vaccine against streptococcosis and examine the immunoprotective mechanism of the vaccine. We constructed a DNA vaccine, pSagE, based on the sagE gene and examined its immunological property in a Japanese flounder (Paralichthys olivaceus) model. The results showed that at 7 days post-vaccination, expression of SagE at transcription and translation levels was detected in the tissues of the vaccinated fish. After challenge with S. iniae at one and two months post-vaccination, pSagE-vaccinated fish exhibited relative percent survival (RPS) of 95% and 88% respectively. Immunological analysis showed that (i) pSagE significantly upregulated the expression of a wide range of immune genes, (ii) pSagE induced the production of specific serum antibodies that bound whole-cell S. iniae, and (iii) treatment of S. iniae with pSagE-induced antibodies blocked bacterial invasion of host cells. To localize the immunoprotective domain of SagE, the ECR-expressing DNA vaccine pSagEECR was constructed. Immunization analysis showed that flounder vaccinated with pSagEECR exhibited a RPS of 68%, and that pSagEECR induced serum antibody production and immune gene expression in a manner similar to, though to lower magnitudes than, those induced by pSagE. We in this study developed a DNA vaccine, pSagE, which induces highly protective immunity against S. iniae. The protective effect of pSagE is probably due to its ability to elicit systemic immune response, in particular that of the humoral branch, which leads to production of specific serum antibodies that impair bacterial infection. These results add insights to the immunoprotective mechanism

  4. Structure of the EMMPRIN N-terminal domain 1: Dimerization via [beta]-strand swapping

    Energy Technology Data Exchange (ETDEWEB)

    Luo, Jinquan; Teplyakov, Alexey; Obmolova, Galina; Malia, Thomas; Wu, Sheng-Jiun; Beil, Eric; Baker, Audrey; Swencki-Underwood, Bethany; Zhao, Yonghong; Sprenkle, Justin; Dixon, Ken; Sweet, Raymond; Gilliland, Gary L.; (Centocor)

    2010-09-27

    Extracellular matrix metalloproteinase inducer (EMMPRIN), also known as Hab18G, CD147, Basigin, M6, and neurothelin, is a membrane glycoprotein expressed on the surface of various cell types and many cancer cells. EMMPRIN stimulates adjacent fibroblasts and tumor cells to produce matrix metalloproteinases and plays an important role in tumor invasion and metastasis, angiogenesis, spermatogensis and fertilization, cell-cell adhesion and communication, and other biological processes (reviewed in Ref. 1 and references therein). It was demonstrated that the EMMPRIN extracellular domain (ECD), which structurally belongs to the IgG superfamily, can form homo-oligomers in a cis dependent manner and the N-terminal domain 1 (residues 22-101) was necessary and sufficient to mediate this interaction. The crystal structure of the ECD of recombinant human EMMPRIN (Hab18G/CD147) expressed in E. coli was reported at 2.8 {angstrom} resolution (Yu et al. 2008). The construct consists of residues 22-205 of the mature protein and has both an N-terminal IgC2 domain (ND1, residues 22-101) and a C-terminal IgC2 domain (ND2, residues 107-205). The two domains are joined by a five amino acid residue linker that constitutes a flexible hinge between the two domains. The crystal form has four copies of the molecule in the asymmetric unit, each of which has a different inter-domain angle that varies from 121{sup o} to 144{sup o}. The two domains each have a conserved disulfide bridge and both are comprised of two {beta}-sheets formed by strands EBA and GFCC, and DEBA and AGFCC for ND1 and ND2, respectively. Based on the crystal packing in this structure, the authors proposed that lateral packing between the two IgG domains of EMMPRIN ECD represents a potential mechanism for cell adhesion. Here we report the 2.0-{angstrom} crystal structure of the N-terminal domain of EMMPRIN ECD (ND1) expressed in mammalian cells. The overall structure of the domain is very similar to that in the full length

  5. Crystallization and preliminary X-ray crystallographic study of the extracellular domain of the 4-1BB ligand, a member of the TNF family

    Energy Technology Data Exchange (ETDEWEB)

    Byun, Jung-Sue; Kim, Dong-Uk [Department of Biology, Yonsei University, Seoul 120-749 (Korea, Republic of); Ahn, Byungchan; Kwon, Byoung Se [Immunomodulation Research Center, Ulsan University, Ulsan 680-749 (Korea, Republic of); Cho, Hyun-Soo, E-mail: hscho8@yonsei.ac.kr [Department of Biology, Yonsei University, Seoul 120-749 (Korea, Republic of)

    2006-01-01

    The extracellular domain of the 4-1BB ligand fused with glutathione-S-transferase was expressed in Escherichia coli (Origami) and purified by using affinity and ion-exchange column chromatographic methods. Crystals of the 4-1BB ligand were obtained at 290 K by the hanging-drop vapour-diffusion method. The 4-1BB ligand, a member of the tumour necrosis factor (TNF) family, is an important co-stimulatory molecule that plays a key role in the clonal expansion and survival of CD8+ T cells. Signalling through binding of the 4-1BB ligand and 4-1BB has been reported to enhance CD8+ T-cell expansion and protect activated CD8+ T cells from death. The 4-1BB ligand is an integral protein expressed on activated antigen-presenting cells. The extracellular domain of the 4-1BB ligand fused with glutathione-S-transferase was expressed in Escherichia coli (Origami) and purified by using affinity and ion-exchange column chromatographic methods. Crystals of the 4-1BB ligand were obtained at 290 K by the hanging-drop vapour-diffusion method. X-ray diffraction data were collected from these crystals to 2.8 Å resolution and the crystals belong to space group C2, with unit-cell parameters a = 114.6, b = 73.8, c = 118.50 Å, β = 115.5°.

  6. Crystallization and preliminary X-ray crystallographic study of the extracellular domain of the 4-1BB ligand, a member of the TNF family

    International Nuclear Information System (INIS)

    Byun, Jung-Sue; Kim, Dong-Uk; Ahn, Byungchan; Kwon, Byoung Se; Cho, Hyun-Soo

    2005-01-01

    The extracellular domain of the 4-1BB ligand fused with glutathione-S-transferase was expressed in Escherichia coli (Origami) and purified by using affinity and ion-exchange column chromatographic methods. Crystals of the 4-1BB ligand were obtained at 290 K by the hanging-drop vapour-diffusion method. The 4-1BB ligand, a member of the tumour necrosis factor (TNF) family, is an important co-stimulatory molecule that plays a key role in the clonal expansion and survival of CD8+ T cells. Signalling through binding of the 4-1BB ligand and 4-1BB has been reported to enhance CD8+ T-cell expansion and protect activated CD8+ T cells from death. The 4-1BB ligand is an integral protein expressed on activated antigen-presenting cells. The extracellular domain of the 4-1BB ligand fused with glutathione-S-transferase was expressed in Escherichia coli (Origami) and purified by using affinity and ion-exchange column chromatographic methods. Crystals of the 4-1BB ligand were obtained at 290 K by the hanging-drop vapour-diffusion method. X-ray diffraction data were collected from these crystals to 2.8 Å resolution and the crystals belong to space group C2, with unit-cell parameters a = 114.6, b = 73.8, c = 118.50 Å, β = 115.5°

  7. Homophilic interactions mediated by receptor tyrosine phosphatases mu and kappa. A critical role for the novel extracellular MAM domain

    DEFF Research Database (Denmark)

    Zondag, G C; Koningstein, G M; Jiang, Y P

    1995-01-01

    and is found in diverse transmembrane proteins, is not known. We previously reported that both RPTP mu and RPTP kappa can mediate homophilic cell interactions when expressed in insect cells. Here we show that despite their striking structural similarity, RPTP mu and RPTP kappa fail to interact...... in a heterophilic manner. To examine the role of the MAM domain in homophilic binding, we expressed a mutant RPTP mu lacking the MAM domain in insect Sf9 cells. Truncated RPTP mu is properly expressed at the cell surface but fails to promote cell-cell adhesion. Homophilic cell adhesion is fully restored...... in a chimeric RPTP mu molecule containing the MAM domain of RPTP kappa. However, this chimeric RPTP mu does not interact with either RPTP mu or RPTP kappa. These results indicate that the MAM domain of RPTP mu and RPTP kappa is essential for homophilic cell-cell interaction and helps determine the specificity...

  8. pH-dependent structural change of the extracellular sensor domain of the DraK histidine kinase from Streptomyces coelicolor

    International Nuclear Information System (INIS)

    Yeo, Kwon Joo; Kim, Eun Hye; Hwang, Eunha; Han, Young-Hyun; Eo, Yumi; Kim, Hyun Jung; Kwon, Ohsuk; Hong, Young-Soo; Cheong, Chaejoon; Cheong, Hae-Kap

    2013-01-01

    Highlights: ► We described the biochemical and biophysical properties of the extracellular sensory domain (ESD) of DraK histidine kinase. ► The ESD of DraK showed a reversible pH-dependent conformational change in a wide pH range. ► The E83 is an important residue for the pH-dependent conformational change. -- Abstract: Recently, the DraR/DraK (Sco3063/Sco3062) two-component system (TCS) of Streptomycescoelicolor has been reported to be involved in the differential regulation of antibiotic biosynthesis. However, it has not been shown that under which conditions and how the DraR/DraK TCS is activated to initiate the signal transduction process. Therefore, to understand the sensing mechanism, structural study of the sensory domain of DraK is highly required. Here, we report the biochemical and biophysical properties of the extracellular sensory domain (ESD) of DraK. We observed a reversible pH-dependent conformational change of the ESD in a pH range of 2.5–10. Size-exclusion chromatography and AUC (analytical ultracentrifugation) data indicated that the ESD is predominantly monomeric in solution and exists in equilibrium between monomer and dimer states in acidic condition. Using NMR (nuclear magnetic resonance) and CD (circular dichroism) spectroscopy, our findings suggest that the structure of the ESD at low pH is more structured than that at high pH. In particular, the glutamate at position 83 is an important residue for the pH-dependent conformational change. These results suggest that this pH-dependent conformational change of ESD may be involved in signal transduction process of DraR/DraK TCS

  9. pH-dependent structural change of the extracellular sensor domain of the DraK histidine kinase from Streptomyces coelicolor

    Energy Technology Data Exchange (ETDEWEB)

    Yeo, Kwon Joo [Division of Magnetic Resonance, Korea Basic Science Institute (KBSI), 16 Yeongudanji-Ro, Ochang, Chungbuk 363-883 (Korea, Republic of); Kim, Eun Hye [Systems and Synthetic Biology Research Center, Korea Research Institute of Bioscience and Biotechnology (KRIBB), 125 Gwahak-Ro, Yuseong-Gu, Daejeon 305-333 (Korea, Republic of); Hwang, Eunha; Han, Young-Hyun; Eo, Yumi; Kim, Hyun Jung [Division of Magnetic Resonance, Korea Basic Science Institute (KBSI), 16 Yeongudanji-Ro, Ochang, Chungbuk 363-883 (Korea, Republic of); Kwon, Ohsuk [Systems and Synthetic Biology Research Center, Korea Research Institute of Bioscience and Biotechnology (KRIBB), 125 Gwahak-Ro, Yuseong-Gu, Daejeon 305-333 (Korea, Republic of); Hong, Young-Soo [Chemical Biology Research Center, KRIBB, 30 Yeongudanji-Ro, Ochang, Chungbuk 363-883 (Korea, Republic of); Cheong, Chaejoon, E-mail: cheong@kbsi.re.kr [Division of Magnetic Resonance, Korea Basic Science Institute (KBSI), 16 Yeongudanji-Ro, Ochang, Chungbuk 363-883 (Korea, Republic of); Cheong, Hae-Kap, E-mail: haekap@kbsi.re.kr [Division of Magnetic Resonance, Korea Basic Science Institute (KBSI), 16 Yeongudanji-Ro, Ochang, Chungbuk 363-883 (Korea, Republic of)

    2013-02-15

    Highlights: ► We described the biochemical and biophysical properties of the extracellular sensory domain (ESD) of DraK histidine kinase. ► The ESD of DraK showed a reversible pH-dependent conformational change in a wide pH range. ► The E83 is an important residue for the pH-dependent conformational change. -- Abstract: Recently, the DraR/DraK (Sco3063/Sco3062) two-component system (TCS) of Streptomycescoelicolor has been reported to be involved in the differential regulation of antibiotic biosynthesis. However, it has not been shown that under which conditions and how the DraR/DraK TCS is activated to initiate the signal transduction process. Therefore, to understand the sensing mechanism, structural study of the sensory domain of DraK is highly required. Here, we report the biochemical and biophysical properties of the extracellular sensory domain (ESD) of DraK. We observed a reversible pH-dependent conformational change of the ESD in a pH range of 2.5–10. Size-exclusion chromatography and AUC (analytical ultracentrifugation) data indicated that the ESD is predominantly monomeric in solution and exists in equilibrium between monomer and dimer states in acidic condition. Using NMR (nuclear magnetic resonance) and CD (circular dichroism) spectroscopy, our findings suggest that the structure of the ESD at low pH is more structured than that at high pH. In particular, the glutamate at position 83 is an important residue for the pH-dependent conformational change. These results suggest that this pH-dependent conformational change of ESD may be involved in signal transduction process of DraR/DraK TCS.

  10. A Novel Interaction of Ecdysoneless (ECD) Protein with R2TP Complex Component RUVBL1 Is Required for the Functional Role of ECD in Cell Cycle Progression.

    Science.gov (United States)

    Mir, Riyaz A; Bele, Aditya; Mirza, Sameer; Srivastava, Shashank; Olou, Appolinaire A; Ammons, Shalis A; Kim, Jun Hyun; Gurumurthy, Channabasavaiah B; Qiu, Fang; Band, Hamid; Band, Vimla

    2015-12-28

    Ecdysoneless (ECD) is an evolutionarily conserved protein whose germ line deletion is embryonic lethal. Deletion of Ecd in cells causes cell cycle arrest, which is rescued by exogenous ECD, demonstrating a requirement of ECD for normal mammalian cell cycle progression. However, the exact mechanism by which ECD regulates cell cycle is unknown. Here, we demonstrate that ECD protein levels and subcellular localization are invariant during cell cycle progression, suggesting a potential role of posttranslational modifications or protein-protein interactions. Since phosphorylated ECD was recently shown to interact with the PIH1D1 adaptor component of the R2TP cochaperone complex, we examined the requirement of ECD phosphorylation in cell cycle progression. Notably, phosphorylation-deficient ECD mutants that failed to bind to PIH1D1 in vitro fully retained the ability to interact with the R2TP complex and yet exhibited a reduced ability to rescue Ecd-deficient cells from cell cycle arrest. Biochemical analyses demonstrated an additional phosphorylation-independent interaction of ECD with the RUVBL1 component of the R2TP complex, and this interaction is essential for ECD's cell cycle progression function. These studies demonstrate that interaction of ECD with RUVBL1, and its CK2-mediated phosphorylation, independent of its interaction with PIH1D1, are important for its cell cycle regulatory function. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

  11. pH-dependent structural change of the extracellular sensor domain of the DraK histidine kinase from Streptomyces coelicolor.

    Science.gov (United States)

    Yeo, Kwon Joo; Kim, Eun Hye; Hwang, Eunha; Han, Young-Hyun; Eo, Yumi; Kim, Hyun Jung; Kwon, Ohsuk; Hong, Young-Soo; Cheong, Chaejoon; Cheong, Hae-Kap

    2013-02-15

    Recently, the DraR/DraK (Sco3063/Sco3062) two-component system (TCS) of Streptomycescoelicolor has been reported to be involved in the differential regulation of antibiotic biosynthesis. However, it has not been shown that under which conditions and how the DraR/DraK TCS is activated to initiate the signal transduction process. Therefore, to understand the sensing mechanism, structural study of the sensory domain of DraK is highly required. Here, we report the biochemical and biophysical properties of the extracellular sensory domain (ESD) of DraK. We observed a reversible pH-dependent conformational change of the ESD in a pH range of 2.5-10. Size-exclusion chromatography and AUC (analytical ultracentrifugation) data indicated that the ESD is predominantly monomeric in solution and exists in equilibrium between monomer and dimer states in acidic condition. Using NMR (nuclear magnetic resonance) and CD (circular dichroism) spectroscopy, our findings suggest that the structure of the ESD at low pH is more structured than that at high pH. In particular, the glutamate at position 83 is an important residue for the pH-dependent conformational change. These results suggest that this pH-dependent conformational change of ESD may be involved in signal transduction process of DraR/DraK TCS. Copyright © 2013 Elsevier Inc. All rights reserved.

  12. Exploring ECD on a Benchtop Q Exactive Orbitrap Mass Spectrometer

    DEFF Research Database (Denmark)

    Fort, Kyle L; Cramer, Christian N; Voinov, Valery G

    2018-01-01

    As the application of mass spectrometry intensifies in scope and diversity, the need for advanced instrumentation addressing a wide variety of analytical needs also increases. To this end, many modern, top-end mass spectrometers are designed or modified to include a wider range of fragmentation...... applications including middle-down proteomics, top-down proteomics, glycoproteomics, and disulfide bond mapping. We describe the modification of the popular Q Exactive Orbitrap mass spectrometer to extend its fragmentation capabilities to include ECD. We show that this modification allows ≥85% matched ion...... intensity to originate from ECD fragment ion types as well as provides high sequence coverage (≥60%) of intact proteins and high fragment identification rates with ∼70% of ion signals matched. Finally, the ECD implementation promotes selective disulfide bond dissociation, facilitating the identification...

  13. Structural characterization by NMR of the natively unfolded extracellular domain of beta-dystroglycan: toward the identification of the binding epitope for alpha-dystroglycan.

    Science.gov (United States)

    Bozzi, Manuela; Bianchi, Marzia; Sciandra, Francesca; Paci, Maurizio; Giardina, Bruno; Brancaccio, Andrea; Cicero, Daniel O

    2003-11-25

    Dystroglycan (DG) is an adhesion molecule playing a crucial role for tissue stability during both early embriogenesis and adulthood and is composed by two tightly interacting subunits: alpha-DG, membrane-associated and highly glycosylated, and the transmembrane beta-DG. Recently, by solid-phase binding assays and NMR experiments, we have shown that the C-terminal domain of alpha-DG interacts with a recombinant extracellular fragment of beta-DG (positions 654-750) independently from glycosylation and that the linear binding epitope is located between residues 550 and 565 of alpha-DG. In order to elucidate which moieties of beta-DG are specifically involved in the complex with alpha-DG, the ectodomain has been recombinantly expressed and purified in a labeled ((13)C,(15)N) form and studied by multidimensional NMR. Although it represents a natively unfolded protein domain, we obtained an almost complete backbone assignment. Chemical shift index, (1)H-(15)N heteronuclear single-quantum coherence and nuclear Overhauser effect (HSQC-NOESY) spectra and (3)J(HN,H)(alpha) coupling constant values confirm that this protein is highly disordered, but (1)H-(15)N steady-state NOE experiments indicate that the protein presents two regions of different mobility. The first one, between residues 659 and 722, is characterized by a limited degree of mobility, whereas the C-terminal portion, containing about 30 amino acids, is highly flexible. The binding of beta-DG(654-750) to the C-terminal region of the alpha subunit, alpha-DG(485-620), has been investigated, showing that the region of beta-DG(654-750) between residues 691 and 719 is involved in the interaction.

  14. The Extracellular Domain of Human High Affinity Copper Transporter (hNdCTR1), Synthesized by E. coli Cells, Chelates Silver and Copper Ions In Vivo.

    Science.gov (United States)

    Sankova, Tatiana P; Orlov, Iurii A; Saveliev, Andrey N; Kirilenko, Demid A; Babich, Polina S; Brunkov, Pavel N; Puchkova, Ludmila V

    2017-11-03

    There is much interest in effective copper chelators to correct copper dyshomeostasis in neurodegenerative and oncological diseases. In this study, a recombinant fusion protein for expression in Escherichia coli cells was constructed from glutathione-S-transferase (GST) and the N-terminal domain (ectodomain) of human high affinity copper transporter CTR1 (hNdCTR1), which has three metal-bound motifs. Several biological properties of the GST-hNdCTR1 fusion protein were assessed. It was demonstrated that in cells, the protein was prone to oligomerization, formed inclusion bodies and displayed no toxicity. Treatment of E. coli cells with copper and silver ions reduced cell viability in a dose- and time-dependent manner. Cells expressing GST-hNdCTR1 protein demonstrated resistance to the metal treatments. These cells accumulated silver ions and formed nanoparticles that contained AgCl and metallic silver. In this bacterial population, filamentous bacteria with a length of about 10 µm were often observed. The possibility for the fusion protein carrying extracellular metal binding motifs to integrate into the cell's copper metabolism and its chelating properties are discussed.

  15. The Extracellular Domain of Human High Affinity Copper Transporter (hNdCTR1, Synthesized by E. coli Cells, Chelates Silver and Copper Ions In Vivo

    Directory of Open Access Journals (Sweden)

    Tatiana P. Sankova

    2017-11-01

    Full Text Available There is much interest in effective copper chelators to correct copper dyshomeostasis in neurodegenerative and oncological diseases. In this study, a recombinant fusion protein for expression in Escherichia coli cells was constructed from glutathione-S-transferase (GST and the N-terminal domain (ectodomain of human high affinity copper transporter CTR1 (hNdCTR1, which has three metal-bound motifs. Several biological properties of the GST-hNdCTR1 fusion protein were assessed. It was demonstrated that in cells, the protein was prone to oligomerization, formed inclusion bodies and displayed no toxicity. Treatment of E. coli cells with copper and silver ions reduced cell viability in a dose- and time-dependent manner. Cells expressing GST-hNdCTR1 protein demonstrated resistance to the metal treatments. These cells accumulated silver ions and formed nanoparticles that contained AgCl and metallic silver. In this bacterial population, filamentous bacteria with a length of about 10 µm were often observed. The possibility for the fusion protein carrying extracellular metal binding motifs to integrate into the cell’s copper metabolism and its chelating properties are discussed.

  16. Synthesis and formulation of 99m Tc-ECD radiopharmaceutical

    International Nuclear Information System (INIS)

    Ocampo G, B.E.

    1998-01-01

    Nuclear medicine is a medical specialty which uses radioactive compounds (radionuclides) for diagnostic and therapeutic purposes. 99m Tc is the more common radionuclide used in many studies in nuclear medicine because its advantages: it has a photopeak of 140 KeV and a half-life of 6 hours; it can be eluted from a Molybdenum 99 generator, so radiopharmaceuticals can be prepared on site. Ethyl cysteine dimer (ECD) labelled with reduced Technetium 99m has been purposed recently as a promising radiopharmaceutical for brain perfusion imaging 99m Tc-ECD is a lipophilic neutral complex which cross the brain blood barrier and show high brain uptake. The objective of this work was synthesize and to design a freeze dried formulation for the instant preparation of 99m Tc-ECD complex useful for brain perfusion imaging. We obtained a freeze dried stable formulation for the preparation of 99m Tc-ECD kit with a radiochemical purity higher than 90 %, which fulfills with the quality control of radiopharmaceuticals. Furthermore, we developed analytic techniques for the determination of the different chemical compounds into the lyophilized kit. (Author)

  17. Alanine rich peptide from Populus trichocarpa inhibit growth of Staphylococcus aureus via targetting its extracellular domain of Sensor Histidine Kinase YycGex protein.

    Science.gov (United States)

    Al Akeel, Raid; Mateen, Ayesha; Syed, Rabbani; Al-Qahtani, Mohammed S; Alqahtani, A

    2018-05-11

    Due to growing concern towards microbial resistance, ongoing search for developing novel bioactive compounds such as peptides is on rise. The aim of this study was to evaluate antimicrobial effect of Populus trichocarpa extract, chemically identify the active peptide fraction and finds its target in Staphylococcus aureus. In this study the active fraction of P. trichocarpa crude extract was purified and characterized using MS/MS. This peptide PT13 antimicrobial activity was confirmed by in-vitro agar based disk diffusion and in-vivo infection model of G. mellonella. The proteomic expression analysis of S. aureus under influence of PT13 was studied using LTQ-Orbitrap-MS in-solution digestion and identity of target protein was acquired with their quantified expression using label-free approach of Progenesis QI software. Docking study was performed with peptide PT13 and its target YycG protein using CABS-dock. The active fraction PT13 sequence was identified as KVPVAAAAAAAAAVVASSMVVAAAK, with 25 amino acid including 13 alanine having M/Z 2194.2469. PT13 was uniformly inhibited growth S. aureus SA91 and MIC was determined 16 μg/mL for SA91 S. aureus strain. Sensor histidine kinase (YycG) was most significant target found differentially expressed under influence of PT13. G. mellonella larvae were killed rapidly due to S aureus infection, whereas death in protected group was insignificant in compare to control. The docking models showed ten docking models with RMSD value 1.89 for cluster 1 and RMSD value 3.95 for cluster 2 which is predicted to be high quality model. Alanine rich peptide could be useful in constructing as antimicrobial peptide for targeting extracellular Domain of Sensor Histidine Kinase YycG from S. aureus used in the study. Copyright © 2018. Published by Elsevier Ltd.

  18. Differential structural properties of GLP-1 and exendin-4 determine their relative affinity for the GLP-1 receptor N-terminal extracellular domain.

    Science.gov (United States)

    Runge, Steffen; Schimmer, Susann; Oschmann, Jan; Schiødt, Christine Bruun; Knudsen, Sanne Möller; Jeppesen, Claus Bekker; Madsen, Kjeld; Lau, Jesper; Thøgersen, Henning; Rudolph, Rainer

    2007-05-15

    Glucagon-like peptide-1 (GLP-1) and exendin-4 (Ex4) are homologous peptides with established potential for treatment of type 2 diabetes. They bind and activate the pancreatic GLP-1 receptor (GLP-1R) with similar affinity and potency and thereby promote insulin secretion in a glucose-dependent manner. GLP-1R belongs to family B of the seven transmembrane G-protein coupled receptors. The N-terminal extracellular domain (nGLP-1R) is a ligand binding domain with differential affinity for Ex4 and GLP-1: low affinity for GLP-1 and high affinity for exendin-4. The superior affinity of nGLP-1R for Ex4 was previously explained by an additional interaction between nGLP-1R and the C-terminal Trp-cage of Ex4. In this study we have combined biophysical and pharmacological approaches thus relating structural properties of the ligands in solution to their relative binding affinity for nGLP-1R. We used both a tracer competition assay and ligand-induced thermal stabilization of nGLP-1R to measure the relative affinity of full length, truncated, and chimeric ligands for soluble refolded nGLP-1R. The ligands in solution and the conformational consequences of ligand binding to nGLP-1R were characterized by circular dichroism and fluorescence spectroscopy. We found a correlation between the helical content of the free ligands and their relative binding affinity for nGLP-1R, supporting the hypothesis that the ligands are helical at least in the segment that binds to nGLP-1R. The Trp-cage of Ex4 was not necessary to maintain a superior helicity of Ex4 compared to GLP-1. The results suggest that the differential affinity of nGLP-1R is explained almost entirely by divergent residues in the central part of the ligands: Leu10-Gly30 of Ex4 and Val16-Arg36 of GLP-1. In view of our results it appears that the Trp-cage plays only a minor role for the interaction between Ex4 and nGLP-1R and for the differential affinity of nGLP-1R for GLP-1 and Ex4.

  19. Diversity of Two-Domain Laccase-Like Multicopper Oxidase Genes in Streptomyces spp.: Identification of Genes Potentially Involved in Extracellular Activities and Lignocellulose Degradation during Composting of Agricultural Waste

    Science.gov (United States)

    Lu, Lunhui; Zhang, Jiachao; Chen, Anwei; Chen, Ming; Jiang, Min; Yuan, Yujie; Wu, Haipeng; Lai, Mingyong; He, Yibin

    2014-01-01

    Traditional three-domain fungal and bacterial laccases have been extensively studied for their significance in various biotechnological applications. Growing molecular evidence points to a wide occurrence of more recently recognized two-domain laccase-like multicopper oxidase (LMCO) genes in Streptomyces spp. However, the current knowledge about their ecological role and distribution in natural or artificial ecosystems is insufficient. The aim of this study was to investigate the diversity and composition of Streptomyces two-domain LMCO genes in agricultural waste composting, which will contribute to the understanding of the ecological function of Streptomyces two-domain LMCOs with potential extracellular activity and ligninolytic capacity. A new specific PCR primer pair was designed to target the two conserved copper binding regions of Streptomyces two-domain LMCO genes. The obtained sequences mainly clustered with Streptomyces coelicolor, Streptomyces violaceusniger, and Streptomyces griseus. Gene libraries retrieved from six composting samples revealed high diversity and a rapid succession of Streptomyces two-domain LMCO genes during composting. The obtained sequence types cluster in 8 distinct clades, most of which are homologous with Streptomyces two-domain LMCO genes, but the sequences of clades III and VIII do not match with any reference sequence of known streptomycetes. Both lignocellulose degradation rates and phenol oxidase activity at pH 8.0 in the composting process were found to be positively associated with the abundance of Streptomyces two-domain LMCO genes. These observations provide important clues that Streptomyces two-domain LMCOs are potentially involved in bacterial extracellular phenol oxidase activities and lignocellulose breakdown during agricultural waste composting. PMID:24657870

  20. Up-regulated Ectonucleotidases in Fas-Associated Death Domain Protein- and Receptor-Interacting Protein Kinase 1-Deficient Jurkat Leukemia Cells Counteract Extracellular ATP/AMP Accumulation via Pannexin-1 Channels during Chemotherapeutic Drug-Induced Apoptosis.

    Science.gov (United States)

    Boyd-Tressler, Andrea M; Lane, Graham S; Dubyak, George R

    2017-07-01

    Pannexin-1 (Panx1) channels mediate the efflux of ATP and AMP from cancer cells in response to induction of extrinsic apoptosis by death receptors or intrinsic apoptosis by chemotherapeutic agents. We previously described the accumulation of extracellular ATP /AMP during chemotherapy-induced apoptosis in Jurkat human leukemia cells. In this study, we compared how different signaling pathways determine extracellular nucleotide pools in control Jurkat cells versus Jurkat lines that lack the Fas-associated death domain (FADD) or receptor-interacting protein kinase 1 (RIP1) cell death regulatory proteins. Tumor necrosis factor- α induced extrinsic apoptosis in control Jurkat cells and necroptosis in FADD-deficient cells; treatment of both lines with chemotherapeutic drugs elicited similar intrinsic apoptosis. Robust extracellular ATP/AMP accumulation was observed in the FADD-deficient cells during necroptosis, but not during apoptotic activation of Panx1 channels. Accumulation of extracellular ATP/AMP was similarly absent in RIP1-deficient Jurkat cells during apoptotic responses to chemotherapeutic agents. Apoptotic activation triggered equivalent proteolytic gating of Panx1 channels in all three Jurkat cell lines. The differences in extracellular ATP/AMP accumulation correlated with cell-line-specific expression of ectonucleotidases that metabolized the released ATP/AMP. CD73 mRNA, and α β -methylene-ADP-inhibitable ecto-AMPase activity were elevated in the FADD-deficient cells. In contrast, the RIP1-deficient cells were defined by increased expression of tartrate-sensitive prostatic acid phosphatase as a broadly acting ectonucleotidase. Thus, extracellular nucleotide accumulation during regulated tumor cell death involves interplay between ATP/AMP efflux pathways and different cell-autonomous ectonucleotidases. Differential expression of particular ectonucleotidases in tumor cell variants will determine whether chemotherapy-induced activation of Panx1 channels

  1. Hyperfixation of Tc-99m ECD in subacute cortical infarction

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Jae Seung; Kweon, Sun Uck; Ryu, Jin Sook; Moon, Dae Hyuk; Lee, Hee Kyung [College of Medicine, Ulsan Univ., Seoul (Korea, Republic of)

    2001-07-01

    It has been known that hyperfixation of Tc-99m ECD (HF) is not shown in subacute cerebral infarction because the brain distribution of Tc-99m ECD reflects not only perfusion but also the metabolic status of brain tissue. However, we observed several cases with HF in the subacute pure cortical infarction. To find out the cause of HF in subacute cortical infarction. We assessed the difference in associated cerebral hemodynamics and clinical findings between the subacute cortical infarctions with and without HF. We reviewed 16 patients (63.8{+-}8.6 yr, M/F: 15/1) with pure cortical infarction not involving adjacent subcortical white matter on MRI. All patients underwent acetazolamide stress brain perfusion SPECT using Tc-99m ECD and MRI at subacute period (7.3{+-}4.4 days from ictus). Uptake of Tc-99m ECD in infarcted cortex was assessed visually comparing the contralateral side. To assess the difference in associate clinical findings between the infarctions with and without HF, rCVR of the cerebral territory including infarcted cortex, extent of Gd-enhancement on MRI. Intervals between SPECT and ictus, and the presence of associated ICA stenosis were evaluated. Infarctions were focal (n=8) or multifocal (n=8) and located in frontoparietal cortices on MRI. Twelve patients were accompanied with ipsilateral ICA stenosis. Resting SPECT showed increased cortical uptake (=HF) in 7 patients and decreased in 9. rCVR of the MCA territory was preserved in all of the 7 patients with HF, compared with 4 of the 9 patients without HF (p=0.03). Gd-enhancement was minimal in all of the 7 patients with HF, compared with of the 0 patients without HF (p=0.03). Presence of ipsilateral ICA stenosis and intervals from ictus were not different (p>0.1) Subacute cerebral cortical infarction with HF was more frequently associated with preserved rCVR and minimal destruction of the blood-brain barrier than that without HF. Our findings suggest that HF may result from luxury perfusion of

  2. Extracellular Membrane-proximal Domain of HAb18G/CD147 Binds to Metal Ion-dependent Adhesion Site (MIDAS) Motif of Integrin β1 to Modulate Malignant Properties of Hepatoma Cells*

    Science.gov (United States)

    Li, Yong; Wu, Jiao; Song, Fei; Tang, Juan; Wang, Shi-Jie; Yu, Xiao-Ling; Chen, Zhi-Nan; Jiang, Jian-Li

    2012-01-01

    Several lines of evidence suggest that HAb18G/CD147 interacts with the integrin variants α3β1 and α6β1. However, the mechanism of the interaction remains largely unknown. In this study, mammalian protein-protein interaction trap (MAPPIT), a mammalian two-hybrid method, was used to study the CD147-integrin β1 subunit interaction. CD147 in human hepatocellular carcinoma (HCC) cells was interfered with by small hairpin RNA. Nude mouse xenograft model and metastatic model of HCC were used to detect the role of CD147 in carcinogenesis and metastasis. We found that the extracellular membrane-proximal domain of HAb18G/CD147 (I-type domain) binds at the metal ion-dependent adhesion site in the βA domain of the integrin β1 subunit, and Asp179 in the I-type domain of HAb18G/CD147 plays an important role in the interaction. The levels of the proteins that act downstream of integrin, including focal adhesion kinase (FAK) and phospho-FAK, were decreased, and the cytoskeletal structures of HCC cells were rearranged bearing the HAb18G/CD147 deletion. Simultaneously, the migration and invasion capacities, secretion of matrix metalloproteinases, colony formation rate in vitro, and tumor growth and metastatic potential in vivo were decreased. These results indicate that the interaction of HAb18G/CD147 extracellular I-type domain with the integrin β1 metal ion-dependent adhesion site motif activates the downstream FAK signaling pathway, subsequently enhancing the malignant properties of HCC cells. PMID:22130661

  3. Republic of Kiribati Early Childhood Development : SABER and NSA-ECD Country Report 2014

    OpenAIRE

    World Bank Group

    2014-01-01

    This report presents an analysis of the Early Childhood Development (ECD) subsector, including programs and policies that affect young children in the Republic of Kiribati. This was a collaborative effort between UNICEF and the World Bank Group; it combines the World Bank Group’s Systems Approach for Better Education Results SABER-ECD framework, which includes analysis of early learningan...

  4. Analysis of Early Childhood Development (ECD) facilities within the city of Cape Town

    CSIR Research Space (South Africa)

    Mokgalaka, H

    2011-10-01

    Full Text Available of the current situation of accessibility of ECD facilities in Cape Town in respect of both location and size. This was followed by modelling of the optimal location of a select number of new ECD Centre’s of Excellence facilities to meet the backlog in current...

  5. Extracellular Signal-Regulated Kinase 2 (ERK2) Phosphorylation Sites and Docking Domain on the Nuclear Pore Complex Protein Tpr Cooperatively Regulate ERK2-Tpr Interaction

    Czech Academy of Sciences Publication Activity Database

    Vomastek, Tomáš; Iwanicky, M. P.; Burack, W. R.; Tiwari, D.; Kumar, D.; Parsons, J. T.; Weber, M. J.; Nandicoori, V. K.

    2008-01-01

    Roč. 28, č. 22 (2008), s. 6954-6966 ISSN 0270-7306 R&D Projects: GA AV ČR IAA500200716 Institutional research plan: CEZ:AV0Z50200510 Keywords : erk * docking domain * cell growth Subject RIV: EE - Microbiology, Virology Impact factor: 5.942, year: 2008

  6. The effectiveness of acupuncture as a treatment for tinnitus: a randomized controlled trial using 99mTc-ECD SPECT

    International Nuclear Information System (INIS)

    Laureano, Maura Regina; Braga Neto, Pedro; Ligia Sanchez, Maura; Cogo Moreira, Hugo; Ribeiro de Almeida, Roberta; Parolin Jackowski, Andrea; Onishi, Ektor Tsuneo; Affonseca Bressan, Rodrigo; Batista, Ilza Rosa; Alves Reis, Marilia; Vieira Castiglioni, Mario Luiz; Vargas Garcia, Michele; Neves de Andrade, Adriana; Jara Garrido, Griselda

    2016-01-01

    Investigate the effect of acupuncture on brain perfusion using ethyl cysteinate dimer single-photon emission computed tomography ( 99m Tc-ECD SPECT) in patients with tinnitus. This randomized, single-blind, sham-control study examined patients (18-60 years old) with normal hearing and chronic, idiopathic, continuous tinnitus. Fifty-seven subjects were randomly assigned to true (n = 30) or sham (n = 27) acupuncture (ACP); 99m Tc-ECD SPECT examinations were performed before and after 12 twice-weekly ACP sessions. Secondary outcomes included changes in the Tinnitus Handicap Inventory (THI), Visual Analog Scale (VAS), Hamilton Anxiety Scale (HAS) and Beck Depression Inventory (BDI). Imaging data were analysed using Statistical Parametric Mapping (SPM8) software. Regression models were used to examine secondary outcomes via two paradigms: intention-to-treat (ITT; where multiple imputations were conducted because of study attrition) and complete cases. No between-group brain perfusion differences were observed. However, a significant improvement in THI scores was observed at the end of true ACP treatment for all domains (all p values < 0.01) except the catastrophic scale. ACP might reduce the effects of tinnitus on daily life; however, additional studies should be conducted to verify the effects of ACP on the neural architecture and brain function of tinnitus patients. (orig.)

  7. Recognition of peptidoglycan and beta-lactam antibiotics by the extracellular domain of the Ser/Thr protein kinase StkP from Streptococcus pneumoniae

    Czech Academy of Sciences Publication Activity Database

    Maestro, B.; Nováková, Linda; Hesek, D.; Lee, M.; Leyva, E.; Mobashery, S.; Sanz, J.M.; Branny, Pavel

    2011-01-01

    Roč. 585, č. 2 (2011), s. 357-363 ISSN 0014-5793 R&D Projects: GA ČR GP204/07/P082; GA ČR GA204/08/0783; GA AV ČR IAA600200801 Institutional research plan: CEZ:AV0Z50200510 Keywords : Signal transduction * Penicillin-binding protein and Ser/Thr protein kinase-associated domain * Peptidoglycan Subject RIV: CE - Biochemistry Impact factor: 3.538, year: 2011

  8. Characterization, cell-surface expression and ligand-binding properties of different truncated N-terminal extracellular domains of the ionotropic glutamate receptor subunit GluR1.

    Science.gov (United States)

    McIlhinney, R A; Molnár, E

    1996-04-01

    To identify the location of the first transmembrane segment of the GluR1 glutamate receptor subunit artificial stop codons have been introduced into the N-terminal domain at amino acid positions 442, 510, and 563, namely just before and spanning the proposed first two transmembrane regions. The resultant truncated N-terminal fragments of GluR1, termed NT1, NT2, and NT3 respectively were expressed in Cos-7 cells and their cellular distribution and cell-surface expression analysed using an N-terminal antibody to GluR1. All of the fragments were fully glycosylated and were found to be associated with cell membranes but none was secreted. Differential extraction of the cell membranes indicated that both NT1 and NT2 behave as peripheral membrane proteins. In contrast NT3, like the full subunit, has integral membrane protein properties. Furthermore only NT3 is expressed at the cell surface as determined by immunofluorescence and cell-surface biotinylation. Protease protection assays indicated that only NT3 had a cytoplasmic tail. Binding studies using the selective ligand [(3)H]alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionate ([(3)H]AMPA) demonstrated that NT3 does not bind ligand. Together these results indicate that the first transmembrane domain of the GluR1 subunit lies between residues 509 and 562, that the N-terminal domain alone cannot form a functional ligand-binding site and that this domain can be targeted to the cell surface provided that it has a transmembrane-spanning region.

  9. Usefulness of 99mTc-ECD SPECT in diseases of central nerve system

    International Nuclear Information System (INIS)

    Ono, Shimato; Yanagimoto, Shinichi; Mimura, Hiroaki

    1992-01-01

    The usefulness of a new cerebral perfusion imaging radiopharmaceutical, 99m Tc-ethyl cysteinate dimer (ECD), was clinically evaluated. The subjects of this study were 14 patients with neurological disorders including 10 patients with cerebral infarction and 4 patients with other diseases. A total of 15 examinations was performed. 99m Tc-hexamethylpropyleneamine oxime ( 99m Tc-HM-PAO) or 123 I-N-isopropyl-p-iodoamphetamine ( 123 I-IMP) SPECTs were performed simultaneously, and the findings from those examinations were compared with 99m Tc-ECD. As to the count ratio of lesions to normal area (L/N), the L/N ratio in severe ischemic patients was lower in 99m Tc-ECD than in 99m Tc-HM-PAO or 123 I-IMP. In mild ischemic patients, on the other hand, the L/N ratio was the lowest in 123 I-IMP. When the relationship between regional cerebral blood flows (rCBFs) obtained from 123 I-IMP and the values of L/N in 99m Tc-ECD or 99m Tc-HM-PAO was compared, the values of L/N in 99m Tc-ECD or 99m Tc-HM-PAO were found to have decreased linearity with increasing rCBF. In a patient showing luxury perfusion, the accumulation pattern of 99m Tc-ECD was different from that of the other two radiopharmaceuticals, and focal defect was revealed in 99m Tc-ECD SPECT. On the dynamic SPECT of 99m Tc-ECD in a patient with meningioma, the tumor showed a change from high to low perfusion with the passage of time. This finding indicated that care should be taken in the evaluation of accumulation of 99m Tc-ECD. Therefore, 99m Tc-ECD was found to be useful as a cerebral perfusion agent. In addition, as accumulation of 99m Tc-ECD might somehow reflect metabolism in some cases, further careful investigation of many cases should be carried out. (author)

  10. A polymorphism of EGFR extracellular domain is associated with progression free-survival in metastatic colorectal cancer patients receiving cetuximab-based treatment

    International Nuclear Information System (INIS)

    Gonçalves, Anthony; Turrini, Olivier; Lelong, Bernard; Viens, Patrice; Borg, Jean-Paul; Birnbaum, Daniel; Olschwang, Sylviane; Viret, Frédéric; Esteyries, Séverine; Taylor-Smedra, Brynn; Lagarde, Arnaud; Ayadi, Mounay; Monges, Geneviève; Bertucci, François; Esterni, Benjamin; Delpero, Jean-Robert

    2008-01-01

    Cetuximab, a monoclonal antibody targeting Epidermal Growth Factor Receptor (EGFR), is currently used in metastatic colorectal cancer (mCRC), but predictive factors for therapeutic response are lacking. Mutational status of KRAS and EGFR, and EGFR copy number are potential determinants of cetuximab activity. We analyzed tumor tissues from 32 EGFR-positive mCRC patients receiving cetuximab/irinotecan combination and evaluable for treatment response. EGFR copy number was quantified by fluorescence in situ hybridization (FISH). KRAS exon 1 and EGFR exons coding for extracellular regions were sequenced. Nine patients experienced an objective response (partial response) and 23 were considered as nonresponders (12 with stable disease and 11 with progressive disease). There was no EGFR amplification found, but high polysomy was noted in 2 patients, both of which were cetuximab responders. No EGFR mutations were found but a variant of exon 13 (R521K) was observed in 12 patients, 11 of which achieved objective response or stable disease. Progression-free and overall survivals were significantly better in patients with this EGFR exon 13 variant. KRAS mutations were found in 14 cases. While there was a trend for an increased KRAS mutation frequency in nonresponder patients (12 mutations out of 23, 52%) as compared to responder patients (2 out of 9, 22%), authentic tumor response or long-term disease stabilization was found in KRAS mutated patients. This preliminary study suggests that: an increase in EGFR copy number may be associated with cetuximab response but is a rare event in CRC, KRAS mutations are associated with low response rate but do not preclude any cetuximab-based combination efficacy and EGFR exon 13 variant (R521K) may predict for cetuximab benefit

  11. Toxic Compounds Analysis With High Performance Liquid Chromatography Detected By Electro Chemical Detector (Ecd)

    OpenAIRE

    Hideharu Shintaniq

    2014-01-01

    The principal area of application of high performance liquid chromatography-electrochemical detector (HPLC-ECD) has been in the analysis of naturally-occurring analytes, such as catecholamines, and pharmaceuticals in biological samples, HPLC-ECD has also applied to the analysis of pesticides and other analytes of interest to the toxicologist. In this paper, toxic area is described. In these, ammatoxins, aromatic amine, nitro-compounds, algal toxins, fungal toxins, pesticides, veterinary drug ...

  12. Evaluation of brain perfusion SPECT imaging using 99mTc-ECD

    International Nuclear Information System (INIS)

    Matsuda, Hiroshi; Kinuya, Keiko; Higashi, Sotaro; Kawasaki, Yasuhiro; Sumiya, Hisashi; Shuke, Noriyuki; Hisada, Kinichi; Yamashita, Junkoh; Yamaguchi, Nariyoshi

    1991-01-01

    Fundamental and clinical evaluation was performed on 99m Tc-ethyl cysteinate dimer ( 99m Tc-ECD), that is a new agent for brain perfusion SPECT. Radiochemical purity reaches a plateau of approximately 98% at 30 min after reconstruction and remains stable up to 24 hours later. A biodistribution study showed approximately 5% injected dose in the brain, very slow brain washout of 5.6% per hour on the average, and rapid washout from the other organ mainly through the urinary system. Brain ECD distribution was determined within 2 min postinjection and remained stable for up to 1 hour. Three hours later, slight but significant changes in brain distribution were observed, that were relative reduction of cerebral cortical activity and gray to white matter activity ratio, and relative elevation of white matter and thalamic activities. Comparative studies of ECD images with 123 I-iodoamphetamine (IMP) and 99m Tc-d, l-hexa-methylpro-pyleneamine oxime (HMPAO) images revealed that radioactivity contrast between affected and unaffected areas was less prominent in ECD than in IMP in cerebral and cerebellar cortical lesions, more prominent in ECD than in IMP in striatal and thalamic lesions, and somewhat more prominent in ECD than in HMPAO in both lesions. Imaging around 1 hour postinjection seems to be more appropriate than immediate postinjection imaging because of the clearance of the extracranial radioactivity and somewhat better radioactivity contrast between affected and unaffected areas. (author)

  13. Respiratory and cardiovascular response during electronic control device (ECD exposure in law enforcement trainees

    Directory of Open Access Journals (Sweden)

    Kirsten M. VanMeenen

    2013-04-01

    Full Text Available Objective: Law enforcement represents a large population of workers who may be exposed to electronic control devices (ECDs. Little is known about the potential effect of exposure to these devices on respiration or cardiovascular response during current discharge. Methods: Participants (N=23 were trainees exposed to 5 seconds of an ECD (Taser X26® as a component of training. Trainees were asked to volitionally inhale during exposure. Respiratory recordings involved a continuous waveform recorded throughout the session including during the exposure period. Heart rate was calculated from a continuous pulse oximetry recording. Results: The exposure period resulted in the cessation of normal breathing patterns in all participants and in particular a decrease in inspiratory activity. No significant changes in heart rate during ECD exposure were found. Conclusions: This is the first study to examine breathing patterns during ECD exposure with the resolution to detect changes over this discrete period of time. In contrast to reports suggesting respiration is unaffected by ECDs, present evidence suggests that voluntary inspiration is severely compromised. There is no evidence of cardiac disruption during ECD exposure.

  14. Mammalian ECD Protein Is a Novel Negative Regulator of the PERK Arm of the Unfolded Protein Response.

    Science.gov (United States)

    Olou, Appolinaire A; Sarkar, Aniruddha; Bele, Aditya; Gurumurthy, C B; Mir, Riyaz A; Ammons, Shalis A; Mirza, Sameer; Saleem, Irfana; Urano, Fumihiko; Band, Hamid; Band, Vimla

    2017-09-15

    Mammalian Ecdysoneless (ECD) is a highly conserved ortholog of the Drosophila Ecd gene product whose mutations impair the synthesis of Ecdysone and produce cell-autonomous survival defects, but the mechanisms by which ECD functions are largely unknown. Here we present evidence that ECD regulates the endoplasmic reticulum (ER) stress response. ER stress induction led to a reduced ECD protein level, but this effect was not seen in PKR-like ER kinase knockout (PERK-KO) or phosphodeficient eukaryotic translation initiation factor 2α (eIF2α) mouse embryonic fibroblasts (MEFs); moreover, ECD mRNA levels were increased, suggesting impaired ECD translation as the mechanism for reduced protein levels. ECD colocalizes and coimmunoprecipitates with PERK and GRP78. ECD depletion increased the levels of both phospho-PERK (p-PERK) and p-eIF2α, and these effects were enhanced upon ER stress induction. Reciprocally, overexpression of ECD led to marked decreases in p-PERK, p-eIF2α, and ATF4 levels but robust increases in GRP78 protein levels. However, GRP78 mRNA levels were unchanged, suggesting a posttranscriptional event. Knockdown of GRP78 reversed the attenuating effect of ECD overexpression on PERK signaling. Significantly, overexpression of ECD provided a survival advantage to cells upon ER stress induction. Taken together, our data demonstrate that ECD promotes survival upon ER stress by increasing GRP78 protein levels to enhance the adaptive folding protein in the ER to attenuate PERK signaling. Copyright © 2017 Olou et al.

  15. Synthesis and formulation of {sup 99m} Tc-ECD radiopharmaceutical; Sintesis y formulacion del radiofarmaco {sup 99m} Tc-ECD

    Energy Technology Data Exchange (ETDEWEB)

    Ocampo G, B E

    1998-06-01

    Nuclear medicine is a medical specialty which uses radioactive compounds (radionuclides) for diagnostic and therapeutic purposes. {sup 99m} Tc is the more common radionuclide used in many studies in nuclear medicine because its advantages: it has a photopeak of 140 KeV and a half-life of 6 hours; it can be eluted from a Molybdenum 99 generator, so radiopharmaceuticals can be prepared on site. Ethyl cysteine dimer (ECD) labelled with reduced Technetium 99m has been purposed recently as a promising radiopharmaceutical for brain perfusion imaging {sup 99m} Tc-ECD is a lipophilic neutral complex which cross the brain blood barrier and show high brain uptake. The objective of this work was synthesize and to design a freeze dried formulation for the instant preparation of {sup 99m} Tc-ECD complex useful for brain perfusion imaging. We obtained a freeze dried stable formulation for the preparation of {sup 99m} Tc-ECD kit with a radiochemical purity higher than 90 %, which fulfills with the quality control of radiopharmaceuticals. Furthermore, we developed analytic techniques for the determination of the different chemical compounds into the lyophilized kit. (Author).

  16. Expression, crystallization and preliminary crystallographic analysis of the extracellular IgV-like domain of the human natural killer cell inhibitory receptor p75/AIRM1.

    Science.gov (United States)

    Dimasi, Nazzareno; Moretta, Lorenzo; Biassoni, Roberto; Mariuzza, Roy A

    2003-10-01

    p75/AIRM1 (Siglec-7) is a sialic acid-binding Ig-like lectin recently identified as an inhibitory receptor on natural killer cells. The expression, in vitro folding, circular-dichroism spectroscopy, crystallization and preliminary X-ray characterization of the Ig-V like domain of p75/AIRM1 are reported. X-ray data were collected from a single crystal at 100 K, with a maximum useful diffraction pattern extending to 1.45 A resolution on a synchrotron source. The crystal belongs to a primitive monoclinic space group, with unit-cell parameters a = 32.65, b = 49.72, c = 39.79 A, alpha = gamma = 90, beta = 113 degrees. The systematic absences indicate that the space group is P2(1). Assuming one molecule per asymmetric unit, V(M) (the Matthews coefficient) was calculated to be 1.879 A(3) Da(-1) and the solvent content was estimated to be 32.01%.

  17. Infection of CD4+ T lymphocytes by the human T cell leukemia virus type 1 is mediated by the glucose transporter GLUT-1: Evidence using antibodies specific to the receptor's large extracellular domain

    International Nuclear Information System (INIS)

    Jin, Qingwen; Agrawal, Lokesh; VanHorn-Ali, Zainab; Alkhatib, Ghalib

    2006-01-01

    To analyze HTLV-1 cytotropism, we developed a highly sensitive vaccinia virus-based assay measuring activation of a reporter gene upon fusion of two distinct cell populations. We used this system in a functional cDNA screening to isolate and confirm that the glucose transporter protein 1 (GLUT-1) is a receptor for HTLV-1. GLUT-1 is a ubiquitously expressed plasma membrane glycoprotein with 12 transmembrane domains and 6 extracellular loops (ECL). We demonstrate for the first time that peptide antibodies (GLUT-IgY) raised in chicken to the large extracellular loop (ECL1) detect GLUT-1 at the cell surface and inhibit envelope (Env)-mediated fusion and infection. Efficient GLUT-IgY staining was detected with peripheral blood CD4 + lymphocytes purified by positive selection. Further, GLUT-IgY caused efficient inhibition of Env-mediated fusion and infection of CD4 + T and significantly lower inhibition of CD8 + T lymphocytes. The specificity of GLUT-IgY antibodies to GLUT-1 was demonstrated by ECL1 peptide competition studies. Grafting ECL1 of GLUT-1 onto the receptor-negative GLUT-3 conferred significant receptor activity. In contrast, grafting ECL1 of GLUT-3 onto GLUT-1 resulted in a significant loss of the receptor activity. The ECL1-mediated receptor activity was efficiently blocked with four different human monoclonal antibody (HMab) to HTLV-1 Env. The ECL1-derived peptide blocked HTLV-1 Env-mediated fusion with several nonhuman mammalian cell lines. The results demonstrate the utilization of cell surface GLUT-1 in HTLV-1 infection of CD4 + T lymphocytes and implicate a critical role for the ECL1 region in viral tropism

  18. Receptor complementation and mutagenesis reveal SR-BI as an essential HCV entry factor and functionally imply its intra- and extra-cellular domains.

    Directory of Open Access Journals (Sweden)

    Marlène Dreux

    2009-02-01

    Full Text Available HCV entry into cells is a multi-step and slow process. It is believed that the initial capture of HCV particles by glycosaminoglycans and/or lipoprotein receptors is followed by coordinated interactions with the scavenger receptor class B type I (SR-BI, a major receptor of high-density lipoprotein (HDL, the CD81 tetraspanin, and the tight junction protein Claudin-1, ultimately leading to uptake and cellular penetration of HCV via low-pH endosomes. Several reports have indicated that HDL promotes HCV entry through interaction with SR-BI. This pathway remains largely elusive, although it was shown that HDL neither associates with HCV particles nor modulates HCV binding to SR-BI. In contrast to CD81 and Claudin-1, the importance of SR-BI has only been addressed indirectly because of lack of cells in which functional complementation assays with mutant receptors could be performed. Here we identified for the first time two cell types that supported HCVpp and HCVcc entry upon ectopic SR-BI expression. Remarkably, the undetectable expression of SR-BI in rat hepatoma cells allowed unambiguous investigation of human SR-BI functions during HCV entry. By expressing different SR-BI mutants in either cell line, our results revealed features of SR-BI intracellular domains that influence HCV infectivity without affecting receptor binding and stimulation of HCV entry induced by HDL/SR-BI interaction. Conversely, we identified positions of SR-BI ectodomain that, by altering HCV binding, inhibit entry. Finally, we characterized alternative ectodomain determinants that, by reducing SR-BI cholesterol uptake and efflux functions, abolish HDL-mediated infection-enhancement. Altogether, we demonstrate that SR-BI is an essential HCV entry factor. Moreover, our results highlight specific SR-BI determinants required during HCV entry and physiological lipid transfer functions hijacked by HCV to favor infection.

  19. [sup 99m]Tc-ECD SPECT study in dementia and aphasia

    Energy Technology Data Exchange (ETDEWEB)

    Kuwabara, Yasuo; Ichiya, Yuichi; Otsuka, Makoto; Sasaki, Masayuki; Akashi, Yuko; Fukumura, Toshimitsu; Yoshida, Tsuyoshi; Masuda, Kouji; Ichimiya, Atsushi (Kyushu Univ., Fukuoka (Japan). Faculty of Medicine)

    1993-09-01

    We studied clinical significance of [sup 99m]Tc-L,L,-ethyl cysteine dimer ([sup 99m]Tc-ECD) SPECT study in dementia and aphasia, and compared it with [sup 99m]Tc-HMPAO SPECT study. The subjects consisted of 13 patients, including 10 patients with dementia and 3 patients with aphasia. Hypoperfusion areas were detected in 5 out of 10 patients with dementia and 2 out of 3 patients with aphasia in [sup 99m]Tc-ECD SPECT, and in 4 out of 10 patients with dementia and all of 3 patients with aphasia in [sup 99m]Tc-HMPAO SPECT. The count rate ratios in [sup 99m]Tc-ECD and [sup 99m]Tc-HMPAO SPECT were correlated well with each other, and the contrast of the [sup 99m]Tc-ECD SPECT image was equivalent or slightly higher as compared with [sup 99m]Tc-HMPAO. Therefore, [sup 99m]Tc-ECD SPECT study was considered to be useful for the evaluation of cerebral perfusion in dementia and aphasia. (author).

  20. 99mTc-ECD SPECT study in dementia and aphasia

    International Nuclear Information System (INIS)

    Kuwabara, Yasuo; Ichiya, Yuichi; Otsuka, Makoto; Sasaki, Masayuki; Akashi, Yuko; Fukumura, Toshimitsu; Yoshida, Tsuyoshi; Masuda, Kouji; Ichimiya, Atsushi

    1993-01-01

    We studied clinical significance of 99m Tc-L,L,-ethyl cysteine dimer ( 99m Tc-ECD) SPECT study in dementia and aphasia, and compared it with 99m Tc-HMPAO SPECT study. The subjects consisted of 13 patients, including 10 patients with dementia and 3 patients with aphasia. Hypoperfusion areas were detected in 5 out of 10 patients with dementia and 2 out of 3 patients with aphasia in 99m Tc-ECD SPECT, and in 4 out of 10 patients with dementia and all of 3 patients with aphasia in 99m Tc-HMPAO SPECT. The count rate ratios in 99m Tc-ECD and 99m Tc-HMPAO SPECT were correlated well with each other, and the contrast of the 99m Tc-ECD SPECT image was equivalent or slightly higher as compared with 99m Tc-HMPAO. Therefore, 99m Tc-ECD SPECT study was considered to be useful for the evaluation of cerebral perfusion in dementia and aphasia. (author)

  1. Identification of an evolutionarily conserved extracellular threonine residue critical for surface expression and its potential coupling of adjacent voltage-sensing and gating domains in voltage-gated potassium channels.

    Science.gov (United States)

    Mckeown, Lynn; Burnham, Matthew P; Hodson, Charlotte; Jones, Owen T

    2008-10-31

    The dynamic expression of voltage-gated potassium channels (Kvs) at the cell surface is a fundamental factor controlling membrane excitability. In exploring possible mechanisms controlling Kv surface expression, we identified a region in the extracellular linker between the first and second of the six (S1-S6) transmembrane-spanning domains of the Kv1.4 channel, which we hypothesized to be critical for its biogenesis. Using immunofluorescence microscopy, flow cytometry, patch clamp electrophysiology, and mutagenesis, we identified a single threonine residue at position 330 within the Kv1.4 S1-S2 linker that is absolutely required for cell surface expression. Mutation of Thr-330 to an alanine, aspartate, or lysine prevented surface expression. However, surface expression occurred upon co-expression of mutant and wild type Kv1.4 subunits or mutation of Thr-330 to a serine. Mutation of the corresponding residue (Thr-211) in Kv3.1 to alanine also caused intracellular retention, suggesting that the conserved threonine plays a generalized role in surface expression. In support of this idea, sequence comparisons showed conservation of the critical threonine in all Kv families and in organisms across the evolutionary spectrum. Based upon the Kv1.2 crystal structure, further mutagenesis, and the partial restoration of surface expression in an electrostatic T330K bridging mutant, we suggest that Thr-330 hydrogen bonds to equally conserved outer pore residues, which may include a glutamate at position 502 that is also critical for surface expression. We propose that Thr-330 serves to interlock the voltage-sensing and gating domains of adjacent monomers, thereby yielding a structure competent for the surface expression of functional tetramers.

  2. Role of 99mTc-ECD SPECT in the Management of Children with Craniosynostosis

    Science.gov (United States)

    Barik, Mayadhar; Bajpai, Minu; Das, Rashmi Ranajn; Malhotra, Arun; Panda, Shasanka Shekhar; Sahoo, Manas Kumar; Dwivedi, Sadanand

    2014-01-01

    Purpose of the Report. There is a paucity of data on correlation of various imaging modalities with clinical findings in craniosynostosis. Moreover, no study has specifically reported the role of 99mTc-ECD SPECT in a large number of subjects with craniosynostosis. Materials and Methods. We prospectively analyzed a cohort of 85 patients with craniosynostosis from year 2007 to 2012. All patients underwent evaluation with 99mTc-ECD SPECT and the results were correlated with radiological and surgical findings. Results. 99mTc-ECD SPECT revealed regional perfusion abnormalities in the cerebral hemisphere corresponding to the fused sutures preoperatively that disappeared postoperatively in all the cases. Corresponding to this, the mean mental performance quotient (MPQ) increased significantly (P < 0.05) postoperatively only in those children with absent perfusion defect postoperatively. Conclusions. Our study suggests that early surgery and release of craniosynostosis in patients with preoperative perfusion defects (absent on 99mTc-ECD SPECT study) are beneficial, as theylead to improved MPQ after surgery. PMID:24987670

  3. Role of 99mTc-ECD SPECT in the Management of Children with Craniosynostosis

    Directory of Open Access Journals (Sweden)

    Mayadhar Barik

    2014-01-01

    Full Text Available Purpose of the Report. There is a paucity of data on correlation of various imaging modalities with clinical findings in craniosynostosis. Moreover, no study has specifically reported the role of Tc99m-ECD SPECT in a large number of subjects with craniosynostosis. Materials and Methods. We prospectively analyzed a cohort of 85 patients with craniosynostosis from year 2007 to 2012. All patients underwent evaluation with Tc99m-ECD SPECT and the results were correlated with radiological and surgical findings. Results. Tc99m-ECD SPECT revealed regional perfusion abnormalities in the cerebral hemisphere corresponding to the fused sutures preoperatively that disappeared postoperatively in all the cases. Corresponding to this, the mean mental performance quotient (MPQ increased significantly P<0.05 postoperatively only in those children with absent perfusion defect postoperatively. Conclusions. Our study suggests that early surgery and release of craniosynostosis in patients with preoperative perfusion defects (absent on Tc99m-ECD SPECT study are beneficial, as theylead to improved MPQ after surgery.

  4. 99mTc-ECD dynamic SPECT in 'luxury perfusion' of subacute stroke

    International Nuclear Information System (INIS)

    Ogasawara, Kuniaki; Fujiwara, Satoru; Yoshimoto, Takashi.

    1995-01-01

    To evaluate the cerebral pharmacokinetics of 99m Tc-ethyl cysteinate dimer ( 99m Tc-ECD) at blood flow levels beyond the normal range, we investigated 'luxury perfusion' in subacute stroke, ictal hyperperfusion in epilepsy and post-decompressive hyperemia in head trauma. All 7 patients showed a hyperactive area on SPECT studies using 99m Tc-HM-PAO. 99m Tc-ECD static image demonstrated a hyperactive area in both epilepsy and head trauma, and a hypoactive area in 'luxury perfusion.' On the dynamic SPECT of 99m Tc-ECD in both epilepsy and head trauma, brain distribution of the tracer was determined within 2 min postinjection and remained stable for up to 1 hour; however, 'luxury perfusion' area showed a change from initial hyperactivity to late hypoactivity with the passage of time. The time activity curve in 'luxury perfusion' area demonstrated a steep decrease of counts/pixel for up to 4-5 minutes postinjection, and a moderate decrease in the following phase. The early wash-out mechanism of 99m Tc-ECD from 'luxury perfusion' area can be described by a biexponential function including an initial steep decrease representing the rapid loss of the lipophilic complexes which were not metabolized in injured brain tissue. (author)

  5. Mismatched uptake of Tc-99m-ECD and Tc-99m-HMPAO in subacute cerebral infarction: Tc-99m-ECD for viability and Tc-99m-HMPAO for flow restoration

    Energy Technology Data Exchange (ETDEWEB)

    Lee, D. S.; Hyun, I. Y.; Kim, S. K. [College of Medicine, Seoul National Univ., Seoul (Korea, Republic of)] [and others

    1997-07-01

    Tc-99m-HMPAO reflects tissue perfusion but Tc-99m-ECD uptake is affected by tissue viability in addition to tissue perfusion which the varied state of cellular retention of Tc-99m-ECD reflects. Luxuriously perfused area on Tc-99m-HMPAO SPECT implies that this cortex was already reperfused either spontaneously or after thrombolysis and that accompanied paralysis of vascular reactivity in those zones warms progressive deterioration. We tried to find out if we can use sequential Tc-99m-ECD/Tc-99m-HMPAO SPECT to reveal cortical perfusion and severity and range of risky areas of cerbral cortex despite reperfusion in sub-acute infarction. In 13 patients (M ; F =7 : 6, mean age 57 (range: 26-84)) with cortical (n=12) and basal ganglia infarction (1), we performed sequential Tc-99m-ECD/Tc-99m-HMPAO SPECT at the same position. At first, 555 MBq of Tc-99m-ECD was injected and imaged and then 1110 MBq of Tc-99m-HMPAO was injected again and imaged with the patients in situ, and the first image (Tc-99m-ECD) and the subtracted image (2nd- 1st : Tc-99m-HMPAO) were compared slice by slice. Study was done from 3 days to 31 days (16{+-}9) after ictus. Tc-99m-ECD uptake was always less than or equal to Tc-99m-HMPAO uptake at the lesion in all cases. Luxury perfusion was prominent in four patients. Mismatched uptake was found in 10 patients. Severity of mismatch showed diverse spectrum and was ranged from total middle cerebral artery territory (1 case) to peripheral thin zones around infarction (2 cases). The other 7 showed intermediate amount of tissues with mismatch , i.e., Tc-99m-ECD defects where Tc-99m-HMPAO uptake is in part increased, normal or decreased. Upon discharge, patients having more uptake with Tc-99m-ECD predicted improvement. Patients having mismatched uptake went dichotomous way. In conclusion, Tc-99m-ECD/Tc-99m-HMPAO sequential SPECT is feasible and reveal both tissue perfusion (Tc-99m-HMPAO ) and discrepant Tc-99m-ECD uptake probably reflecting viability in acute

  6. Biomimetic synthesis and HPLC–ECD analysis of the isomers of dracocephins A and B

    Directory of Open Access Journals (Sweden)

    Viktor Ilkei

    2016-11-01

    Full Text Available Starting from racemic naringenin ((±-1, a mixture of dracocephin A stereoisomers 6-(2”-pyrrolidinone-5”-ylnaringenin (±-2a–d and its regioisomer, dracocephin B 8-(2”-pyrrolidinone-5”-ylnaringenin (±-3a–d originally isolated from Dracocephalum rupestre, have been synthesized in a one-pot reaction. The separation of 2a–d and 3a–d was achieved by preparative HPLC. The four stereoisomers of each natural product were separated by analytical chiral HPLC and their absolute configuration was studied by the combination of HPLC–ECD measurements and TDDFT–ECD calculations. The synthesized flavonoid alkaloids were further characterized by physicochemical and in vitro pharmacological studies.

  7. Comparison of Tc-99m ECD brain SPECT between patients with delayed development and cerebral palsy

    International Nuclear Information System (INIS)

    Cho, I.; Chun, K.; Won, K.; Lee, H.; Jang, S.; Lee, J.

    2002-01-01

    Purpose: In previous study, thalamic or cerebellar hypoperfusion were reported in patients with cerebral palsy. This study was performed to evaluate cerebral perfusion abnormalities using Tc-99m ECD brain SPECT in patients with delayed motor development. Methods: Nineteen patients (9 boys, 10 girls, mean age 25.5 months) with delayed development underwent brain SPECT after injection of 185∼370 MBq of Tc-99m ECD. The imaging was obtained between 30 minutes and 1hr after injection. The patients were divided clinically as follows, patients with delayed development (n=5) and patients with cerebral palsy (n=14) who has delayed development and abnormal movement. The clinical subtypes of cerebral palsy were spastic quadriplegia (n=5), spastic diplegia (n=6) and spastic hemiplegia (n=3). In each group, decrease of cerebral perfusion was evaluated visually as mild, moderate and severe and quantitation of cerebral perfusion after Lassen's correction was also obtained. Results: SPECT findings showed normal or mildly decreased thalamic perfusion in patients with delayed development and severe decrease of thalamic or cerebellar perfusion in patients with spastic quadriplegia. In patients with spastic diplegia, mild decrease of perfusion was observed in thalamus. In quantified data, thalamic perfusion was lowest in patients with spastic quadriplegia and highest in patients with delayed development, but there were no statistically significant differences. Conclusion: Brain SPECT with Tc-99m ECD has a role in the detection of perfusion abnormalities in patients with delayed development and cerebral palsy

  8. Early and delayed Tc-99m ECD brain SPECT in SLE patients with CNS involvement

    International Nuclear Information System (INIS)

    Kikukawa, Kaoru; Toyama, Hiroshi; Katayama, Masao

    2000-01-01

    We compared early and delayed Tc-99m ECD SPECT scans in 32 SLE patients (Group 1, definite neuropsychiatric disorders; Group 2, minor neurologic symptoms or normal) with those of normal controls by visual inspection and semi-quantitative evaluation. With visual interpretation, 13 out of 14 patients in Group 1 (93%) and 7 out of 18 patients in Group 2 (39%) had diffuse uneven decrease in early scans. Seven patients in Group 2 (39%) who had normal early scans demonstrated focal decrease in the medial frontal lobe in delayed scans. With cerebral region to cerebellar ratios, in early scans, the medial frontal lobe in Group 1 and Group 2 was significantly lower than in normal controls, and lateral frontal lobe and occipital lobes in Group 1 were significantly lower than in normal controls. Nevertheless, in delayed scans, every cortical region except for the parietal lode in Groups 1 and 2 was significantly lower than in normal controls. The retention rates in all regions in SLE patients were significantly lower than in normal controls. No case showed SPECT improvement on follow-up studies in either group in spite of clinical improvement. Delayed Tc-99m ECD brain SPECT of high sensitivity might be useful in detecting CNS involvement. Although the SPECT findings did not correlate with the neuropsychiatric symptoms, early and delayed Tc-99m ECD SPECT seems to provide useful objective diagnostic information in SLE patients. (author)

  9. Incorporation of the KERN ECDS-PC software into a project oriented software environment

    International Nuclear Information System (INIS)

    Oren, W.; Pushor, R.; Ruland, R.

    1986-11-01

    The Stanford Linear Accelerator Center (SLAC) is in the process of building a new particle collider, the Stanford Linear Collider (SLC). The tunnel which houses the SLC is about 3 km long and contains approximately 1000 magnets. Besides a very precise absolute positioning of these magnets, the alignment of adjacent magnet ends is of particular importance to the success of the whole project. Because of this and the limited time frame, a survey method which was not only reliable and self-checking but also fast had to be developed. Therefore, the concept of MAS (Magnet Alignment System) was developed. This system utilizes the on-line data collection and the rigorous least-squares bundle adjustment of the KERN ECDS-PC system to fulfill these requirements. The ECDS software is embedded in a project tailored software system with modules which take care of: fixture and magnet calibration corrections, the calculation of ideal coordinates and their comparison to measured coordinates, the translation of detected misalignments into the coordinate system of the mechanical adjustments and the control of the adjustments with on-line electronic dial-gauges. This paper gives a brief introduction to the SLC project and some of the survey problems which are unique to this machine. The basic ideas of the KERN ECDS-PC system are explained and a discussion of the practical aspects, such as targeting and set-ups, are given. MAS and its modules are explained in detail

  10. The Impact of Seed Layer Structure on the Recrystallization of ECD Cu and its Alloys

    Science.gov (United States)

    O'Brien, Brendan B.

    Despite the significant improvements originally offered by the use of Cu over Al as the interconnect material for semiconductor devices, the continued down-scaling of interconnects has presented significant challenges for semiconductor engineers. As the metal line widths shrink, both the conductivity and reliability of lines decrease due to a stubbornly fine-grained microstructure in narrow lines. Understanding microstructural transformation of the ECD Cu in narrow features which leads to this polygranular microstructure is the first focus of this dissertation. As in the case of Cu films, the underlying seed layer strongly influences progress of transformation. Unlike films, however, the seed layer is not homogenous in patterned substrates, but differs according to the size of the trench and the location within the trench (field, bottom, and sidewall). Based on these findings, and the known influence of texture on the transformation of ECD Cu, a rapid trench initiated transformation process was posited for narrow interconnect lines. Time-resolved TEM observation of the ECD Cu in 48 nm lines during the transformation process confirmed the hypothesis. In fact, the TEM images revealed that the transformation was even faster than anticipated, and that the microstructure of the Cu inside the lines was stagnant after a mere 1.5 hours at room temperature. Studies of the transformation at elevated temperatures found that, despite anneals at 250°C for up to an hour, the grain size distribution for the Cu in narrow lines for all times converged, whether annealed at room temperature or 250°C. These data suggest that process was being driven by the 'consumable' internal energy stored in the as-plated microstructure. This is different than the transformation of the overburden, which is driven by a competition between surface energy and internal stress buildup due to film densification and relief due to the secondary growth of a 200 texture component. Based on these findings

  11. Extracellular DNA metabolism in Haloferax volcanii

    Directory of Open Access Journals (Sweden)

    Scott eChimileski

    2014-02-01

    Full Text Available Extracellular DNA is found in all environments and is a dynamic component of the micro-bial ecosystem. Microbial cells produce and interact with extracellular DNA through many endogenous mechanisms. Extracellular DNA is processed and internalized for use as genetic information and as a major source of macronutrients, and plays several key roles within prokaryotic biofilms. Hypersaline sites contain some of the highest extracellular DNA con-centrations measured in nature–a potential rich source of carbon, nitrogen and phosphorus for halophilic microorganisms. We conducted DNA growth studies for the halophilic archaeon Haloferax volcanii DS2 and show that this model Halobacteriales strain is capable of using exogenous double-stranded DNA as a nutrient. Further experiments with varying medium composition, DNA concentration and DNA types revealed that DNA is utilized primarily as a phosphorus source, that growth on DNA is concentration-dependent and that DNA isolated from different sources is metabolized selectively, with a bias against highly divergent methylated DNA sources. Additionally, fluorescence microscopy experiments showed that labeled DNA colocalized with Haloferax volcanii cells. The gene Hvo_1477 was also identified using a comparative genomic approach as a factor likely to be involved in extracellular DNA processing at the cell surface, and deletion of Hvo_1477 created an H. volcanii strain deficient in its ability to grow on extracellular DNA. Widespread distribution of Hvo_1477 homologs in archaea suggests metabolism of extracellular DNA may be of broad ecological and physiological relevance in this domain of life.

  12. Evaluation of {sup 99m}Tc-ECD SPECT for the detection of brain tumor. Comparison with {sup 201}Tl SPECT

    Energy Technology Data Exchange (ETDEWEB)

    Oshima, Motoo; Sasaki, Yasushi; Kikuchi, Yoshirou; Kaminaga, Tatsuro; Furui, Shigeru [Teikyo Univ., Tokyo (Japan). Faculty of Medicine; Konoeda, Kouichi; Karigome, Masato; Yoshida, Katsuhiko

    1997-01-01

    For the evaluation of brain tumor (n=15), we performed both dynamic and static {sup 99m}Tc-ECD (ECD) SPECT studies. {sup 201}Tl SPECT was also used for comparison with the results of ECD SPECT. Dynamic ECD SPECT was obtained following the injection of 600 MBq of ECD. Five min after the injection of ECD, static ECD SPECT was performed. {sup 201}Tl SPECT was obtained 10 min after the injection of 74 MBq. Abnormal uptake was recognized in 7 of 15 tumors with dynamic ECD; 5 of 7 meningiomas, 1 of 1 glioblastoma and 1 of 1 astrocytoma. However, no abnormal uptake was seen in 3 of 3 benign tumors (1 low grade astrocytoma, 1 hemangioma, 1 craniopharyngioma) and in 2 of 2 brain metastases. In contrast abnormal uptake was seen in 11 of 15 tumors with {sup 201}Tl; 7 of 7 meningiomas, 2 of 2 brain metastases, 1 of 1 glioblastoma and 1 of 1 craniopharyngioma. No abnormal uptake was seen in 3 of 3 benign tumors (1 hemangioma and 2 low grade astrocytomas). Equivocal uptake was seen in 1 low grade astrocytoma with dynamic ECD and {sup 201}Tl. The mechanism of the accumulation of dynamic ECD to brain tumor is unclear. However, it may reflect not only blood flow, but also metabolism. (author)

  13. Value of 99mTc-ECD SPET for the diagnosis of dementia with Lewy bodies

    International Nuclear Information System (INIS)

    Pasquier, Jacques; Brenot-Rossi, Isabelle; Hassan-Sebbag, Nathalie; Sauvan, Richard; Michel, Bernard F.; Gastaut, Jean Louis

    2002-01-01

    Despite improved diagnostic accuracy, differentiation of dementia with Lewy bodies (DLB) and Alzheimer's disease (AD) on the basis of clinical findings remains problematic. The purpose of this retrospective study was to evaluate the utility of technetium-99m ethyl cysteinate dimer (ECD) single-photon emission tomography (SPET) as a potential tool for the diagnosis of DLB and discrimination from AD. Cerebral perfusion patterns detected by 99m Tc-ECD SPET were compared in patients presenting with a probable diagnosis of DLB (n=34) or AD (n=28). Tracer distribution was quantified using the region of interest technique in eight symmetrical paired zones and expressed as a perfusion index (ratio of mean uptake in a brain region to that in the cerebellum). Comparison of findings in the DLB and AD groups demonstrated significant differences in mean perfusion indexes in the right occipital region (P=0.004), left occipital region (P=0.005) and left medial temporal region (P=0.013). Mean perfusion indexes in the right and left occipital regions were lower in DLB than in AD patients. Conversely, the mean perfusion index in the left medial temporal region was lower in AD than in DLB patients. DLB was correctly identified in 22 patients (sensitivity, 65%) while AD was correctly identified in 20 patients (specificity, 71%). In the DLB group, right and left occipital perfusion indexes were 0.95 or more in all eight non-hallucinating patients, and bilateral occipital hypoperfusion was observed in 15 of the 26 patients with visual hallucinations (57.7%). To our knowledge, this is the first study in which 99m Tc-ECD SPET has been used exclusively for the diagnosis of DLB. The results suggest that brain perfusion scintigraphy could be helpful in distinguishing DLB from AD if diagnosis based on clinical criteria alone is difficult. The findings also support a link between visual hallucinations and structural/functional changes in the occipital region in DLB patients. (orig.)

  14. Study of organohalogen contaminants in yogurt by NAA and GC-ECD

    International Nuclear Information System (INIS)

    Zhang, H.; Chai, Z.F.; Shenzen Univ.; Sun, H.B.; Zhang, J.L.; Ouyang, H.; Xin, L.; Chuai, Y.D.

    2007-01-01

    The concentrations and distributions of total halogen (TX), extractable organohalogen (EOX) and extractable persistent organohalogen (EPOX) were determined in 20 kinds of yogurt specimens collected from Chinese supermarkets using neutron activation analysis (NAA) and gas chromatography equipped with a 63 Ni electron capture detector (GC-ECD). The results indicated that the halogens in yogurt mainly existed as non-extractable organohalogen compounds. About 25-30% of EOX was EPOX. EOCl and EPOCl were the main organohalogen species in yogurt. The average concentration of the identified organochlorine, such as organochlorine pesticides (OCPs) and polychlorinated biphenyls (PCBs), was below 4% of EPOCl. (author)

  15. Structure dependent antioxidant capacity of phlorotannins from Icelandic Fucus vesiculosus by UHPLC-DAD-ECD-QTOFMS

    DEFF Research Database (Denmark)

    Hermund, Ditte Baun; Plaza, Merichel; Turner, Charlotta

    2018-01-01

    Brown algae are rich in polyphenolic compounds, phlorotannins, which have been found to possess high in vitro antioxidant capacity, especially DPPH radical scavenging activity, due to the high number of hydroxyl groups. Whereas, the overall antioxidant capacity of brown algae extracts has been...... was determined by an on-line method using liquid chromatography and an electrochemical detector followed by quadrupole Time of Flight mass spectrometry (UHPLC-DAD-ECD-QTOFMS). Tentative structural elucidation of 13 phlorotannin isomers from EAF was obtained by LC-DAD-QTOFMS, ranging from 374 to 870 Da. On...

  16. Experimental Setup for Diamond Grinding Using Electrochemical InProcess Controlled Dressing (ECD of Grinding Wheel

    Directory of Open Access Journals (Sweden)

    M. A. Shavva

    2014-01-01

    Full Text Available The most effective method for finish machining of hard-metals and alloys is to use the diamond grinding wheels for grinding. An application of diamond wheels significantly increases the employee output, reduces costs, and raises manufacturing efficiency with achieving the high performance properties of treated surfaces.During grinding a working surface of diamond wheel wears out. It adversely affects the cutting capability of the diamond grains, and depending on the grinding conditions can occur through different mechanisms. Wear of diamond wheel causes distortion of its shape and reduces cutting properties. However, dressing of diamond wheels is a complicated and time-consuming operation in terms of manufacturing technique.Methods to make dressing of diamond grinding wheel have different types of classification. Classification of dressing methods by the type of energy used is as follows: mechanical, chemical, electrophysical, electromechanical, and electrochemical. All these methods have their advantages and disadvantages.Electrochemical method of dressing is the most productive and efficient. Electrochemical method comprises anode-mechanical dressing and electrochemical (electrolytic one. The paper presents the electrochemical in-process dressing (ECD and the electrolytic in-process dressing (ELID.The source of energy, grinding a wheel with metal bond, and an electrode are necessary for providing ELID. The ELID consists of several stages. The first stage is preliminary electrolytic dressing of diamond wheel. The electrolyte is placed into the gap between the wheel and electrode. The bond of the wheel is oxidized. An insulating layer is formed. It reduces an electrical conductivity of the wheel and controls consumption of diamond grains, as well as polishes the surface of the work piece. Further, the insulating layer is destroyed. The cycle of dressing begins anew.The ECD proceeds in the same way as ELID. However during the ECD-process there

  17. Tc-99m ECD brain SPECT in MELAS syndrome and mitochondrial myopathy: comparison with MR findings

    International Nuclear Information System (INIS)

    Park, Sang Joon; Ryu, Young Hoon; Jeon, Tae Joo; Kim, Jai Keun; Nam, Ji Eun; Yoon, Pyeong Ho; Yoon, Choon Sik; Lee, Jong Doo

    1998-01-01

    We evaluated brain perfusion SPECT findings of MELAS syndrome and mitochondrial myopathy in correlation with MR imaging in search of specific imaging features. Subjects were five patients (four females and one male; age range, 1 to 25 year) who presented with repeated stroke like episodes, seizures or developmental delay or asymptomatic but had elevated lactic acid in CSF and serum. Conventional non-contrast MR imaging and Tc-99m-ethyl cysteinate dimer (ECD) brain perfusion SPECT were performed and imaging features were analyzed. MRI demonstrated increased T2 signal intensities in the affected areas of gray and white matters mainly in the parietal (4/5) and occipital lobes (4/5) and in the basal ganglia (1/5), which were not restricted to a specific vascular territory. SPECT demonstrated decreased perfusion in the corresponding regions of MRI lesions. In addition, there were perfusion defects in parietal (1 patient), temporal (2), and frontal (1) lobes and basal ganglia (1) and thalami (2). In a patient with mitochondrial myopathy who had normal MRI, decreased perfusion was noted in left parietal area and bilateral thalami. Tc-99m ECD SPECT imaging in patients with MELAS syndrome and mitochondrial myopathy showed hypoperfusion of parieto-occipital cortex, basal ganglia, thalamus and temporal cortex, which were not restricted to a specific vascular territory. There were no specific imaging features on SPECT. The significance of abnormal perfusion on SPECT without corresponding MR abnormalities needs to be evaluated further in larger number of patients

  18. The Extracellular Protein Factor Epf from Streptococcus pyogenes Is a Cell Surface Adhesin That Binds to Cells through an N-terminal Domain Containing a Carbohydrate-binding Module*

    Science.gov (United States)

    Linke, Christian; Siemens, Nikolai; Oehmcke, Sonja; Radjainia, Mazdak; Law, Ruby H. P.; Whisstock, James C.; Baker, Edward N.; Kreikemeyer, Bernd

    2012-01-01

    Streptococcus pyogenes is an exclusively human pathogen. Streptococcal attachment to and entry into epithelial cells is a prerequisite for a successful infection of the human host and requires adhesins. Here, we demonstrate that the multidomain protein Epf from S. pyogenes serotype M49 is a streptococcal adhesin. An epf-deficient mutant showed significantly decreased adhesion to and internalization into human keratinocytes. Cell adhesion is mediated by the N-terminal domain of Epf (EpfN) and increased by the human plasma protein plasminogen. The crystal structure of EpfN, solved at 1.6 Å resolution, shows that it consists of two subdomains: a carbohydrate-binding module and a fibronectin type III domain. Both fold types commonly participate in ligand receptor and protein-protein interactions. EpfN is followed by 18 repeats of a domain classified as DUF1542 (domain of unknown function 1542) and a C-terminal cell wall sorting signal. The DUF1542 repeats are not involved in adhesion, but biophysical studies show they are predominantly α-helical and form a fiber-like stalk of tandem DUF1542 domains. Epf thus conforms with the widespread family of adhesins known as MSCRAMMs (microbial surface components recognizing adhesive matrix molecules), in which a cell wall-attached stalk enables long range interactions via its adhesive N-terminal domain. PMID:22977243

  19. The extracellular protein factor Epf from Streptococcus pyogenes is a cell surface adhesin that binds to cells through an N-terminal domain containing a carbohydrate-binding module.

    Science.gov (United States)

    Linke, Christian; Siemens, Nikolai; Oehmcke, Sonja; Radjainia, Mazdak; Law, Ruby H P; Whisstock, James C; Baker, Edward N; Kreikemeyer, Bernd

    2012-11-02

    Streptococcus pyogenes is an exclusively human pathogen. Streptococcal attachment to and entry into epithelial cells is a prerequisite for a successful infection of the human host and requires adhesins. Here, we demonstrate that the multidomain protein Epf from S. pyogenes serotype M49 is a streptococcal adhesin. An epf-deficient mutant showed significantly decreased adhesion to and internalization into human keratinocytes. Cell adhesion is mediated by the N-terminal domain of Epf (EpfN) and increased by the human plasma protein plasminogen. The crystal structure of EpfN, solved at 1.6 Å resolution, shows that it consists of two subdomains: a carbohydrate-binding module and a fibronectin type III domain. Both fold types commonly participate in ligand receptor and protein-protein interactions. EpfN is followed by 18 repeats of a domain classified as DUF1542 (domain of unknown function 1542) and a C-terminal cell wall sorting signal. The DUF1542 repeats are not involved in adhesion, but biophysical studies show they are predominantly α-helical and form a fiber-like stalk of tandem DUF1542 domains. Epf thus conforms with the widespread family of adhesins known as MSCRAMMs (microbial surface components recognizing adhesive matrix molecules), in which a cell wall-attached stalk enables long range interactions via its adhesive N-terminal domain.

  20. {sup 99m}Tc-ECD: Comparison of radiochemical purity evaluation techniques; {sup 99m}Tc-ECD: Comparaison de techniques d`evaluation de la purete radiochimique

    Energy Technology Data Exchange (ETDEWEB)

    Dumont, A.; De Beco, V.; Ait Ben Ali, S.; Goudou-Sinha, C.; Izembart, M.; Jourdain, J.R.; Lemercier, V.; Linsker, S.; Lours, S.; Moati, F.; Pajard, D.; Piketty, M.L.; Rizzo, N.; Schlageter, M.H.; Moretti, J.L. [Groupe de travail `Radiopharmaceutiques - Ile de France` (France)

    1997-12-31

    The aim of this study was to validate a testing method for the radiochemical purity (RCP) of the preparations of {sup 99m}Tc - ECD that is to be reproducible and easy to realize in services of Nuclear Medicine. After a review of literature, four thin-layer chromatography techniques allowing to evidence the TcO{sub 4}{sup -} were evaluated: no.1 - Papier Whatman 31ET / ethyl acetate; no.2 - Papier Whatman 3MM chr / ethyl acetate; no.3 - ITLC Silica gel / ethyl acetate; no.4 - Baker Flex silica gel aluminium oxide IB-F / ethyl acetate (the method proposed by the laboratory). The technique no.1 has presented a bad reproducibility, as well as percentages of RCP very different from those obtained by the other techniques. The techniques no.2 and no.3, although rapid, are characterized by lower reproducibilities in comparison with technique no.4, with, some times, peaks of undetermined nature on the radio-chromatograms no.3. So, in spite of a slower migration (10 min.) the technique no.4 has been selected from the group as the most reliable technique. For this technique, the comparison between the two modes of reading the chromatography (by means of a radio-chromatograph or by measuring the activity of the two halves of the plate by an activity-meter), has shown no significant difference in RCP. Consequently, this method (Baker Flex / ethyl acetate) may by adapted in any service of nuclear medicine, no matter of its equipment

  1. Native MS and ECD Characterization of a Fab-Antigen Complex May Facilitate Crystallization for X-ray Diffraction

    Science.gov (United States)

    Zhang, Ying; Cui, Weidong; Wecksler, Aaron T.; Zhang, Hao; Molina, Patricia; Deperalta, Galahad; Gross, Michael L.

    2016-07-01

    Native mass spectrometry (MS) and top-down electron-capture dissociation (ECD) combine as a powerful approach for characterizing large proteins and protein assemblies. Here, we report their use to study an antibody Fab (Fab-1)-VEGF complex in its near-native state. Native ESI with analysis by FTICR mass spectrometry confirms that VEGF is a dimer in solution and that its complex with Fab-1 has a binding stoichiometry of 2:2. Applying combinations of collisionally activated dissociation (CAD), ECD, and infrared multiphoton dissociation (IRMPD) allows identification of flexible regions of the complex, potentially serving as a guide for crystallization and X-ray diffraction analysis.

  2. Tc-99m-bicisate (ECD)-brain-SPECT in rapidly progressive dementia; Hirn-SPECT mit Tc-99m-Bicisat (ECD) bei rasch progredientem dementiellen Syndrom

    Energy Technology Data Exchange (ETDEWEB)

    Marienhagen, J.; Eilles, C. [Regensburg Univ. (Germany). Abt. fuer Nuklearmedizin; Weingaertner, U.; Blaha, L. [Bezirkskrankenhaus Mainkofen (Germany). Psychiatrische Klinik; Zerr, I.; Poser, S. [Goettingen Univ. (Germany). Klinik und Poliklinik fuer Neurologie

    1999-07-01

    We present a 61-year-old male patient with progressive dementia. A brain SPECT with Tc-99m-bicisate was performed for confirmation of clinically suspected Alzheimer-dementia. At the time of the SPECT-investigation marked apraxia and aphasia besides severe dementia were present. Electrophysiological as well as anatomical neuroimaging findings showed non-diagnostic alterations. SPECT revealed distinct perfusion defects, which made Alzheimer Dementia unlikely. The further course of the patient was determined by rapidly progressive deterioration with development of akinetic mutism. Thereafter, increased levels of neuron-specific enolase as well as 14-3-3 proteins were found in the cerebro-spinal fluid (CSF). The patient finally died with signs of cerebral decortication. Due to the clinical course and the CSF-findings the patient's final diagnosis was Creutzfeld-Jakob-disease, nevertheless no autopsy was performed. The presented case report underscores the clinical utility of perfusion brain SPECT in the differential diagnosis of dementias. (orig.) [German] Wir berichten ueber einen 61jaehrigen Patienten mit progredientem dementiellen Syndrom, der unter der Verdachtsdiagnose einer Demenz vom Alzheimer-Typ (DAT) zur Hirn-SPECT-Untersuchung mit TC-99m-Bicisat (ECD) vorgestellt wurde. Zum Untersuchungszeitpunkt bestanden neben dem Vollbild einer Demenz eine ausgepraegte Apraxie und Aphasie bei unspezifischen Veraenderungen im EEG sowie der neuroradiologischen Bildgebung. In der Hirn-SPECT-Untersuchung fanden sich fuer eine DAT untypische ausgedehnte, vorwiegend rechtshemisphaerische Perfusionsstoerungen. Im weiteren Verlauf rasche Progredienz des Krankheitsbildes mit Entwicklung eines akinetischen Mutismus sowie Nachweis erhoehter Werte der neuronspezifischen Enolase und des 14-3-3-Proteins im Liquor. Der Patient verstarb schliesslich unter dem Bild einer Decortication. Aufgrund des klinischen Verlaufs sowie der Liquorbefunde wurde, da eine autoptische Befundsicherung

  3. A comparative study of 99mTc-ECD and X-CT in the evaluation of patients with cerebropathy

    International Nuclear Information System (INIS)

    Sun Da

    1995-01-01

    The clinical value of functional brain imaging was evaluated preliminarily by comparing 99m Tc-ECD SPECT with X-CT in patients with cerebropathy. 219 patients underwent 99m Tc-ECD SPECT and X-CT. Both procedures were performed within two weeks in each patient. The data showed that while X-CT was an excellent technique in detecting structural lesions of brain, such as cerebral hemorrhage or infarct, cerebrum and cephalophyma, it is not capable of demonstrating a metabolic and functional lesions without structural abnormalities. The functional imaging had great advantage in this regard. Results indicated that 99m Tc-ECD SPECT can easily detect cerebral blood flow change of intracerebral foci not detectable by traditional morphological imaging. This advantage was obvious especially in patients with early stage of acute cerebral infarct, TIA, hemicrania, cerebral arteriosclerosis, dementia, extraphyramidal disorders, sequelae of head trauma and epilepsy. In some of the patients with sequelae of head trauma or epilepsy, satisfactory effect was obtained when conservative treatments were performed to improve rCBF. 99 Tc-ECD SPECT may play a useful role in the diagnosis and therapy of patients with cerebropathy

  4. On Instructional Utility, Statistical Methodology, and the Added Value of ECD: Lessons Learned from the Special Issue

    Science.gov (United States)

    Nelson, Brian; Nugent, Rebecca; Rupp, Andre A.

    2012-01-01

    This special issue of "JEDM" was dedicated to bridging work done in the disciplines of "educational and psychological assessment" and "educational data mining" (EDM) via the assessment design and implementation framework of "evidence-centered design" (ECD). It consisted of a series of five papers: one…

  5. Dry Kit Development And Clinical Test Of 99mTc-L,L-ECD Radiopharmaceutical For Vrain Perfusion Imaging

    International Nuclear Information System (INIS)

    Kartini, Nani; Sofyan, Rohestri; Rukmini; Iswahyudi

    2000-01-01

    Technetium- 99m ethyl cysteinate dimer ( 99m Tc-L,L-ECD) has been synthesized and formulated based on ligand exchanged reaction with 99mTc-glucoheptonate. Considering the low stability of the dry kit produced, it was fairly necessary to develop the manufacture of L,L-ECD dry kit in order to obtain a more stable one that meet the requirements as brain perfusion imaging radiopharmaceutical. Experiment was done by modifying the packing of dry kit components of the previous formulation. Characteristics of 99m Tc-L,L-ECD produced from this formulation were studied by carrying out the sterility and toxicity test, then followed with clinical test to some volunteers. Evaluation of brain perfusion was done by tomography technique using gamma camera at Nuclear Medicine Installation, Hasan Sadikin Hospital. The modified formulation obtained consist of three components i.e. main ligand L,L-ECD; exchange ligand Ca-glucoheptonate and HaOH. Its stability could reach 8 months at-10 derajat C of storage. Effects of toxicity on mice did not appear. The result of clinical study shows that the radiopharmaceutical was distributed very rapidly in the blood brain circulations and retained in the brain for about one hour. The body scanning indicates that the substance was excreted in the brain for about one hour. The body scanning indicates that the substance was excreted though kidney, liver and spleen. Based on the results obtained the radiopharmaceutical could be promoted to be used for brain perfusion imaging

  6. 99mTc-ECD brain perfusion SPECT in hyperalgesic fibromyalgia

    International Nuclear Information System (INIS)

    Guedj, Eric; Taieb, David; Cammilleri, Serge; Lussato, David; Laforte, Catherine de; Mundler, Olivier; Niboyet, Jean

    2007-01-01

    Neuro-imaging studies with 99m Tc-HMPAO SPECT in fibromyalgia (FM) patients have reported only limited subcortical hypoperfusion. 99m Tc-ECD SPECT is known to provide better evaluation of areas of high cerebral blood flow and regional metabolic rate. We evaluated a homogeneous group of hyperalgesic patients with FM using 99m Tc-ECD SPECT. The aim of this study was to investigate brain processing associated with spontaneous pain in FM patients. Eighteen hyperalgesic FM women (mean age 49 years, range 25-63 years; American College of Rheumatology criteria) and ten healthy women matched for age were enrolled in the study. A voxel-by-voxel group analysis was performed using SPM2 (p<0.05, corrected for multiple comparisons). Visual Analogue Scale score for pain was 82±4 at the time of the SPECT study. Compared with control subjects, we observed individual brain SPECT abnormalities in FM patients, confirmed by SPM2 analysis, with hyperperfusion of the somatosensory cortex and hypoperfusion of the frontal, cingulate, medial temporal and cerebellar cortices. In the present study, performed without noxious stimuli in hyperalgesic FM patients, we found significant hyperperfusion in regions of the brain known to be involved in the sensory dimension of pain processing and significant hypoperfusion in areas assumed to be associated with the affective-attentional dimension. As current pharmacological and non-pharmacological therapies act differently on the two components of pain, we hypothesise that SPECT could be a valuable and readily available tool to guide individual therapeutic strategy and provide objective follow-up of pain processing recovery under treatment. (orig.)

  7. Analysis of iodine content in seaweed by GC-ECD and estimation of iodine intake

    Directory of Open Access Journals (Sweden)

    Tai Sheng Yeh

    2014-06-01

    Full Text Available Edible seaweed products have been consumed in many Asian countries. Edible seaweeds accumulate iodine from seawater, and are therefore a good dietary source of iodine. An adequate consumption of seaweed can eliminate iodine deficiency disorders, but excessive iodine intake is not good for health. The recommended dietary reference intake of 0.15 mg/d and 0.14 mg/d for iodine has been established in the United States and Taiwan, respectively. In this study, 30 samples of seaweed were surveyed for iodine content. The samples included 10 nori (Porphyra, 10 wakame (Undaria, and 10 kombu (Laminaria products. The iodine in seaweed was derivatized with 3-pentanone and detected by gas chromatography-electron capture detector (GC-ECD. The method detection limit was 0.5 mg/kg. The iodine content surveyed for nori was 29.3–45.8 mg/kg, for wakame 93.9–185.1 mg/kg, and for kombu 241–4921 mg/kg. Kombu has the highest average iodine content 2523.5 mg/kg, followed by wakame (139.7 mg/kg and nori (36.9 mg/kg. The GC-ECD method developed in this study is a low-cost alternative to inductively coupled plasma-optical emission spectroscopy for iodine detection in seaweeds. The iodine intake from seaweed in the current survey was calculated and compared with the iodine dietary reference intake of Taiwan. The risk and benefit of seaweed consumption is also discussed.

  8. Tc-99m ECD brain SPET in the evaluation of dementia for institutionalized elderly patients

    International Nuclear Information System (INIS)

    Lee, Myoung Hoon; Park, Chan H.; Yoon, Seok Na.; Hwang, Kyung Hoon

    2001-01-01

    Dementia is one of the clinically recognized indications of regional cerebral blood flow (rCBF) measurement by Tc-99m ECD brain SPET (Single Photon Emission Tomography). There is only limited number of institutions for elderly demented patients who are institutionalized in Korea and SPET is nor available at these institutions. The aim of the study is to evaluate rCBF SPET findings of the patients from such an institution. Thirty-one patients were reffered for rCBF SPET from Yongin Hyoja Hospital, Yongin. They were screened using NINCDS-ADRDA criteria for probable Alzheimers disease (AD) and dementia severity was assumed by the Mini-Mental State examination. In a quite, dim light room, patients were injected with 740 mBq (20mCi) Tc-99m ethyl cysteinate dimmer (ECD), Neurolite R, Dupont Pharmaceuticals, Billerica, MA, USA). SPET was acquired using fanbeam collimators and triple-head gamma camera (MultiSPECT III, Siemens medical systems. Inc. Hoffman Estates, III.USA). SPET was done one hour after the tracer injection and most of the patients needed sedation 30 minutes before the scan. SPET was evaluated visually by 2 nuclear medicine physicians blinded to clinical information. The SPET scans of 31 patients revealed 3 typical AD, 9 atypical AD patterns. Other dementia patterns were 4 cases of frontotemporal lobe dementia, 5 cases of frontal lobe dementia and 2 multifocal infarctions. Only cerebral atrophy is depicted in 8 patients and normal SPECT findings was noted in one patient. Patients who are institutionalized for dementia have varying SPET patterns as expected and SPET findings are useful in the management of these patients with more clearer clinical insight

  9. {sup 99m}Tc-ECD brain perfusion SPECT in hyperalgesic fibromyalgia

    Energy Technology Data Exchange (ETDEWEB)

    Guedj, Eric; Taieb, David; Cammilleri, Serge; Lussato, David; Laforte, Catherine de; Mundler, Olivier [Assistance Publique des Hopitaux de Marseille, Centre Hospitalo-Universitaire de la Timone, Service Central de Biophysique et de Medecine Nucleaire, Marseille Cedex 05 (France); Niboyet, Jean [Clinique La Phoceanne, Unite d' Etude et de Traitement de la Douleur, Marseille (France)

    2007-01-15

    Neuro-imaging studies with {sup 99m}Tc-HMPAO SPECT in fibromyalgia (FM) patients have reported only limited subcortical hypoperfusion. {sup 99m}Tc-ECD SPECT is known to provide better evaluation of areas of high cerebral blood flow and regional metabolic rate. We evaluated a homogeneous group of hyperalgesic patients with FM using {sup 99m}Tc-ECD SPECT. The aim of this study was to investigate brain processing associated with spontaneous pain in FM patients. Eighteen hyperalgesic FM women (mean age 49 years, range 25-63 years; American College of Rheumatology criteria) and ten healthy women matched for age were enrolled in the study. A voxel-by-voxel group analysis was performed using SPM2 (p<0.05, corrected for multiple comparisons). Visual Analogue Scale score for pain was 82{+-}4 at the time of the SPECT study. Compared with control subjects, we observed individual brain SPECT abnormalities in FM patients, confirmed by SPM2 analysis, with hyperperfusion of the somatosensory cortex and hypoperfusion of the frontal, cingulate, medial temporal and cerebellar cortices. In the present study, performed without noxious stimuli in hyperalgesic FM patients, we found significant hyperperfusion in regions of the brain known to be involved in the sensory dimension of pain processing and significant hypoperfusion in areas assumed to be associated with the affective-attentional dimension. As current pharmacological and non-pharmacological therapies act differently on the two components of pain, we hypothesise that SPECT could be a valuable and readily available tool to guide individual therapeutic strategy and provide objective follow-up of pain processing recovery under treatment. (orig.)

  10. Effect of the mobile phase on the RP-HPLC analysis of 99mTc-ECD

    International Nuclear Information System (INIS)

    Almeida, Erika V.; Monteiro, Elisiane G.; Mengatti, Jair; Fukumori, Neuza T.O.; Silva, Constancia P.G. da; Matsuda, Margareth M.N.

    2009-01-01

    Technetium-99m labeled L,L-ethylene cysteine dimer ( 99m Tc-ECD) is a neutral, lipophilic complex and brain perfusion imaging agent. The aim of this study was to investigate the effect of the mobile phase on the reversed phase high performance liquid chromatography (RP-HPLC) analysis of 99m Tc-ECD. The HPLC system was LC20AT Prominence model and a Shim-Pack VP-ODS column (250 x 4.6 mm i.d., 5 μm). 99m Tc-ECD was prepared by adding 1 mL of 0.9% NaCl, 1 mL of phosphate buffer (pH 7.5) and 1 mL of Na 99m TcO 4 . The radioactive concentration was 55.5 MBq mL -1 . 20 μL sample volume was injected and 1.0 mL min -1 flow rate was applied. A linear gradient was performed separately with a mixture of ethanol with three different solvents: 12.5 mmol L -1 phosphate buffer (pH 2.5) (solvent A), 0.2 % PIC A (w/v) (pH 6.0) (solvent B) and 0.2 % PIC B5 (w/v) (pH 4.0) (solvent C). 99m Tc-ECD retention times for the mixture of ethanol with solvent A, B and C were 17.38, 17.65 and 17.60 minutes, respectively. These results suggested that the ion pairing reagents (PIC A and PIC B5) did not influence the 99m Tc-ECD analysis, but affected the retention time of impurities as 99m Tc-EC (ethylene dicysteine) and 99m TcO 4 -. 99m Tc-ECD radiochemical purity determined by RP-HPLC was higher than 96% for the mixture of ethanol with the three solvents. The results indicated that the retention time of 99m Tc-ECD was not influenced by the nature of the mobile phase. (author)

  11. Characterization of epithelial domains in the nasal passages of chick embryos: spatial and temporal mapping of a range of extracellular matrix and cell surface molecules during development of the nasal placode.

    Science.gov (United States)

    Croucher, S J; Tickle, C

    1989-07-01

    The formation of the nasal passages involves complex morphogenesis and their lining develops a spatially ordered pattern of differentiation, with distinct domains of olfactory and respiratory epithelium. Using antibodies to the neural cell adhesion molecule (N-CAM), keratan sulphate and heparan sulphate proteoglycan (HSPG) and a panel of lectins (agglutinins of Canavalia ensiformis (ConA), Dolichos biflorus (DBA), peanut (PNA), Ricinis communis (RCA1), soybean (SBA), Ulex europaeus (UEA1), and wheatgerm (WGA], we have documented cell surface characteristics of each epithelial domain. Binding of antibodies to N-CAM and to keratan sulphate, and the lectins ConA, PNA, RCA1, SBA and WGA marks the olfactory epithelial domain only. The restriction of N-CAM to the sensory region of the epithelium has also been reported in the developing ear. This striking similarity is consistent with the idea that N-CAM may be involved in the division of functionally and histologically distinct cell groups within an epithelium. We traced the olfactory-specific cell markers during development to gain insights into the origin of the epithelial lining of the nasal passages. All reagents bind at early stages to the thickened nasal placode and surrounding head ectoderm and then become progressively restricted to the olfactory domain. The expression of these characteristics appears to be modulated during development rather than being cell autonomous. The distribution of keratan sulphate was compared with collagen type II in relation to the specification of the chondrocranium. Keratan sulphate and collagen type II are only colocalized at the epithelial-mesenchymal interface during early nasal development. At later stages, only collagen type II is expressed at the interface throughout the nasal passages, whereas keratan sulphate is absent beneath the respiratory epithelium.

  12. Domains and domain loss

    DEFF Research Database (Denmark)

    Haberland, Hartmut

    2005-01-01

    politicians and in the media, especially in the discussion whether some languages undergo ‘domain loss’ vis-à-vis powerful international languages like English. An objection that has been raised here is that domains, as originally conceived, are parameters of language choice and not properties of languages...

  13. Power recovery method for testing the efficiency of the ECD of an integrated generation unit for offshore wind power and ocean wave energy

    Institute of Scientific and Technical Information of China (English)

    CHEN WeiXing; GAO Feng; MENG XiangDun; REN AnYe; HU Yan

    2017-01-01

    Offshore wind power and ocean wave energy are clean,renewable and rich resources.The integrated generation unit for the two kinds of energy is introduced.The energy conversion device (ECD) is utilized to convert the mechanical energy absorbed from the wind power and wave energy into the hydraulic energy,the conversion efficiency of which is significant.In this paper,a power recovery method for testing the efficiency of the ECD is proposed.A simulation desktop is developed to validate the proposed method.The efficiency of the ECD is influenced by the hydraulic cylinders and the mechanical transmission.Here,the static efficiency of the hydraulic cylinders of the ECD is tested first.The results show that the static mechanical efficiency is about 95 % and that the volumetric efficiency is over 99%.To test the effects induced by the mechanical transmission of the ECD,each hydraulic cylinder of the ECD is substituted with two springs.Then the power loss of the ECDM under different rotational speeds is obtained.Finally,a test platform is built and the efficiency of the ECD under different rotational speeds and pressures is obtained.The results show that the efficiency is about 80%.

  14. Discrepant 99mTc-ECD images of CBF in patients with subacute cerebral infarction. A comparison of CBF, CMRO2 and 99mTc-HMPAO imaging

    International Nuclear Information System (INIS)

    Shishido, Fumio; Uemura, Kazuo; Inugami, Atsushi; Ogawa, Toshihide; Fujita, Hideaki; Shimosegawa, Eku; Nagata, Ken.

    1995-01-01

    Three patients with subacute ischemic cerebral infarction examined by SPECT with 99m Tc-ECD and PET within the same day showed signs of luxury perfusion in the subacute phase, which is between 9 to 20 days after the onset. A 99m Tc-HMPAO SPECT study was also performed within 2 days of the ECD-SPECT study. ECD-SPECT images of three patients displayed a focal decreased uptake in the infarcted lesions, while in infarcted foci, there was almost equivalent or increased CBF compared to normal and unaffected areas, decreased CMRO 2 , and high HMPAO uptake. The ECD-SPECT results were similar to those of CMRO 2 rather than CBF, though the HMPAO-SPECT image was similar to that of CBF. In one patient, HMPAO images revealed hyperfixation of the tracer. In the chronic phase and in the acute phase before 5 days after the onset, there were no discrepancies among the ECD-SPECT, CBF, HMPAO-SPECT, and CMRO 2 images. These observations indicated that 99m Tc-ECD is a good indicator of damaged brain tissues in subacute ischemic infarction. They also suggested that 99m Tc-ECD is a potential agent with which to evaluate cerebral tissue viability in some pathological states of cerebrovascular disease. The characteristics may be suitable for confirming the effects of thrombolytic therapy in acute ischemia, because these conditions often show signs of luxury perfusion when the therapy is successful. (author)

  15. Expression, purification, and analysis of three recombinant ECD disintegrins (r-colombistatins) from P-III class snake venom metalloproteinases affecting platelet aggregation and SK-MEL-28 cell adhesion.

    Science.gov (United States)

    Suntravat, Montamas; Helmke, Thomas J; Atphaisit, Chairat; Cuevas, Esteban; Lucena, Sara E; Uzcátegui, Nestor L; Sánchez, Elda E; Rodriguez-Acosta, Alexis

    2016-11-01

    Crotalid venoms are rich sources of components that affect the hemostatic system. Snake venom metalloproteinases are zinc-dependent enzymes responsible for hemorrhage that also interfere with hemostasis. The disintegrin domain is a part of snake venom metalloproteinases, which involves the binding of integrin receptors. Integrins play an essential role in cancer survival and invasion, and they have been major targets for drug development and design. Both native and recombinant disintegrins have been widely investigated for their anti-cancer activities in biological systems as well as in vitro and in vivo systems. Here, three new cDNAs encoding ECD disintegrin-like domains of metalloproteinase precursor sequences obtained from a Venezuelan mapanare (Bothrops colombiensis) venom gland cDNA library have been cloned. Three different N- and C-terminal truncated ECD disintegrin-like domains of metalloproteinases named colombistatins 2, 3, and 4 were amplified by PCR, cloned into a pGEX-4T-1 vector, expressed in Escherichia coli BL21, and tested for inhibition of platelet aggregation and inhibition of adhesion of human skin melanoma (SK-Mel-28) cancer cell lines on collagen I. Purified recombinant colombistatins 2, 3, and 4 were able to inhibit ristocetin- and collagen-induced platelet aggregation. r-Colombistatins 2 showed the most potent inhibiting SK-Mel-28 cancer cells adhesion to collagen. These results suggest that colombistatins may have utility in the development of therapeutic tools in the treatment of melanoma cancers and also thrombotic diseases. Copyright © 2016. Published by Elsevier Ltd.

  16. Technetium-99m-ECD SPECT in antiphospholipid antibody syndrome: a drastic improvement in brain perfusion by antiplatelet therapy

    Energy Technology Data Exchange (ETDEWEB)

    Tokumaru, Sunao; Yoshikai, Tomonori; Uchino, Akira; Kudo, Sho [Dept. of Radiology, Saga Medical School (Japan); Matsui, Makoto; Kuroda, Yasuo [Dept. of Neurology, Saga Medical School (Japan)

    2001-12-01

    We present a case of antiphospholipid antibody syndrome (APS) with repeated transient ischemic attacks (TIAs). Magnetic resonance imaging showed multiple cerebral infarcts and ischemic changes in the cerebral white matter. Cerebral angiographies showed no abnormalities. Technetium-99m-ethyl cysteinate dimer (Tc-99m-ECD) brain SPECT showed multiple decreased perfusion areas, which were more extensive than the lesions demonstrated on MRI. After treatment with an antiplatelet agent, the patient subsequently recovered from the TIAs. Although no interval changes were observed by MRI after therapy, follow-up Tc-99m-ECD SPECT revealed a marked improvement in brain perfusion. This is the first imaging report of remarkable post-therapy improvement in brain perfusion in APS cases. (orig.)

  17. Technetium-99m-ECD SPECT in antiphospholipid antibody syndrome: a drastic improvement in brain perfusion by antiplatelet therapy

    International Nuclear Information System (INIS)

    Tokumaru, Sunao; Yoshikai, Tomonori; Uchino, Akira; Kudo, Sho; Matsui, Makoto; Kuroda, Yasuo

    2001-01-01

    We present a case of antiphospholipid antibody syndrome (APS) with repeated transient ischemic attacks (TIAs). Magnetic resonance imaging showed multiple cerebral infarcts and ischemic changes in the cerebral white matter. Cerebral angiographies showed no abnormalities. Technetium-99m-ethyl cysteinate dimer (Tc-99m-ECD) brain SPECT showed multiple decreased perfusion areas, which were more extensive than the lesions demonstrated on MRI. After treatment with an antiplatelet agent, the patient subsequently recovered from the TIAs. Although no interval changes were observed by MRI after therapy, follow-up Tc-99m-ECD SPECT revealed a marked improvement in brain perfusion. This is the first imaging report of remarkable post-therapy improvement in brain perfusion in APS cases. (orig.)

  18. The clinical usefulness of Tc-99m ECD brain SPECT in acute measles encephalitis

    International Nuclear Information System (INIS)

    Lim, Seok Tae; Sohn, Myung Hee

    2003-01-01

    Since the prognosis of measles encephalitis is poor, early diagnosis and proper management are very important to improve clinical outcomes. We compared Tc-99m ECD brain SPECT (SPECT) with MR imaging (MRI) for the detection of acute measles encephalitis. Eleven patients (M : F=4 : 7, age range 18 months-14 yrs) with acute measles encephalitis were enrolled in this studies. All of them underwent both MRI and SPECT. The results of SPECT were scored from 0 (normal) to 3 (most severe defect) according to perfusion state. We compared two image modalities for the detection of brain abnormality in acute measles encephalitis. Seven of 11 patients (63.6%) revealed high signal intensity in the white matter on T2WI of MRI, on the other hand all patients (100%) showed hypoperfusion on SPECT. Severe perfusion deficits above score 2 were located with decreasing frequencies in the frontal lobe (81.8%), temporal lobe (72.7%), occipital lobe (27.3%), basal ganglia (27.3%), and parietal lobe (9.1%). We conclude that SPECT is more useful than MRI for the detection of brain involvement in patients with acute measles encephalitis

  19. PCBs determination in water, soils and biota by perchlorination and GC-ECD

    Energy Technology Data Exchange (ETDEWEB)

    Feshin, D. [Institute of Ecology and Evolution RAS, Moscow (Russian Federation); Klyuev, N.; Brodsky, E.; Kalinkevich, G.

    2004-09-15

    Most of the PCB pollutions are caused by industrial application. PCB-containing mixtures whose compositions are well known and constant should be rapidly and precisely analyzable as total PCB to monitor real pollution exposure. Consequently, a simple but reproducible and cheap method for operative control is needed which should be periodically controlled by standard analytical methods. So it is advisable to have a rapid method that will allow PCBs determination in wide range of quantities by simple instrumentation with internal standards. The method based on perchlorination of PCBs considerably simplifies the detection of PCB congeners as compared to other techniques. In this practice all polychlorinated derivatives are converted into decachlorobiphenyl (DCB). This is followed by gas chromatography with electron capture detector (GC-ECD), which is highly sensitive for the substances of interest. Currently EPA procedure 508a (1989) is the only official screening method for detection of PCBs using perchlorination. This uses SbCl5 and powdered Fe mixture as reagent at 270 C without internal standard. We reported about using of perchlorination reagent based on mixture of powdered duralumin, sulphur and SO2Cl2 heated at 105 C for 2 h for detection of PCBs as decachlorobiphenyls in a wide range of concentration: 16.4 ng/sample to 164 {mu}/sample. 4,4'-difluorobiphenyl was proposed as internal standard that allowed to control all steps of sample preparation including perchlorination reaction. In the present work we extended our experience of perchlorination to detect the PCBs.

  20. The clinical usefulness of Tc-99m ECD brain SPECT in acute measles encephalitis

    Energy Technology Data Exchange (ETDEWEB)

    Lim, Seok Tae; Sohn, Myung Hee [School of Medicine, Chonbuk National Univ., Chonju (Korea, Republic of)

    2003-08-01

    Since the prognosis of measles encephalitis is poor, early diagnosis and proper management are very important to improve clinical outcomes. We compared Tc-99m ECD brain SPECT (SPECT) with MR imaging (MRI) for the detection of acute measles encephalitis. Eleven patients (M : F=4 : 7, age range 18 months-14 yrs) with acute measles encephalitis were enrolled in this studies. All of them underwent both MRI and SPECT. The results of SPECT were scored from 0 (normal) to 3 (most severe defect) according to perfusion state. We compared two image modalities for the detection of brain abnormality in acute measles encephalitis. Seven of 11 patients (63.6%) revealed high signal intensity in the white matter on T2WI of MRI, on the other hand all patients (100%) showed hypoperfusion on SPECT. Severe perfusion deficits above score 2 were located with decreasing frequencies in the frontal lobe (81.8%), temporal lobe (72.7%), occipital lobe (27.3%), basal ganglia (27.3%), and parietal lobe (9.1%). We conclude that SPECT is more useful than MRI for the detection of brain involvement in patients with acute measles encephalitis.

  1. Structure dependent antioxidant capacity of phlorotannins from Icelandic Fucus vesiculosus by UHPLC-DAD-ECD-QTOFMS.

    Science.gov (United States)

    Hermund, Ditte B; Plaza, Merichel; Turner, Charlotta; Jónsdóttir, Rosa; Kristinsson, Hordur G; Jacobsen, Charlotte; Nielsen, Kristian Fog

    2018-02-01

    Brown algae are rich in polyphenolic compounds, phlorotannins, which have been found to possess high in vitro antioxidant capacity, especially DPPH radical scavenging activity, due to the high number of hydroxyl groups. Whereas, the overall antioxidant capacity of brown algae extracts has been widely studied, the antioxidant capacity of individual phlorotannins has been rarely explored. The aim of this study was to determine the structure dependant antioxidant capacity of phlorotannins from Icelandic brown algae, Fucus vesiculosus. The antioxidant capacity of individual phlorotannins was determined by an on-line method using liquid chromatography and an electrochemical detector followed by quadrupole Time of Flight mass spectrometry (UHPLC-DAD-ECD-QTOFMS). Tentative structural elucidation of 13 phlorotannin isomers from EAF was obtained by LC-DAD-QTOFMS, ranging from 374 to 870Da. On-line determination of antioxidant capacity of the individual phlorotannins generally showed that low molecular phlorotannins exhibited higher antioxidant capacity and that the capacity decreased with polymerisation. Copyright © 2017 Elsevier Ltd. All rights reserved.

  2. 99mTc-ECD: Comparison of radiochemical purity evaluation techniques

    International Nuclear Information System (INIS)

    Dumont, A.; De Beco, V.; Ait Ben Ali, S.; Goudou-Sinha, C.; Izembart, M.; Jourdain, J.R.; Lemercier, V.; Linsker, S.; Lours, S.; Moati, F.; Pajard, D.; Piketty, M.L.; Rizzo, N.; Schlageter, M.H.; Moretti, J.L.

    1997-01-01

    The aim of this study was to validate a testing method for the radiochemical purity (RCP) of the preparations of 99m Tc - ECD that is to be reproducible and easy to realize in services of Nuclear Medicine. After a review of literature, four thin-layer chromatography techniques allowing to evidence the TcO 4 - were evaluated: no.1 - Papier Whatman 31ET / ethyl acetate; no.2 - Papier Whatman 3MM chr / ethyl acetate; no.3 - ITLC Silica gel / ethyl acetate; no.4 - Baker Flex silica gel aluminium oxide IB-F / ethyl acetate (the method proposed by the laboratory). The technique no.1 has presented a bad reproducibility, as well as percentages of RCP very different from those obtained by the other techniques. The techniques no.2 and no.3, although rapid, are characterized by lower reproducibilities in comparison with technique no.4, with, some times, peaks of undetermined nature on the radio-chromatograms no.3. So, in spite of a slower migration (10 min.) the technique no.4 has been selected from the group as the most reliable technique. For this technique, the comparison between the two modes of reading the chromatography (by means of a radio-chromatograph or by measuring the activity of the two halves of the plate by an activity-meter), has shown no significant difference in RCP. Consequently, this method (Baker Flex / ethyl acetate) may by adapted in any service of nuclear medicine, no matter of its equipment

  3. Tc-99m ECD brain SPECT in MELAS syndrome: comparison with MR finding

    International Nuclear Information System (INIS)

    Park, Sang Joon; Ryu, Young Hoon; Yoon, Pyeong Ho; Jeon, Tae Joo; Kim, Jai Keun; Nam, Ji Eun; Lee, Jong Doo; Lee, Byung Hee; Shin, Hyung Cheol

    1998-01-01

    The purpose of this study was to evaluate SPECT findings of MELAS syndrome and mitochondrial myopathy and correlate them with MR findings in search of specific imaging features and to assess the role of SPECT in MELAS syndrome. Five patients (four females and one male; age range, 1 to 25 years) who presented with repeated stroke-like episodes or seizures or developmental delay or were asymptomatic but had elevated lactic acid in CSF and serum were evaluated with conventional noncontrast MR imaging and SPECT. MRI demonstrated increased T2 signal intensities in the affected areas of gray and white matters mainly on the parietal (4/5) and occipital lobes (4/5) and in the basal ganglias (1/5), which were not restricted to a specific vascular territory. SPECT demonstrated decreased uptake of Tc-99m ECD on parietal (5/5) and occipital (4/5) and temporal (2/5) and frontal (1/5) lobe and basal ganglia (2/5) and thalami (2/5). In a patient with mitochondrial myopathy who had normal MRI, decreased perfusion is noted on left parietal area and bilateral thalami. Comparison of the numbers of abnormal findings revealed that decreased perfusion seen on SPECT were more numerous than anatomical abnormalities seen on MRI. SPECT may be a sensitive method for pathophysiological study of metabolic disturbances in MELAS. Moreover, in patients with mitochondrial myopathy without clinical encephalopathy, SPECT may play a role in evaluating subclinical encephalopathy even with normal conventional MR findings

  4. Relationship between 99mTc-ECD SPECT images and performance of the Wisconsin card sorting test in healthy humans

    International Nuclear Information System (INIS)

    Zhao Jinhua; Lin Xiangtong; Guan Yihui; Xu Lianqin; Zhang Guangming; Xue Fangpeng

    1998-01-01

    Purpose: To investigate the relationship between 99m Tc-ECD SPECT and performance of the Wisconsin Card Sorting Test (WCST) in normal young and aged volunteers through the semi-quantitative analysis of the baseline and cognition activated rCBF images. Methods: 18 young and 17 aged volunteers were enrolled. The education level of the two groups was matched. All subjects were healthy and CT results of all the aged volunteers were normal. Both baseline and cognition activated 99m Tc-ECD SPECT were performed on all the subjects. The cognition activated brain SPECT: The cognitive task was WCST. 1110 MBq 99m Tc-ECD was administered by intravenous bolus injection 5 minutes after the onset of the WCST. The images were reconstructed and semiquantitatively analyzed. WCST were scored through 9 items: 1) period, 2) categories, 3) total corrects, 4) conceptual level (%), 5) perseverative responses, 6) total errors, 7) perseverative errors, 8) perseverative error (%), 9) non-perseverative errors. Results: 1) Compared with the baseline, the activated rCBF values of frontal, left temporal and left parietal lobes were higher in young subjects. 2) The activated rCBF values of right frontal lobe is slightly higher than those of baseline in aged group. 3) WCST scores: the young had significantly better scores than the aged in 7 items. 4) The baseline and activated rCBF values of the frontal lobes and anterior cingule were closely related to the scores of categories, perseverative responses, total errors and non-perseverative errors. Conclusions: 1) The distribution patterns of 99m Tc-ECD are different between young and aged volunteers. 2) The performance of WCST mainly relates to frontal lobes. The mild hypoperfusion of the frontal lobe in aged group may be the cause of poor conceptual and thinking ability. 3) First found the significant correlation between anterior cingule and performance of WCST

  5. Analysis of neonicotinoids by gas chromatography coupled to nuclide 63Ni - Electron Capture Detector - GC/ECD

    International Nuclear Information System (INIS)

    Amaral, Priscila O.; Leao, Claudio; Redigolo, Marcelo M.; Crepaldi, Caike; Bustillos, Oscar V.

    2015-01-01

    Recently, several reports have been published discussing reduction in bee population which polymerizes cultures around the world this phenomenon is known as Colony Collapse Disorder (CCD). The phenomenon describes the lack of worker honeybees in the colony despite having pups and food. The causes of this problem are unknown but there are studies that claim that reduction of population of bees is linked to poisoning through insecticides specifically neonicotinoids. Among this type of pesticide are imidacloprid (C 9 H 10 ClN 5 O 2 ), clothianidin (C 6 H 8 ClN 5 O 2 S) and thiamethoxam (C 8 H 10 ClN 5 O 3 S). This paper presents the analysis of neonicotinoids - clothianidin, imidacloprid and thiamethoxam - by the technique of gas chromatography coupled to nuclide 63 Ni electron capture detector (GC/ECD). The electron capture detector (ECD) is a gas chromatography detector that has been used for the detection of organic halogens, nitriles, nitrates and organometallic compounds. The ECD detector ionizes the analytes by the beta particles from the nuclide sources 63 Ni within carrier gas N 2 . The electrons produced in this process are collected and create a current that are amplified and generates a chromatographic peak. Methodology and details of the analysis are present in this work. (author)

  6. Analysis of neonicotinoids by gas chromatography coupled to nuclide {sup 63}Ni - Electron Capture Detector - GC/ECD

    Energy Technology Data Exchange (ETDEWEB)

    Amaral, Priscila O.; Leao, Claudio; Redigolo, Marcelo M.; Crepaldi, Caike; Bustillos, Oscar V., E-mail: priscilaoamaral@gmail.com, E-mail: claudio.leao@usp.br, E-mail: marceloredigolo@gmail.com, E-mail: caike1995@gmail.com, E-mail: ovega@ipen.bremails [Instituto de Pesquisas Energeticas e Nucleares (IPEN/CNEN-SP), Sao Paulo, SP (Brazil)

    2015-07-01

    Recently, several reports have been published discussing reduction in bee population which polymerizes cultures around the world this phenomenon is known as Colony Collapse Disorder (CCD). The phenomenon describes the lack of worker honeybees in the colony despite having pups and food. The causes of this problem are unknown but there are studies that claim that reduction of population of bees is linked to poisoning through insecticides specifically neonicotinoids. Among this type of pesticide are imidacloprid (C{sub 9}H{sub 10}ClN{sub 5}O{sub 2}), clothianidin (C{sub 6}H{sub 8}ClN{sub 5}O{sub 2}S) and thiamethoxam (C{sub 8}H{sub 10}ClN{sub 5}O{sub 3}S). This paper presents the analysis of neonicotinoids - clothianidin, imidacloprid and thiamethoxam - by the technique of gas chromatography coupled to nuclide {sup 63}Ni electron capture detector (GC/ECD). The electron capture detector (ECD) is a gas chromatography detector that has been used for the detection of organic halogens, nitriles, nitrates and organometallic compounds. The ECD detector ionizes the analytes by the beta particles from the nuclide sources {sup 63}Ni within carrier gas N{sub 2}. The electrons produced in this process are collected and create a current that are amplified and generates a chromatographic peak. Methodology and details of the analysis are present in this work. (author)

  7. Investigation of olfactory function in normal volunteers by Tc-99m ECD Brain SPECT: Analysis using statistical parametric mapping

    International Nuclear Information System (INIS)

    Chung, Y.A.; Kim, S.H.; Park, Y.H.; Lee, S.Y.; Sohn, H.S.; Chung, S.K.

    2002-01-01

    The purpose of this study was to investigate olfactory function according to Tc-99m ECD uptake pattern in brain perfusion SPET of normal volunteer by means of statistical parametric mapping (SPM) analysis. The study population was 8 healthy volunteer subjects (M:F = 6:2, age range: 22-54 years, mean 34 years). We performed baseline brain perfusion SPET using 555 MBq of Tc-99m ECD in a silent dark room. Two hours later, we obtained brain perfusion SPET using 1110 MBq of Tc-99m ECD after 3% butanol solution under the same condition. All SPET images were spatially transformed to standard space smoothed and globally normalized. The differences between the baseline and odor-identification SPET images were statistically analyzed using SPM-99 software. The difference between two sets of brain perfusion SPET was considered significant at a threshold of uncorrected p values less than 0.01. SPM analysis revealed significant hyper-perfusion in both cingulated gyri, right middle temporal gyrus, right superior and inferior frontal gyri, right lingual gyrus and right fusiform gyrus on odor-identification SPET. This study shows that brain perfusion SPET can securely support other diagnostic techniques in the evaluation of olfactory function

  8. PENGEMBANGAN MEDIA EDUKASI GIZI MELALUI BUKU MEWARNAI UNTUK PESERTA PENDIDIKAN ANAK USIA DINI (PAUD) (MEDIA DEVELOPMENT OF NUTRITION EDUCATION THROUGH COLORING BOOKS FOR EARLY CHILDHOOD EDUCATION (ECD))

    OpenAIRE

    Yurista Permanasari; Erna Luciasari; Aditianti Aditianti

    2013-01-01

    ABSTRACT Early childhood education (ECD) is a right time to provide nutrition education. The education can be given through playing patterns suitable for age 2-6 years. Currently there is no active nutrition education media for ECD participants. Therefore it is necessary to develop an active media of nutrition education through colouring book. Colouring activitities can stimulate both the cognitive and motor development of children. This study aimed to develop a colouring book as a nutrition ...

  9. Extracellular Gd-CA

    DEFF Research Database (Denmark)

    Thomsen, Henrik S; Marckmann, Peter

    2008-01-01

    Until recently it was believed that extracellular gadolinium-based contrast agents were safe for both the kidneys and all other organs within the dose range up to 0.3 mmol/kg body weight. However, in 2006, it was demonstrated that some gadolinium-based contrast agents may trig the development...... gadolinium-based agent (3-7% versus 0-1% per injection) in patients with reduced renal function. Prevalence after exposure to two gadodiamide injections is as high as 36% in patients with chronic kidney disease (CKD) stage 5. No report of NSF after the most stable agents has been reported in the peer...

  10. Brain 99Tcm-ECD SPECT imaging in patients with obsessive-compulsive disorder

    International Nuclear Information System (INIS)

    Sun Da; Zhan Hongwei; Ye Xiaojuan; He Gangqiang

    2010-01-01

    Purpose Obsessive-compulsive disorder (OCD) is a chronic anxiety disorder of unknown aetiology. The purpose of the study is to evaluate the changes of brain function in patients with OCD. Methods: Regional cerebral perfusion was investigated using SPECT in 25 patients with OCD. The mean ages of the patients were 29.04 (1 8-46) years old. The clinical symptom consists of repeating thinking (suspect, worry, nervous) and repeating action (checking, washing, counting, making telephone calls) principally. After administration of 740-925 MBq (20-25 mCi) 99mTc-ECD a single photon emission tomography study was performed and then transaxial, sagittal and coronal slices were obtained. For the semiquantitative analysis of the data Results: 92 per cent of patients (23125) had relative hypoperfusions in some cerebral regions. The patients had a significant decrease of rCBF in the frontal lobes, temporal lobes, basal ganglia, thalamus, and cingulate gyrus. There were no correlation between the change of rCBF and age of age or course of disease. But there were some correlation with clinical symptom. Conclusion: Obsessive-compulsive disorder (OCD) has been linked to a dysfunction of brain orbitofrontal-striatum-pallidum-thalamus networks that were confirmed by PET SPECT functional imaging studies. These study indicated hypoperfusion in frontal lobes, basal ganglion, thalamus in OCD patients, and suggests a reduced serotonergic input into the fronto-subcortical circuits in OCD, thereby diminishing the inhibitory regulation of serotonin on these circuits. According to our results, patients with OCD had profound dysfunction of the frontal and temporal cortices, and basal ganglia. These may reflect a fundamental feature of clinical neuropathophysiology in OCD, and support previous findings about dysfunction of frontal-subcortical circuits in this disorder. (authors)

  11. Study of cellular retention of HMPAO and ECD in a model simulating the blood-brain barrier; Etude de la retention cellulaire de l`HMPAO et de l`ECD dans un modele simulant la barriere hematoencephalique

    Energy Technology Data Exchange (ETDEWEB)

    Ponce, C.; Pittet, N.; Slosman, D.O. [HUG, 1211 Geneve 14, (Switzerland)

    1997-12-31

    The HMPAO and ECD are two technetium-labelled lipophilic agents clinically used in the imagery of cerebral perfusion. These molecules cross the membranes and are retained inside the cell after being converted to a hydrophilic form. The aim of this study is to establish the distribution of this retention at the level of blood-brain barrier (BBB) and nerve cells. The incorporation of HMPAO or ECD was studied on a model of co-culture simulating the BBB by means of a T84 single-cell layer of tight junction separated from another layer of U373 astrocyte cells. The cell quality and tight junction permeability were evaluated by the cellular retention of 111-indium chloride and by para-cellular diffusion of {sup 14}C mannitol,d-1. The values reported below were obtained at 180 minutes when the radiotracers were added near the `T84 layer`. The cell quality is validated by the low cellular retention of the indium chloride(2.3{+-}0.3 {mu}g{sup -1} for the T84 cells and 8.2{+-}5.8 {mu}g{sup -1} for the U373 cells). The activity of {sup 14}C mannitol,d-1 diminishes by 23 {+-} 5 % in the added compartment. The retention of ECD by the U373 cells is significantly higher (20.7 {+-}4.5 g{sup -1}) than that of T84 cells (2.9 {+-} 0.2 {mu}g{sup -1}). For HMPAO a non-significant tendency could be observed (49 {+-} 34 {mu}g{sup -1} for the U373 cells and 38 {+-} 25 {mu}g{sup -1} for the T84 cells)> The results of cellular retention of indium by HMPAO or ECD when added near `U373 layer` are not significantly different.In conclusion, independently of the side exposed to the radiotracers, one observes an enhanced incorporation of the U373 cells. The ensemble of these results represent additional arguments in favour of a specific cellular incorporation of the radiotracers, independent of the BBB permittivity

  12. Regional differences between 99mTc-ECD and 99mTc-HMPAO SPET in perfusion changes with age and gender in healthy adults

    International Nuclear Information System (INIS)

    Inoue, Kentaro; Nakagawa, Manabu; Goto, Ryoi; Kinomura, Shigeo; Sato, Tachio; Sato, Kazunori; Fukuda, Hiroshi

    2003-01-01

    A number of studies using single-photon emission tomography (SPET) have shown perfusion changes with age in several cortical and subcortical areas, which might distort the results of perfusion imaging studies of neuropsychiatric disorders. Technetium-99m labelled ethyl cysteinate dimer (ECD) and hexamethylpropylene amine oxime (HMPAO) are both used as markers of cerebral perfusion, but have different pharmacokinetics and retention patterns. The aim of this study was to determine whether age and gender effects on perfusion SPET differ depending on whether 99m Tc-HMPAO or 99m Tc-ECD is used. Forty-five subjects (20 male and 25 female, mean age 52.8±6.6 years) were assigned to 99m Tc-HMPAO SPET (HMPAO group), and 39 subjects (24 male and 15 female, mean age 52.6±6.7 years) to 99m Tc-ECD SPET (ECD group). SPET images were obtained about 10 min after intravenous injection of approximately 800 MBq 99m Tc-HMPAO or 99m Tc-ECD using the same SPET scanner. Three-dimensional volumetric magnetic resonance imaging was performed to as7sess morphological changes in the grey matter. All image processing and statistical analyses were performed using SPM99 software. An area in the right anterior frontal lobe showed an increase in perfusion with age only in the HMPAO group, whereas areas in the bilateral retrosplenial cortex showed decreases in perfusion with age only in the ECD group; neither group showed corresponding changes in the grey matter. The present study shows that different effects of age on perfusion are observed depending on whether 99m Tc-HMPAO and 99m Tc-ECD is used. This suggests that the results of perfusion SPET are differently confounded depending on the tracer used, and that perfusion SPET with these tracers has limitations when used in research on subtle perfusion changes. (orig.)

  13. Research priority setting for integrated early child development and violence prevention (ECD+) in low and middle income countries: An expert opinion exercise.

    Science.gov (United States)

    Tomlinson, Mark; Jordans, Mark; MacMillan, Harriet; Betancourt, Theresa; Hunt, Xanthe; Mikton, Christopher

    2017-10-01

    Child development in low and middle income countries (LMIC) is compromised by multiple risk factors. Reducing children's exposure to harmful events is essential for early childhood development (ECD). In particular, preventing violence against children - a highly prevalent risk factor that negatively affects optimal child development - should be an intervention priority. We used the Child Health and Nutrition Initiative (CHNRI) method for the setting of research priorities in integrated Early Childhood Development and violence prevention programs (ECD+). An expert group was identified and invited to systematically list and score research questions. A total of 186 stakeholders were asked to contribute five research questions each, and contributions were received from 81 respondents. These were subsequently evaluated using a set of five criteria: answerability; effectiveness; feasibility and/or affordability; applicability and impact; and equity. Of the 400 questions generated, a composite group of 50 were scored by 55 respondents. The highest scoring research questions related to the training of Community Health Workers (CHW's) to deliver ECD+ interventions effectively and whether ECD+ interventions could be integrated within existing delivery platforms such as HIV, nutrition or mental health platforms. The priority research questions can direct new research initiatives, mainly in focusing on the effectiveness of an ECD+ approach, as well as on service delivery questions. To the best of our knowledge, this is the first systematic exercise of its kind in the field of ECD+. The findings from this research priority setting exercise can help guide donors and other development actors towards funding priorities for important future research related to ECD and violence prevention. Copyright © 2017 The Authors. Published by Elsevier Ltd.. All rights reserved.

  14. Regional differences between {sup 99m}Tc-ECD and {sup 99m}Tc-HMPAO SPET in perfusion changes with age and gender in healthy adults

    Energy Technology Data Exchange (ETDEWEB)

    Inoue, Kentaro; Nakagawa, Manabu; Goto, Ryoi; Kinomura, Shigeo; Sato, Tachio; Sato, Kazunori; Fukuda, Hiroshi [Department of Nuclear Medicine and Radiology, Institute of Development, Aging and Cancer, Tohoku University, 4-1 Seiryomachi Aoba-ku, 980-8575, Sendai (Japan)

    2003-11-01

    A number of studies using single-photon emission tomography (SPET) have shown perfusion changes with age in several cortical and subcortical areas, which might distort the results of perfusion imaging studies of neuropsychiatric disorders. Technetium-99m labelled ethyl cysteinate dimer (ECD) and hexamethylpropylene amine oxime (HMPAO) are both used as markers of cerebral perfusion, but have different pharmacokinetics and retention patterns. The aim of this study was to determine whether age and gender effects on perfusion SPET differ depending on whether {sup 99m}Tc-HMPAO or {sup 99m}Tc-ECD is used. Forty-five subjects (20 male and 25 female, mean age 52.8{+-}6.6 years) were assigned to {sup 99m}Tc-HMPAO SPET (HMPAO group), and 39 subjects (24 male and 15 female, mean age 52.6{+-}6.7 years) to {sup 99m}Tc-ECD SPET (ECD group). SPET images were obtained about 10 min after intravenous injection of approximately 800 MBq {sup 99m}Tc-HMPAO or {sup 99m}Tc-ECD using the same SPET scanner. Three-dimensional volumetric magnetic resonance imaging was performed to as7sess morphological changes in the grey matter. All image processing and statistical analyses were performed using SPM99 software. An area in the right anterior frontal lobe showed an increase in perfusion with age only in the HMPAO group, whereas areas in the bilateral retrosplenial cortex showed decreases in perfusion with age only in the ECD group; neither group showed corresponding changes in the grey matter. The present study shows that different effects of age on perfusion are observed depending on whether {sup 99m}Tc-HMPAO and {sup 99m}Tc-ECD is used. This suggests that the results of perfusion SPET are differently confounded depending on the tracer used, and that perfusion SPET with these tracers has limitations when used in research on subtle perfusion changes. (orig.)

  15. Noninvasive regional cerebral blood flow measurements at pre- and post-acetazolamide test using 99mTc-ECD

    International Nuclear Information System (INIS)

    Matsuda, Hiroshi; Nakano, Seigo; Tanaka, Masaaki.

    1996-01-01

    A technique for serial noninvasive cerebral blood flow measurements at pre- and post-acetazolamide (Diamox) test was newly developed using 99m Tc-ECD without blood sampling. Baseline mean cerebral blood flow (mCBF) was measured from graphical analysis of time activity curves for brain and aortic arch obtained from radionuclide angiography by injection of 370-555 MBq 99m Tc-ECD. The first SPECT study was performed immediately after intravenous administration of 1 g of Diamox, then baseline regional cerebral blood flow (rCBF) was calculated using Lassen's correction algorithm. Immediately after the stop of the first SPECT study, additional 555-740 MBq of 99m Tc-ECD was administered, thereafter the second SPECT study was started. Post-Diamox SPECT images were obtained by subtraction of the first baseline images from the second images. Using Lassen's algorithm, post-Diamox mCBF was estimated from the baseline mCBF, the baseline mean SPECT counts, and post-Diamox mean SPECT counts corrected for administered dose and imaging time. Post-Diamox rCBF was obtained from the post-Diamox mCBF and the post-Diamox mean SPECT counts using Lassen's algorithm. Coefficient variation was shown 2.7% and 3.5%: mCBF and rCBF, respectively in test-retest results in six patients without Diamox administration. Nine demented patients without vascular disorders showed significant mCBF increase of 35.7% on the average by post-Diamox. In conclusion, this simplified method is practically useful for measuring CBF at pre- and post-Diamox test within short period of time without any blood sample. (author)

  16. {sup 99m}Tc-ECD brain perfusion SPET: variability, asymmetry and effects of age and gender in healthy adults

    Energy Technology Data Exchange (ETDEWEB)

    Van Laere, K.; Versijpt, J.; Goethals, I.; Dierckx, R. [Div. of Nuclear Medicine, Ghent University Hospital (Belgium); Audenaert, K. [Dept. of Psychiatry and Medical Psychology, Ghent University Hospital, Ghent (Belgium); Koole, M. [Medical Signal and Image Processing Department (MEDISIP), Ghent University, Ghent (Belgium); Achten, E. [Division of Neuroradiology, Radiology Department, Ghent University Hospital, Ghent (Belgium)

    2001-07-01

    Reliable and high-resolution reference data for regional cerebral blood flow measured with single-photon emission tomography (SPET) are necessary for optimal clinical and research use. Therefore, a large dataset of normal technetium-99m labelled ethylene cysteine dimer (ECD) perfusion SPET in carefully screened healthy volunteers with an age range spanning six decades was created, with correction for non-uniform attenuation and scatter and based on an anatomically standardised analysis. Eighty-nine healthy volunteers, stratified for gender (46 females, 43 males; age 20-81 years), were included. Twelve volunteers underwent repeated {sup 99m}Tc-ECD SPET after 2.5{+-}2.3 weeks. An automated whole-brain volume of interest analysis with MANOVA as well as voxelwise analysis using SPM99 was conducted. Average intersubject variability was 4.8% while intrasubject reproducibility was 3.0%. An age-related decline in tracer uptake was found in the anterior cingulate gyrus, bilateral basal ganglia, left prefrontal, left lateral frontal and left superior temporal and insular cortex (all P=0.001-0.02). There was an overall increase in right/left asymmetry with age, which was most pronounced in the frontal and temporal neocortex. The most significant correlations between AI and age decade were found in the prefrontal (R=0.35, P=0.001) and superior temporal neocortex (R=0.43, P<0.001). Women had significantly higher uptake in the right parietal cortex (P<0.001), while men showed higher uptake in the cerebellum and the left anterior temporal and orbitofrontal cortex (all P<0.01). This normative dataset allows age- and gender-specific patient and group assessment of {sup 99m}Tc-ECD perfusion SPET under a wide variety of clinical circumstances in relation to normal variations and highlights the importance of both age- and gender-specific normal datasets for optimal analysis sensitivity. (orig.)

  17. 99mTc-ECD brain perfusion SPET: variability, asymmetry and effects of age and gender in healthy adults

    International Nuclear Information System (INIS)

    Van Laere, K.; Versijpt, J.; Goethals, I.; Dierckx, R.; Audenaert, K.; Koole, M.; Achten, E.

    2001-01-01

    Reliable and high-resolution reference data for regional cerebral blood flow measured with single-photon emission tomography (SPET) are necessary for optimal clinical and research use. Therefore, a large dataset of normal technetium-99m labelled ethylene cysteine dimer (ECD) perfusion SPET in carefully screened healthy volunteers with an age range spanning six decades was created, with correction for non-uniform attenuation and scatter and based on an anatomically standardised analysis. Eighty-nine healthy volunteers, stratified for gender (46 females, 43 males; age 20-81 years), were included. Twelve volunteers underwent repeated 99m Tc-ECD SPET after 2.5±2.3 weeks. An automated whole-brain volume of interest analysis with MANOVA as well as voxelwise analysis using SPM99 was conducted. Average intersubject variability was 4.8% while intrasubject reproducibility was 3.0%. An age-related decline in tracer uptake was found in the anterior cingulate gyrus, bilateral basal ganglia, left prefrontal, left lateral frontal and left superior temporal and insular cortex (all P=0.001-0.02). There was an overall increase in right/left asymmetry with age, which was most pronounced in the frontal and temporal neocortex. The most significant correlations between AI and age decade were found in the prefrontal (R=0.35, P=0.001) and superior temporal neocortex (R=0.43, P 99m Tc-ECD perfusion SPET under a wide variety of clinical circumstances in relation to normal variations and highlights the importance of both age- and gender-specific normal datasets for optimal analysis sensitivity. (orig.)

  18. Study of cellular retention of HMPAO and ECD in a model simulating the blood-brain barrier

    International Nuclear Information System (INIS)

    Ponce, C.; Pittet, N.; Slosman, D.O.

    1997-01-01

    The HMPAO and ECD are two technetium-labelled lipophilic agents clinically used in the imagery of cerebral perfusion. These molecules cross the membranes and are retained inside the cell after being converted to a hydrophilic form. The aim of this study is to establish the distribution of this retention at the level of blood-brain barrier (BBB) and nerve cells. The incorporation of HMPAO or ECD was studied on a model of co-culture simulating the BBB by means of a T84 single-cell layer of tight junction separated from another layer of U373 astrocyte cells. The cell quality and tight junction permeability were evaluated by the cellular retention of 111-indium chloride and by para-cellular diffusion of 14 C mannitol,d-1. The values reported below were obtained at 180 minutes when the radiotracers were added near the 'T84 layer'. The cell quality is validated by the low cellular retention of the indium chloride(2.3±0.3 μg -1 for the T84 cells and 8.2±5.8 μg -1 for the U373 cells). The activity of 14 C mannitol,d-1 diminishes by 23 ± 5 % in the added compartment. The retention of ECD by the U373 cells is significantly higher (20.7 ±4.5 g -1 ) than that of T84 cells (2.9 ± 0.2 μg -1 ). For HMPAO a non-significant tendency could be observed (49 ± 34 μg -1 for the U373 cells and 38 ± 25 μg -1 for the T84 cells)> The results of cellular retention of indium by HMPAO or ECD when added near 'U373 layer' are not significantly different.In conclusion, independently of the side exposed to the radiotracers, one observes an enhanced incorporation of the U373 cells. The ensemble of these results represent additional arguments in favour of a specific cellular incorporation of the radiotracers, independent of the BBB permittivity

  19. Domain analysis

    DEFF Research Database (Denmark)

    Hjørland, Birger

    2017-01-01

    The domain-analytic approach to knowledge organization (KO) (and to the broader field of library and information science, LIS) is outlined. The article reviews the discussions and proposals on the definition of domains, and provides an example of a domain-analytic study in the field of art studies....... Varieties of domain analysis as well as criticism and controversies are presented and discussed....

  20. Usefulness of {sup 99m}Tc-ECD brain SPECT in acute onset pediatric CNS diseases. In comparison with CT and MRI

    Energy Technology Data Exchange (ETDEWEB)

    Hashimoto, Teisuke; Chikatsu, Hiroko; Nishiyama, Hiromune; Endo, Hiroko; Kono, Tatsuo; Iimura, Fumitoshi; Kuwashima, Shigeko; Saiki, Natoru; Fujioka, Mutsuhisa [Dokkyo Univ., Mibu, Tochigi (Japan). School of Medicine

    2001-07-01

    The purpose of this study was to assess the usefulness of regional cerebral blood flow (rCBF) measured by {sup 99m}Tc-L, L-ethyl cysteinate dimer (ECD) brain SPECT in the acute onset type of pediatric central nervous system (CNS) diseases. Thirteen children (7 girls, 6 boys, 4 month-12 years of age) who were diagnosed with 9 cases of viral encephalitis, two cases of febrile convulsion and one each of migraine and metabolic disorder underwent {sup 99m}Tc-ECD brain SPECT, CT and/or MRI within one week interval. The incidence of abnormal findings in the 13 patients was 96.4% (30/31) on {sup 99m}Tc-ECD brain SPECT, 17.6% (3/17) on CT and 63.6% (14/22) on MRI. The positive detection rate of {sup 99m}Tc-ECD brain SPECT was statistically (P<0.01 by a {chi}{sup 2} and/or Fisher's exact probability test) higher than those of CT and MRI. And the changes in rCBF were demonstrated. {sup 99m}Tc-ECD brain SPECT is a useful examination for the diagnosis and follow up management in patients with the acute onset type of pediatric CNS diseases. (author)

  1. Extracellular matrix structure.

    Science.gov (United States)

    Theocharis, Achilleas D; Skandalis, Spyros S; Gialeli, Chrysostomi; Karamanos, Nikos K

    2016-02-01

    Extracellular matrix (ECM) is a non-cellular three-dimensional macromolecular network composed of collagens, proteoglycans/glycosaminoglycans, elastin, fibronectin, laminins, and several other glycoproteins. Matrix components bind each other as well as cell adhesion receptors forming a complex network into which cells reside in all tissues and organs. Cell surface receptors transduce signals into cells from ECM, which regulate diverse cellular functions, such as survival, growth, migration, and differentiation, and are vital for maintaining normal homeostasis. ECM is a highly dynamic structural network that continuously undergoes remodeling mediated by several matrix-degrading enzymes during normal and pathological conditions. Deregulation of ECM composition and structure is associated with the development and progression of several pathologic conditions. This article emphasizes in the complex ECM structure as to provide a better understanding of its dynamic structural and functional multipotency. Where relevant, the implication of the various families of ECM macromolecules in health and disease is also presented. Copyright © 2015 Elsevier B.V. All rights reserved.

  2. Donor-estimated GFR as an appropriate criterion for allocation of ECD kidneys into single or dual kidney transplantation.

    Science.gov (United States)

    Snanoudj, R; Rabant, M; Timsit, M O; Karras, A; Savoye, E; Tricot, L; Loupy, A; Hiesse, C; Zuber, J; Kreis, H; Martinez, F; Thervet, E; Méjean, A; Lebret, T; Legendre, C; Delahousse, M

    2009-11-01

    It has been suggested that dual kidney transplantation (DKT) improves outcomes for expanded criteria donor (ECD) kidneys. However, no criteria for allocation to single or dual transplantation have been assessed prospectively. The strategy of DKT remains underused and potentially eligible kidneys are frequently discarded. We prospectively compared 81 DKT and 70 single kidney transplant (SKT) receiving grafts from ECD donors aged >65 years, allocated according to donor estimated glomerular filtration rate (eGFR): DKT if eGFR between 30 and 60 mL/min, SKT if eGFR greater than 60 mL/min. Patient and graft survival were similar in the two groups. In the DKT group, 13/81 patients lost one of their two kidneys due to hemorrhage, arterial or venous thrombosis. Mean eGFR at month 12 was similar in the DKT and SKT groups (47.8 mL/min and 46.4 mL/min, respectively). Simulated allocation of kidneys according to criteria based on day 0 donor parameters such as those described by Remuzzi et al., Andres et al. and UNOS, did not indicate an improvement in 12-month eGFR compared to our allocation based on donor eGFR.

  3. Comparison between CT perfusion and Tc-99m ECD SPECT in the assessment of cerebrovascular reserve: a case study

    International Nuclear Information System (INIS)

    Crouch, J.; Wood, C.; Campbell, A.; McCarthy, M.; Dunne, M.; Bynevelt, M.; Lenzo, N.

    2003-01-01

    Full text: Brain perfusion is sensitively assessed by cerebral SPECT imaging utilising perfusion agents such as Tc-99m HMPAO and Tc-99m ethyl cysteinate dimer (ECD). Positron emission tomography can accurately assess and quantify brain perfusion and MRI can also be used for perfusion assessment. Both MRI and PET however are currently limited by cost and availability. A new technique utilising CT with contrast has been developed to assess and quantitate cerebral perfusion. The technique utilises arterial input information and deconvolution analysis to develop quantifiable measures of perfusion and contrast transit. The technique has been validated for acute stroke assessment and is being assessed for other possible applications. We present a case study comparison of this technique with cerebral SPECT perfusion using Tc-99m ECD in the assessment of cerebrovasular reserve. In each case, the CT and SPECT studies were performed pre- and post-acetazolamide and the SPECT study was statistically compared with a normal database utilising an automated brain perfusion statistical analysis package (NeurostatT). We discuss the correlation found between techniques, their strengths, weaknesses and possible future roles. Copyright (2003) The Australian and New Zealand Society of Nuclear Medicine Inc

  4. Brain perfusion abnormalities in Rett syndrome: a qualitative and quantitative SPET study with 99Tc(m)-ECD.

    Science.gov (United States)

    Burroni, L; Aucone, A M; Volterrani, D; Hayek, Y; Bertelli, P; Vella, A; Zappella, M; Vattimo, A

    1997-06-01

    Rett syndrome is a progressive neurological paediatric disorder associated with severe mental deficiency, which affects only girls. The aim of this study was to determine if brain blood flow abnormalities detected with 99Tc(m)-ethyl-cysteinate-dimer (99Tc[m]-ECD) single photon emission tomography (SPET) can explain the clinical manifestation and progression of the disease. Qualitative and quantitative global and regional brain blood flow was evaluated in 12 girls with Rett syndrome and compared with an aged-matched reference group of children. In comparison with the reference group, SPET revealed a considerable global reduction in cerebral perfusion in the groups of girls with Rett syndrome. A large statistical difference was noted, which was more evident when comparing the control group with girls with stage IV Rett syndrome than girls with stage III Rett syndrome. The reduction in cerebral perfusion reflects functional disturbance in the brain of children with Rett syndrome. These data confirm that 99Tc(m)-ECD brain SPET is sensitive in detecting hypoperfused areas in girls with Rett syndrome that may be associated with brain atrophy, even when magnetic resonance imaging appears normal.

  5. 99mTc-ECD and 99mTc-HM-PAO SPECT in five patients with MELAS

    International Nuclear Information System (INIS)

    Katagiri, Shinako; Nishimaki, Hiroshi; Kitano, Masashi; Horiike, Shigeharu; Kan, Shinichi; Ishii, Katsumi; Matsubayashi, Takashi; Sakai, Fumihiko

    1998-01-01

    Cerebral perfusion was studied in five patients with mitochondrial encephalomyopathy with lactic acidosis and stroke-like episodes syndrome (MELAS), using single photon emission computed tomography (SPECT) with 99m Tc-ethyl cysteinate dimer ( 99m Tc-ECD) or 99m Tc-hexamethyl propyleneamine oxime ( 99m Tc-HM-PAO). In four cases, regional cerebral blood flow (rCBF) was evaluated by the method reported by Mastuda et al. Immediately after the stroke-like episodes, accumulation of the tracer was relatively increased in the temporooccipital lobe, and also increased rCBF was shown in the same area. However, the region showed decreased radioactivity at the chronic stage, and rCBF decreased also. These findings are consistent with positron emission tomography (PET) at the acute stage and autopsy. 99m Tc-ECD SPECT and 99m Tc-HM-PAO SPECT may be useful in the diagnosis and assessment of the progress of the MELAS. (author)

  6. Concordance between (99m)Tc-ECD SPECT and 18F-FDG PET interpretations in patients with cognitive disorders diagnosed according to NIA-AA criteria.

    Science.gov (United States)

    Ito, Kimiteru; Shimano, Yasumasa; Imabayashi, Etsuko; Nakata, Yasuhiro; Omachi, Yoshie; Sato, Noriko; Arima, Kunimasa; Matsuda, Hiroshi

    2014-10-01

    The purpose of this study was to clarify the concordance of diagnostic abilities and interobserver agreement between 18F-fluorodeoxyglucose (FDG) positron emission tomography (PET) and brain perfusion single photon-emission computed tomography (SPECT) in patients with Alzheimer's disease (AD) who were diagnosed according to the research criteria of the National Institute of Aging-Alzheimer's Association Workshop. Fifty-five patients with "AD and mild cognitive impairment (MCI)" (n = 40) and "non-AD" (n = 15) were evaluated with 18F-FDG PET and (99m)Tc-ethyl cysteinate dimer (ECD) SPECT during an 8-week period. Three radiologists independently graded the regional uptake in the frontal, temporal, parietal, and occipital lobes as well as the precuneus/posterior cingulate cortex in both images. Kappa values were used to determine the interobserver reliability regarding regional uptake. The regions with better interobserver reliability between 18F-FDG PET and (99m)Tc-ECD SPECT were the frontal, parietal, and temporal lobes. The (99m)Tc-ECD SPECT agreement in the occipital lobes was not significant. The frontal, temporal, and parietal lobes showed good correlations between 18F-FDG PET and (99m)Tc-ECD SPECT in the degree of uptake, but the occipital lobe and precuneus/posterior cingulate cortex did not show good correlations. The diagnostic accuracy rates of "AD and MCI" ranged from 60% to 70% in both of the techniques. The degree of uptake on 18F-FDG PET and (99m)Tc-ECD SPECT showed significant correlations in the frontal, temporal, and parietal lobes. The diagnostic abilities of 18F-FDG PET and (99m)Tc-ECD SPECT for "AD and MCI," when diagnosed according to the National Institute of Aging-Alzheimer's Association Workshop criteria, were nearly identical. Copyright © 2014 John Wiley & Sons, Ltd.

  7. Concrete domains

    OpenAIRE

    Kahn, G.; Plotkin, G.D.

    1993-01-01

    This paper introduces the theory of a particular kind of computation domains called concrete domains. The purpose of this theory is to find a satisfactory framework for the notions of coroutine computation and sequentiality of evaluation.

  8. Different uptake of 99mTc-ECD adn 99mTc-HMPAO in the same brains: analysis by statistical parametric mapping.

    Science.gov (United States)

    Hyun, Y; Lee, J S; Rha, J H; Lee, I K; Ha, C K; Lee, D S

    2001-02-01

    The purpose of this study was to investigate the differences between technetium-99m ethyl cysteinate dimer (99mTc-ECD) and technetium-99m hexamethylpropylene amine oxime (99mTc-HMPAO) uptake in the same brains by means of statistical parametric mapping (SPM) analysis. We examined 20 patients (9 male, 11 female, mean age 62+/-12 years) using 99mTc-ECD and 99mTc-HMPAO single-photon emission tomography (SPET) and magnetic resonance imaging (MRI) of the brain less than 7 days after onset of stroke. MRI showed no cortical infarctions. Infarctions in the pons (6 patients) and medulla (1), ischaemic periventricular white matter lesions (13) and lacunar infarction (7) were found on MRI. Split-dose and sequential SPET techniques were used for 99mTc-ECD and 99mTc-HMPAO brain SPET, without repositioning of the patient. All of the SPET images were spatially transformed to standard space, smoothed and globally normalized. The differences between the 99mTc-ECD and 99mTc-HMPAO SPET images were statistically analysed using statistical parametric mapping (SPM) 96 software. The difference between two groups was considered significant at a threshold of uncorrected P values less than 0.01. Visual analysis showed no hypoperfused areas on either 99mTc-ECD or 99mTc-HMPAO SPET images. SPM analysis revealed significantly different uptake of 99mTc-ECD and 99mTc-HMPAO in the same brains. On the 99mTc-ECD SPET images, relatively higher uptake was observed in the frontal, parietal and occipital lobes, in the left superior temporal lobe and in the superior region of the cerebellum. On the 99mTc-HMPAO SPET images, relatively higher uptake was observed in the medial temporal lobes, thalami, periventricular white matter and brain stem. These differences in uptake of the two tracers in the same brains on SPM analysis suggest that interpretation of cerebral perfusion is possible using SPET with 99mTc-ECD and 99mTc-HMPAO.

  9. Different uptake of {sup 99m}Tc-ECD and {sup 99m}Tc-HMPAO in the same brains: analysis by statistical parametric mapping

    Energy Technology Data Exchange (ETDEWEB)

    Hyun, I.Y. [Dept. of Nuclear Medicine, Inha University College of Medicine, Incheon (Korea); Lee, J.S.; Lee, D.S. [Dept. of Nuclear Medicine, Seoul National University College of Medicine, Seoul (Korea); Rha, J.H.; Lee, I.K.; Ha, C.K. [Dept. of Neurology, Inha University College of Medicine, Incheon (Korea)

    2001-02-01

    The purpose of this study was to investigate the differences between technetium-99m ethyl cysteinate dimer ({sup 99m}Tc-ECD) and technetium-99m hexamethylpropylene amine oxime ({sup 99m}Tc-HMPAO) uptake in the same brains by means of statistical parametric mapping (SPM) analysis. We examined 20 patients (9 male, 11 female, mean age 62{+-}12 years) using {sup 99m}Tc-ECD and {sup 99m}Tc-HMPAO single-photon emission tomography (SPET) and magnetic resonance imaging (MRI) of the brain less than 7 days after onset of stroke. MRI showed no cortical infarctions. Infarctions in the pons (6 patients) and medulla (1), ischaemic periventricular white matter lesions (13) and lacunar infarction (7) were found on MRI. Split-dose and sequential SPET techniques were used for {sup 99m}Tc-ECD and {sup 99m}Tc-HMPAO brain SPET, without repositioning of the patient. All of the SPET images were spatially transformed to standard space, smoothed and globally normalized. The differences between the {sup 99m}Tc-ECD and {sup 99m}Tc-HMPAO SPET images were statistically analysed using statistical parametric mapping (SPM) 96 software. The difference between two groups was considered significant at a threshold of uncorrected P values less than 0.01. Visual analysis showed no hypoperfused areas on either {sup 99m}Tc-ECD or {sup 99m}Tc-HMPAO SPET images. SPM analysis revealed significantly different uptake of {sup 99m}Tc-ECD and {sup 99m}Tc-HMPAO in the same brains. On the {sup 99m}Tc-ECD SPET images, relatively higher uptake was observed in the frontal, parietal and occipital lobes, in the left superior temporal lobe and in the superior region of the cerebellum. On the {sup 99m}Tc-HMPAO SPET images, relatively higher uptake was observed in the medial temporal lobes, thalami, periventricular white matter and brain stem. These differences in uptake of the two tracers in the same brains on SPM analysis suggest that interpretation of cerebral perfusion is possible using SPET with {sup 99m}Tc-ECD and

  10. Domain Engineering

    Science.gov (United States)

    Bjørner, Dines

    Before software can be designed we must know its requirements. Before requirements can be expressed we must understand the domain. So it follows, from our dogma, that we must first establish precise descriptions of domains; then, from such descriptions, “derive” at least domain and interface requirements; and from those and machine requirements design the software, or, more generally, the computing systems.

  11. Dermal extracellular lipid in birds.

    Science.gov (United States)

    Stromberg, M W; Hinsman, E J; Hullinger, R L

    1990-01-01

    A light and electron microscopic study of the skin of domestic chickens, seagulls, and antarctic penguins revealed abundant extracellular dermal lipid and intracellular epidermal lipid. Dermal lipid appeared ultrastructurally as extracellular droplets varying from less than 1 micron to more than 25 microns in diameter. The droplets were often irregularly contoured, sometimes round, and of relatively low electron density. Processes of fibrocytes were often seen in contact with extracellular lipid droplets. Sometimes a portion of such a droplet was missing, and this missing part appeared to have been "digested away" by the cell process. In places where cells or cell processes are in contact with fact droplets, there are sometimes extracellular membranous whorls or fragments which have been associated with the presence of fatty acids. Occasionally (in the comb) free fat particles were seen in intimate contact with extravasated erythrocytes. Fat droplets were seen in the lumen of small dermal blood and lymph vessels. We suggest that the dermal extracellular lipid originates in the adipocyte layer and following hydrolysis the free fatty acids diffuse into the epidermis. Here they become the raw material for forming the abundant neutral lipid contained in many of the epidermal cells of both birds and dolphins. The heretofore unreported presence and apparently normal utilization of abundant extracellular lipid in birds, as well as the presence of relatively large droplets of neutral lipid in dermal vessels, pose questions which require a thorough reappraisal of present concepts of the ways in which fat is distributed and utilized in the body.

  12. PENGEMBANGAN MEDIA EDUKASI GIZI MELALUI BUKU MEWARNAI UNTUK PESERTA PENDIDIKAN ANAK USIA DINI (PAUD (MEDIA DEVELOPMENT OF NUTRITION EDUCATION THROUGH COLORING BOOKS FOR EARLY CHILDHOOD EDUCATION (ECD

    Directory of Open Access Journals (Sweden)

    Yurista Permanasari

    2013-06-01

    Full Text Available ABSTRACT Early childhood education (ECD is a right time to provide nutrition education. The education can be given through playing patterns suitable for age 2-6 years. Currently there is no active nutrition education media for ECD participants. Therefore it is necessary to develop an active media of nutrition education through colouring book. Colouring activitities can stimulate both the cognitive and motor development of children. This study aimed to develop a colouring book as a nutrition education media for ECD participants and assess the acceptance of the media. This research is a formative study using qualitative design. The sample in this study was children aged 2-6 years, their parents, and teachers from four selected ECD in the city and district of Bogor. The research was conducted in 2 steps. The first step was baseline data collection through observations, group interviews and in-depth interviews to the children, the parents, and the teachers of ECD. Based on the baseline data, the nutrition education materials was prepared and the colouring book was designed. The second step was the assessment of the acceptance and the application of educational media by observing the use of educational media while teachers provided the material in the ECD classroom and the use of colouring books by ECD participants. A group interview was also conducted to see the teacher’s response of the nutrition education media through colouring books, and to find qualitative change in nutrition knowledge of ECD participants. Education materials through colouring books were made in two versions. First version is the books purposed for group A of ECD (< 4 years and second version is for group B of ECD (4-6 years. After the teacher provided the nutrition education materials, nutritional knowledge of ECD participants was better than before. In addition, the media play as an atractive teaching material that stimulated children’s interest. Colouring books can be

  13. Regulation of corneal stroma extracellular matrix assembly.

    Science.gov (United States)

    Chen, Shoujun; Mienaltowski, Michael J; Birk, David E

    2015-04-01

    The transparent cornea is the major refractive element of the eye. A finely controlled assembly of the stromal extracellular matrix is critical to corneal function, as well as in establishing the appropriate mechanical stability required to maintain corneal shape and curvature. In the stroma, homogeneous, small diameter collagen fibrils, regularly packed with a highly ordered hierarchical organization, are essential for function. This review focuses on corneal stroma assembly and the regulation of collagen fibrillogenesis. Corneal collagen fibrillogenesis involves multiple molecules interacting in sequential steps, as well as interactions between keratocytes and stroma matrix components. The stroma has the highest collagen V:I ratio in the body. Collagen V regulates the nucleation of protofibril assembly, thus controlling the number of fibrils and assembly of smaller diameter fibrils in the stroma. The corneal stroma is also enriched in small leucine-rich proteoglycans (SLRPs) that cooperate in a temporal and spatial manner to regulate linear and lateral collagen fibril growth. In addition, the fibril-associated collagens (FACITs) such as collagen XII and collagen XIV have roles in the regulation of fibril packing and inter-lamellar interactions. A communicating keratocyte network contributes to the overall and long-range regulation of stromal extracellular matrix assembly, by creating micro-domains where the sequential steps in stromal matrix assembly are controlled. Keratocytes control the synthesis of extracellular matrix components, which interact with the keratocytes dynamically to coordinate the regulatory steps into a cohesive process. Mutations or deficiencies in stromal regulatory molecules result in altered interactions and deficiencies in both transparency and refraction, leading to corneal stroma pathobiology such as stromal dystrophies, cornea plana and keratoconus. Copyright © 2014 Elsevier Ltd. All rights reserved.

  14. Parieto-occipital hypoperfusion in late whiplash syndrome: first quantitative SPET study using technetium-99m bicisate (ECD)

    International Nuclear Information System (INIS)

    Otte, A.; Ettlin, T.; Fierz, L.; Mueller-Brand, J.

    1996-01-01

    Brain single-photon emission tomography (SPET) with N,N''-1,2-ethylene-diylbis-L-cysteine diethyl ester dihydrochloride (ECD) was performed on ten patients with a clinically high grade late whiplash syndrome and on 11 controls. Two independent readers blinded to the clinical diagnosis were able to separate the ten patients from normal controls. All these patients had qualitative bilateral parieto-occipital hypoperfusion. To confirm this, the perfusion rate of parieto-occipital over global (perfusion index) was calculated after drawing elliptical regions of interest in transversal-oblique slices. The perfusion indices in patients were significantly lower than in controls as tested by the Mann-Whitney U test. This quantitative study proves our recent qualitatively analysed observation. (orig./MG)

  15. Parieto-occipital hypoperfusion in late whiplash syndrome: first quantitative SPET study using technetium-99m bicisate (ECD)

    Energy Technology Data Exchange (ETDEWEB)

    Otte, A. [Inst. of Nuclear Medicine, Univ. Hospital Basel (Switzerland); Ettlin, T. [Rehabilitation Clinic, Rheinfelden (Switzerland); Fierz, L.; Mueller-Brand, J.

    1996-01-01

    Brain single-photon emission tomography (SPET) with N,N``-1,2-ethylene-diylbis-L-cysteine diethyl ester dihydrochloride (ECD) was performed on ten patients with a clinically high grade late whiplash syndrome and on 11 controls. Two independent readers blinded to the clinical diagnosis were able to separate the ten patients from normal controls. All these patients had qualitative bilateral parieto-occipital hypoperfusion. To confirm this, the perfusion rate of parieto-occipital over global (perfusion index) was calculated after drawing elliptical regions of interest in transversal-oblique slices. The perfusion indices in patients were significantly lower than in controls as tested by the Mann-Whitney U test. This quantitative study proves our recent qualitatively analysed observation. (orig./MG)

  16. A preliminary report of 99Tcm-ECD brain SPECT imaging in patients with acquired immunodeficiency syndrome

    International Nuclear Information System (INIS)

    Xu Hao; Tong Yuwei; Luo Jinxiang; Chen Jian; Wu Qiulian

    2001-01-01

    Objective: To investigate the changes of regional cerebral blood flow (rCBF) in patients with acquired immunodeficiency syndrome (AIDS). Methods: 99 Tc m -ECD brain SPECT imaging was performed on 5 patients with AIDS and 16 sex- and age-matched normal controls. The rCBF percentages compared to the cerebellum were calculated using a semi-quantitative processing software. Results: Hypo-perfusions in the right and left frontal, temporal, parietal lobe, basal ganglia and left thalamus were seen in 1 patient with dementia. Hypo-perfusions in the right and left frontal and temporal lobe were seen in 4 patients without dementia. The rCBF in the right and left frontal, temporal, parietal lobe, basal ganglia and thalamus, straight gyri and pons decreased significantly in patients with AIDS than those of the control subjects (P < 0.01). Conclusion: There is reduced cortico-subcortical rCBF in patients with AIDS

  17. A quantitative study of brain perfusion patterns of 99mTc-ECD SPECT in children with developmental disabilities

    International Nuclear Information System (INIS)

    Hirano, Keiko; Aiba, Hideo; Oguro, Katsuhiko

    2004-01-01

    The aim of this study was to investigate the relationship between developmental disabilities and brain perfusion patterns. We performed technetium-99m-ethylcysteinate dimer ( 99m Tc-ECD) single photon emission computed tomography (SPECT) in 30 children with neurological disorders using the Patlak plot method. In children without developmental disabilities, the distribution of regional cortical perfusion evolved in relation to brain maturation. At one month of age, there was a predominant uptake in the perirolandic cortex. Radionuclide uptake in both the parietal and occipital cortices became evident by three months. Uptake in the temporal and frontal cortex increased by 6 and 11 months, respectively. Brain perfusion showed a pattern similar to that of adults by two years of age at the latest. In children with developmental disabilities, developmental changes of brain perfusion were delayed compared to normally developing children. Brain SPECT is a useful tool to assess the brain maturation in children with developmental disabilities. (author)

  18. A quantitative study of cerebral blood flow in childhood using {sup 99m}Tc-ECD patlak-plot method

    Energy Technology Data Exchange (ETDEWEB)

    Yamamori, Hiroyuki; Kohira, Ryutaro; Fujita, Yukihiko [Nihon Univ., Tokyo (Japan). School of Medicine] (and others)

    2001-12-01

    Mean cerebral blood flow (mCBF) was measured by SPECT using the {sup 99m}Tc-ECD Patlak-Plot method in 54 subjects (age: 1 to 14 years). Regional cerebral blood flow (rCBF) values were obtained using Lassen's correction algorithm. The mCBF values were 52.2 ml/100 g/min in those aged 1-4 years, 49.3 ml/100 g/min in those aged 5-9 years, and 48.1 ml/100 g/min in those aged 10-14 years. There was a slight decrease with advancing age. The rCBF in the frontal region was lower than in other regions in those aged 1-4 years. The Patlak-Plot method is suitable for examination of children, because it does not require arterial blood sampling. (author)

  19. Extracellular glycosylphosphatidylinositol-anchored mannoproteins and proteases of Cryptococcus neoformans.

    Science.gov (United States)

    Eigenheer, Richard A; Jin Lee, Young; Blumwald, Eduardo; Phinney, Brett S; Gelli, Angie

    2007-06-01

    Extracellular proteins of Cryptococcus neoformans are involved in the pathogenesis of cryptococcosis, and some are immunoreactive antigens that may potentially serve as candidates for vaccine development. To further study the extracellular proteome of the human fungal pathogen Cry. neoformans, we conducted a proteomic analysis of secreted and cell wall-bound proteins with an acapsular strain of Cry. neoformans. Proteins were identified from both intact cells and cell walls. In both cases, extracellular proteins were removed with trypsin or beta-glucanase, and then all proteins/peptides were purified by solid-phase extraction, spin dialysis, and HPLC, and identified by liquid chromatography-mass spectrometry. This study identified 29 extracellular proteins with a predicted N-terminal signal sequence and also a predicted glycosylphosphatidylinositol anchor motif in more than half. Among the novel proteins identified were five glycosylphosphatidylinositol-anchored proteins with extensive Ser/Thr-rich regions but no apparent functional domains, a glycosylphosphatidylinositol-anchored aspartic protease, and a metalloprotease with structural similarity to an elastinolytic metalloprotease of Aspergillus fumigatus. This study suggests that Cry. neoformans has the machinery required to target glycosylphosphatidylinositol-anchored proteins to the cell wall, and it confirms the extracellular proteolytic ability of Cry. neoformans.

  20. Analysis of persistent organic pollutants (POPs) in sediments by GC/ECD generated by nuclide {sup 63}Ni

    Energy Technology Data Exchange (ETDEWEB)

    Mesquita, Katia A.; Amaral, Priscila O.; Bustillos, José O. V., E-mail: ovega@ipen.br [Instituto de Pesquisas Energéticas e Nucleares (IPEN/CNEN-SP), São Paulo, SP (Brazil)

    2017-07-01

    Persistent Organic Pollutants (POPs) are toxic organic compounds resistant to environmental degradation. Besides, POPs bioaccumulation with potential significant impacts on human health and the environment. The Global Monitoring Plan (GMP) under the Stockholm Convention on POPs is a program that enables collection of comparable monitoring data from all regions of the world to assess the effectiveness in minimizing human and environmental exposure to POPs. The scope of this work is to develop and validate a method for the extraction and determination of POPs in sediments collected at Billings dam locate in São Paulo Metropolitan City. The compounds studied in this work are: Aldrin (C{sub 12}H{sub 8}C{sub l6}), DDD (C{sub 14}H{sub 10}C{sub l4}), DDE (C{sub 14}H{sub 8}Cl{sub l4)}, DDT (C{sub 14}H{sub 9}C{sub l5}), Dieldrin (C{sub 12}H{sub 8}C{sub l6}O), Endrin (C{sub 12}H{sub 8}Cl{sub 6}O) and Heptachlor (C{sub 10}H{sub 5}C{sub l7}). This work use the QuEChERS extraction method (Quick, Easy, Cheap, Effective, Rugged, and Safe) for sediment samples and the analysis of the compounds were carried out by gas chromatography with the electron capture detector (GC/ECD). The ECD detector ionizes the analytes by the beta particles from the nuclide sources {sup 63}Ni within carrier gas N{sub 2}. The electrons produced in this process are collected, create an amplified current, and generating the chromatographic peak. The recovery of this method obtained values between 57% and 65% and the Relative Standard Deviation (RSD) values are between 1 and 4%. Details of the analytical method beside quantitative analyses data are presented in this work. (author)

  1. Advance prediction of mild cognitive impairment (MCI) using 99mTc-ECD SPECT brain blood flow imaging

    International Nuclear Information System (INIS)

    Kawasaki, Yohsuke

    2008-01-01

    Mild Cognitive Impairment (MCI) is considered as a precursor state of Alzheimer disease (AD). Single photon emission computed tomography (SPECT) brain blood flow imaging was investigated in MCI and it's relevance to the prognosis of MCI was evaluated in an attempt define the characteristics of brain blood flow imaging of MCI (amnestic MCI; aMCI) converting to AD. Ninety-two patients over 60 years old with amnesia were studied. 99m Tc-ethyl cysteinate dimer (ECD) SPECT brain blood flow examinations of the subject under drug-free conditions were conducted and imaging was analyzed according to the first clinical diagnosis. Patients given a diagnosis of MCI on the first clinical diagnosis, were examined again after 2 years and the SPECT imaging before 2 years previously was classified and analyzed. Of them, there were 35 MCI patients, converting of 13 AD patients (37.1%; aMCI), 10 MCI patients (28.6%; non-converter), 4 depression patients (11.4%; Depression type MCI (dMCI)), 1 Geriatric psychosis patient, but 7 patients dropped out. In the aMCI group, relative hypoperfusion was recognized in the posterior cingulate and the precuneus. In the dMCI group, relative hypoperfusion was recognized in the dorsolateral prefrontal cortex (DLPFC) and the anterior cingulate. In the non-converter group, relative hypoperfusion was recognized in the basal forebrain. The hypoperfusion of the precuneus in aMCI, and the hypoperfusion of the right frontal lobe (DLPFC, dorsal-anterior cingulate) in dMCI were characteristic brain blood-flow abnormalities. We believe 99m Tc-ECD SPECT brain blood flow imaging to be useful in the diagnosis of aMCI and in the early detection of depression. (author)

  2. Analysis of persistent organic pollutants (POPs) in sediments by GC/ECD generated by nuclide 63Ni

    International Nuclear Information System (INIS)

    Mesquita, Katia A.; Amaral, Priscila O.; Bustillos, José O. V.

    2017-01-01

    Persistent Organic Pollutants (POPs) are toxic organic compounds resistant to environmental degradation. Besides, POPs bioaccumulation with potential significant impacts on human health and the environment. The Global Monitoring Plan (GMP) under the Stockholm Convention on POPs is a program that enables collection of comparable monitoring data from all regions of the world to assess the effectiveness in minimizing human and environmental exposure to POPs. The scope of this work is to develop and validate a method for the extraction and determination of POPs in sediments collected at Billings dam locate in São Paulo Metropolitan City. The compounds studied in this work are: Aldrin (C 12 H 8 C l6 ), DDD (C 14 H 10 C l4 ), DDE (C 14 H 8 Cl l4) , DDT (C 14 H 9 C l5 ), Dieldrin (C 12 H 8 C l6 O), Endrin (C 12 H 8 Cl 6 O) and Heptachlor (C 10 H 5 C l7 ). This work use the QuEChERS extraction method (Quick, Easy, Cheap, Effective, Rugged, and Safe) for sediment samples and the analysis of the compounds were carried out by gas chromatography with the electron capture detector (GC/ECD). The ECD detector ionizes the analytes by the beta particles from the nuclide sources 63 Ni within carrier gas N 2 . The electrons produced in this process are collected, create an amplified current, and generating the chromatographic peak. The recovery of this method obtained values between 57% and 65% and the Relative Standard Deviation (RSD) values are between 1 and 4%. Details of the analytical method beside quantitative analyses data are presented in this work. (author)

  3. Incorporation of Tenascin-C into the Extracellular Matrix by Periostin Underlies an Extracellular Meshwork Architecture*

    Science.gov (United States)

    Kii, Isao; Nishiyama, Takashi; Li, Minqi; Matsumoto, Ken-ichi; Saito, Mitsuru; Amizuka, Norio; Kudo, Akira

    2010-01-01

    Extracellular matrix (ECM) underlies a complicated multicellular architecture that is subjected to significant forces from mechanical environment. Although various components of the ECM have been enumerated, mechanisms that evolve the sophisticated ECM architecture remain to be addressed. Here we show that periostin, a matricellular protein, promotes incorporation of tenascin-C into the ECM and organizes a meshwork architecture of the ECM. We found that both periostin null mice and tenascin-C null mice exhibited a similar phenotype, confined tibial periostitis, which possibly corresponds to medial tibial stress syndrome in human sports injuries. Periostin possessed adjacent domains that bind to tenascin-C and the other ECM protein: fibronectin and type I collagen, respectively. These adjacent domains functioned as a bridge between tenascin-C and the ECM, which increased deposition of tenascin-C on the ECM. The deposition of hexabrachions of tenascin-C may stabilize bifurcations of the ECM fibrils, which is integrated into the extracellular meshwork architecture. This study suggests a role for periostin in adaptation of the ECM architecture in the mechanical environment. PMID:19887451

  4. Incorporation of tenascin-C into the extracellular matrix by periostin underlies an extracellular meshwork architecture.

    Science.gov (United States)

    Kii, Isao; Nishiyama, Takashi; Li, Minqi; Matsumoto, Ken-Ichi; Saito, Mitsuru; Amizuka, Norio; Kudo, Akira

    2010-01-15

    Extracellular matrix (ECM) underlies a complicated multicellular architecture that is subjected to significant forces from mechanical environment. Although various components of the ECM have been enumerated, mechanisms that evolve the sophisticated ECM architecture remain to be addressed. Here we show that periostin, a matricellular protein, promotes incorporation of tenascin-C into the ECM and organizes a meshwork architecture of the ECM. We found that both periostin null mice and tenascin-C null mice exhibited a similar phenotype, confined tibial periostitis, which possibly corresponds to medial tibial stress syndrome in human sports injuries. Periostin possessed adjacent domains that bind to tenascin-C and the other ECM protein: fibronectin and type I collagen, respectively. These adjacent domains functioned as a bridge between tenascin-C and the ECM, which increased deposition of tenascin-C on the ECM. The deposition of hexabrachions of tenascin-C may stabilize bifurcations of the ECM fibrils, which is integrated into the extracellular meshwork architecture. This study suggests a role for periostin in adaptation of the ECM architecture in the mechanical environment.

  5. Ictal 99mTc-ECD brain SPECT imaging: localization of seizure foci and correlation with semiology in temporal lobe epilepsy

    International Nuclear Information System (INIS)

    Kang, Do Young; Ryu, Jin Sook; Lee, Hee Kyung; Ma, Hyeo Il; Lee, Sang Ahm; Lee, Jung Kyo; Kang, Joong Koo

    1997-01-01

    The purpose of this study was to evaluate the usefulness of ictal 99m Tc-ECD brain SPECT in temporal lobe epilepsy (TLE) patients for presurgical localization of seizure foci, and to correlate ictal SPECT patterns with the semiology of seizure. ictal 99m Tc-ECD Brain SPECT was performed in 23 TLE patients whose MRI showed unilateral hippocampal atrophy (18 patients), other focal temporal lesions (4 patients) and normal finding (1 patient). Under CCTV monitoring, injection was done during ictal period in all patients with the mean delay of 38.5±17.3 sec (mean seizure duration : 90.5±35.9 sec). Ictal 99m Tc-ECD Brain SPECT was visually analysed by three blinded observers. All patients underwent temporal lobectomy with a minimum 3 months follow-up (range 3-29 months) ; all had good post-surgical seizure control (Engel's calssification class I). Ictal 99m Tc-ECD Brain SPECT showed unilateral temporal hyperperfusion concordant with epileptic foci in 22/23 (95.7%), whereas non-lateralization in 1/23 (4.3%). The hyperperfusion of the ipsilateral basal ganglia was present in 72.7% (16/22) of patients with dystonic/tonic posture of the contralateral hand. The contralateral cerebellar hyperperfusion was observed in the 7/22 (32%). The group with secondary generalized tonic clonic seizure (GTC) had brain stem and bilateral thalamic hyperperfusion in 4/7 (57.1%) while the group without secondary GTC had the same hyperperfusion in 1/16 (6.3%). There was statistically significant difference in brain stem and bilateral thalamic perfusion between two groups. Ictal 99m Tc-ECD Brain SPECT is a useful modality in pre-surgical localization of the epileptic foci and well correlated with the semiology of seizure

  6. {sup 99m}Tc-HMPAO and {sup 99m}Tc-ECD perform differently in typically hypoperfused areas in Alzheimer's disease

    Energy Technology Data Exchange (ETDEWEB)

    Koulibaly, Pierre Malick [Nuclear Medicine Department, Centre Antoine Lacassagne, University of Nice-Sophia Antipolis (France); Laboratoire de Biophysique, Universite de Nice-Sophia Antipolis, UFR de Medecine, 28 Avenue de Valombrose, 06107, Nice Cedex 2 (France); Nobili, Flavio; Vitali, Paolo; Girtler, Nicola; Rodriguez, Guido [Clinical Neurophysiology, Department of Internal Medicine, University of Genoa (Italy); Migneco, Octave; Darcourt, Jacques [Nuclear Medicine Department, Centre Antoine Lacassagne, University of Nice-Sophia Antipolis (France); Robert, Philippe H. [Memory Center, Federation of Clinical Neuroscience, Centre Hospitalier Universitaire, University of Nice-Sophia Antipolis (France)

    2003-07-01

    Technetium-99m hexamethylpropylene amine oxime (HMPAO) and {sup 99m}Tc-N,N''-1,2-ethylene diylbis-l-cysteine diethyl ester dihydrochloride (ECD) yield significantly different images of cerebral perfusion owing to their particular pharmacokinetics. The aim of this study was to assess the topography, extension and statistical significance of these differences in Alzheimer's disease (AD). Sixty-four patients with mild to moderate AD were retrospectively selected by two European centres. Two series of patients, including 32 studied with {sup 99m}Tc-HMPAO single-photon emission tomography (SPET) and 32 studied with {sup 99m}Tc-ECD SPET, were matched for sex, age ({+-}3 years) and severity of cognitive impairment as assessed by the Mini-Mental State Examination (MMSE) ({+-}2 points), following a case-control procedure. SPET data were processed using SPM99 software (uncorrected height threshold: P=0.001). {sup 99m}Tc-ECD SPET gave significantly higher uptake ratio values than {sup 99m}Tc-HMPAO SPET in several symmetrical clusters, including the right and left occipital cuneus, the left occipital and parietal precuneus, and the left superior and middle temporal gyri. {sup 99m}Tc-HMPAO SPET gave significantly higher uptake ratio values than ECD in two smaller clusters, including the hippocampus in both hemispheres. In AD, relative brain uptake of {sup 99m}Tc-HMPAO and {sup 99m}Tc-ECD is different in several brain regions, some of which are typically involved in AD, such as the precuneus and the hippocampus. These differences confirm the need for specific normal databases, but their impact on routine SPET reports in AD is not known and deserves an ad hoc investigation. (orig.)

  7. Extracellular vesicles: Exosomes, microvesicles, and friends

    NARCIS (Netherlands)

    Raposo, G.; Stoorvogel, W.|info:eu-repo/dai/nl/074352385

    2013-01-01

    Cells release into the extracellular environment diverse types of membrane vesicles of endosomal and plasma membrane origin called exosomes and microvesicles, respectively. These extracellular vesicles (EVs) represent an important mode of intercellular communication by serving as vehicles for

  8. Extracellular secretion of recombinant proteins

    Science.gov (United States)

    Linger, Jeffrey G.; Darzins, Aldis

    2014-07-22

    Nucleic acids encoding secretion signals, expression vectors containing the nucleic acids, and host cells containing the expression vectors are disclosed. Also disclosed are polypeptides that contain the secretion signals and methods of producing polypeptides, including methods of directing the extracellular secretion of the polypeptides. Exemplary embodiments include cellulase proteins fused to secretion signals, methods to produce and isolate these polypeptides, and methods to degrade lignocellulosic biomass.

  9. Follow-up of pain processing recovery after ketamine in hyperalgesic fibromyalgia patients using brain perfusion ECD-SPECT

    Energy Technology Data Exchange (ETDEWEB)

    Guedj, Eric; Cammilleri, Serge; Colavolpe, Cecile; Laforte, Catherine de; Mundler, Olivier [Assistance Publique des Hopitaux de Marseille, Centre Hospitalo-Universitaire de la Timone, Service Central de Biophysique et de Medecine Nucleaire, Marseille, Cedex 5 (France); Niboyet, Jean [Clinique La Phoceanne, Unite d' Etude et de Traitement de la Douleur, Marseille (France)

    2007-12-15

    The aim of this study was to determine whether the follow-up of pain processing recovery in hyperalgesic fibromyalgia (FM) could be objectively evaluated with brain perfusion ethyl cysteinate dimer single photon computerized tomography (ECD-SPECT) after administration of ketamine. We enrolled 17 hyperalgesic FM women patients (48.5 {+-} 11 years, range 25-63). After treatment with subcutaneous ketamine, 11 patients were considered as 'good responders', with a decrease in pain intensity, evaluated by visual analog scale (VAS), greater than 50%. On the other hand, six patients were considered as 'poor responders'. A voxel-based analysis of regional cerebral blood flow (rCBF) was conducted (p{sub voxel} < 0.001uc), in the two subgroups of patients, before and after treatment, in comparison to a group of ten healthy subjects, matched for age and gender. In comparison to baseline brain SPECT, midbrain rCBF showed a greater increase after ketamine in the responder group than in the nonresponder group (p{sub cluster} = 0.016c). In agreement with the clinical response, the change in midbrain rCBF after ketamine was highly correlated with the reduction of VAS pain score (r = 0.7182; p = 0.0041). This prospective study suggests that blockade of facilitatory descending modulation of pain with ketamine can be evaluated in the periaqueductal grey with brain perfusion SPECT. (orig.)

  10. Follow-up of pain processing recovery after ketamine in hyperalgesic fibromyalgia patients using brain perfusion ECD-SPECT

    International Nuclear Information System (INIS)

    Guedj, Eric; Cammilleri, Serge; Colavolpe, Cecile; Laforte, Catherine de; Mundler, Olivier; Niboyet, Jean

    2007-01-01

    The aim of this study was to determine whether the follow-up of pain processing recovery in hyperalgesic fibromyalgia (FM) could be objectively evaluated with brain perfusion ethyl cysteinate dimer single photon computerized tomography (ECD-SPECT) after administration of ketamine. We enrolled 17 hyperalgesic FM women patients (48.5 ± 11 years, range 25-63). After treatment with subcutaneous ketamine, 11 patients were considered as ''good responders'', with a decrease in pain intensity, evaluated by visual analog scale (VAS), greater than 50%. On the other hand, six patients were considered as ''poor responders''. A voxel-based analysis of regional cerebral blood flow (rCBF) was conducted (p voxel cluster = 0.016c). In agreement with the clinical response, the change in midbrain rCBF after ketamine was highly correlated with the reduction of VAS pain score (r = 0.7182; p = 0.0041). This prospective study suggests that blockade of facilitatory descending modulation of pain with ketamine can be evaluated in the periaqueductal grey with brain perfusion SPECT. (orig.)

  11. Comparison of acetazolamide-enhanced brain SPECT using Tc-99m ECD with cerebral angiography in patients with cerebrovascular disease

    International Nuclear Information System (INIS)

    Choi, Y. Y.; Moon, D. H.; Ryu, J. S.; Yang, S. H.; Lee, H. K.; Lee, J. H.; Kim, J. S.; Kim, K. A.

    1997-01-01

    Cerebral vascular reserve can be assessed by development of collateral channels (DCC) on cerebral angiography(CA) or vasoreactivity (VR) on acetazolamide-enhanced brain SPECT (ACZ-SPECT). The purpose of this study was to compare Tc-99m ECD ACZ-SPECT with CA in the evaluation of vascular reserve in patients (pts) with cerebrovascular disease(CVD). Twenty seven patients with CVD, including TIA (n=13), infarction (n=11) and asymptomatic pts (AS, n=3), underwent CA and ACZ-SPECT. Basal and ACZ-SPECT was obtained consecutively, and image subtraction was performed. On CA, degree of DCC was scored 0-3 (0: normal, 3: poor) according to parenchymal staining on delayed film. In ACZ-SPECT, decrease of VR was graded 0-3 (0: normal, 3: more than 30% decrease). The correlation between degree of stenosis, DCC and VR were analyzed. 1) Variable degree of VR or DCC was observed in totally occluded or stenotic cerebral arterial territories. 2) In arterial territories with poor DCC, ACZ-SPECT showed poor VR. However, in 5 out of 11 TIA or AS with good DCC, poor VR was observed. These data suggests that 1) cerebral hemodynamic status cannot be assessed by the degree of stenosis on CA alone. 2) DCC may overestimate the cerebral vascular reserve in patients with TIA or AS. 3) ACZ-SPECT plays a complementary role to CA for evaluation of cerebral hemodynamic status in pts with CVD

  12. Evaluation of regional cerebral blood flow changes in normal aging using 99mTc-ECD SPECT and Patlak method

    International Nuclear Information System (INIS)

    Mizuno, Shinji; Nandate, Yuka; Enya, Mayumi; Manabe, Tomoko; Goto, Hiroo; Hoshi, Hiroaki; Takahashi, Yukitoshi; Kato, Zenichiro; Kondo, Naomi

    1998-01-01

    Changes in regional cerebral blood flow (rCBF) were studied using 99m Tc-ECD SPECT and Patlak method in 47 subjects, aged 1 to 79 years (mean 28.5 years). No abnormal finding was found on MR imaging in the subjects. We obtained mean cerebral blood flow (mCBF) by means of Patlak method. rCBF was measured by positioning regions of interest (ROIs) in frontal lobe, parietal lobe, temporal lobe, occipital lobe, cerebellum and thalamus. The correlation coefficient between ages and mCBF was r=-0.897 (p<0.0001). mCBF of children was significantly higher (p<0.05), compared with adults. By dispersion analysis according to the groups of ages, significantly higher rCBF values were available in the frontal, parietal, temporal and occipital lobes during childhood than adulthood (p<0.05), but there were almost no significant differences of rCBF during childhood than adulthood in cerebellum and thalamus. (author)

  13. Reduction in mean cerebral blood flow measurements using {sup 99m}Tc-ECD-SPECT during normal aging

    Energy Technology Data Exchange (ETDEWEB)

    Kawahata, Nobuya; Daitoh, Nobuyuki; Shirai, Fumie; Hara, Shigeru [Narita Memorial Hospital, Toyohashi, Aichi (Japan)

    1997-10-01

    Mean cerebral blood flow (mCBF) was measured by SPECT using the {sup 99m}Tc-ECD-Patlak-Plot method in a selected group of 61 normal non-hospitalized subjects aged 51 to 91 years. The mCBF values showed 48.4{+-}4.7 ml/100 g/min in aged 50-59 years group, 49.9{+-}5.9 ml/100 g/min in aged 60-69 years group, 46.4 {+-}6.5 ml/100 g/min in aged 70-79 group, 38.0{+-}3.7 ml/100 g/min in aged 80-89 years group, 38.9 ml/100 g/min in aged 90-99 years group. There was a statistically significant reduction of mCBF with advancing age (R=-0.41; p=0.001). Women have significantly higher mCBF values than men up to aged 70 years group. In this study, there was no significant laterality in the mCBF between right and left hemispheres in all decade groups. The history of hypertension, alcohol consumption, and cigarette smoking failed to show significant difference in the mCBF values. The present study shows that normal aging is associated with mCBF reduction. (author)

  14. The study on regional brain blood flow in the patients with Parkinson's disease using 99Tcm-ECD SPECT

    International Nuclear Information System (INIS)

    Sun Da; Ye Xiaojuan; Zhan Hongwei; Xu Wei; Bao Chengkan

    2010-01-01

    Purpose The aim of this study is to evaluate the changes of brain blood floe in patients with Parkinson's disease (PD) and to investigate the clinical characteristics of the patients with PD correlate with rCBF. Methods: Regional cerebral perfusion was investigated using SPECT in 34 patients with PD . The mean ages of the patients were 56.61±11.04 Years old. The course of disease in most patients was from 1 to over 20 years. Results: 94.1 per cent of patients (32/3) had a significant decrease of rCBF in the basal ganglia, frontal lobes, temporal lobes and thalamus. Parietal and occipital cortex were involved in some patients. The decrease of rCBF in the basal ganglia is unilateral in most patients with PD. There were over 3 brain regions that Conclusion: According to our results, patients with PD had decreased rCBF in the basal ganglia, frontal and temporal cortices. These may reflect a fundamental feature of clinical neuropathophysiology in PD. 99 Tc m -ECD SPECT imaging is helpful to the diagnosis of PD and may help investigate the potential pathophysiology of PD. (authors)

  15. Environmental contaminants of honeybee products in Uganda detected using LC-MS/MS and GC-ECD.

    Directory of Open Access Journals (Sweden)

    Deborah Ruth Amulen

    Full Text Available Pollinator services and the development of beekeeping as a poverty alleviating tool have gained considerable focus in recent years in sub-Saharan Africa. An improved understanding of the pervasive environmental extent of agro-chemical contaminants is critical to the success of beekeeping development and the production of clean hive products. This study developed and validated a multi-residue method for screening 36 pesticides in honeybees, honey and beeswax using LC-MS/MS and GC-ECD. Of the 36 screened pesticides, 20 were detected. The highest frequencies occurred in beeswax and in samples from apiaries located in the proximity of citrus and tobacco farms. Fungicides were the most prevalent chemical class. Detected insecticides included neonicotinoids, organophosphates, carbamates, organophosphorus, tetrazines and diacylhydrazines. All detected pesticide levels were below maximum residue limits (according to EU regulations and the lethal doses known for honeybees. However, future risk assessment is needed to determine the health effects on the African genotype of honeybees by these pesticide classes and combinations of these. In conclusion, our data present a significant challenge to the burgeoning organic honey sector in Uganda, but to achieve this, there is an urgent need to regulate the contact routes of pesticides into the beehive products. Interestingly, the "zero" detection rate of pesticides in the Mid-Northern zone is a significant indicator of the large potential to promote Ugandan organic honey for the export market.

  16. Reduction in mean cerebral blood flow measurements using 99mTc-ECD-SPECT during normal aging

    International Nuclear Information System (INIS)

    Kawahata, Nobuya; Daitoh, Nobuyuki; Shirai, Fumie; Hara, Shigeru

    1997-01-01

    Mean cerebral blood flow (mCBF) was measured by SPECT using the 99m Tc-ECD-Patlak-Plot method in a selected group of 61 normal non-hospitalized subjects aged 51 to 91 years. The mCBF values showed 48.4±4.7 ml/100 g/min in aged 50-59 years group, 49.9±5.9 ml/100 g/min in aged 60-69 years group, 46.4 ±6.5 ml/100 g/min in aged 70-79 group, 38.0±3.7 ml/100 g/min in aged 80-89 years group, 38.9 ml/100 g/min in aged 90-99 years group. There was a statistically significant reduction of mCBF with advancing age (R=-0.41; p=0.001). Women have significantly higher mCBF values than men up to aged 70 years group. In this study, there was no significant laterality in the mCBF between right and left hemispheres in all decade groups. The history of hypertension, alcohol consumption, and cigarette smoking failed to show significant difference in the mCBF values. The present study shows that normal aging is associated with mCBF reduction. (author)

  17. The anchorless adhesin Eap (extracellular adherence protein) from Staphylococcus aureus selectively recognizes extracellular matrix aggregates but binds promiscuously to monomeric matrix macromolecules

    NARCIS (Netherlands)

    Hansen, Uwe; Hussain, Muzaffar; Villone, Daniela; Herrmann, Mathias; Robenek, Horst; Peters, Georg; Sinha, Bhanu; Bruckner, Peter

    Besides a number of cell wall-anchored adhesins, the majority of Staphylococcus aureus strains produce anchorless, cell wall-associated proteins, such as Eap (extracellular adherence protein). Eap contains four to six tandem repeat (EAP)-domains. Eap mediates diverse biological functions, including

  18. A limitation of the split-dose method for evaluating rCBF changes using {sup 99m}Tc-ECD and SPECT

    Energy Technology Data Exchange (ETDEWEB)

    Odano, Ikuo; Takahashi, Makoto; Noguchi, Eikichi; Ohtaki, Hiro; Shibaki, Mitsurou; Kasahara, Tosifumi; Hatano, Masayoshi [Niigata Univ. (Japan). School of Medicine; Ohkubo, Masaki

    1997-05-01

    The purpose of the study is to validate the split-dose method corrected with dose ratio of {sup 99m}Tc-ECD for brain perfusion scan. A dose of 600 MBq of {sup 99m}Tc-ECD was divided into two with various dose ratios from 1 : 1 to 1 : 4, and injected to eleven patients with various cerebral diseases. A lesser dose of {sup 99m}Tc-ECD was injected under a control state for the first SPECT scan, and 15 min SPECT scan was performed 10 min after injection with a triple-head high resolution gamma camera. After the scan, the other dose of {sup 99m}Tc-ECD was injected under the same control state and the second SPECT scan was performed as same as above. A ratio of the activity of the first scan to the net activity of the second scan corrected by dose ratio, defined as K, was measured in brain regions of each subject. Expected value of K was 1, but the value was distributed with large variations in each subject. The mean % error of the K value was 10.4{+-}4.9%. Hence it is considered that activity changes by more than 20% from the control values should be required to detect a significant rCBF change in an activation SPECT study. Then, we proposed a new method in which the activity of both two SPECT scans was normalized by cerebellar or occipital activity and compared. The ratio obtained by the proposed method came closer to 1 with less variations and with less mean % error in comparison with those of K value obtained by the dose-correction method. Although the proposed method has a limitation in the use of an activation study loaded with Diamox, it may be useful to evaluate an alteration of rCBF in the study such as postural testing or finger-moving test. (author)

  19. A limitation of the split-dose method for evaluating rCBF changes using 99mTc-ECD and SPECT

    International Nuclear Information System (INIS)

    Odano, Ikuo; Takahashi, Makoto; Noguchi, Eikichi; Ohtaki, Hiro; Shibaki, Mitsurou; Kasahara, Tosifumi; Hatano, Masayoshi; Ohkubo, Masaki.

    1997-01-01

    The purpose of the study is to validate the split-dose method corrected with dose ratio of 99m Tc-ECD for brain perfusion scan. A dose of 600 MBq of 99m Tc-ECD was divided into two with various dose ratios from 1 : 1 to 1 : 4, and injected to eleven patients with various cerebral diseases. A lesser dose of 99m Tc-ECD was injected under a control state for the first SPECT scan, and 15 min SPECT scan was performed 10 min after injection with a triple-head high resolution gamma camera. After the scan, the other dose of 99m Tc-ECD was injected under the same control state and the second SPECT scan was performed as same as above. A ratio of the activity of the first scan to the net activity of the second scan corrected by dose ratio, defined as K, was measured in brain regions of each subject. Expected value of K was 1, but the value was distributed with large variations in each subject. The mean % error of the K value was 10.4±4.9%. Hence it is considered that activity changes by more than 20% from the control values should be required to detect a significant rCBF change in an activation SPECT study. Then, we proposed a new method in which the activity of both two SPECT scans was normalized by cerebellar or occipital activity and compared. The ratio obtained by the proposed method came closer to 1 with less variations and with less mean % error in comparison with those of K value obtained by the dose-correction method. Although the proposed method has a limitation in the use of an activation study loaded with Diamox, it may be useful to evaluate an alteration of rCBF in the study such as postural testing or finger-moving test. (author)

  20. The quantification of short-chain chlorinated paraffins in sediment samples using comprehensive two-dimensional gas chromatography with μECD detection.

    Science.gov (United States)

    Muscalu, Alina M; Morse, Dave; Reiner, Eric J; Górecki, Tadeusz

    2017-03-01

    The analysis of persistent organic pollutants in environmental samples is a challenge due to the very large number of compounds with varying chemical and physical properties. Chlorinated paraffins (CPs) are complex mixtures of chlorinated n-alkanes with varying chain lengths (C 10 to C 30 ) and degree of chlorination (30 to 70% by weight). Their physical-chemical properties make these compounds persistent in the environment and able to bioaccumulate in living organisms. Comprehensive two-dimensional gas chromatography (GC × GC) coupled with micro-electron capture detection (μECD) was used to separate and quantify short-chain chlorinated paraffins (SCCP) in sediment samples. Distinct ordered bands were observed in the GC × GC chromatograms pointing to group separation. Using the Classification function of the ChromaTOF software, summary tables were generated to determine total area counts to set up multilevel-calibration curves for different technical mixes. Fortified sediment samples were analyzed by GC × GC-μECD with minimal extraction and cleanup. Recoveries ranged from 120 to 130%. To further validate the proposed method for the analysis of SCCPs, the laboratory participated in interlaboratory studies for the analysis of standards and sediment samples. The results showed recoveries between 75 and 95% and z-score values <2, demonstrating that the method is suitable for the analysis of SCCPs in soil/sediment samples. Graphical abstract Quantification of SCCPs by 2D-GC-μECD.

  1. SPM analysis of cerebrovascular reserve capacity after stimulation with acetazolamide measured by Tc-99m ECD SPECT in normal brain MRI patient

    Energy Technology Data Exchange (ETDEWEB)

    Lee, M. H.; Yoon, S. N.; Yoon, J. K.; Cho, C. W. [College of Medicine, Univ. of Ajou, Suwon (Korea, Republic of)

    2003-07-01

    This study was undertaken to evaluate normal response of acetazolamide in normal individuals, whose brain MRI is normal, using SPM99. In total, 10 Tc- 99m ECD brain SPECT were evaluated retrospectively. The half of the patients were male. Their mean age was 47.1 years old with a range of 33-61 years. They all visited our neurology department to evaluate stroke symptom. Their brain MRI was normal. Rest/acetazolamide brain SPECT was perfomed using Tc-99m ECD and the sequential injection and subtraction method. SPECT was acquired using fanbeam collimators and triple-head gamma camera (MultiSPECT III, Siemens medical systems, Inc. Hoffman Estates, III, USA). Chang's attenuation correction was applied their brain SPECT revealed normal rCBF pattern in visual analysis by two nuclear physician and they were diagnosed clinically normal. Using SPM method, we compared rest brain SPECT images with those of acetazolamide brain SPECT and measured the extent of the area with significant perfusion change (P<0.05) in predefined 34 cerebral regions. Acetazolamide brain SPECT showed no significant decreased region in comparison to rest brain SPECT. Only small portion of left mid temporal gyrus revealed increased rCBF on acetazolamide brain SPECT in comparison to rest brain SPECT. It apperas that there is no significant change in rCBF between rest and acetazolamide brain SPECT using Tc-99m ECD. The small number of this study is limitation of our study.

  2. Presurgical evaluation of cerebral perfusion reserve in patients for cardiovascular surgery using {sup 99m}Tc-ECD SPECT with diamox enhancement

    Energy Technology Data Exchange (ETDEWEB)

    Matsumura, Kaname; Nakashima, Hiromichi; Tanaka, Kuniyoshi; Kitano, Tokio; Murashima, Shuichi; Takeda, Kan; Yuasa, Hiroshi; Yada, Isao; Nakagawa, Tsuyoshi [Mie Univ., Tsu (Japan). School of Medicine

    1996-03-01

    Cerebrovascular stroke is one of the major complications in cardiovascular surgery with cardiopulmonary bypass. The purpose of this study was to evaluate the usefulness of preoperative {sup 99m}Tc-ethyl cysteinate dimer (ECD) SPECT and acetazolamide (diamox) enhancement to predict neurological complications in cardiovascular surgery. Eighteen patients with coronary disease, valvular disease or aortic aneurysm were studied before the operations. Regional cerebral blood flow and perfusion reserve were evaluated using ECD SPECT before and after the intravenous administration of diamox (1 g). Three cases with moderate to severe baseline abnormalities and poor perfusion reserve had cerebral infarction postoperatively. Twelve cases with good to fair perfusion reserve had no neurological complication. Three cases having poor perfusion reserve had the operations with more intensive brain protection, in which higher perfusion pressure to the brain was maintained during cardiopulmonary bypass, and no neurological complication was observed. In conclusion, patients who have moderately or markedly abnormal baseline flow with poor perfusion reserve may have some risk of neurological complications in cardiovascular surgery. ECD SPECT with diamox enhancement may give information useful for selection of operation procedures. (author).

  3. Domain crossing

    DEFF Research Database (Denmark)

    Schraefel, M. C.; Rouncefield, Mark; Kellogg, Wendy

    2012-01-01

    In CSCW, how much do we need to know about another domain/culture before we observe, intersect and intervene with designs. What optimally would that other culture need to know about us? Is this a “how long is a piece of string” question, or an inquiry where we can consider a variety of contexts a...

  4. Evaluation of time dependency of the acetazolamide effect on cerebral hemodynamics as measured by {sup 99m}Tc-ECD single-photon emission computed tomography

    Energy Technology Data Exchange (ETDEWEB)

    Takaki, Akihiro; Urata, Joji; Okada, Kazuhiro; Takaki, Rie [Saiseikai Kumamoto Hospital (Japan). Diagnostic Imaging Center; Mizuta, Yoshihiko; Murakami, Masaji; Yonehara, Toshiroh; Hirano, Teruyuki; Fujioka, Shodo

    2001-01-01

    Kuwabara et al. have examined the cerebral artery dilation with acetazolamide (ACZ) challenge test using PET. And, they reported that ACZ reaction came out time dependently. We have developed a unique SPECT's method using Technetium-99m ethyl cysteinate dimer ({sup 99m}Tc-ECD) to verify the results obtained by Kuwabara et al. One thousand MBq of {sup 99m}Tc-ECD was exactly divided into three syringes. Each of which was intravenous infused (IV) at rest, 7.5, and 20 minutes after ACZ administration. Data collection was started using dynamic SPECT immediately after {sup 99m}Tc-ECD IV at rest. Using necessary data only, SPECT images representing each of the three {sup 99m}Tc-ECD IV was reconstructed. SPECT counts were obtained by the ROI method from each images to calculate relative CBF from rest to 7.5 and 20 minutes after ACZ administration. The 18 hemispheres of nine patients in the negative control group in whom ACZ was not loaded, CBF was stable during the three evaluation. The measurement error our method was estimated as small. The 18 hemispheres of nine patients in the positive control group who has normal vasodilatory reserve, CBF was increased by 26.2{+-}8.1% at 7.5 minutes and 29.3{+-}13.1% at 20 minutes after ACZ administration. Seven patients with and chronic stage unilateral internal carotid artery severe stenosis and/or occlusion were evaluated as the test group. Case of unaffected side, CBF was increased by 17.6{+-}6.9% at 7.5 minutes and 24.8{+-}11.3% 20 minutes after ACZ administration. And, increase rate of CBF in the affected side was 2.8{+-}1.6% at 7.5 minutes and 17.3{+-}5.0% at 20 minutes after ACZ administration. In the affected side, timing of the maximum CBF increase caused by ACZ was remarkably delayed. Our method based on {sup 99m}Tc-ECD SPECT also revealed delayed cerebral artery dilation in the affected side. It was suggested that ACZ reaction came out time dependently, as reported by Kuwabara et al. (author)

  5. Voxel-by-voxel analysis of ECD-brain SPECT can separate penumbra from irreversibly damaged tissue at the acute phase of stroke

    International Nuclear Information System (INIS)

    Darcourt, J.; Migneco, O.; David, O.; Bussiere, F.; Mahagne, M.H.; Dunac, A.; Baron, J.C.

    2002-01-01

    Aim. At the acute phase of ischemic stroke, the target of treatment is still salvageable hypoperfused cerebral tissue; so called penumbra. We tested the possibility of separating on early ECD brain SPECT penumbral voxels (P) from irreversibly damaged damaged tissue (IDT). We used ECD which is not only a perfusion tracer but also a metabolic marker. Materials and methods. We prospectively studied 18 patients who underwent ECD-SPECT within the 12 hours following a first-ever acute middle cerebral artery stroke. Neurological evaluation was performed using the Orgogozo's scale at admission and 3 months later in order to calculate and evolution index (IE%) (Martinez-Vila et al.). SPECT data were obtained using a triple head camera equipped with fan beam collimators one hour after injection of 1000 MBq of 99mTc-ECD. On reconstructed images gray matter voxels were automatically segmented. Contralateral healthy hemisphere was used as reference leading to the identification of 3 cortical voxel types: normal (N-SPECT) above 80%; penumbra (P-SPECT) between 80% and 40% and IDT (IDT-SPECT) below 40%. 10 patients also underwent a T2 weighted 3D MRI study at 3 months. Cortical voxels with hypersignal served as reference for IDT (IDT-MRI) the others were considered normal (N-MRI). SPECT and MRI data were co-registered. Therefore each voxel belonged to one of 6 categories (3 SPECT x 2 MRI). Results. (1) The SPECT thresholds were validated on the MRI subgroup. 99% of the N-SPECT voxels were normal on late MRI. 84% of IDT-SPECT voxels corresponded to IDT-MRI. 89% of P-SPECT voxels were normal on late MRI and 11% corresponded to IDT on late MRI. Other categories of voxels (N-SPECT IDT-MRI and IDT-SPECT N-MRI) represented less than 5%. (2) Percentages of each voxel SPECT type was correlated with the EI% on the entire population (Spearman test). P-SPECT extent correlated with EI% improvement (p<0.001) and IDT-SPECT with EI% worsening (p<0.001). Conclusion. Analysis of ECD cortical

  6. Insights into function of PSI domains from structure of the Met receptor PSI domain

    International Nuclear Information System (INIS)

    Kozlov, Guennadi; Perreault, Audrey; Schrag, Joseph D.; Park, Morag; Cygler, Miroslaw; Gehring, Kalle; Ekiel, Irena

    2004-01-01

    PSI domains are cysteine-rich modules found in extracellular fragments of hundreds of signaling proteins, including plexins, semaphorins, integrins, and attractins. Here, we report the solution structure of the PSI domain from the human Met receptor, a receptor tyrosine kinase critical for proliferation, motility, and differentiation. The structure represents a cysteine knot with short regions of secondary structure including a three-stranded antiparallel β-sheet and two α-helices. All eight cysteines are involved in disulfide bonds with the pattern consistent with that for the PSI domain from Sema4D. Comparison with the Sema4D structure identifies a structurally conserved core comprising the N-terminal half of the PSI domain. Interestingly, this part links adjacent SEMA and immunoglobulin domains in the Sema4D structure, suggesting that the PSI domain serves as a wedge between propeller and immunoglobulin domains and is responsible for the correct positioning of the ligand-binding site of the receptor

  7. Extracellular Vesicles in Hematological Disorders

    Directory of Open Access Journals (Sweden)

    Anat Aharon

    2014-10-01

    Full Text Available Extracellular vesicles (EVs, comprised of exosomes, microparticles, apoptotic bodies, and other microvesicles, are shed from a variety of cells upon cell activation or apoptosis. EVs promote clot formation, mediate pro-inflammatory processes, transfer proteins and miRNA to cells, and induce cell signaling that regulates cell differentiation, proliferation, migration, invasion, and apoptosis. This paper will review the contribution of EVs in hematological disorders, including hemoglobinopathies (sickle cell disease, thalassemia, paroxysmal nocturnal hemoglobinuria, and hematological malignancies (lymphomas, myelomas, and acute and chronic leukemias.

  8. Blood extracellular DNA after irradiation

    International Nuclear Information System (INIS)

    Vladimirov, V.G.; Tishchenko, L.I.; Surkova, E.A.; Vasil'eva, I.N.

    1993-01-01

    It has been shown that blood extracellular DNA of irradiated rats largely consists of the low-molecular DNA and its oligomers. Molecular masses of oligomers are multiple to molecular mass of monomer fragment with nucleosome size. The low-molecular DNA has linear form. The average content of GC-pairs in low-molecular DNA is higher than in total rat's DNA (48.5% against 41.5%). The low-molecular DNA is a part of complex containing RNA, acidic proteins and lipids. It is assumed that the formation of low-molecular DNA is a result of Ca/Mg - dependent nuclear endonuclease action

  9. Ex-post evaluation by bibliometric method. Institutional comparison among 9 resembled foreign research institutes by using the energy citation database (ECD)

    International Nuclear Information System (INIS)

    Yanagisawa, Kazuaki

    2007-09-01

    From a viewpoint of ex-post evaluation, research papers published from nine resembled nuclear research institutes located in Japan, the U.S.A., Germany and France were compared by a bibliometric method. A research database used was the Energy Citation Database (ECD) owned by USDOE. ECD is a database run by USDOE and has a high frequency of research paper acquisition assembled in the U.S. Response speed of ECD on the Website is quick and all logged data can be handled easily. INIS database is run by the International Atomic Energy Agency (IAEA) and contains a lot of nuclear research papers collected from the member countries such as the U.S.A., Japan, Germany and France. INIS underestimates about 20% of the U.S.A. data than that of ECD. I. Institutional Comparison. (1) ECD shows that a total number of research papers published during 25 years (1978-2002) was of the order of the ORNL (34, 149 papers)>SNL>ANL>BNL>Idaho (>Karlsruhe>JAERI>Jeulich>Cadarache). Where, INIS shows it as ORNL>JAERI. (2) ECD can show a long-term data comparison with a time span more than 50 years (1953-2002). Disclosed research papers were of the order of the ORNL (55,857)>ANL (37,129)>SNL (24,628)>BNL (24,829)> Idaho (2,398). There were many records loaded without publication dates-over 50,000. Because of this, any searches which use dates are not finding these documents. Typically, the author found over 5,000 SNL items in the NSA range of records. SNL also kept a lot of defense reports, those are not disclosed yet. One had better know a historical background of each cite as to the case for long-range dates comparison. (3) ECD founds that research papers at a five-year period varied those numbers. At past (10), thus 1988-1922, paper reduction occurred sharply at most US-institutes. This might be attributed to lay-offs, funding shifts or complete elimination of programs, a policy change in reporting requirements for contract reporting deliverables. Definitions of what constituted STI (science

  10. Voxel-based analysis of Tc-99 m ECD brain perfusion SPECT in patients with normal pressure hydrocephalus

    Energy Technology Data Exchange (ETDEWEB)

    Yoon, Bora [Department of Neurology, College of Medicine, Catholic University of Korea, Seoul 137-701 (Korea, Republic of); Yang, Dong-Won [Department of Neurology, College of Medicine, Catholic University of Korea, Seoul 137-701 (Korea, Republic of)], E-mail: neuroman@catholic.ac.kr; Shim, Yong-Soo; Chung, Sung-Woo [Department of Neurology, College of Medicine, Catholic University of Korea, Seoul 137-701 (Korea, Republic of); Ahn, Kook-Jin; O, Joo-Hyun; Kim, Sung-Hoon; Sohn, Hyung-Sun; Chung, Soo-Kyo [Department of Radiology, College of Medicine, Catholic University of Korea, Seoul 137-701 (Korea, Republic of); Chung, Yong-An [Department of Radiology, College of Medicine, Catholic University of Korea, Seoul 137-701 (Korea, Republic of); East-West Research Institute of Translational Medicine (EWTM), Incheon St. Mary' s Hospital, Incheon 403-720 (Korea, Republic of)], E-mail: nm@catholic.ac.kr

    2009-07-15

    Idiopathic normal pressure hydrocephalus (iNPH) is a reversible dementia characterized by gait disturbance, incontinence and dementia. This study investigates the neuropsychological characteristics and changes of regional cerebral blood flow (rCBF) in patients with iNPH. Ten patients who met the criteria of probable iNPH and 13 normal control subjects were evaluated. The general cognitive function and detailed neuropsychological functions were measured by K-MMSE and comprehensive neuropsychological battery. Tc-99m-ethyl cysteinate dimmer (Tc-99m-ECD) single photon emission computed tomography (SPECT) was performed to measure the rCBF and statistical parametric mapping (SPM) and statistical probabilistic brain anatomic map (SPAM) was applied to the objective analysis of SPECT data. On the neuropsychological examination, all the patients showed abnormality in memory, psychomotor speed and frontal executive function. SPM analysis of SPECT images revealed that rCBF in bilateral thalami, right prefrontal area, bilateral anterior and posterior cingulate gyri, right caudate nucleus, and left parahippocampal gyrus was significantly decreased in patients with iNPH compared to normal controls (uncorrected P<0.005). In SPAM analysis, rCBF reduction was observed in bilateral prefrontal area, anterior, posterior cingulate gyri and caudate nuclei. We have found that rCBF changes occurred predominantly in prefrontal and subcortical areas, the changes were associated with frontal subcortical circuit, and the affected frontal subcortical circuit may contribute to the cognitive decline seen in the iNPH patients. The reduction of rCBF and clinical cognitive impairment are closely connected in patients with iNPH.

  11. Improvement of the 99mTc-ECD brain uptake ratio (BUR) method for measurement of cerebral blood flow

    International Nuclear Information System (INIS)

    Ito, Shigeki; Takaki, Akihiro; Inoue, Shinya

    2012-01-01

    The brain uptake ratio (BUR) method for the 99m Tc-ethylcysteinate dimer ( 99m Tc-ECD) single photon emission computed tomography (SPECT), a non-invasive measurement method of regional cerebral blood flow (rCBF), has been used in clinical practice in Japan, because it is simple to use. However, the accuracy of this method is limited, as it has problems in the determination of input function and the regression equation. The purpose of this study is to improve the BUR method by reconstructing the determination process of the input function and regression equation based on measurement of the rCBF by H 2 15 O positron emission tomography (PET). The input function was obtained by setting the region of interest on the ascending aorta instead of the aortic arch. The 3DSRT algorithm was used to obtain the anatomically standardized rCBF, and developed a semi-automatic analyzing software using C++ in order to stabilize the repeatability of the improved BUR (IBUR) method. The regression equation for the IBUR method was obtained by the H 2 15 O PET rCBFs in 15 patients with the arterial blood sampling method. All the measurements in this study were performed with the patient in the resting state. A good correlation was observed between the rCBF values measured by H 2 15 O PET and the regional BURs measured by the IBUR method (r=0.86, p 2 15 O PET. This finding indicates the potential clinical usefulness of this method. (author)

  12. Voxel-based analysis of Tc-99 m ECD brain perfusion SPECT in patients with normal pressure hydrocephalus

    International Nuclear Information System (INIS)

    Yoon, Bora; Yang, Dong-Won; Shim, Yong-Soo; Chung, Sung-Woo; Ahn, Kook-Jin; O, Joo-Hyun; Kim, Sung-Hoon; Sohn, Hyung-Sun; Chung, Soo-Kyo; Chung, Yong-An

    2009-01-01

    Idiopathic normal pressure hydrocephalus (iNPH) is a reversible dementia characterized by gait disturbance, incontinence and dementia. This study investigates the neuropsychological characteristics and changes of regional cerebral blood flow (rCBF) in patients with iNPH. Ten patients who met the criteria of probable iNPH and 13 normal control subjects were evaluated. The general cognitive function and detailed neuropsychological functions were measured by K-MMSE and comprehensive neuropsychological battery. Tc-99m-ethyl cysteinate dimmer (Tc-99m-ECD) single photon emission computed tomography (SPECT) was performed to measure the rCBF and statistical parametric mapping (SPM) and statistical probabilistic brain anatomic map (SPAM) was applied to the objective analysis of SPECT data. On the neuropsychological examination, all the patients showed abnormality in memory, psychomotor speed and frontal executive function. SPM analysis of SPECT images revealed that rCBF in bilateral thalami, right prefrontal area, bilateral anterior and posterior cingulate gyri, right caudate nucleus, and left parahippocampal gyrus was significantly decreased in patients with iNPH compared to normal controls (uncorrected P<0.005). In SPAM analysis, rCBF reduction was observed in bilateral prefrontal area, anterior, posterior cingulate gyri and caudate nuclei. We have found that rCBF changes occurred predominantly in prefrontal and subcortical areas, the changes were associated with frontal subcortical circuit, and the affected frontal subcortical circuit may contribute to the cognitive decline seen in the iNPH patients. The reduction of rCBF and clinical cognitive impairment are closely connected in patients with iNPH.

  13. Mechanism study of recovery from aphasia with 99Tcm-ECD SPECT brain imaging after oral reading test

    International Nuclear Information System (INIS)

    Chen Jian; Xu Hao; Wang Hong; Chen Zhuoming; Wu Qiulian

    2007-01-01

    Objective: There may be two mechanisms for recovery from aphasia----repair of dam- aged left hemisphere language network and(or) activation of compensatory areas in right hemisphere. It is, however, still controversial on which mechanism plays a more important role. The goal of this study was to compare the regional cerebral blood flow (rCBF) between baseline and during oral reading in aphasic patients who had shown definite recovery from a focal lesion in left hemisphere. It also aimed at exploring the role of the two hemispheres in recovery from aphasia, thus providing theoretic basis for rehabilitation therapy. Methods: Baseline and oral reading 99 Tc m -ethylcysteinate dimer (ECD) brain SPECT imaging were performed in 7 patients with aphasia separately in one-day interval. Semi-quantitative analysis of rCBF was conducted on 26 symmetrical ROIs in bilateral hemispheres on the transverse slices. The rCBF was estimated by the ratio of the counts per pixel of each ROI to the average counts per pixel of the whole encephalon. The change in blood flow was represented by the difference in rCBF between oral reading and baseline. Results: The activated perfusion pattern of brain region was different in all 7 aphasic patients. In 5 of the 7 patients, whose reading capacity were relatively preserved, the brain perfusion pattern was mainly left lateralized, while the other two patients who had poor performance in reading showed bilateral activation foci with fight-sided dominance. Conclusion: Oral reading activated SPECT brain imaging may be a useful tool for monitoring the progress of speech recovery in the treatment of aphasic patients. (authors)

  14. Trusted Domain

    DEFF Research Database (Denmark)

    Hjorth, Theis Solberg; Torbensen, Rune

    2012-01-01

    remote access via IP-based devices such as smartphones. The Trusted Domain platform fits existing legacy technologies by managing their interoperability and access controls, and it seeks to avoid the security issues of relying on third-party servers outside the home. It is a distributed system...... of wireless standards, limited resources of embedded systems, etc. Taking these challenges into account, we present a Trusted Domain home automation platform, which dynamically and securely connects heterogeneous networks of Short-Range Wireless devices via simple non-expert user. interactions, and allows......In the digital age of home automation and with the proliferation of mobile Internet access, the intelligent home and its devices should be accessible at any time from anywhere. There are many challenges such as security, privacy, ease of configuration, incompatible legacy devices, a wealth...

  15. Extracellular nucleotide signaling in plants

    Energy Technology Data Exchange (ETDEWEB)

    Stacey, Gary [Univ. of Missouri, Columbia, MO (United States)

    2016-09-08

    Over the life of this funded project, our research group identified and characterized two key receptor proteins in plants; one mediating the innate immunity response to chitin and the other elucidating the key receptor for extracellular ATP. In the case of chitin recognition, we recently described the quaternary structure of this receptor, shedding light on how the receptor functions. Perhaps more importantly, we demonstrated that all plants have the ability to recognize both chitin oligomers and lipochitooligosacchardes, fundamentally changing how the community views the evolution of these systems and strategies that might be used, for example, to extend symbiotic nitrogen fixation to non-legumes. Our discovery of DORN1 opens a new chapter in plant physiology documenting conclusively that eATP is an important extracellular signal in plants, as it is in animals. At this point, we cannot predict just how far reaching this discovery may prove to be but we are convinced that eATP signaling is fundamental to plant growth and development and, hence, we believe that the future will be very exciting for the study of DORN1 and its overall function in plants.

  16. Binding of matrix metalloproteinase inhibitors to extracellular matrix: 3D-QSAR analysis.

    Science.gov (United States)

    Zhang, Yufen; Lukacova, Viera; Bartus, Vladimir; Nie, Xiaoping; Sun, Guorong; Manivannan, Ethirajan; Ghorpade, Sandeep R; Jin, Xiaomin; Manyem, Shankar; Sibi, Mukund P; Cook, Gregory R; Balaz, Stefan

    2008-10-01

    Binding to the extracellular matrix, one of the most abundant human protein complexes, significantly affects drug disposition. Specifically, the interactions with extracellular matrix determine the free concentrations of small molecules acting in tissues, including signaling peptides, inhibitors of tissue remodeling enzymes such as matrix metalloproteinases, and other drug candidates. The nature of extracellular matrix binding was elucidated for 63 matrix metalloproteinase inhibitors, for which the association constants to an extracellular matrix mimic were reported here. The data did not correlate with lipophilicity as a common determinant of structure-nonspecific, orientation-averaged binding. A hypothetical structure of the binding site of the solidified extracellular matrix surrogate was analyzed using the Comparative Molecular Field Analysis, which needed to be applied in our multi-mode variant. This fact indicates that the compounds bind to extracellular matrix in multiple modes, which cannot be considered as completely orientation-averaged and exhibit structural dependence. The novel comparative molecular field analysis models, exhibiting satisfactory descriptive and predictive abilities, are suitable for prediction of the extracellular matrix binding for the untested chemicals, which are within applicability domains. The results contribute to a better prediction of the pharmacokinetic parameters such as the distribution volume and the tissue-blood partition coefficients, in addition to a more imminent benefit for the development of more effective matrix metalloproteinase inhibitors.

  17. Analysis of extracellular RNA by digital PCR

    Directory of Open Access Journals (Sweden)

    Kenji eTakahashi

    2014-06-01

    Full Text Available The transfer of extracellular RNA is emerging as an important mechanism for intracellular communication. The ability for the transfer of functionally active RNA molecules from one cell to another within vesicles such as exosomes enables a cell to modulate cellular signaling and biological processes within recipient cells. The study of extracellular RNA requires sensitive methods for the detection of these molecules. In this methods article, we will describe protocols for the detection of such extracellular RNA using sensitive detection technologies such as digital PCR. These protocols should be valuable to researchers interested in the role and contribution of extracellular RNA to tumor cell biology.

  18. Extracellular Molecules Involved in Cancer Cell Invasion

    International Nuclear Information System (INIS)

    Stivarou, Theodora; Patsavoudi, Evangelia

    2015-01-01

    Nowadays it is perfectly clear that understanding and eradicating cancer cell invasion and metastasis represent the crucial, definitive points in cancer therapeutics. During the last two decades there has been a great interest in the understanding of the extracellular molecular mechanisms involved in cancer cell invasion. In this review, we highlight the findings concerning these processes, focusing in particular on extracellular molecules, including extracellular matrix proteins and their receptors, growth factors and their receptors, matrix metalloproteinases and extracellular chaperones. We report the molecular mechanisms underlying the important contribution of this pool of molecules to the complex, multi-step phenomenon of cancer cell invasion

  19. Extracellular Molecules Involved in Cancer Cell Invasion

    Directory of Open Access Journals (Sweden)

    Theodora Stivarou

    2015-01-01

    Full Text Available Nowadays it is perfectly clear that understanding and eradicating cancer cell invasion and metastasis represent the crucial, definitive points in cancer therapeutics. During the last two decades there has been a great interest in the understanding of the extracellular molecular mechanisms involved in cancer cell invasion. In this review, we highlight the findings concerning these processes, focusing in particular on extracellular molecules, including extracellular matrix proteins and their receptors, growth factors and their receptors, matrix metalloproteinases and extracellular chaperones. We report the molecular mechanisms underlying the important contribution of this pool of molecules to the complex, multi-step phenomenon of cancer cell invasion.

  20. The International Society for Extracellular Vesicles launches the first massive open online course on extracellular vesicles

    OpenAIRE

    L?sser, Cecilia; Th?ry, Clotilde; Buz?s, Edit I.; Mathivanan, Suresh; Zhao, Weian; Gho, Yong Song; L?tvall, Jan

    2016-01-01

    The International Society for Extracellular Vesicles (ISEV) has organised its first educational online course for students and beginners in the field of extracellular vesicles (EVs). This course, “Basics of Extracellular Vesicles,” uses recorded lectures from experts in the field and will be open for an unlimited number of participants. The course is divided into 5 modules and can be accessed at www.coursera.org/learn/extracellular-vesicles. The first module is an introduction to the field co...

  1. Identification by [{sup 99m}Tc]ECD SPECT of anterior cingulate hypoperfusion in progressive supranuclear palsy, in comparison with Parkinson's disease

    Energy Technology Data Exchange (ETDEWEB)

    Varrone, Andrea [University Federico II, Biostructure and Bioimaging Institute, National Research Council/Department of Biomorphological and Functional Sciences, Napoli (Italy); Karolinska Hospital, Department of Clinical Neuroscience, Psychiatry Section, Stockholm (Sweden); Pagani, Marco; Salmaso, Dario [National Research Council, Institute of Cognitive Sciences and Technologies, Rome and Padua (Italy); Salvatore, Elena; Amboni, Marianna; De Michele, Giuseppe; Filla, Alessandro; Barone, Paolo [University Federico II, Department of Neurological Sciences, Napoli (Italy); Sansone, Valeria; Pappata, Sabina; Salvatore, Marco [University Federico II, Biostructure and Bioimaging Institute, National Research Council/Department of Biomorphological and Functional Sciences, Napoli (Italy); Nobili, Flavio [University of Genoa, Clinical Neurophysiology, Department of Endocrinological and Metabolic Sciences, Genoa (Italy)

    2007-07-15

    Progressive supranuclear palsy (PSP) is an akinetic-rigid syndrome that can be difficult to differentiate from Parkinson's disease (PD), particularly at an early stage. [{sup 99m}Tc]ethyl cysteinate dimer (ECD) SPECT could represent a widely available tool to assist in the differential diagnosis. In this study we used voxel-based analysis and Computerised Brain Atlas (CBA)-based principal component analysis (PCA) of [{sup 99m}Tc]ECD SPECT data to test whether: (1) specific patterns of rCBF abnormalities can differentiate PSP from controls and PD; (2) networks of dysfunctional brain regions can be found in PSP vs controls and PD. Nine PD patients, 16 PSP patients and ten controls were studied with [{sup 99m}Tc]ECD SPECT using a brain-dedicated device (Ceraspect). Voxel-based analysis was performed with statistical parametric mapping. PCA was applied to volume of interest data after spatial normalisation to CBA. The voxel-based analysis showed hypoperfusion of the anterior cingulate and medial frontal cortex in PSP compared with controls and PD. In PSP patients the rCBF impairment extended to the pre-supplementary motor area and prefrontal cortex, areas involved in executive function and motor networks. Compared with PSP patients, PD patients showed a mild rCBF decrease in associative visual areas which could be related to the known impairment of visuospatial function. The PCA identified three principal components differentiating PSP patients from controls and/or PD patients that included groups of cortical and subcortical brain regions with relatively decreased (cingulate cortex, prefrontal cortex and caudate) or increased (parietal cortex) rCBF, representing distinct functional networks in PSP. Anterior cingulate hypoperfusion seems to be an early, distinct brain abnormality in PSP as compared with PD. (orig.)

  2. Assessment of organochlorine pesticides and atrazine residues in Maize Produced in Ghana using GC-ECD/ GC-MS

    International Nuclear Information System (INIS)

    Miensah, E.D.

    2015-07-01

    Maize is consumed by over 95% of the Ghanaian populace and the practice of using atrazine, lindane and other organochlorine pesticides in its production has raised concerns about potential adverse effects on human health and the environment. A field survey was conducted in this study to assess farmers’ knowledge of safe handling and use of these chemicals as well as toxicity awareness and symptoms among farmers and traders. Residues of the pesticides in maize samples as well as the potential health risk associated with exposure to them were also evaluated. Data obtained from the field survey indicate that a large proportion of farmers are at risk of pesticide poisoning through occupational exposure. Over 80 % of applicators do not apply safety precautions during pesticide mixing and application; resulting in considerable prevalence of pesticide related illnesses in the study area. Residues of lindane and atrazine were below detection limits of 0.005 mg/kg and 0.010 mg/kg respectively of the GC-ECD/MS used for the analysis; except in the Ashanti region where atrazine was 0.05 mg/kg which is within the EU-MRL of 0.1 mg/kg. However, minute concentrations of other organochlorines (endrin- 0.007 mg/kg; heptachlor- 0.023 mg/kg; alpha-endosulfan- 0.013 mg/kg) were found in maize samples from Brong Ahafo, Eastern, Central and Upper west regions. Though endrin and alpha-endosulfan were below the EU-MRL of 0.01 mg/kg and 0.10 mg/kg respectively, heptachlor was slightly above the EU-MRL of 0.01 mg/kg. The estimated doses for all these other organochlorines pesticides do not pose direct hazard to human health because their estimated dose values were lower than toxic thresholds and reference doses. However, due to the unsafe application practices by applicators, there exists a potential risk for systemic and carcinogenic health effects by these chemicals in the study area. (au)

  3. Early effects of treatment in first episode schizophrenia on brain perfusion evaluated with 99mTc-ECD-SPECT

    International Nuclear Information System (INIS)

    Milcinski, M.; Grmek, M.; Novak, B.; Kocmur, M.

    2002-01-01

    Aim of our study was to evaluate regional cerebral blood flow (r-CBF) in acute first-episode schizophrenia and the early effects of antipsychotic drugs on r-CBF. Methods: Clinical criteria for schizophrenia were met according to International Classification of Diseases - 10th Edition (ICD-10). Psychic status and severity of disease of each patient were evaluated with a semi-structured psychiatric interview, Positive and Negative Syndrome Scale (PANSS) and Clinical Global Impression (CGI) on the same day as the scintigraphic study. R-CBF was evaluated using 99m-Tc-ECD. Until now 9 first-episode schizophrenic patients 2-7 (average 5.2) days after the beginning of antipsychotic treatment and 7 pts 8-15 (average 10.4) weeks later were investigated. R-CBF was evaluated in every patient in comparison to cerebellar blood flow in frontal and temporal regions in 4 slices. Both studies were compared. Homogeneity of brain perfusion was assessed visually as homogenous, moderately and severely non-homogenous. Results: In acute patients, perfusion was non-homogenous and decreased in left frontal lobe (90.1±4.4) in comparison to the right side (93.3±±4.6, p<0.05). Increase in perfusion was significant (p<0.05) between first and second scan in left (90.1±4.4 to 94.1±5.2, p<0.05) and right (93.4±4.6 to 96.9±5.8, p<0.05) frontal lobes. Significant difference in perfusion between right and left posterior region of temporal lobe found on the first scan was no longer present on the second scan. Perfusion was significantly more homogenous on the second scan. Significant decrease in PANSS (p<0.05) and CGI (p<0.001) scores was noted. Conclusions: Despite the low number of patients included so far, our findings implicate that patients with first-episode schizophrenia have significant bilateral frontal hypoperfusion, more pronounced on the left side. After average 10 weeks of medication, bilateral increase in blood flow was observed in frontal lobes and left-right difference was no

  4. The use of experimental design in the development of an HPLC-ECD method for the analysis of captopril.

    Science.gov (United States)

    Khamanga, Sandile M; Walker, Roderick B

    2011-01-15

    An accurate, sensitive and specific high performance liquid chromatography-electrochemical detection (HPLC-ECD) method that was developed and validated for captopril (CPT) is presented. Separation was achieved using a Phenomenex(®) Luna 5 μm (C(18)) column and a mobile phase comprised of phosphate buffer (adjusted to pH 3.0): acetonitrile in a ratio of 70:30 (v/v). Detection was accomplished using a full scan multi channel ESA Coulometric detector in the "oxidative-screen" mode with the upstream electrode (E(1)) set at +600 mV and the downstream (analytical) electrode (E(2)) set at +950 mV, while the potential of the guard cell was maintained at +1050 mV. The detector gain was set at 300. Experimental design using central composite design (CCD) was used to facilitate method development. Mobile phase pH, molarity and concentration of acetonitrile (ACN) were considered the critical factors to be studied to establish the retention time of CPT and cyclizine (CYC) that was used as the internal standard. Twenty experiments including centre points were undertaken and a quadratic model was derived for the retention time for CPT using the experimental data. The method was validated for linearity, accuracy, precision, limits of quantitation and detection, as per the ICH guidelines. The system was found to produce sharp and well-resolved peaks for CPT and CYC with retention times of 3.08 and 7.56 min, respectively. Linear regression analysis for the calibration curve showed a good linear relationship with a regression coefficient of 0.978 in the concentration range of 2-70 μg/mL. The linear regression equation was y=0.0131x+0.0275. The limits of detection (LOQ) and quantitation (LOD) were found to be 2.27 and 0.6 μg/mL, respectively. The method was used to analyze CPT in tablets. The wide range for linearity, accuracy, sensitivity, short retention time and composition of the mobile phase indicated that this method is better for the quantification of CPT than the

  5. Determinação de herbicidas usados no cultivo de arroz irrigado na região sul do estado de Santa Catarina através da SPME-GC-ECD Determination of herbicides used in irrigated rice cultivation in the south of Santa Catarina using SPME-GC-ECD

    Directory of Open Access Journals (Sweden)

    Léa L. F. Costa

    2008-01-01

    Full Text Available Evaluation of the pollution by the herbicides alachlor, propanil and atrazine in water samples from four rivers in the cities of Turvo and Meleiro, south of Santa Catarina State, was made using the SPME-GC-ECD method. The proposed method was optimized and validated. The correlation coefficients were higher than 0.997 and linear ranges of the analytical curves were 0.1-4; 0.1-2.5 and 0.1-5 µg L-1 for atrazine, alachlor and propanil, respectively. The herbicides were quantified by GC-ECD and identified by GC-MS. Both of the selected rivers presented contamination by at least one of the studied herbicides.

  6. Effect of sumatriptan on cerebral blood flow during migraine headache. Measurement by sequential SPECT used 99mTc-ECD background subtraction method

    International Nuclear Information System (INIS)

    Ueda, Takashi; Torihara, Yoshito; Tsuneyoshi, Noritaka; Ikeda, Yoshitomo

    2001-01-01

    The present study was designed to examine the effect of sumatriptan on regional cerebral blood flow (CBF) during migraine headache. Nine cases were examined by 99m Tc-ECD background subtraction method for the absolute value measurement of regional CBF before and after sumatriptan injection. rCBF except for occipital and perioccipital lobes, were increased 10-20% during migraine headache and significant decreases were observed by sumatriptan injection. Two cases of nine had transiently increased systemic blood pressure and cardiac pulse rate, however, all cases improved migraine headache after injection of sumatriptan. (author)

  7. Regional cerebral blood flow of the patients with schizophrenia. A study using 99mTc-ECD SPECT at rest and activation

    International Nuclear Information System (INIS)

    Hu Ping; Wu Kening; Zeng Shiquan; Lin Zengtao; Yu Jinlong

    1996-01-01

    Regional cerebral blood flow (rCBF) changes of the patients with schizophrenia were observed. 99m Tc-ECD SPECT was performed on 22 patients with schizophrenia and 10 healthy volunteers at rest and activation with a cognitive task: a modified Wisconsin Card Sorting Test. At rest state, only 4 patients have abnormal rCBF pattern: left hemisphere over-perfusion relative to the right. A significant relative activation deficit in the left inferior prefrontal region was revealed in the patients during activation. The patients with schizophrenia may have frontal lobe dysfunction

  8. Effect of sumatriptan on cerebral blood flow during migraine headache. Measurement by sequential SPECT used {sup 99m}Tc-ECD background subtraction method

    Energy Technology Data Exchange (ETDEWEB)

    Ueda, Takashi; Torihara, Yoshito; Tsuneyoshi, Noritaka; Ikeda, Yoshitomo [Miyazaki Social Insurance Hospital (Japan)

    2001-07-01

    The present study was designed to examine the effect of sumatriptan on regional cerebral blood flow (CBF) during migraine headache. Nine cases were examined by {sup 99m}Tc-ECD background subtraction method for the absolute value measurement of regional CBF before and after sumatriptan injection. rCBF except for occipital and perioccipital lobes, were increased 10-20% during migraine headache and significant decreases were observed by sumatriptan injection. Two cases of nine had transiently increased systemic blood pressure and cardiac pulse rate, however, all cases improved migraine headache after injection of sumatriptan. (author)

  9. 99mTc-ECD brain SPECT in patients with Moyamoya disease: a reflection of cerebral perfusion status at tissue level in the disease process

    International Nuclear Information System (INIS)

    Kashyap, Raghava; Mittal, Bhagwant Rai; Sunil, Hejjaji Venkataramarao; Bhattacharya, Anish; Singh, Baljinder; Mukherjee, Kanchan Kumar; Gupta, Sunil Kumar

    2011-01-01

    Moyamoya disease is a rare, progressive cerebrovascular disorder caused by intracranial stenosis of the circle of Willis, resulting in successive ischemic events. Computed tomography (CT) and magnetic resonance imaging (MRI) play a major role in diagnosis. The aim of the study was to describe the spectrum of findings on brain SPECT in patients with Moyamoya disease and to compare the findings with other investigations. 99m Tc-ECD SPECT scans of seventeen patients (7 children and 10 adults) were analysed to study the brain perfusion. Features of Moyamoya disease were detected on DSA in 11 patients, CTA in one, MR angiography in one patient. Brain perfusion SPECT analysis showed unilateral perfusion defects in 11 patients, normal perfusion in 2 and bilateral defects in 4 patients. No perfusion defects despite bilateral vascular changes were noted in one patient. Cerebral infarcts were detected on MRI unilaterally in three subjects while multiple infarcts were identified in one. 99m Tc-ECD Brain SPECT showed perfusion defects that were more extensive compared to those detected on MRI. Post acetazolamide studies for assessment of cerebrovascular reserve were done in three patients. Two of them showed good cerebrovascular reserve (>1). Follow-up studies post-surgical procedures (Myo-dura synangiosis) done in two patients showed partial resolution of perfusion defects in the involved areas. Brain perfusion scintigraphy is an important adjunct in evaluation of patients with Moyamoya disease yielding information about the direct end results of the pathology in the vessels and also prognostic information. (author)

  10. Perfusion impairments in children with reactive attachment disorder (RAD) on 99mTc-ECD brain SPECT: comparison with MR findings

    International Nuclear Information System (INIS)

    Yoo, Y. H.; Sin, E. J.; Cheon, K. A.; Yoon, M. J.; Lee, J. D.; Jeon, T. J.

    2002-01-01

    This study aimed to reveal that severe disturbance of attachment relationship with primary care-giver can affect functional and anatomical brain development by measuring cerebral perfusion on 99m Tc-ECD brain SPECT and correlative MRI. We included 18 children aged 31 to 76 months who met the diagnostic criteria of RAD as defined in DSM-IV and ICD-10 and SSP and CARS. 99m Tc-ECD SPECT was performed using CERASPECT. MRI was performed in all patients. SPECT data were visually assessed. 15 of 18 children had abnormal perfusion on SPECT, revealing decreased perfusion of Lt.thalamus (7/15) and Rt.thalamus (3/15), and bilateral thalami (5/15). Perfusion of basal ganglia was decreased in 8 children. Decreased perfusion of Lt. parietal area was seen in 2. Whereas, all patients had normal MR findings. Perfusion abnormalities involving thalamus, BG in most children with RAD were found in this study. These results suggest that brain development of infant could be impeded by severe pathologic care and early nurturing environment would be important for normal development

  11. Defining structural and functional dimensions of the extracellular thyrotropin receptor region.

    Science.gov (United States)

    Kleinau, Gunnar; Mueller, Sandra; Jaeschke, Holger; Grzesik, Paul; Neumann, Susanne; Diehl, Anne; Paschke, Ralf; Krause, Gerd

    2011-06-24

    The extracellular region of the thyrotropin receptor (TSHR) can be subdivided into the leucine-rich repeat domain (LRRD) and the hinge region. Both the LRRD and the hinge region interact with thyrotropin (TSH) or autoantibodies. Structural data for the TSHR LRRD were previously determined by crystallization (amino acids Glu(30)-Thr(257), 10 repeats), but the structure of the hinge region is still undefined. Of note, the amino acid sequence (Trp(258)-Tyr(279)) following the crystallized LRRD comprises a pattern typical for leucine-rich repeats with conserved hydrophobic side chains stabilizing the repeat fold. Moreover, functional data for amino acids between the LRRD and the transmembrane domain were fragmentary. We therefore investigated systematically these TSHR regions by mutagenesis to reveal insights into their functional contribution and potential structural features. We found that mutations of conserved hydrophobic residues between Thr(257) and Tyr(279) cause TSHR misfold, which supports a structural fold of this peptide, probably as an additional leucine-rich repeat. Furthermore, we identified several new mutations of hydrophilic amino acids in the entire hinge region leading to partial TSHR inactivation, indicating that these positions are important for intramolecular signal transduction. In summary, we provide new information regarding the structural features and functionalities of extracellular TSHR regions. Based on these insights and in context with previous results, we suggest an extracellular activation mechanism that supports an intramolecular agonistic unit as a central switch for activating effects at the extracellular region toward the serpentine domain.

  12. .Gov Domains API

    Data.gov (United States)

    General Services Administration — This dataset offers the list of all .gov domains, including state, local, and tribal .gov domains. It does not include .mil domains, or other federal domains outside...

  13. Bacterial binding to extracellular proteins - in vitro adhesion

    DEFF Research Database (Denmark)

    Schou, C.; Fiehn, N.-E.

    1999-01-01

    Viridans streptococci, bacterial adherence, extracellular matrix proteins, surface receptors, endocarditis......Viridans streptococci, bacterial adherence, extracellular matrix proteins, surface receptors, endocarditis...

  14. Detection of extracellular enzymatic activity in microorganisms ...

    African Journals Online (AJOL)

    Detection of extracellular enzymatic activity in microorganisms isolated from waste vegetable oil contaminated soil using plate methodologies. Eugenia G. Ortiz Lechuga, Isela Quintero Zapata, Katiushka Arévalo Niño ...

  15. Purification and characterization of extracellular amylolytic enzyme ...

    African Journals Online (AJOL)

    DOSS

    2012-10-16

    Oct 16, 2012 ... Available online at http://www.academicjournals.org/AJB ... characterization of extracellular amylases from four ... Somogyi-Nelson's method (Nelson, 1944; Somogyi, 1952). ... The mycelia dry weight of currently studied four.

  16. Neutrophil Extracellular Traps in Ulcerative Colitis

    DEFF Research Database (Denmark)

    Bjerg Bennike, Tue; Carlsen, Thomas Gelsing; Ellingsen, Torkell

    2015-01-01

    microscopy and confocal microscopy. RESULTS: We identified and quantified 5711 different proteins with proteomics. The abundance of the proteins calprotectin and lactotransferrin in the tissue correlated with the degree of tissue inflammation as determined by histology. However, fecal calprotectin did...... not correlate. Forty-six proteins were measured with a statistically significant differences in abundances between the UC colon tissue and controls. Eleven of the proteins with increased abundances in the UC biopsies were associated with neutrophils and neutrophil extracellular traps. The findings were...... validated by microscopy, where an increased abundance of neutrophils and the presence of neutrophil extracellular traps by extracellular DNA present in the UC colon tissue were confirmed. CONCLUSIONS: Neutrophils, induced neutrophil extracellular traps, and several proteins that play a part in innate...

  17. Sources of extracellular tau and its signaling.

    Science.gov (United States)

    Avila, Jesús; Simón, Diana; Díaz-Hernández, Miguel; Pintor, Jesús; Hernández, Félix

    2014-01-01

    The pathology associated with tau protein, tauopathy, has been recently analyzed in different disorders, leading to the suggestion that intracellular and extracellular tau may itself be the principal agent in the transmission and spreading of tauopathies. Tau pathology is based on an increase in the amount of tau, an increase in phosphorylated tau, and/or an increase in aggregated tau. Indeed, phosphorylated tau protein is the main component of tau aggregates, such as the neurofibrillary tangles present in the brain of Alzheimer's disease patients. It has been suggested that intracellular tau could be toxic to neurons in its phosphorylated and/or aggregated form. However, extracellular tau could also damage neurons and since neuronal death is widespread in Alzheimer's disease, mainly among cholinergic neurons, these cells may represent a possible source of extracellular tau. However, other sources of extracellular tau have been proposed that are independent of cell death. In addition, several ways have been proposed for cells to interact with, transmit, and spread extracellular tau, and to transduce signals mediated by this tau. In this work, we will discuss the role of extracellular tau in the spreading of the tau pathology.

  18. Transcriptome of extracellular vesicles released by hepatocytes.

    Directory of Open Access Journals (Sweden)

    Felix Royo

    Full Text Available The discovery that the cells communicate through emission of vesicles has opened new opportunities for better understanding of physiological and pathological mechanisms. This discovery also provides a novel source for non-invasive disease biomarker research. Our group has previously reported that hepatocytes release extracellular vesicles with protein content reflecting the cell-type of origin. Here, we show that the extracellular vesicles released by hepatocytes also carry RNA. We report the messenger RNA composition of extracellular vesicles released in two non-tumoral hepatic models: primary culture of rat hepatocytes and a progenitor cell line obtained from a mouse foetal liver. We describe different subpopulations of extracellular vesicles with different densities and protein and RNA content. We also show that the RNA cargo of extracellular vesicles released by primary hepatocytes can be transferred to rat liver stellate-like cells and promote their activation. Finally, we provide in vitro and in vivo evidence that liver-damaging drugs galactosamine, acetaminophen, and diclofenac modify the RNA content of these vesicles. To summarize, we show that the extracellular vesicles secreted by hepatocytes contain various RNAs. These vesicles, likely to be involved in the activation of stellate cells, might become a new source for non-invasive identification of the liver toxicity markers.

  19. Regional Cerebral Blood-Flow with 99mTc-ECD Brain Perfusion SPECT in Landau-Kleffner Syndrome: Report of Two Cases

    Directory of Open Access Journals (Sweden)

    Reza Nemati

    2014-01-01

    Full Text Available Landau-Kleffner syndrome (LKS is a rare childhood disorder characterized by acquired aphasia and epilepsy. 99mTc-ECD SPECT imaging was performed in two right-handed children with LKS. A relative decrease in perfusion was found in the left frontal-temporal cortices of both patients as well as in the left and right parietal cortices of one patient with aphasia, without clinical epilepsy. The degree of regional cerebral perfusion impairment did not correlate with the severity of the clinical and EEG abnormalities, but the area of hypoperfusion was compatible with the speech area of the brain. Overall, although asymmetrical temporoparietal perfusion appears as a common finding in LKS, SPECT findings in LKS alone cannot elucidate the pathogenic features of the disorder in the brain. Here, we present two cases of LKS in which we investigated SPECT perfusion scans.

  20. Development of method for determination of Sodium monochloroacetate and sodium dichloroacetate in Cocoamido Propyl Betaine by gas chromatography: FID, ECD and MS

    International Nuclear Information System (INIS)

    Leao, Claudio; Redigolo, Marcelo Miyada; Amaral, Priscila Oliveira; Bustillos, Oscar Vega

    2015-01-01

    Cocoamide Propyl Betaine is a feedstock for consumer products of cosmetic and household segments. Sodium monochloroacetate and sodium dichloroacetate impurities are toxic, irritating and harmful to the environment and lower concentrations - parts per million level - are required in process control of producers and regulatory affairs. Regarding analytical test method, two conditions should be met: quantification limit, precision and accuracy should be appropriate; different techniques for gas chromatography - ECD, FID and MS - should be available, since for manufacturer is not so easy to keep electron capture detector by radioactive source Ni 63 due to government control and need of qualified radiological protection. The samples are obtained at manufacturers; for the analyte separation, treatment methodologies are employed by liquid-liquid extraction and solid phase extraction. Alternative detectors used in this study are: Flame Ionization and Mass Spectrometer with Electron Ionization. The validation process will be applied to methodology to ensure a selective, robust, accurate and reproducible analytical determination. (author)

  1. Development of method for determination of Sodium monochloroacetate and sodium dichloroacetate in Cocoamido Propyl Betaine by gas chromatography: FID, ECD and MS

    Energy Technology Data Exchange (ETDEWEB)

    Leao, Claudio; Redigolo, Marcelo Miyada; Amaral, Priscila Oliveira; Bustillos, Oscar Vega, E-mail: claudio.leao@usp.br [Instituto de Pesquisas Energeticas e Nucleares (IPEN/CNEN-SP), Sao Paulo, SP (Brazil)

    2015-07-01

    Cocoamide Propyl Betaine is a feedstock for consumer products of cosmetic and household segments. Sodium monochloroacetate and sodium dichloroacetate impurities are toxic, irritating and harmful to the environment and lower concentrations - parts per million level - are required in process control of producers and regulatory affairs. Regarding analytical test method, two conditions should be met: quantification limit, precision and accuracy should be appropriate; different techniques for gas chromatography - ECD, FID and MS - should be available, since for manufacturer is not so easy to keep electron capture detector by radioactive source Ni 63 due to government control and need of qualified radiological protection. The samples are obtained at manufacturers; for the analyte separation, treatment methodologies are employed by liquid-liquid extraction and solid phase extraction. Alternative detectors used in this study are: Flame Ionization and Mass Spectrometer with Electron Ionization. The validation process will be applied to methodology to ensure a selective, robust, accurate and reproducible analytical determination. (author)

  2. Role of extracellular histones in the cardiomyopathy of sepsis.

    Science.gov (United States)

    Kalbitz, Miriam; Grailer, Jamison J; Fattahi, Fatemeh; Jajou, Lawrence; Herron, Todd J; Campbell, Katherine F; Zetoune, Firas S; Bosmann, Markus; Sarma, J Vidya; Huber-Lang, Markus; Gebhard, Florian; Loaiza, Randall; Valdivia, Hector H; Jalife, José; Russell, Mark W; Ward, Peter A

    2015-05-01

    The purpose of this study was to define the relationship in polymicrobial sepsis (in adult male C57BL/6 mice) between heart dysfunction and the appearance in plasma of extracellular histones. Procedures included induction of sepsis by cecal ligation and puncture and measurement of heart function using echocardiogram/Doppler parameters. We assessed the ability of histones to cause disequilibrium in the redox status and intracellular [Ca(2+)]i levels in cardiomyocytes (CMs) (from mice and rats). We also studied the ability of histones to disturb both functional and electrical responses of hearts perfused with histones. Main findings revealed that extracellular histones appearing in septic plasma required C5a receptors, polymorphonuclear leukocytes (PMNs), and the Nacht-, LRR-, and PYD-domains-containing protein 3 (NLRP3) inflammasome. In vitro exposure of CMs to histones caused loss of homeostasis of the redox system and in [Ca(2+)]i, as well as defects in mitochondrial function. Perfusion of hearts with histones caused electrical and functional dysfunction. Finally, in vivo neutralization of histones in septic mice markedly reduced the parameters of heart dysfunction. Histones caused dysfunction in hearts during polymicrobial sepsis. These events could be attenuated by histone neutralization, suggesting that histones may be targets in the setting of sepsis to reduce cardiac dysfunction. © FASEB.

  3. Influence of a {sup 99m}TcN core on the biological and physicochemical behavior of {sup 99m}Tc complexes of L,L-EC and L,L-ECD

    Energy Technology Data Exchange (ETDEWEB)

    Mangera, Kennedy O; Vanbilloen, Hubert P.; Bellande, Emmanuel; Pasqualini, Roberto; Verbruggen, Alfons M

    1996-11-01

    {sup 99m}Tc-nitrido complexes of L,L-ethylene dicysteine ({sup 99m}TcN-L,L-EC) and {sup 99m}TcN-L,L-ethylene dicysteine diethylester ({sup 99m}TcN-L,L-ECD) were prepared and their characteristics compared to those of the respective {sup 99m}Tc-oxo complexes. {sup 99m}TcN-L,L-EC and {sup 99m}TcO-L,L-EC migrate to similar extents during electrophoresis at pH 12, but, at pH 6, {sup 99m}TcN-L,L-EC migrates further than {sup 99m}TcO-L,L-EC. Renal excretion of {sup 99m}TcN-L,L-EC is inferior to that of {sup 99m}TcO-L,L-EC, indicating that the TcN-glycine sequence has lower affinity for the renal tubular system. Both {sup 99m}TcO-L,L-ECD and {sup 99m}TcN-L,L-ECD are neutral, but {sup 99m}TcN-L,L-ECD is hydrophilic and shows minimal brain uptake in both mice and the baboon.

  4. Noninvasive quantitative assessment of cerebral blood flow (CBF) using Tc-99m ECD SPECT with adjunctive radionuclide angiography in ischemic stroke

    International Nuclear Information System (INIS)

    Yim, Jun Sung; Choi, Yun Young; Kim, Seung Hyun; Kim, Myung Ho; Cho, Suk Shin

    1999-01-01

    Quantitative CBF measurements are essential for diagnosing ischemic lesion, evaluating the therapeutic effects and predicting the prognosis of cerebral ischemia. Even though several methods have been introduced, these techniques are too cumbersome and invasive to be applied to routine studies. In this study, a non-invasive simple method for the quantitative angiography. Fifteen normal controls and 27 patients with unilateral carotid ischemic stoke were selected. Brain perfusion index (BPI) of each hemisphere was measured in each subject by acquisition of serial radionuclide angiography after injection of 20mCi of Tc-99m ECD. With Lassen's correction algorithm of curve-linear relationship between the brain activity and blood flow, rCBF on transaxial SPECT slice corresponding with MRI lesion sites (ischemic core, border zone and contralateral mirror locus) were calculated. BPI values for normal controls showed a significant negative correlation with advantage age (r=-0.64, p=0.021) and hemisphric BPI were 11.02±1.6 and 7.8±1.4 for normal controls and patient, respectively. Significant differences were observed between two groups (p=0.0012). rCBF obtained from core zone (12±2.5 ml/100/min), boneder zone (29.2±8.1) and contralateral mirror locus (52.1±15.1) were clearly defined in each subject of patient group. Measurement of BPI and rCBF using Tc-99m ECD SPECT with adjunctive radionuclide angiography could be an useful, simple and non-invasive method in evaluation of the cerebral flood in the ischemic stroke

  5. Interictal Spect with Tc-99m ECD studies in migraine patients. Role of Spect in the diagnosis of migraine with ura in headache-free periods

    International Nuclear Information System (INIS)

    Gonzalez, F.M.; Martinez, J.J.; Bermudez, M.C.; Fernandez, B.; Delgado, J.; Martin, A.; Padilla, O.; Nieto, R.

    2002-01-01

    Aim: The migraine is one of the neurological alterations more frequent provoking a discharge morbility and big inconveniences the one that endures it. The presence of brain perfusion abnormalities during the pain-free intervals of migraine even is fueling of controversies being so numerous the supporters of its existence as whom they deny it. This study was aimed at assessing the presence of cerebral perfusion abnormalities also during the interictal phase and the role of SPECT with Tc99 ECD on diagnosis of this migrainous patients. Materials And Methods: We studied 20 patients (16 woman and 4 man) with diagnose of migraine with aura during a periods without headache and 20 apareaded controls for age and sex of this patients.Was realized brain spect with 925 MBq Tc99m-ECD on a two heads rotating gamma camera (Piker Axis) equipped with a fan-beam collimator. The images were reconstructed using a filter Mezt and presented in courts in the three axes and reconstruction in three dimensions and later they were analyzed by two observers. Results: We found 14 of the patients (70%) with images of focal hypoperfusion with clear interhemispheric asymmetry while on control groups only two have this find (10%) being this difference statistically significant. The part of the brain more affected was the occipital one, though also they predominated over the more widespread hipocaptations. Conclusion: The brain Spect is a useful technology to identify the patients with migraine, being able to be used in the periods without pain. The boss of perfusion in the patients is the located hypoperfusion and is significantly different from that of the controls, It can be explained by an impaired regional cerebral vascular autoregulation may exist even during headache-free intervals in patients suffering from migraine. Future studies must be realized to determine if it exists relation between the zone of hypoperfusion and the type of present symptomatology in the aura

  6. The mechanism of reduced IgG/IgE-binding of β-lactoglobulin by pulsed electric field pretreatment combined with glycation revealed by ECD/FTICR-MS.

    Science.gov (United States)

    Yang, Wenhua; Tu, Zongcai; Wang, Hui; Zhang, Lu; Kaltashov, Igor A; Zhao, Yunlong; Niu, Chendi; Yao, Honglin; Ye, Wenfeng

    2018-01-24

    Bovine β-lactoglobulin (β-Lg) is a major allergen existing in milk and causes about 90% of IgE-mediated cow's milk allergies. Previous studies showed that pulsed electric field (PEF) treatment could partially unfold the protein, which may contribute to the improvement of protein glycation. In this study, the effect of PEF pretreatment combined with glycation on the IgG/IgE-binding ability and the structure of β-Lg was investigated. The result showed that PEF pretreatment combined with glycation significantly reduced the IgG and IgE binding abilities, which was attributed to the changes of secondary and tertiary structure and the increase in glycation sites and degree of substitution per peptide (DSP) value determined by electron capture dissociation Fourier transform ion cyclotron resonance mass spectrometry (ECD/FTICR-MS). Unexpectedly, glycation sites (K47, K91 and K135) added by two mannose molecules were identified in glycated β-Lg with PEF pretreatment. Moreover, the results indicated that PEF pretreatment at 25 kV cm -1 for 60 μs promoted the reduction of IgG/IgE-binding capacity by increasing the glycation degree of β-Lg, whereas single PEF treatment under the same conditions markedly enhanced the IgG/IgE-binding ability by partially unfolding the structure of β-Lg. The results suggested that ECD/FTICR-MS could help us to understand the mechanism of reduction in the IgG/IgE-binding of β-Lg by structural characterization at the molecular level. Therefore, PEF pretreatment combined with glycation may provide an alternative method for β-Lg desensitization.

  7. Labelled Preformed liposomes with 99MTC-DTPA, 99 MTC-ECD, 99MTC-MDP and 99MTC-MIBI : Labelling procedures and stability studies

    International Nuclear Information System (INIS)

    Savio, E.O.; Teran, M.A.; Vales, M.E.; Frier, M.

    2004-01-01

    Liposomes labelled with gamma e miters like 99mTc, can be used for scintigraphic imaging to non-invasively track and quantify the distribution of liposomes in the body. In vitro studies were done to choose a suitable radiopharmaceutical (RF) to be attached to performed liposomes. 99mTc-Complexes (DTPA, ECD, MDP, MIBI) were used to label collagen liposomes. Commercial kits were labelled with 99mTc04-(TechnoNuclear). Quality controls of the RF were performed. Collagen liposomes suspended in saline 0.9% were incubated at 4.25.37 and 60 for 30 min. Efficiency of the labelling procedure was determined by gel filtration using Sephadex G25 (Pharmacia) and NaC10.9%. Samples of 100mL (74MBq), were seeded and fractions of 0.5mL were colleted and measured in an ionisation chamber (Capintec CRC). Stability of the labelled liposomes was assessed incubating 0.5mL, of the suspension with 1mL of human serum during 30 min at 37 . Dialysis was performed using dialysis bags of 64 K pore size and NaCI 0.9% at room temperature. Samples of the saline bath were collected at 30.60 and 90 min. and measured in a solid scintillation counter Ortec.Liposomes labelled with 99mTc-DTPA and 99mTcMIBI showed a labelling efficiency of 80%; liposomes incubated with 99mTc-MDP were labelled in a 50% and 99mTc-ECD did not bind to liposomes in the conditions of study. Incubation of labelled liposomes with human serum showed 50% of strong binding to the plasmatic proteins for 99mTc-DTPA but low values (5%) for the other specimens. Labelled liposomes were achieved, with different RF, showing a suitable in vitro stability to perform in vivo studies

  8. Role of 99mTc-ECD brain SPECT in the detection of cerebral perfusion abnormality in cases of Attention Deficit Hyperactivity Disorder

    International Nuclear Information System (INIS)

    Kumar, A.; Phom, H.; Thomas, E.J.; Tripathi, M.; Chandrashekar, N.; Bal, C.S.; Zamir, H.; Gulati, S.; Kalra, V.

    2002-01-01

    Aim: A randomized placebo controlled drug trial with Mentat, a herbal pharmacological agent, was initiated in January 2000 in the department of pediatrics at AIIMS, New Delhi, to compare the efficacy of Mentat with a placebo in school children with Attention Deficit Hyperactivity Disorder (ADHD). Materials and Methods: Contact was established with 12 Public schools in Delhi, to identify poor performers in classes I-V (age 6-12 yrs.). About 195 children with poor school performance were recruited in the study. They were screened for causes of poor performance, which included attention problems, hyperactivity, behavior problems, emotional problems, mental sub-normality and learning disability. ADHD suspected children were identified using the Malin's WISC, Connor's rating scale, Problem Behavior checklist, Bender Gestalt test and some sub tests of the Kaufman's Assessment Battery for Children (K-ABC). Sixty children diagnosed as ADHD (using DSM-IV criteria), with an IQ between 90-110, were enrolled into the study. Of the 60 children randomized in the study, 30 received Mentat and 30 received an identical looking placebo. The drug/Placebo was given for a six-month period. 99mTC-ECD brain SPECT was performed in a subset of 34 children with ADHD. Results: Abnormal cerebral perfusion was seen in 23/34; thalamic hypoperfusion in 11, basal ganglia hypoperfusion in 9, thalamus and basal ganglia hypoperfusion in 2 and basifrontal hypoperfusion in 1. So far, in ten children with abnormal pretreatment scans, post treatment scans have also been done. In mentat group, 2/5 children showed normalization of perfusion abnormality after treatment whereas in placebo group, 1/5. Conclusion: This study suggests that there is selective focal cerebral hypoperfusion in cases of ADHD and 99mTc-ECD brain SPECT can be used for evaluation and further monitoring of therapeutic outcome in such cases

  9. Importance of the extracellular loops in G protein-coupled receptors for ligand recognition and receptor activation.

    Science.gov (United States)

    Peeters, M C; van Westen, G J P; Li, Q; IJzerman, A P

    2011-01-01

    G protein-coupled receptors (GPCRs) are the major drug target of medicines on the market today. Therefore, much research is and has been devoted to the elucidation of the function and three-dimensional structure of this large family of membrane proteins, which includes multiple conserved transmembrane domains connected by intra- and extracellular loops. In the last few years, the less conserved extracellular loops have garnered increasing interest, particularly after the publication of several GPCR crystal structures that clearly show the extracellular loops to be involved in ligand binding. This review will summarize the recent progress made in the clarification of the ligand binding and activation mechanism of class-A GPCRs and the role of extracellular loops in this process. Copyright © 2010 Elsevier Ltd. All rights reserved.

  10. Assessment of extracellular dehydration using saliva osmolality.

    Science.gov (United States)

    Ely, Brett R; Cheuvront, Samuel N; Kenefick, Robert W; Spitz, Marissa G; Heavens, Kristen R; Walsh, Neil P; Sawka, Michael N

    2014-01-01

    When substantial solute losses accompany body water an isotonic hypovolemia (extracellular dehydration) results. The potential for using blood or urine to assess extracellular dehydration is generally poor, but saliva is not a simple ultra-filtrate of plasma and the autonomic regulation of salivary gland function suggests the possibility that saliva osmolality (Sosm) may afford detection of extracellular dehydration via the influence of volume-mediated factors. This study aimed to evaluate the assessment of extracellular dehydration using Sosm. In addition, two common saliva collection methods and their effects on Sosm were compared. Blood, urine, and saliva samples were collected in 24 healthy volunteers during paired euhydration and dehydration trials. Furosemide administration and 12 h fluid restriction were used to produce extracellular dehydration. Expectoration and salivette collection methods were compared in a separate group of eight euhydrated volunteers. All comparisons were made using paired t-tests. The diagnostic potential of body fluids was additionally evaluated. Dehydration (3.1 ± 0.5% loss of body mass) decreased PV (-0.49 ± 0.12 L; -15.12 ± 3.94% change), but Sosm changes were marginal ( 0.05). Extracelluar dehydration was not detectable using plasma, urine, or saliva measures. Salivette and expectoration sampling methods produced similar, consistent results for Sosm, suggesting no methodological influence on Sosm.

  11. Extracellular histones in tissue injury and inflammation.

    Science.gov (United States)

    Allam, Ramanjaneyulu; Kumar, Santhosh V R; Darisipudi, Murthy N; Anders, Hans-Joachim

    2014-05-01

    Neutrophil NETosis is an important element of host defense as it catapults chromatin out of the cell to trap bacteria, which then are killed, e.g., by the chromatin's histone component. Also, during sterile inflammation TNF-alpha and other mediators trigger NETosis, which elicits cytotoxic effects on host cells. The same mechanism should apply to other forms of regulated necrosis including pyroptosis, necroptosis, ferroptosis, and cyclophilin D-mediated regulated necrosis. Beyond these toxic effects, extracellular histones also trigger thrombus formation and innate immunity by activating Toll-like receptors and the NLRP3 inflammasome. Thereby, extracellular histones contribute to the microvascular complications of sepsis, major trauma, small vessel vasculitis as well as acute liver, kidney, brain, and lung injury. Finally, histones prevent the degradation of extracellular DNA, which promotes autoimmunization, anti-nuclear antibody formation, and autoimmunity in susceptible individuals. Here, we review the current evidence on the pathogenic role of extracellular histones in disease and discuss how to target extracellular histones to improve disease outcomes.

  12. Extracellular matrix organization in developing muscle: correlation with acetylcholine receptor aggregates.

    Science.gov (United States)

    Bayne, E K; Anderson, M J; Fambrough, D M

    1984-10-01

    organization over myotubes and fibroblasts suggest that each of these cell types uses somewhat different means to regulate the assembly of extracellular matrix components within its domain. (c) The limited co-distribution of different components within the extracellular matrix in vitro and the selective immune precipitation of each antigen from conditioned medium suggest that each extracellular matrix component is secreted in a form that is not complexed with other matrix constituents.

  13. A collagen-binding EGFR antibody fragment targeting tumors with a collagen-rich extracellular matrix

    OpenAIRE

    Hui Liang; Xiaoran Li; Bin Wang; Bing Chen; Yannan Zhao; Jie Sun; Yan Zhuang; Jiajia Shi; He Shen; Zhijun Zhang; Jianwu Dai

    2016-01-01

    Many tumors over-express collagen, which constitutes the physical scaffold of tumor microenvironment. Collagen has been considered to be a target for cancer therapy. The collagen-binding domain (CBD) is a short peptide, which could bind to collagen and achieve the sustained release of CBD-fused proteins in collagen scaffold. Here, a collagen-binding EGFR antibody fragment was designed and expressed for targeting the collagen-rich extracellular matrix in tumors. The antibody fragment (Fab) of ...

  14. The Staphylococcus aureus extracellular adherence protein (Eap) adopts an elongated but structured conformation in solution

    OpenAIRE

    Hammel, Michal; Němeček, Daniel; Keightley, J. Andrew; Thomas, George J.; Geisbrecht, Brian V.

    2007-01-01

    The extracellular adherence protein (Eap) of Staphylococcus aureus participates in a wide range of protein–protein interactions that facilitate the initiation and dissemination of Staphylococcal disease. In this report, we describe the use of a multidisciplinary approach to characterize the solution structure of full-length Eap. In contrast to previous reports suggesting that a six-domain isoform of Eap undergoes multimerization, sedimentation equilibrium analytical ultracentrifugation data r...

  15. Shaping Synapses by the Neural Extracellular Matrix

    Directory of Open Access Journals (Sweden)

    Maura Ferrer-Ferrer

    2018-05-01

    Full Text Available Accumulating data support the importance of interactions between pre- and postsynaptic neuronal elements with astroglial processes and extracellular matrix (ECM for formation and plasticity of chemical synapses, and thus validate the concept of a tetrapartite synapse. Here we outline the major mechanisms driving: (i synaptogenesis by secreted extracellular scaffolding molecules, like thrombospondins (TSPs, neuronal pentraxins (NPs and cerebellins, which respectively promote presynaptic, postsynaptic differentiation or both; (ii maturation of synapses via reelin and integrin ligands-mediated signaling; and (iii regulation of synaptic plasticity by ECM-dependent control of induction and consolidation of new synaptic configurations. Particularly, we focused on potential importance of activity-dependent concerted activation of multiple extracellular proteases, such as ADAMTS4/5/15, MMP9 and neurotrypsin, for permissive and instructive events in synaptic remodeling through localized degradation of perisynaptic ECM and generation of proteolytic fragments as inducers of synaptic plasticity.

  16. A Conserved Salt Bridge between Transmembrane Segment 1 and 10 Constitutes an Extracellular Gate in the Dopamine Transporter

    DEFF Research Database (Denmark)

    Pedersen, Anders Vingborg; Andreassen, Thorvald Faurschou; Løland, Claus Juul

    2014-01-01

    into the structural and dynamic requirements for substrate transport. These structures support the existence of gating domains controlling access to a central binding site. On the extracellular side, access is controlled by the "thin gate" formed by an interaction between Arg-30 and Asp-404. In the human dopamine...

  17. Extracellular enzyme kinetics scale with resource availability

    Science.gov (United States)

    Sinsabaugh, Robert L.; Belnap, Jayne; Findlay, Stuart G.; Follstad Shah, Jennifer J.; Hill, Brian H.; Kuehn, Kevin A.; Kuske, Cheryl; Litvak, Marcy E.; Martinez, Noelle G.; Moorhead, Daryl L.; Warnock, Daniel D.

    2014-01-01

    Microbial community metabolism relies on external digestion, mediated by extracellular enzymes that break down complex organic matter into molecules small enough for cells to assimilate. We analyzed the kinetics of 40 extracellular enzymes that mediate the degradation and assimilation of carbon, nitrogen and phosphorus by diverse aquatic and terrestrial microbial communities (1160 cases). Regression analyses were conducted by habitat (aquatic and terrestrial), enzyme class (hydrolases and oxidoreductases) and assay methodology (low affinity and high affinity substrates) to relate potential reaction rates to substrate availability. Across enzyme classes and habitats, the scaling relationships between apparent Vmax and apparent Km followed similar power laws with exponents of 0.44 to 0.67. These exponents, called elasticities, were not statistically distinct from a central value of 0.50, which occurs when the Km of an enzyme equals substrate concentration, a condition optimal for maintenance of steady state. We also conducted an ecosystem scale analysis of ten extracellular hydrolase activities in relation to soil and sediment organic carbon (2,000–5,000 cases/enzyme) that yielded elasticities near 1.0 (0.9 ± 0.2, n = 36). At the metabolomic scale, the elasticity of extracellular enzymatic reactions is the proportionality constant that connects the C:N:P stoichiometries of organic matter and ecoenzymatic activities. At the ecosystem scale, the elasticity of extracellular enzymatic reactions shows that organic matter ultimately limits effective enzyme binding sites. Our findings suggest that one mechanism by which microbial communities maintain homeostasis is regulating extracellular enzyme expression to optimize the short-term responsiveness of substrate acquisition. The analyses also show that, like elemental stoichiometry, the fundamental attributes of enzymatic reactions can be extrapolated from biochemical to community and ecosystem scales.

  18. MR imaging of intracellular and extracellular deoxyhemoglobin

    International Nuclear Information System (INIS)

    Janick, P.A.; Grossman, R.I.; Asakura, T.

    1989-01-01

    MR imaging was performed on varying concentrations of intracellular and extracellular deoxyhemoglobin as well as varying proportions of deoxyhemoglobin and oxyhemoglobin in vitro at 1.5T with use of standard spin-echo and gradient-refocused spin sequences. This study indicates that susceptibility-induced T2 shortening occurs over a broad range of intracellular deoxyhemoglobin concentrations (maximal at hematocrits between 20% and 45%), reflecting diffusional effects at the cellular level. T2* gradient-echo imaging enhances the observed hypointensity in images of intracellular deoxyhemoglobin. The characteristic MR appearance of acute hemotomas can be modeled by the behavior of intracellular and extracellular deoxyhemoglobin and oxyhemoglobin

  19. Optimization of extracellular catalase production from Aspergillus ...

    African Journals Online (AJOL)

    The studies of the effect of each variable and the establishment of a correlation between the response of enzyme activity and variables revealed that the link is a multiple linear regression form. The optimization was carried out through a simplex algorithm. The amount of extracellular catalase produced by the strain in the ...

  20. Managing Brain Extracellular K(+) during Neuronal Activity

    DEFF Research Database (Denmark)

    Larsen, Brian Roland; Stoica, Anca; MacAulay, Nanna

    2016-01-01

    characteristics required to fulfill their distinct physiological roles in clearance of K(+) from the extracellular space in the face of neuronal activity. Understanding the nature, impact and effects of the various Na(+)/K(+)-ATPase isoform combinations in K(+) management in the central nervous system might...... understanding of the pathological events occurring during disease....

  1. Extracellular vesicles: fundamentals and clinical relevance

    Directory of Open Access Journals (Sweden)

    Wael Nassar

    2015-01-01

    Full Text Available All types of cells of eukaryotic organisms produce and release small nanovesicles into their extracellular environment. Early studies have described these vesicles as ′garbage bags′ only to remove obsolete cellular molecules. Valadi and colleagues, in 2007, were the first to discover the capability of circulating extracellular vesicles (EVs to horizontally transfer functioning gene information between cells. These extracellular vesicles express components responsible for angiogenesis promotion, stromal remodeling, chemoresistance, genetic exchange, and signaling pathway activation through growth factor/receptor transfer. EVs represent an important mode of intercellular communication by serving as vehicles for transfer between cells of membrane and cytosolic proteins, lipids, signaling proteins, and RNAs. They contribute to physiology and pathology, and they have a myriad of potential clinical applications in health and disease. Moreover, vesicles can pass the blood-brain barrier and may perhaps even be considered as naturally occurring liposomes. These cell-derived EVs not only represent a central mediator of the disease microenvironment, but their presence in the peripheral circulation may serve as a surrogate for disease biopsies, enabling real-time diagnosis and disease monitoring. In this review, we′ll be addressing the characteristics of different types of extracellular EVs, as well as their clinical relevance and potential as diagnostic markers, and also define therapeutic options.

  2. Extracellular space diffusion and extrasynaptic transmission

    Czech Academy of Sciences Publication Activity Database

    Vargová, Lýdia; Syková, Eva

    2008-01-01

    Roč. 57, Suppl.3 (2008), S89-S99 ISSN 0862-8408 R&D Projects: GA MŠk 1M0538; GA MŠk(CZ) LC554 Institutional research plan: CEZ:AV0Z50390512 Keywords : Diffusion * Extracellular volume * Tortuosity Subject RIV: FH - Neurology Impact factor: 1.653, year: 2008

  3. Integrins and extracellular matrix in mechanotransduction

    Directory of Open Access Journals (Sweden)

    Ramage L

    2011-12-01

    Full Text Available Lindsay RamageQueen’s Medical Research Institute, University of Edinburgh, Edinburgh, UKAbstract: Integrins are a family of cell surface receptors which mediate cell–matrix and cell–cell adhesions. Among other functions they provide an important mechanical link between the cells external and intracellular environments while the adhesions that they form also have critical roles in cellular signal-transduction. Cell–matrix contacts occur at zones in the cell surface where adhesion receptors cluster and when activated the receptors bind to ligands in the extracellular matrix. The extracellular matrix surrounds the cells of tissues and forms the structural support of tissue which is particularly important in connective tissues. Cells attach to the extracellular matrix through specific cell-surface receptors and molecules including integrins and transmembrane proteoglycans. Integrins work alongside other proteins such as cadherins, immunoglobulin superfamily cell adhesion molecules, selectins, and syndecans to mediate cell–cell and cell–matrix interactions and communication. Activation of adhesion receptors triggers the formation of matrix contacts in which bound matrix components, adhesion receptors, and associated intracellular cytoskeletal and signaling molecules form large functional, localized multiprotein complexes. Cell–matrix contacts are important in a variety of different cell and tissue properties including embryonic development, inflammatory responses, wound healing, and adult tissue homeostasis. This review summarizes the roles and functions of integrins and extracellular matrix proteins in mechanotransduction.Keywords: ligand binding, α subunit, ß subunit, focal adhesion, cell differentiation, mechanical loading, cell–matrix interaction

  4. Interaction of acetamiprid with extracellular polymeric substances ...

    African Journals Online (AJOL)

    Extracellular polymeric substances (EPS) are important components of activated sludge and it plays an important role in removing pollutants. The interaction between EPS and organic pollutants is still little known. In the present study, the interaction of soluble/bound EPS with acetamiprid, a neonicotinoid insecticide, was ...

  5. Optimization of extracellular catalase production from Aspergillus ...

    African Journals Online (AJOL)

    aghomotsegin

    extracellular catalase produced by the strain in the optimized medium was about four times higher than ... celial and unicellular fungi in synthetic media (Kurakov et .... covering the appropriate range and the broad calibration kit ... This optimization allowed us to define new cultural con- ..... Ann. New York Academy Sci.

  6. Production of extracellular aspartic protease in submerged ...

    African Journals Online (AJOL)

    Fungal milk-clotting enzymes have gained value as bovine Chymosin substitutes in the cheese industry. In this work, the effects of culture conditions on the production of extracellular milk clotting enzymes from Mucor mucedo DSM 809 in submerged fermentation were studied. The maximum activity was observed after 48 h ...

  7. Extracellular matrix and tissue engineering applications

    NARCIS (Netherlands)

    Fernandes, H.A.M.; Moroni, Lorenzo; van Blitterswijk, Clemens; de Boer, Jan

    2009-01-01

    The extracellular matrix is a key component during regeneration and maintenance of tissues and organs, and it therefore plays a critical role in successful tissue engineering as well. Tissue engineers should recognise that engineering technology can be deduced from natural repair processes. Due to

  8. DPP6 domains responsible for its localization and function.

    Science.gov (United States)

    Lin, Lin; Long, Laura K; Hatch, Michael M; Hoffman, Dax A

    2014-11-14

    Dipeptidyl peptidase-like protein 6 (DPP6) is an auxiliary subunit of the Kv4 family of voltage-gated K(+) channels known to enhance channel surface expression and potently accelerate their kinetics. DPP6 is a single transmembrane protein, which is structurally remarkable for its large extracellular domain. Included in this domain is a cysteine-rich motif, the function of which is unknown. Here we show that this cysteine-rich domain of DPP6 is required for its export from the ER and expression on the cell surface. Disulfide bridges formed at C349/C356 and C465/C468 of the cysteine-rich domain are necessary for the enhancement of Kv4.2 channel surface expression but not its interaction with Kv4.2 subunits. The short intracellular N-terminal and transmembrane domains of DPP6 associates with and accelerates the recovery from inactivation of Kv4.2, but the entire extracellular domain is necessary to enhance Kv4.2 surface expression and stabilization. Our findings show that the cysteine-rich domain of DPP6 plays an important role in protein folding of DPP6 that is required for transport of DPP6/Kv4.2 complexes out of the ER. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

  9. The SH2 domain interaction landscape.

    Science.gov (United States)

    Tinti, Michele; Kiemer, Lars; Costa, Stefano; Miller, Martin L; Sacco, Francesca; Olsen, Jesper V; Carducci, Martina; Paoluzi, Serena; Langone, Francesca; Workman, Christopher T; Blom, Nikolaj; Machida, Kazuya; Thompson, Christopher M; Schutkowski, Mike; Brunak, Søren; Mann, Matthias; Mayer, Bruce J; Castagnoli, Luisa; Cesareni, Gianni

    2013-04-25

    Members of the SH2 domain family modulate signal transduction by binding to short peptides containing phosphorylated tyrosines. Each domain displays a distinct preference for the sequence context of the phosphorylated residue. We have developed a high-density peptide chip technology that allows for probing of the affinity of most SH2 domains for a large fraction of the entire complement of tyrosine phosphopeptides in the human proteome. Using this technique, we have experimentally identified thousands of putative SH2-peptide interactions for more than 70 different SH2 domains. By integrating this rich data set with orthogonal context-specific information, we have assembled an SH2-mediated probabilistic interaction network, which we make available as a community resource in the PepspotDB database. A predicted dynamic interaction between the SH2 domains of the tyrosine phosphatase SHP2 and the phosphorylated tyrosine in the extracellular signal-regulated kinase activation loop was validated by experiments in living cells. Copyright © 2013 The Authors. Published by Elsevier Inc. All rights reserved.

  10. The SH2 Domain Interaction Landscape

    Directory of Open Access Journals (Sweden)

    Michele Tinti

    2013-04-01

    Full Text Available Members of the SH2 domain family modulate signal transduction by binding to short peptides containing phosphorylated tyrosines. Each domain displays a distinct preference for the sequence context of the phosphorylated residue. We have developed a high-density peptide chip technology that allows for probing of the affinity of most SH2 domains for a large fraction of the entire complement of tyrosine phosphopeptides in the human proteome. Using this technique, we have experimentally identified thousands of putative SH2-peptide interactions for more than 70 different SH2 domains. By integrating this rich data set with orthogonal context-specific information, we have assembled an SH2-mediated probabilistic interaction network, which we make available as a community resource in the PepspotDB database. A predicted dynamic interaction between the SH2 domains of the tyrosine phosphatase SHP2 and the phosphorylated tyrosine in the extracellular signal-regulated kinase activation loop was validated by experiments in living cells.

  11. Extracellular nucleotide derivatives protect cardiomyctes against hypoxic stress

    DEFF Research Database (Denmark)

    Golan, O; Issan, Y; Isak, A

    2011-01-01

    assures cardioprotection. Treatment with extracellular nucleotides, or with tri/di-phosphate, administered under normoxic conditions or during hypoxic conditions, led to a decrease in reactive oxygen species production. CONCLUSIONS: Extracellular tri/di-phosphates are apparently the molecule responsible...

  12. Involvement of extracellular matrix constituents in breast cancer

    Energy Technology Data Exchange (ETDEWEB)

    Lochter, Andre; Bissell, Mina J

    1995-06-01

    It has recently been established that the extracellular matrix is required for normal functional differentiation of mammary epithelia not only in culture, but also in vivo. The mechanisms by which extracellular matrix affects differentiation, as well as the nature of extracellular matrix constituents which have major impacts on mammary gland function, have only now begun to be dissected. The intricate variety of extracellular matrix-mediated events and the remarkable degree of plasticity of extracellular matrix structure and composition at virtually all times during ontogeny, make such studies difficult. Similarly, during carcinogenesis, the extracellular matrix undergoes gross alterations, the consequences of which are not yet precisely understood. Nevertheless, an increasing amount of data suggests that the extracellular matrix and extracellular matrix-receptors might participate in the control of most, if not all, of the successive stages of breast tumors, from appearance to progression and metastasis.

  13. Ligand binding and activation mechanism og the glucagon-like peptide-1 receptor

    DEFF Research Database (Denmark)

    Underwood, Christina Rye

    GLP-1R interacts with receptor agonists. The thesis includes four studies, which investigate different aspects of these interactions. The first study elucidates GLP-1 binding to the extracellular domain of GLP-1R (ECD) (Study I), whereas the second study identifies receptor domains important for small...

  14. Motor cortex stimulation(MCS) for intractable complex regional pain syndrome (CRPS) type II: PSM analysis of Tc-99m ECD brain perfusion SPECT

    Energy Technology Data Exchange (ETDEWEB)

    Chung, Y. A.; Son, B. C.; Yoo, I. R.; Kim, S. H.; Kim, E. N.; Park, Y. H.; Lee, S. Y.; Sohn, H. S.; Chung, S. K. [College of Medicine, The Catholic Univ. of Korea, Seoul (Korea, Republic of)

    2001-07-01

    We had experienced a patient with intractable CRPS in whom statistical parametric mapping (SPM) analysis of cerebral perfusion explained the mechanism of pain control by MCS. A 43-year-old man presented spontaneous severe burning pain in his left hand and forearm and allodynia over the left arm and left hemibody. After the electrodes for neuromodulation therapy were inserted in the central sulcus, the baseline and stimulation brain perfusion SPECT using Tc-99m ECD were obtained within two days. The differences between the baseline and stimulation SPECT images, estimated at every voxel using t-statistics using SPM-99 software, were considered significant at a threshold of uncorrected P values less than 0.01. Among several areas significantly activated following pain relief with MCS, ipsilateral pyramidal tract in the cerebral peduncle might be related to the mechanism of pain control with MCS through efferent motor pathway. The result suggested that corticospinal neurons themselves or motor cortex efferent pathway maintained by the presence of intact corticospinal neurons could play an important role in producing pain control after MCS. This study would helpful in understanding of neurophysiology.

  15. Determination of pesticide residues in leafy vegetables at parts per billion levels by a chemometric study using GC-ECD in Cameron Highlands, Malaysia.

    Science.gov (United States)

    Farina, Yang; Abdullah, Md Pauzi; Bibi, Nusrat; Khalik, Wan Mohd Afiq Wan Mohd

    2017-06-01

    A simple and sensitive analytical method has been developed employing gas chromatography coupled with electron capture detector (GC-ECD), and validated for screening and quantification of 15 pesticide residues at trace levels in cabbage, broccoli, cauliflower, lettuce, celery, spinach, and mustard. The method consists of two steps, first, to determine the significance of each factor by Pareto chart followed by optimization of these significant factors using central composite design (CCD). Minitab statistical software was used for these multivariate experiments for the generation of 2 4-1 design and CCD matrices. The method evaluation was done by external standard calibration with linearity range between 0.5 and 3mg/kg, with correlation coefficient 0.99, limit of detection (LOD) ranges between 0.02 and 4.5ng/g, and limit of quantification (LOQ) ranges between 0.2 and 45ng/g. The average recovery was between 60% and 128%, with RSD 0.2-19.8%. The method was applied on real vegetable samples from Cameron Highlands. Copyright © 2016 Elsevier Ltd. All rights reserved.

  16. Cerebral perfusion abnormalities in therapy-resistant epilepsy in childhood: comparison between EEG, MRI and 99Tcm-ECD brain SPET.

    Science.gov (United States)

    Vattimo, A; Burroni, L; Bertelli, P; Volterrani, D; Vella, A

    1996-01-01

    We performed 99Tcm-ethyl cysteinate dimer (ECD) interictal single photon emission tomography (SPET) in 26 children with severe therapy-resistant epilepsy. All the children underwent a detailed clinical examination, an electroencephalogram (EEG) investigation and brain magnetic resonance imaging (MRI). In 21 of the 26 children, SPET demonstrated brain blood flow abnormalities, in 13 cases in the same territories that showed EEG alterations. MRI showed structural lesions in 6 of the 26 children, while SPET imaging confirmed these abnormalities in only 5 children. The lesion not detected on SPET was shown to be 3 mm thick on MRI. Five symptomatic patients had normal SPET. In one of these patients, the EEG findings were normal and MRI revealed a small calcific nodule (4 mm thick); in the others, the EEG showed non-focal but diffuse abnormalities. These data confirm that brain SPET is sensitive in detecting and localizing hypoperfused areas that could be associated with epileptic foci in this group of patients, even when the MRI image is normal.

  17. Motor cortex stimulation(MCS) for intractable complex regional pain syndrome (CRPS) type II: PSM analysis of Tc-99m ECD brain perfusion SPECT

    International Nuclear Information System (INIS)

    Chung, Y. A.; Son, B. C.; Yoo, I. R.; Kim, S. H.; Kim, E. N.; Park, Y. H.; Lee, S. Y.; Sohn, H. S.; Chung, S. K.

    2001-01-01

    We had experienced a patient with intractable CRPS in whom statistical parametric mapping (SPM) analysis of cerebral perfusion explained the mechanism of pain control by MCS. A 43-year-old man presented spontaneous severe burning pain in his left hand and forearm and allodynia over the left arm and left hemibody. After the electrodes for neuromodulation therapy were inserted in the central sulcus, the baseline and stimulation brain perfusion SPECT using Tc-99m ECD were obtained within two days. The differences between the baseline and stimulation SPECT images, estimated at every voxel using t-statistics using SPM-99 software, were considered significant at a threshold of uncorrected P values less than 0.01. Among several areas significantly activated following pain relief with MCS, ipsilateral pyramidal tract in the cerebral peduncle might be related to the mechanism of pain control with MCS through efferent motor pathway. The result suggested that corticospinal neurons themselves or motor cortex efferent pathway maintained by the presence of intact corticospinal neurons could play an important role in producing pain control after MCS. This study would helpful in understanding of neurophysiology

  18. Determination of Chlorinated Pesticides and Polychlorinated Biphenyls in Edible Oils and Fats using MSPD Extraction and Freeze Lipids-Filtration Cleanup with GC-ECD Measurement

    International Nuclear Information System (INIS)

    Pla Jauregui, Daniela; Valcarcel Rojas, Lino; Estevez Alvarez, Juan

    2015-01-01

    Persistent organic pollutants (POPs) such as organo chlorine pesticides (OCPs) and polychlorinated biphenyls (PCBs) have numerous adverse effects on human health. Although their use have been banned or restricted, residues of these compounds can be found in foods, especially those with high fat content. The purpose of this study was to establish a rapid and simple analytical method for the determination of these compounds in edible oils and fats. Matrix solid-phase dispersion (MSPD) was used for the simultaneous extraction and purification of analytes from samples using an acetonitrile - acetone (19:1, v/v) mixture as eluting solvent. Lipids remaining in the extract were removed by freezing-lipid filtration, prior to analytes re-extraction with n-hexane. The measurement was performed using a gas chromatograph with electron capture detector (GC-ECD). Spiking experiments were carried out to determine the recovery, precision and limits of detection (LODs) of the method. Recoveries were higher than 70% and detection limits were in the range of 20 ng/g to 90 ng/g. These LOD values are lower than the maximum permissible limit which makes this method useful for the purpose intended. This method was successfully applied to the analysis of vegetable oils and butters sampled from the market. (Author)

  19. Age-dependence of local cerebral blood flow - investigations of 4 to 15 years old children and adults with Tc-99m-ECD

    International Nuclear Information System (INIS)

    Barthel, H.; Wiener, M.; Dannenberg, C.; Sattler, B.; Martin, W.G.; Knapp, W.H.

    1997-01-01

    Aim: This present study deals with the question whether normal distribution of local cerebral blood flow (ICBF) agents in adults can be transferred to the age group of 4 to 15 years old children. Methods: 23 children (age: 4-15 years, mean 11±3 y) (group I) and 10 adults (age: 27-56 years, mean 45±10 y) (group II) without evidence of cerebrovascular disease or other brain diseases underwent Tc-99m-ECD-SPECT imaging. Counts in the cortical ROIs were related to those of cerebellar ROIs (=100%). Results: In group I, relative cortical activity exceeded that of group II, particularly in parietal (107.6±9.8 vs. 84.1±12.4%), frontal (97.7±6.7 vs. 79.4±8.9%), left temporal areas (99.7±7.4 vs. 84.9±10.1%) and in the cingular cortex (112.1±9.1 vs. 95.9±10.1%, p [de

  20. Evidence for cooperative signal triggering at the extracellular loops of the TSH receptor.

    Science.gov (United States)

    Kleinau, Gunnar; Jaeschke, Holger; Mueller, Sandra; Raaka, Bruce M; Neumann, Susanne; Paschke, Ralf; Krause, Gerd

    2008-08-01

    The mechanisms governing transition of the thyroid stimulating hormone (TSH) receptor (TSHR) from basal to active conformations are poorly understood. Considering that constitutively activating mutations (CAMs) and inactivating mutations in each of the extracellular loops (ECLs) trigger only partial TSHR activation or inactivation, respectively, we hypothesized that full signaling occurs via multiple extracellular signal propagation events. Therefore, individual CAMs in the extracellular region were combined to create double and triple mutants. In support of our hypothesis, combinations of mutants in the ECLs are in some cases additive, while in others they are even synergistic, with triple mutant I486A/I568V/V656F exhibiting a 70-fold increase in TSH-independent signaling. The proximity but likely different spatial orientation of the residues of activating and inactivating mutations in each ECL supports a dual functionality to facilitate signal induction and conduction, respectively. This is the first report for G-protein coupled receptors, suggesting that multiple and cooperative signal propagating events at all three ECLs are required for full receptor activation. Our findings provide new insights concerning molecular signal transmission from extracellular domains toward the transmembrane helix bundle of the glycoprotein hormone receptors.

  1. Extracellular signaling and multicellularity in Bacillus subtilis.

    Science.gov (United States)

    Shank, Elizabeth Anne; Kolter, Roberto

    2011-12-01

    Bacillus subtilis regulates its ability to differentiate into distinct, co-existing cell types in response to extracellular signaling molecules produced either by itself, or present in its environment. The production of molecules by B. subtilis cells, as well as their response to these signals, is not uniform across the population. There is specificity and heterogeneity both within genetically identical populations as well as at the strain-level and species-level. This review will discuss how extracellular signaling compounds influence B. subtilis multicellularity with regard to matrix-producing cannibal differentiation, germination, and swarming behavior, as well as the specificity of the quorum-sensing peptides ComX and CSF. It will also highlight how imaging mass spectrometry can aid in identifying signaling compounds and contribute to our understanding of the functional relationship between such compounds and multicellular behavior. Copyright © 2011 Elsevier Ltd. All rights reserved.

  2. Biotechnological Aspects of Microbial Extracellular Electron Transfer

    Science.gov (United States)

    Kato, Souichiro

    2015-01-01

    Extracellular electron transfer (EET) is a type of microbial respiration that enables electron transfer between microbial cells and extracellular solid materials, including naturally-occurring metal compounds and artificial electrodes. Microorganisms harboring EET abilities have received considerable attention for their various biotechnological applications, in addition to their contribution to global energy and material cycles. In this review, current knowledge on microbial EET and its application to diverse biotechnologies, including the bioremediation of toxic metals, recovery of useful metals, biocorrosion, and microbial electrochemical systems (microbial fuel cells and microbial electrosynthesis), were introduced. Two potential biotechnologies based on microbial EET, namely the electrochemical control of microbial metabolism and electrochemical stimulation of microbial symbiotic reactions (electric syntrophy), were also discussed. PMID:26004795

  3. Methods to isolate extracellular vesicles for diagnosis

    Science.gov (United States)

    Kang, Hyejin; Kim, Jiyoon; Park, Jaesung

    2017-12-01

    Extracellular vesicles (EVs) are small membrane-bound bodies that are released into extracellular space by diverse cells, and are found in body fluids like blood, urine and saliva. EVs contain RNA, DNA and proteins, which can be biomarkers for diagnosis. EVs can be obtained by minimally-invasive biopsy, so they are useful in disease diagnosis. High yield and purity contribute to precise diagnosis of disease, but damaged EVs and impurities can cause confu sed results. However, EV isolation methods have different yields and purities. Furthermore, the isolation method that is most suitable to maximize EV recovery efficiency depends on the experimental conditions. This review focuses on merits and demerits of several types of EV isolation methods, and provides examples of how to diagnose disease by exploiting information obtained by analysis of EVs.

  4. Functional Elements on SIRPα IgV domain Mediate Cell Surface Binding to CD47

    OpenAIRE

    Liu, Yuan; Tong, Qiao; Zhou, Yubin; Lee, Hsiau-Wei; Yang, Jenny J.; Bühring, Hans-Jörg; Chen, Yi-Tien; Ha, Binh; Chen, Celia X-J.; Zen, Ke

    2006-01-01

    SIRPα and SIRPβ1, the two major isoforms of the signal regulatory protein (SIRP) family, are co-expressed in human leukocytes but mediate distinct extracellular binding interactions and divergent cell signaling responses. Previous studies have demonstrated that binding of SIRPα with CD47, another important cell surface molecule, through the extracellular IgV domain regulates important leukocyte functions including macrophage recognition, leukocyte adhesion and transmigration. Although SIRPβ1 ...

  5. Solution structure of leptospiral LigA4 Big domain

    Energy Technology Data Exchange (ETDEWEB)

    Mei, Song; Zhang, Jiahai [Hefei National Laboratory for Physical Sciences at Microscale, School of Life Sciences, University of Science and Technology of China, Hefei, Anhui 230026 (China); Zhang, Xuecheng [School of Life Sciences, Anhui University, Hefei, Anhui 230039 (China); Tu, Xiaoming, E-mail: xmtu@ustc.edu.cn [Hefei National Laboratory for Physical Sciences at Microscale, School of Life Sciences, University of Science and Technology of China, Hefei, Anhui 230026 (China)

    2015-11-13

    Pathogenic Leptospiraspecies express immunoglobulin-like proteins which serve as adhesins to bind to the extracellular matrices of host cells. Leptospiral immunoglobulin-like protein A (LigA), a surface exposed protein containing tandem repeats of bacterial immunoglobulin-like (Big) domains, has been proved to be involved in the interaction of pathogenic Leptospira with mammalian host. In this study, the solution structure of the fourth Big domain of LigA (LigA4 Big domain) from Leptospira interrogans was solved by nuclear magnetic resonance (NMR). The structure of LigA4 Big domain displays a similar bacterial immunoglobulin-like fold compared with other Big domains, implying some common structural aspects of Big domain family. On the other hand, it displays some structural characteristics significantly different from classic Ig-like domain. Furthermore, Stains-all assay and NMR chemical shift perturbation revealed the Ca{sup 2+} binding property of LigA4 Big domain. - Highlights: • Determining the solution structure of a bacterial immunoglobulin-like domain from a surface protein of Leptospira. • The solution structure shows some structural characteristics significantly different from the classic Ig-like domains. • A potential Ca{sup 2+}-binding site was identified by strains-all and NMR chemical shift perturbation.

  6. Solution structure of leptospiral LigA4 Big domain

    International Nuclear Information System (INIS)

    Mei, Song; Zhang, Jiahai; Zhang, Xuecheng; Tu, Xiaoming

    2015-01-01

    Pathogenic Leptospiraspecies express immunoglobulin-like proteins which serve as adhesins to bind to the extracellular matrices of host cells. Leptospiral immunoglobulin-like protein A (LigA), a surface exposed protein containing tandem repeats of bacterial immunoglobulin-like (Big) domains, has been proved to be involved in the interaction of pathogenic Leptospira with mammalian host. In this study, the solution structure of the fourth Big domain of LigA (LigA4 Big domain) from Leptospira interrogans was solved by nuclear magnetic resonance (NMR). The structure of LigA4 Big domain displays a similar bacterial immunoglobulin-like fold compared with other Big domains, implying some common structural aspects of Big domain family. On the other hand, it displays some structural characteristics significantly different from classic Ig-like domain. Furthermore, Stains-all assay and NMR chemical shift perturbation revealed the Ca"2"+ binding property of LigA4 Big domain. - Highlights: • Determining the solution structure of a bacterial immunoglobulin-like domain from a surface protein of Leptospira. • The solution structure shows some structural characteristics significantly different from the classic Ig-like domains. • A potential Ca"2"+-binding site was identified by strains-all and NMR chemical shift perturbation.

  7. Stem cell extracellular vesicles and kidney injury

    OpenAIRE

    Grange, Cristina; Iampietro, Corinne; Bussolati, Benedetta

    2017-01-01

    Extracellular vesicles (EVs) appear as a new promising cell-free therapy for acute and chronic renal diseases. EVs retain characteristics of the cell of origin and those derived from stem cells may mimic their regenerative properties per se. In fact, EVs contain many active molecules such as proteins and RNA species that act on target cells through different mechanisms, stimulating proliferation and angiogenesis and reducing apoptosis and inflammation. There are several reports that demonstra...

  8. Extracellular deoxyribonuclease production by periodontal bacteria.

    Science.gov (United States)

    Palmer, L J; Chapple, I L C; Wright, H J; Roberts, A; Cooper, P R

    2012-08-01

    Whilst certain bacteria have long been known to secrete extracellular deoxyribonuclease (DNase), the purpose in microbial physiology was unclear. Recently, however, this enzyme has been demonstrated to confer enhanced virulence, enabling bacteria to evade the host's immune defence of extruded DNA/chromatin filaments, termed neutrophil extracellular traps (NETs). As NETs have recently been identified in infected periodontal tissue, the aim of this study was to screen periodontal bacteria for extracellular DNase activity. To determine whether DNase activity was membrane bound or secreted, 34 periodontal bacteria were cultured in broth and on agar plates. Pelleted bacteria and supernatants from broth cultures were analysed for their ability to degrade DNA, with relative activity levels determined using an agarose gel electrophoresis assay. Following culture on DNA-supplemented agar, expression was determined by the presence of a zone of hydrolysis and DNase activity related to colony size. Twenty-seven bacteria, including red and orange complex members Porphyromonas gingivalis, Tannerella forsythia, Fusobacterium nucleatum, Parvimonas micra, Prevotella intermedia, Streptococcus constellatus, Campylobacter rectus and Prevotella nigrescens, were observed to express extracellular DNase activity. Differences in DNase activity were noted, however, when bacteria were assayed in different culture states. Analysis of the activity of secreted DNase from bacterial broth cultures confirmed their ability to degrade NETs. The present study demonstrates, for the first time, that DNase activity is a relatively common property of bacteria associated with advanced periodontal disease. Further work is required to determine the importance of this bacterial DNase activity in the pathogenesis of periodontitis. © 2011 John Wiley & Sons A/S.

  9. Extracellular matrix component signaling in cancer

    DEFF Research Database (Denmark)

    Multhaupt, Hinke A. B.; Leitinger, Birgit; Gullberg, Donald

    2016-01-01

    Cell responses to the extracellular matrix depend on specific signaling events. These are important from early development, through differentiation and tissue homeostasis, immune surveillance, and disease pathogenesis. Signaling not only regulates cell adhesion cytoskeletal organization and motil...... as well as matrix constitution and protein crosslinking. Here we summarize roles of the three major matrix receptor types, with emphasis on how they function in tumor progression. [on SciFinder(R)]...

  10. Development and validation of method for the determination of organochlorine pesticides and trihalomethanes in the water by HRGC-ECD = Desenvolvimento e validação de método para a determinação de pesticidas organoclorados e trihalometanos em água usando HRGC-ECD

    Directory of Open Access Journals (Sweden)

    Maria Isabel Ribeiro Alves

    2010-10-01

    Full Text Available The development and validation of a simultaneous liquid-liquid extraction method for organochlorine pesticides and trihalomethanes in surface and drinking water by HRGC-ECD is described. The method presents acceptable recovery, with detection ranging from 2.7 to 49.0 ng L-1 for organochlorine pesticides and from 18.0 to 860.0 ng L-1for trihalomethanes. The extraction method also presents excellent linearity for all the analytes, with excellent repeatability. Extraction is simple, fast, and low cost, uses small amounts of solvent and aqueous sample, and is suitable for routine analyses.O presente trabalho trata do desenvolvimento e validação de um método de extração para a determinação simultânea de trihalometanos e pesticidas organoclorados em água superficial e água potável por HRGC-ECD. O método apresenta recuperação aceitável, com limites dedetecção que variam de 2,7 a 49,0 ng L-1 para pesticidas organoclorados e de 18,0 a 860,0 ng L-1 para trihalometanos. O método de extração apresenta também excelente linearidade para todos os analitos e boa repetibilidade. A extração é simples, rápida, de baixo custo, além de utilizar pequenas quantidades de solvente e de amostra aquosa, sendo, portanto, de alta aplicabilidade em análises de rotina.

  11. Autocrine signal transmission with extracellular ligand degradation

    International Nuclear Information System (INIS)

    Muratov, C B; Posta, F; Shvartsman, S Y

    2009-01-01

    Traveling waves of cell signaling in epithelial layers orchestrate a number of important processes in developing and adult tissues. These waves can be mediated by positive feedback autocrine loops, a mode of cell signaling where binding of a diffusible extracellular ligand to a cell surface receptor can lead to further ligand release. We formulate and analyze a biophysical model that accounts for ligand-induced ligand release, extracellular ligand diffusion and ligand–receptor interaction. We focus on the case when the main mode for ligand degradation is extracellular and analyze the problem with the sharp threshold positive feedback nonlinearity. We derive expressions that link the speed of propagation and other characteristics of traveling waves to the parameters of the biophysical processes, such as diffusion rates, receptor expression level, etc. Analyzing the derived expressions we found that traveling waves in such systems can exhibit a number of unusual properties, e.g. non-monotonic dependence of the speed of propagation on ligand diffusivity. Our results for the fully developed traveling fronts can be used to analyze wave initiation from localized perturbations, a scenario that frequently arises in the in vitro models of epithelial wound healing, and guide future modeling studies of cell communication in epithelial layers

  12. Extracellular proteases of Trichoderma species. A review.

    Science.gov (United States)

    Kredics, L; Antal, Zsuzsanna; Szekeres, A; Hatvani, L; Manczinger, L; Vágvölgyi, Cs; Nagy, Erzsébet

    2005-01-01

    Cellulolytic, xylanolytic, chitinolytic and beta-1,3-glucanolytic enzyme systems of species belonging to the filamentous fungal genus Trichoderma have been investigated in details and are well characterised. The ability of Trichoderma strains to produce extracellular proteases has also been known for a long time, however, the proteolytic enzyme system is relatively unknown in this genus. Fortunately, in the recent years more and more attention is focused on the research in this field. The role of Trichoderma proteases in the biological control of plant pathogenic fungi and nematodes has been demonstrated, and it is also suspected that they may be important for the competitive saprophytic ability of green mould isolates and may represent potential virulence factors of Trichoderma strains as emerging fungal pathogens of clinical importance. The aim of this review is to summarize the information available about the extracellular proteases of Trichoderma. Numerous studies are available about the extracellular proteolytic enzyme profiles of Trichoderma strains and about the effect of abiotic environmental factors on protease activities. A number of protease enzymes have been purified to homogeneity and some protease encoding genes have been cloned and characterized. These results will be reviewed and the role of Trichoderma proteases in biological control as well as their advantages and disadvantages in biotechnology will be discussed.

  13. EXTRACELLULAR VESICLES: CLASSIFICATION, FUNCTIONS AND CLINICAL RELEVANCE

    Directory of Open Access Journals (Sweden)

    A. V. Oberemko

    2014-12-01

    Full Text Available This review presents a generalized definition of vesicles as bilayer extracellular organelles of all celular forms of life: not only eu-, but also prokaryotic. The structure and composition of extracellular vesicles, history of research, nomenclature, their impact on life processes in health and disease are discussed. Moreover, vesicles may be useful as clinical instruments for biomarkers, and they are promising as biotechnological drug. However, many questions in this area are still unresolved and need to be addressed in the future. The most interesting from the point of view of practical health care represents a direction to study the effect of exosomes and microvesicles in the development and progression of a particular disease, the possibility of adjusting the pathological process by means of extracellular vesicles of a particular type, acting as an active ingredient. Relevant is the further elucidation of the role and importance of exosomes to the surrounding cells, tissues and organs at the molecular level, the prospects for the use of non-cellular vesicles as biomarkers of disease.

  14. Autocrine signal transmission with extracellular ligand degradation

    Science.gov (United States)

    Muratov, C B; Posta, F; Shvartsman, S Y

    2009-03-01

    Traveling waves of cell signaling in epithelial layers orchestrate a number of important processes in developing and adult tissues. These waves can be mediated by positive feedback autocrine loops, a mode of cell signaling where binding of a diffusible extracellular ligand to a cell surface receptor can lead to further ligand release. We formulate and analyze a biophysical model that accounts for ligand-induced ligand release, extracellular ligand diffusion and ligand-receptor interaction. We focus on the case when the main mode for ligand degradation is extracellular and analyze the problem with the sharp threshold positive feedback nonlinearity. We derive expressions that link the speed of propagation and other characteristics of traveling waves to the parameters of the biophysical processes, such as diffusion rates, receptor expression level, etc. Analyzing the derived expressions we found that traveling waves in such systems can exhibit a number of unusual properties, e.g. non-monotonic dependence of the speed of propagation on ligand diffusivity. Our results for the fully developed traveling fronts can be used to analyze wave initiation from localized perturbations, a scenario that frequently arises in the in vitro models of epithelial wound healing, and guide future modeling studies of cell communication in epithelial layers.

  15. Toward a general mixed quantum/classical method for the calculation of the vibronic ECD of a flexible dye molecule with different stable conformers: Revisiting the case of 2,2,2-trifluoro-anthrylethanol.

    Science.gov (United States)

    Cerezo, Javier; Aranda, Daniel; Avila Ferrer, Francisco J; Prampolini, Giacomo; Mazzeo, Giuseppe; Longhi, Giovanna; Abbate, Sergio; Santoro, Fabrizio

    2018-06-01

    We extend a recently proposed mixed quantum/classical method for computing the vibronic electronic circular dichroism (ECD) spectrum of molecules with different conformers, to cases where more than one hindered rotation is present. The method generalizes the standard procedure, based on the simple Boltzmann average of the vibronic spectra of the stable conformers, and includes the contribution of structures that sample all the accessible conformational space. It is applied to the simulation of the ECD spectrum of (S)-2,2,2-trifluoroanthrylethanol, a molecule with easily interconvertible conformers, whose spectrum exhibits a pattern of alternating positive and negative vibronic peaks. Results are in very good agreement with experiment and show that spectra averaged over all the sampled conformational space can deviate significantly from the simple average of the contributions of the stable conformers. The present mixed quantum/classical method is able to capture the effect of the nonlinear dependence of the rotatory strength on the molecular structure and of the anharmonic couplings among the modes responsible for molecular flexibility. Despite its computational cost, the procedure is still affordable and promises to be useful in all cases where the ECD shape arises from a subtle balance between vibronic effects and conformational variety. © 2018 Wiley Periodicals, Inc.

  16. Single Cell Responses to Spatially Controlled Photosensitized Production of Extracellular Singlet Oxygen

    DEFF Research Database (Denmark)

    Pedersen, Brian Wett; Sinks, Louise E.; Breitenbach, Thomas

    2011-01-01

    The response of individual HeLa cells to extracellularly produced singlet oxygen was examined. The spatial domain of singlet oxygen production was controlled using the combination of a membrane-impermeable Pd porphyrin-dendrimer, which served as a photosensitizer, and a focused laser, which served...... to localize the sensitized production of singlet oxygen. Cells in close proximity to the domain of singlet oxygen production showed morphological changes commonly associated with necrotic cell death. The elapsed post-irradiation “waiting period” before necrosis became apparent depended on (a) the distance...... between the cell membrane and the domain irradiated, (b) the incident laser fluence and, as such, the initial concentration of singlet oxygen produced, and (c) the lifetime of singlet oxygen. The data imply that singlet oxygen plays a key role in this process of light-induced cell death. The approach...

  17. EVALUACIÓN DE PLAGUICIDAS 0RGAN0CL0RAD0S POR HS-SPME - GC/ECD EN LECHE PASTEURIZADA COMERCIALIZADA EN LA CIUDAD DE CARTAGENA (COLOMBIA A AVALIAQÁO DOS PESTICIDAS ORGANOCLORADOS POR HS-SPME - GC/ECD EM LEITE PASTEURIZADO COMERCIALIZADAS NA CIDADE CARTAGENA (COLOMBIA EVALUATION OF PESTICIDES BY HS-SPME ORGANOCHLORINE - GC / ECD IN PASTEURIZED MILK MARKETED IN THE CITY CARTAGENA (COLOMBIA

    Directory of Open Access Journals (Sweden)

    DENILES DEL CARMEN DE ARCO R

    2011-12-01

    Full Text Available Los plaguicidas han sido utilizados en todo el mundo desde mediados del siglo XX, en campañas de Salud Pública y en prácticas agrícolas. Los Plaguicidas Organoclorados (POCs presentan mayor impacto sobre el ambiente porque no son biodegradables, son lipofílicos, tienen gran movilidad por todo el planeta y gran tendencia a la bioacumulación, através de la cadena trófica. Con el objetivo de identificar y cuantificar plaguicidas organoclorados en leche entera pasteurizada de dos marcas (A y B, que se comercializan en la ciudad de Cartagena de Indias, se realizó la presente investigación, utilizando Microextracción en Fase Sólida en Espacio de Cabeza y Cromatografía de Gases con Detector de Captura de Electrones (HS-SPME-GC/ECD. Fueron analizadas 36 muestras (n=36, 18 de cada marca. El 100% de las muestras (n=36 resultaron positivas para el plaguicida lindano (γ-HCH, superando el Límite Máximo Residual (LMR de 0,01 mg/kg, establecido por la Food and Agriculture Organization y la Organización Mundial de la Salud (FAO/OMS. Las concentraciones promedio de lindano obtenidas para la marca A fueron de 0,042 mg/kg ± 0,003 y 0,062 ± 0,0016 mg/kg (base grasa para la marca B, no superando la Ingesta Diaria Admisible (IDA de 0,001 mg/kg de peso corporal.Os pesticidas têm sido usados em todo o mundo desde meados do século XX, as campanhas de saúde pública e práticas agrícolas. Os inseticidas organoclorados (AP têm maior impacto sobre o meio ambiente porque não são biodegradáveis, são lipofílicas, têm grande mobilidade ao redor do mundo e maior tendencia para a bioacumulação através da cadeia alimentar. A fim de identificar e quantificar pesticidas organoclorados em leite integral, duas marcas (A e B, comercializada na cidade de Cartagena das Índias, a presente investigação foi realizada por cromatografia em Fase Sólida, Microextração Headspace Gás com Detector de Captura de Elétrons(HS-SPME-GC/ECD. 36 amostras foram

  18. Halogenated pesticide analysis in orange juice by gas chromatography with electron capture detector (GC-ECD) with "6"3Ni nuclide

    International Nuclear Information System (INIS)

    Manhani, Kelly C.; Amaral, Priscila O.; Bustillos, José O.V.; Mendes, Cristiana de A.; Lacerda, João P.A. de; Silva, Jorge L. da; Instituto de Pesquisas Tecnológicas do Estado de São Paulo

    2017-01-01

    Brazil has been gain space in the market of orange juice in the last years. For the exportation of this product to keep growing, its quality of this product must be ensured by putting in force more strict legislations and custom barriers, in order to improve the well-being and health of the population. In this work were analyzed four orange juices brands produced in the State of São Paulo. It was quantified the acaricide known as Dicofol (2,2,2-trichloro-1,1-bis(4-chlorophenyl) ethanol) widely used to combat the Citrusleprosis virus, transmitted by mites to the citrus culture. This pesticide was chosen due to its importance in the production of orange in large scale and their indiscriminate use may pose risks to humans and of environment. The analytical technique applied was gas chromatography coupled with electron capture detector (GC-ECD) using the "6"3Ni nuclide. This beta (β) radiation source ionizes the carrier gas (N_2), generating an electron current that forms the baseline. The analites pass through the detector and capture electrons, generating the analytical signal that is proportional to the concentration of analite. The sample preparation was done by QuEChERS. The limits of detection (LOD) and quantification (LOQ) found were 0.005 and 0.025 mg kg"-"1 respectively. The applied methodology was efficient and presented excellent analytical sensitivity for the pesticide Dicofol, being that of four samples analyzed, only in one was found concentration of 0.03 mg kg"-"1, above the LOQ, however below the Maximum Residue Limit (MRL) for fruits (0.1 mg kg"-"1) established by Codex alimentarius. (author)

  19. Effect of acupuncture on regional cerebral blood flow at acupoints GV 20, GV. 26, LI, 4. ST. 36, SP. 6 evaluated by Tc-99m ECD brain SPECT

    Energy Technology Data Exchange (ETDEWEB)

    Song, Ho Chun; Bom, Hee Seung; Kang, Hwa Jeong; Kim, Seong Min; Jeong, Hwan Jeong; Kim, Ji Yeul [College of Medicine, Dongshin Univ., Naju (Korea, Republic of); Ahn, Soo Gi [College of Medicine, Wonkwang Univ., Iksan (Korea, Republic of)

    2000-12-01

    To evaluate the effect of acupuncture on regional cerebral blood flow (rCBF) at acupoints suggested by oriental medicine to be related to the treatment of cerebrovascular diseases. Rest/acupuncture-stimulation Tc-99m ECD brain SPECT using a same-dose subtraction method was performed on 54 normal volunteers (34 males, 20 females, age range from 18 to 62 years) using six paradigms: acupuncture at acupoints GV. 20, GV. 26, LI. 4, ST. 36 and SP. 6. In the control study, needle location was chosen on a non-meridian focus 1 cm posterior to the right fibular head. All images were spatially normalized, and the differences between rest and acupuncture stimulation were statistically analyzed using SPM for Windows. Acupuncture applied at acupoint GV. 20 increased rCBF in both the anterior frontal lobes, the right frontotemporal lobes, and the left anterior temporal lobe and the left cerebellar hemisphere. Acupuncture at GV. 26 increased rCBF in the left prefrontal cortex. Acupuncture at LI. 4 increased rCBF in the left prefrontal and both the inferior frontal lobes, and the left anterior temporal lobe and the left cerebellar hemisphere. Acupuncture at ST. 36 increased rCBF in the left anterior temporal lobe, the right inferior frontal lobes, and the left cerebellum. Acupuncture at SP 6 increased rCBF in the left inferior frontal and anterior temporal lobes. In the control stimulation, no significant rCBF increase was observed. The results demonstrated a correlation between stimuation at each acupoint with increase in rCBF to the corresponding brain areas.

  20. Effect of acupuncture on regional cerebral blood flow at acupoints GV 20, GV. 26, LI, 4. ST. 36, SP. 6 evaluated by Tc-99m ECD brain SPECT

    International Nuclear Information System (INIS)

    Song, Ho Chun; Bom, Hee Seung; Kang, Hwa Jeong; Kim, Seong Min; Jeong, Hwan Jeong; Kim, Ji Yeul; Ahn, Soo Gi

    2000-01-01

    To evaluate the effect of acupuncture on regional cerebral blood flow (rCBF) at acupoints suggested by oriental medicine to be related to the treatment of cerebrovascular diseases. Rest/acupuncture-stimulation Tc-99m ECD brain SPECT using a same-dose subtraction method was performed on 54 normal volunteers (34 males, 20 females, age range from 18 to 62 years) using six paradigms: acupuncture at acupoints GV. 20, GV. 26, LI. 4, ST. 36 and SP. 6. In the control study, needle location was chosen on a non-meridian focus 1 cm posterior to the right fibular head. All images were spatially normalized, and the differences between rest and acupuncture stimulation were statistically analyzed using SPM for Windows. Acupuncture applied at acupoint GV. 20 increased rCBF in both the anterior frontal lobes, the right frontotemporal lobes, and the left anterior temporal lobe and the left cerebellar hemisphere. Acupuncture at GV. 26 increased rCBF in the left prefrontal cortex. Acupuncture at LI. 4 increased rCBF in the left prefrontal and both the inferior frontal lobes, and the left anterior temporal lobe and the left cerebellar hemisphere. Acupuncture at ST. 36 increased rCBF in the left anterior temporal lobe, the right inferior frontal lobes, and the left cerebellum. Acupuncture at SP 6 increased rCBF in the left inferior frontal and anterior temporal lobes. In the control stimulation, no significant rCBF increase was observed. The results demonstrated a correlation between stimuation at each acupoint with increase in rCBF to the corresponding brain areas

  1. Halogenated pesticide analysis in orange juice by gas chromatography with electron capture detector (GC-ECD) with {sup 63}Ni nuclide

    Energy Technology Data Exchange (ETDEWEB)

    Manhani, Kelly C.; Amaral, Priscila O.; Bustillos, José O.V.; Mendes, Cristiana de A.; Lacerda, João P.A. de; Silva, Jorge L. da, E-mail: ovega@ipen.br, E-mail: kmanhani@gmail.com [Instituto de Pesquisas Energeticas e Nucleares (CQMA/IPEN-CNEN-SP), Sao Paulo, SP (Brazil). Centro de Química e Meio Ambiente; Instituto de Pesquisas Tecnológicas do Estado de São Paulo (LAQ- CQuim/IPT-SP), SP (Brazil). Lab. de Análises Químicas

    2017-11-01

    Brazil has been gain space in the market of orange juice in the last years. For the exportation of this product to keep growing, its quality of this product must be ensured by putting in force more strict legislations and custom barriers, in order to improve the well-being and health of the population. In this work were analyzed four orange juices brands produced in the State of São Paulo. It was quantified the acaricide known as Dicofol (2,2,2-trichloro-1,1-bis(4-chlorophenyl) ethanol) widely used to combat the Citrusleprosis virus, transmitted by mites to the citrus culture. This pesticide was chosen due to its importance in the production of orange in large scale and their indiscriminate use may pose risks to humans and of environment. The analytical technique applied was gas chromatography coupled with electron capture detector (GC-ECD) using the {sup 63}Ni nuclide. This beta (β) radiation source ionizes the carrier gas (N{sub 2}), generating an electron current that forms the baseline. The analites pass through the detector and capture electrons, generating the analytical signal that is proportional to the concentration of analite. The sample preparation was done by QuEChERS. The limits of detection (LOD) and quantification (LOQ) found were 0.005 and 0.025 mg kg{sup -1} respectively. The applied methodology was efficient and presented excellent analytical sensitivity for the pesticide Dicofol, being that of four samples analyzed, only in one was found concentration of 0.03 mg kg{sup -1}, above the LOQ, however below the Maximum Residue Limit (MRL) for fruits (0.1 mg kg{sup -1}) established by Codex alimentarius. (author)

  2. Brain perfusion abnormalities associated to drug abuse in recent abstinent patients using SPECT 99m Tc-ethylen-cysteinate-dimer (ECD)

    Energy Technology Data Exchange (ETDEWEB)

    Massardo, Teresa [University of Chile Clinical Hospital Nuclear Medicine Section, Department of Medicine, Santiago (Chile); Pallavicini, Julio [Addiction Unit, Psychiatric Clinic. University of Chile Clinical Hospital (Chile); Gonzalez, Patricio; Jaimovich, Rodrigo [University of Chile Clinical Hospital Nuclear Medicine Section, Department of Medicine, Santiago (Chile); Servat, Monica [Addiction Unit, Psychiatric Clinic. University of Chile Clinical Hospital (Chile); Lavados, Hugo [University of Chile Clinical Hospital Nuclear Medicine Section, Department of Medicine, Santiago (Chile); Arancibia, Pablo [Addiction Unit, Psychiatric Clinic. University of Chile Clinical Hospital (Chile); Padilla, Pamela [University of Chile Clinical Hospital Nuclear Medicine Section, Department of Medicine, Santiago (Chile)

    2009-04-15

    Several substances may produce brain perfusion abnormalities in drug-dependent patients. Their mechanism is unclear and several causes might be involved, especially vasospasm in cocaine consumption. Goal: To characterize residual brain perfusion abnormalities in substance-dependent population. We analyzed brain perfusion in 100 dependant patients (DSM-IV criteria) following a month of strict in-hospital abstinence (age:35{+-}12 y.o.; 86% men); 55% corresponded to poly-drug dependents, mainly to cocaine, alcohol and cannabis; 44% mono-drug users, mostly to alcohol. Results: Single Photon Emission Computed Tomography (SPECT) with 99mTc-ethylen-cysteinate-dimer (ECD) was abnormal in 54% of the cases, with bilateral cortical hypo-perfusion in 89%, focal in 54% and diffuse in 46% of them, with moderate or severe intensity in 61%. The abnormal perfusion group's age was 38{+-}12 versus 31{+-}10 years in the normal SPECT group (P=0.005) with a consumption period of 16{+-}11 versus 11{+-}8 years, respectively (P=0.043). Only 29% of women had abnormal perfusion versus 58% of men (P=0.047). Abnormal brain perfusion in 64% of mono and 45% in poly-drug dependents (P=0.07). Psychometric tests performed in 25 patients demonstrated association between perfusion defects and cognitive abnormalities. Relative risk for abnormal psychometric test was 2.5 [95%;CI=1.1-5.6] for abnormal SPECT. Conclusion: Dependent population after a month of abstinence persists with cortical brain perfusion abnormalities, associated to age, sex and type of drug consumption.

  3. Compromised Cerebral Blood Flow(CBF) in Congestive Heart Failure (CHB): non-invasive quantification with {sup 99m}Tc-ECD radionuclide angiography

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Jae Seung; Kim, Jae Joong; Lim, Ki Chun; Lee, Hee Kyung; Moon, Dae Hyuk [Ulsan University College of Medicine, Seoul (Korea, Republic of)

    2002-07-01

    Recent reports revealed that cerebral metabolism in CHF was abnormally deranged and proposed as a potential marker of disease severity. Since deranged cerebral metabolism in CHF may result from compromised cerebral perfusion, quantification of CHF may be useful for accurate risk stratification of CHF. Therefore, we investigated whether CHF in patients with CHF is compromised and correlated with clinical parameters. Fifteen patients (M/F:11/5, 45{+-}9yr) with CHF (LVEF<40%) and 7 healthy controls (M/F:5/2, 41{+-}8yr) were prospectively studied. All patients underwent radionuclide angiography including cerebral hemispheres and aortic arch using {sup 99m}Tc-ECD. Global CBF was measured non-invasively by the application of Patlak graphical plot analysis. All patients were also evaluated using a standardized protocol that included echocardiography and clinical evaluation. Global CBF (40.3{+-}5.2 ml/min/100g) of the patients with CHF were significantly lower than those (49.7{+-}2.4 ml/min/100g) of controls (p<0.01). Global CBF were correlated with NYHA functional class (r=-0.617, p=0.43), but not correlated with other clinical parameters such as age (r=-0.463, p=0.082), duration (r=0.237, p>0.1), systolic BP (r=-0.063, p>0.5), LVEF (r=-0.13, p>0.1), LV dimension(r=0.139, p>0.5), and PV pressure gradients (r=0.072, p>0.5). Cerebral perfusion of the patients with CHF was compromised and not correlated with cardiopulmonary hemodynamic parameters.

  4. Visual and SPM analysis of regional cerebral perfusion with Tc-99m ECD brain SPECT in patients with developmental language disorder

    International Nuclear Information System (INIS)

    Yoon, Joon Kee; Lee, Myung Hoon; Joh, Chul Woo; Yoon, Seok Nam; Oh, Eun Young

    2003-01-01

    Developmental language disorder (DLD) refers to inadequate language acquisition at the expected age in children with otherwise normal development. However, language delay can be observed in patients with other developmental disoder (ODD). We, therefore, evaluated regional cerebral perfusion pattern in patients with DLD and ODD by means of visual and SPM analysis. Twelve patients, who underwent Tc-99m ECD brain SPECT within 3 weeks of their first visit, were included in the study. Psychological and language tests classified the patients into 2 groups ; 6 with DLD (3-7 yr, 5 male and I female) and 6 with ODD (2-6 yr, 6 male). Visual analysis for regional cerebral perfusion was done in each patient. SPM with 7 controls (age=7) was performed to evaluate difference between 2 groups using t-test. P value of less than 0.005 was considered to be significant. All patients had significant language delay for their age (9 month 3.5 yr). Among 6 patients with ODD, 4 had pervasive developmental disorder, 1 mental retardation and 1 attachment disorder. Visual analysis revealed significant perfusion decrease in only 1 patient with DLD and 2 with ODD ; the regions were left parieto-temporal cortex, both frontal and cerebellar cortices, and right temporal cortex respectively. Nine of 12 patients showed normal perfusion. SPM demonstrated perfusion decrease in left inferior frontal cortex and left superior parietal cortex (Wernicke's area) in patients with DLD, while, in patients with ODD, perfusion decrease was mostly located in the right hemisphere (lateral frontoorbital gyrus, occipitotemporal gyrus, cuneus and cerebellum). Corpus callosum showed no significant perfusion abnormality in both groups. Regional cerebral perfusion of patients with DLD, which was mainly located in the speech area, is quite different from that of ODD-patients with language delay. While SPM successfully revealed this difference in perfusion pattern, visual analysis had limited value

  5. Changes in regional cerebral blood flow correlate with symptoms of depression and severity of fatigue in 99mTc-ECD SPECT study in 45 workers

    International Nuclear Information System (INIS)

    Koyama, Fumihiko; Hojo, Kei; Otsuki, Kenro; Yamamoto, Haruyoshi

    2008-01-01

    We performed single photon emission computed tomography ( 99m Tc-ECD SPECT) of the brain in 25 workers suffering from major depression (22 males and 3 females at the average age of 47.5±7.7 standard deviation (S.D.) years and all patients were right handed) and identified frontal lobe hypoperfusion in 18 cases using the easy Z-score Imaging System (eZIS). During the remission period, which was within ten months of therapy, we observed normalization of the frontal cerebral perfusion in 75% of the patients. In further evaluation of the cerebral blood flow after the addition of 20 healthy volunteer workers (18 males and 2 females at the average age 47.1±9.8 S.D. years and all were right handed) as a control group, reduced cerebral blood flow in the dorsal area of the frontal lobe, which is supposed to be correlated to the severity of the feeling of fatigue estimated based on the self-rating depression scale (SDS), was demonstrated by statistical parametric mapping (SPM). The severity of the fatigue of all workers was estimated using a self-rating checklist disseminated by the Japan Ministry of Health, Labor, and Welfare, which pertains to the accumulation of fatigue due to labor. In the SPM analysis, limbic hypoperfusion, which is correlated to the total severity of fatigue in all worker cases, was estimated based on the reduction in the cerebral blood flow in the lower part of the temporal lobe. Accordingly, the accumulation of fatigue due to, for instance, over work may result in easy fatigue and a pre-depressive state, which are associated with a reduction in the cerebral blood flow. This research is a part of the research and development and the dissemination projects related to the 13 fields of occupational injuries and illnesses of the Japan Labor Health and Welfare Organization. (author)

  6. Tc-99m ECD brain SPECT in patients with traumatic brain injury: evaluating distribution of hypoperfusion and assessment of cognitive and behavioral impairment in relation to thalamic hypoperfusion

    Energy Technology Data Exchange (ETDEWEB)

    Park, Soon Ah; Lim, Seok Tae; Sohn, Myung Hee [College of Medicine, Chonbuk National Univ., Chonju (Korea, Republic of)

    2000-12-01

    We evaluated the distribution of hypoperfusion in patients with traumatic brain injury (TBI) and the relationship of thalamic hypoperfusion to severity of cognitive and behavioral sequelae. Tc-99m ECD SPECT and MRI were performed in 103 patients (M/F=81/22, mean age 34.7{+-} 15.4 yrs) from 0.5 to 55 months (mean 10.3 months) after TBI. The patients were divided into three groups showing no abnormalities (G1), focal (G2) and diffuse injury (G3) on MRI. Psychometric tests assessed 11 cognitive or behavioral items. In all patients, we evaluated the distribution of hypoperfused areas in SPECT, and in 57/103 patients, neuropsychological (NP) abnormalities in patients with thalamic hypoperfusion were compared with those of patients without thalamic hypoperfusion. The perfusion dificits were most frequently located in the frontal lobe (G1, 42.3%: G2 34.5%: G3 33.3%), temporal lobe (24{approx}26%) thalami (21{approx}22.4%), parietal and occipital lobe ({<=}10%). Numbers of NP abnormalities in the cases of cortical hypoperfusion with or without concomitant thalamic hypoperfusion were following: the former 4.7{+-}1.5 and the latter 3.2{+-}1.4 in G1, 5.0{+-}1.1 and 4.8{+-}1.2 in G2, 6.8{+-}1.8 and 6.3{+-}1.1 in G3, respectively. This difference according to thalamic hypoperfusion was significant in G1 (p=0.002), but was not significant in G2 or G3. SPECT in patients with TBI had demonstrated hypoperfusion mostly involving the frontal, temporal and thalami. In normal group on MRI, frontal hypoperfusion was more prominent than that of any other group, Furthermore in this group, SPECT could predict severity of NP outcome by concomitant thalamic hypoperfusion with cerebral cortical abnormalities.

  7. Tc-99m ECD brain SPECT in patients with traumatic brain injury: evaluating distribution of hypoperfusion and assessment of cognitive and behavioral impairment in relation to thalamic hypoperfusion

    International Nuclear Information System (INIS)

    Park, Soon Ah; Lim, Seok Tae; Sohn, Myung Hee

    2000-01-01

    We evaluated the distribution of hypoperfusion in patients with traumatic brain injury (TBI) and the relationship of thalamic hypoperfusion to severity of cognitive and behavioral sequelae. Tc-99m ECD SPECT and MRI were performed in 103 patients (M/F=81/22, mean age 34.7± 15.4 yrs) from 0.5 to 55 months (mean 10.3 months) after TBI. The patients were divided into three groups showing no abnormalities (G1), focal (G2) and diffuse injury (G3) on MRI. Psychometric tests assessed 11 cognitive or behavioral items. In all patients, we evaluated the distribution of hypoperfused areas in SPECT, and in 57/103 patients, neuropsychological (NP) abnormalities in patients with thalamic hypoperfusion were compared with those of patients without thalamic hypoperfusion. The perfusion dificits were most frequently located in the frontal lobe (G1, 42.3%: G2 34.5%: G3 33.3%), temporal lobe (24∼26%) thalami (21∼22.4%), parietal and occipital lobe (≤10%). Numbers of NP abnormalities in the cases of cortical hypoperfusion with or without concomitant thalamic hypoperfusion were following: the former 4.7±1.5 and the latter 3.2±1.4 in G1, 5.0±1.1 and 4.8±1.2 in G2, 6.8±1.8 and 6.3±1.1 in G3, respectively. This difference according to thalamic hypoperfusion was significant in G1 (p=0.002), but was not significant in G2 or G3. SPECT in patients with TBI had demonstrated hypoperfusion mostly involving the frontal, temporal and thalami. In normal group on MRI, frontal hypoperfusion was more prominent than that of any other group, Furthermore in this group, SPECT could predict severity of NP outcome by concomitant thalamic hypoperfusion with cerebral cortical abnormalities

  8. Evaluation of therapeutic effects of cognitive-behavioral therapy in patients with panic disorder using serial 99mTc-ECD brain perfusion

    International Nuclear Information System (INIS)

    Kim, Jung Hee; Song, Ho Chun; Yang, Jong Chul; Lee, Byeong Il; Heo, Young Jun; Bom, Hee Seung; Min, Jung Joon; Park, Tae Jin

    2006-01-01

    Although several neuroanatomical models of panic disorder have been proposed, little is known regarding the neurological mechanisms underlying cognitive-behavioral therapy (CBT) in patients with panic disorder. This study was performed to identify the brain structures that show changes of regional cerebral blood flow (rCBF) after CBT in patients with panic disorder. Seven patients who were diagnosed as panic disorder by DSM-IV were treated with CBT for 8 weeks and twelve healthy volunteers joined in this study. Serial 99m Tc-ECD brain perfusion SPECT images were aquisited and PDSS-SR (Self-Report version of Panic Disorder Severity Scale) and ACQ (Agoraphobic Cognitive Question) scores were measured just before and after CBT in all patients. Data were analyzed using SPM2. Subjective symptoms were improved, and PDSS-SR and ACQ scores were significantly reduced (14.9 ± 3.9 vs. 7.0 ± 1.8, ρ < 0.05; 30.3 ± 8.5 vs. 21.6 ± 3.4, ρ < 0.05, respectively) after CBT in panic patients. Before CBT, a significant increase of rCBF was found in the cingulate gylus, thalamus, midbrain, both medial frontal and temporal lobes of the panic patients compared to the normal volunteers. After CBT, we observed a significant rCBF decrease in the left parahippocamus, right insula and cingulate gyrus, both frontal and temporal lobes, and a significant rCBF increase in both the occipital lobes, left insula, both frontal and left parietal lobes. These data suggested that CBT is effective for panic disorder and diminish the activity of the brain areas associated with fear in panic disorder

  9. Evaluation of therapeutic effects of cognitive-behavioral therapy in patients with panic disorder using serial {sup 99m}Tc-ECD brain perfusion

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Jung Hee; Song, Ho Chun; Yang, Jong Chul; Lee, Byeong Il; Heo, Young Jun; Bom, Hee Seung; Min, Jung Joon [Chonnam National University Hospital, Gwangju (Korea, Republic of); Park, Tae Jin [Chonnam National University, Gwangju (Korea, Republic of)

    2006-12-15

    Although several neuroanatomical models of panic disorder have been proposed, little is known regarding the neurological mechanisms underlying cognitive-behavioral therapy (CBT) in patients with panic disorder. This study was performed to identify the brain structures that show changes of regional cerebral blood flow (rCBF) after CBT in patients with panic disorder. Seven patients who were diagnosed as panic disorder by DSM-IV were treated with CBT for 8 weeks and twelve healthy volunteers joined in this study. Serial {sup 99m}Tc-ECD brain perfusion SPECT images were aquisited and PDSS-SR (Self-Report version of Panic Disorder Severity Scale) and ACQ (Agoraphobic Cognitive Question) scores were measured just before and after CBT in all patients. Data were analyzed using SPM2. Subjective symptoms were improved, and PDSS-SR and ACQ scores were significantly reduced (14.9 {+-} 3.9 vs. 7.0 {+-} 1.8, {rho} < 0.05; 30.3 {+-} 8.5 vs. 21.6 {+-} 3.4, {rho} < 0.05, respectively) after CBT in panic patients. Before CBT, a significant increase of rCBF was found in the cingulate gylus, thalamus, midbrain, both medial frontal and temporal lobes of the panic patients compared to the normal volunteers. After CBT, we observed a significant rCBF decrease in the left parahippocamus, right insula and cingulate gyrus, both frontal and temporal lobes, and a significant rCBF increase in both the occipital lobes, left insula, both frontal and left parietal lobes. These data suggested that CBT is effective for panic disorder and diminish the activity of the brain areas associated with fear in panic disorder.

  10. Validation of QuEChERS analytical technique for organochlorines and synthetic pyrethroids in fruits and vegetables using GC-ECD.

    Science.gov (United States)

    Dubey, J K; Patyal, S K; Sharma, Ajay

    2018-03-19

    In the present day scenario of increasing awareness and concern about the pesticides, it is very important to ensure the quality of data being generated in pesticide residue analysis. To impart confidence in the products, terms like quality assurance and quality control are used as an integral part of quality management. In order to ensure better quality of results in pesticide residue analysis, validation of analytical methods to be used is extremely important. Keeping in view the importance of validation of method, the validation of QuEChERS (quick, easy, cheap, effective, rugged, and safe) a multiresidue method for extraction of 13 organochlorines and seven synthetic pyrethroids in fruits and vegetables followed by GC ECD for quantification was done so as to use this method for analysis of samples received in the laboratory. The method has been validated as per the Guidelines issued by SANCO (French words Sante for Health and Consommateurs for Consumers) in accordance with their document SANCO/XXXX/2013. Various parameters analyzed, viz., linearity, specificity, repeatability, reproducibility, and ruggedness were found to have acceptable values with a per cent RSD of less than 10%. Limit of quantification (LOQ) for the organochlorines was established to be 0.01 and 0.05 mg kg -1 for the synthetic pyrethroids. The uncertainty of the measurement (MU) for all these compounds ranged between 1 and 10%. The matrix-match calibration was used to compensate the matrix effect on the quantification of the compounds. The overall recovery of the method ranged between 80 and 120%. These results demonstrate the applicability and acceptability of this method in routine estimation of pesticide residues of these 20 pesticides in the fruits and vegetables by the laboratory.

  11. Visual and SPM analysis of regional cerebral perfusion with Tc-99m ECD brain SPECT in patients with developmental language disorder

    Energy Technology Data Exchange (ETDEWEB)

    Yoon, Joon Kee; Lee, Myung Hoon; Joh, Chul Woo; Yoon, Seok Nam; Oh, Eun Young [College of Medicine, Univ. of Ajou, Suwon (Korea, Republic of)

    2003-07-01

    Developmental language disorder (DLD) refers to inadequate language acquisition at the expected age in children with otherwise normal development. However, language delay can be observed in patients with other developmental disoder (ODD). We, therefore, evaluated regional cerebral perfusion pattern in patients with DLD and ODD by means of visual and SPM analysis. Twelve patients, who underwent Tc-99m ECD brain SPECT within 3 weeks of their first visit, were included in the study. Psychological and language tests classified the patients into 2 groups ; 6 with DLD (3-7 yr, 5 male and I female) and 6 with ODD (2-6 yr, 6 male). Visual analysis for regional cerebral perfusion was done in each patient. SPM with 7 controls (age=7) was performed to evaluate difference between 2 groups using t-test. P value of less than 0.005 was considered to be significant. All patients had significant language delay for their age (9 month 3.5 yr). Among 6 patients with ODD, 4 had pervasive developmental disorder, 1 mental retardation and 1 attachment disorder. Visual analysis revealed significant perfusion decrease in only 1 patient with DLD and 2 with ODD ; the regions were left parieto-temporal cortex, both frontal and cerebellar cortices, and right temporal cortex respectively. Nine of 12 patients showed normal perfusion. SPM demonstrated perfusion decrease in left inferior frontal cortex and left superior parietal cortex (Wernicke's area) in patients with DLD, while, in patients with ODD, perfusion decrease was mostly located in the right hemisphere (lateral frontoorbital gyrus, occipitotemporal gyrus, cuneus and cerebellum). Corpus callosum showed no significant perfusion abnormality in both groups. Regional cerebral perfusion of patients with DLD, which was mainly located in the speech area, is quite different from that of ODD-patients with language delay. While SPM successfully revealed this difference in perfusion pattern, visual analysis had limited value.

  12. The extracellular domain of Lrp5/6 inhibits noncanonical Wnt signaling in vivo

    Czech Academy of Sciences Publication Activity Database

    Bryja, Vítězslav; Andersson, E.R.; Schambony, A.; Esner, M.; Bryjová, Lenka; Biris, K.K.; Hall, A.C.; Kraft, B.; Čajánek, L.; Yamaguchi, T.P.; Buckingham, M.; Arenas, E.

    2009-01-01

    Roč. 20, č. 3 (2009), s. 924-936 ISSN 1059-1524 Institutional research plan: CEZ:AV0Z50040507; CEZ:AV0Z50040702 Keywords : non-canonical Wnt signaling * Lrp5/6 * mouse Subject RIV: BO - Biophysics Impact factor: 5.979, year: 2009

  13. Binding of N-methylscopolamine to the extracellular domain of muscarinic acetylcholine receptors

    Czech Academy of Sciences Publication Activity Database

    Jakubík, Jan; Randáková, Alena; Zimčík, Pavel; El-Fakahany, E. E.; Doležal, Vladimír

    2017-01-01

    Roč. 7, Jan 16 (2017), č. článku 40381. ISSN 2045-2322 R&D Projects: GA ČR(CZ) GBP304/12/G069 Institutional support: RVO:67985823 Keywords : muscarinic acetylcholine receptors * N-methylscopolamine * ligand binding * molecular dynamics Subject RIV: ED - Physiology OBOR OECD: Physiology (including cytology) Impact factor: 4.259, year: 2016

  14. Role of an Absolutely Conserved Tryptophan Pair in the Extracellular Domain of Cys-Loop Receptors

    DEFF Research Database (Denmark)

    Braun, Nina; Lynagh, Timothy; Yu, Rilei

    2016-01-01

    Cys-loop receptors mediate fast synaptic transmission in the nervous system, and their dysfunction is associated with a number of diseases. While some sequence variability is essential to ensure specific recognition of a chemically diverse set of ligands, other parts of the underlying amino acid...... sequences show a high degree of conservation, possibly to preserve the overall structural fold across the protein family. In this study, we focus on the only two absolutely conserved residues across the Cys-loop receptor family, two Trp side chains in the WXD motif of Loop D and in the WXPD motif of Loop A...

  15. Extracellular Signatures as Indicators of Processing Methods

    Energy Technology Data Exchange (ETDEWEB)

    Wahl, Karen L.

    2012-01-09

    As described in other chapters within this volume, many aspects of microbial cells vary with culture conditions and therefore can potentially be analyzed as forensic signatures of growth conditions. In addition to changes or variations in components of the microbes themselves, extracellular materials indicative of production processes may remain associated with the final bacterial product. It is well recognized that even with considerable effort to make pure products such as fine chemicals or pharmaceuticals, trace impurities from components or synthesis steps associated with production processes can be detected in the final product. These impurities can be used as indicators of production source or methods, such as to help connect drugs of abuse to supply chains. Extracellular residue associated with microbial cells could similarly help to characterize production processes. For successful growth of microorganisms on culture media there must be an available source of carbon, nitrogen, inorganic phosphate and sulfur, trace metals, water and vitamins. The pH, temperature, and a supply of oxygen or other gases must also be appropriate for a given organism for successful culture. The sources of these components and the range in temperature, pH and other variables has adapted over the years with currently a wide range of possible combinations of media components, recipes and parameters to choose from for a given organism. Because of this wide variability in components, mixtures of components, and other parameters, there is the potential for differentiation of cultured organisms based on changes in culture conditions. The challenge remains how to narrow the field of potential combinations and be able to attribute variations in the final bacterial product and extracellular signatures associated with the final product to information about the culture conditions or recipe used in the production of that product.

  16. Light Regimes Shape Utilization of Extracellular Organic C and N in a Cyanobacterial Biofilm

    Energy Technology Data Exchange (ETDEWEB)

    Stuart, Rhona K.; Mayali, Xavier; Boaro, Amy A.; Zemla, Adam; Everroad, R. Craig; Nilson, Daniel; Weber, Peter K.; Lipton, Mary; Bebout, Brad M.; Pett-Ridge, Jennifer; Thelen, Michael P.

    2016-06-28

    Although it is becoming clear that many microbial primary producers can also play a role as organic consumers, we know very little about the metabolic regulation of photoautotroph organic matter consumption. Cyanobacteria in phototrophic biofilms can reuse extracellular organic carbon, but the metabolic drivers of extracellular processes are surprisingly complex. We investigated the metabolic foundations of organic matter reuse by comparing exoproteome composition and incorporation of13C-labeled and15N-labeled cyanobacterial extracellular organic matter (EOM) in a unicyanobacterial biofilm incubated using different light regimes. In the light and the dark, cyanobacterial direct organic C assimilation accounted for 32% and 43%, respectively, of all organic C assimilation in the community. Under photosynthesis conditions, we measured increased excretion of extracellular polymeric substances (EPS) and proteins involved in micronutrient transport, suggesting that requirements for micronutrients may drive EOM assimilation during daylight hours. This interpretation was supported by photosynthesis inhibition experiments, in which cyanobacteria incorporated N-rich EOM-derived material. In contrast, under dark, C-starved conditions, cyanobacteria incorporated C-rich EOM-derived organic matter, decreased excretion of EPS, and showed an increased abundance of degradative exoproteins, demonstrating the use of the extracellular domain for C storage. Sequence-structure modeling of one of these exoproteins predicted a specific hydrolytic activity that was subsequently detected, confirming increased EOM degradation in the dark. Associated heterotrophic bacteria increased in abundance and upregulated transport proteins under dark relative to light conditions. Taken together, our results indicate that biofilm cyanobacteria are successful competitors for organic C and N and that cyanobacterial nutrient and energy requirements control the use of EOM.

  17. Nanostructured gold microelectrodes for extracellular recording

    Energy Technology Data Exchange (ETDEWEB)

    Brueggemann, Dorothea; Wolfrum, Bernhard; Maybeck, Vanessa; Offenhaeusser, Andreas [CNI Center of Nanoelectronic Systems for Information Technology and Institute of Bio- and Nanosystems 2, Forschungszentrum Juelich (Germany)

    2010-07-01

    Electrophysiological activity of electrogenic cells is currently recorded with planar bioelectronic interfaces such as microelectrode arrays (MEAs). In this work, a novel concept of biocompatible nanostructured gold MEAs for extracellular signal recording is presented. MEAs were fabricated using clean room technologies, e.g. photolithography and metallization. Subsequently, they were modified with gold nanopillars of approximately 300 to 400 nm in height and 60 nm width. The nanostructuring process was carried out with a template-assisted approach using nanoporous aluminium oxide. Impedance spectroscopy of the resulting nanostructures showed higher capacitances compared to planar gold. This confirmed the expected increase of the surface area via nanostructuring. We used the nanostructured microelectrodes to record extracellular potentials from heart muscle cells (HL1), which were plated onto the chips. Good coupling between the HL1 cells and the nanostructured electrodes was observed. The resulting signal-to-noise ratio of nanopillar-MEAs was increased by a factor of 2 compared to planar MEAs. In future applications this nanopillar concept can be adopted for distinct interface materials and coupling to cellular and molecular sensing components.

  18. Secretory proteins of the pulmonary extracellular lining

    International Nuclear Information System (INIS)

    Gupta, R.P.; Patton, S.E.; Eddy, M.; Smits, H.L.; Jetten, A.M.; Nettesheim, P.; Hook, G.E.R.

    1986-01-01

    The objective of this investigation was to identify proteins in the pulmonary extracellular lining (EL) that are secreted by cells of the pulmonary epithelium. Pulmonary lavage effluents from the lungs of rabbits were centrifuged to remove all cells and particulate materials. Serum proteins were removed by repeatedly passing concentrated lavage effluent fluid through an affinity column containing IgG fraction of goat anti-rabbit (whole serum) antiserum bound to Sepharose-4B. Nonserum proteins accounted for 21.3 +/- 10.3% of the total soluble proteins in pulmonary lavage effluents. Serum free lavage effluents (SFL) contained 25 identifiable proteins as determined by using SDS-PAGE under reducing conditions. Of these proteins approximately 73% was accounted for by a single protein with MW of 66 kd. The secretory nature of the proteins present in SFL was investigated by studying the incorporation of 35 S-methionine into proteins released by lung slices and trachea followed by SDS-PAGE and autoradiography. Many, but not all proteins present in SFL were identified as proteins secreted by pulmonary tissues. The major secretory proteins appeared to have MWs of 59, 53, 48, 43, 24, 14, and 6 kd under reducing conditions. These data demonstrate the presence of several proteins in the pulmonary extracellular lining that appear to be secreted by the pulmonary epithelium

  19. Extracellular histones induce erythrocyte fragility and anemia.

    Science.gov (United States)

    Kordbacheh, Farzaneh; O'Meara, Connor H; Coupland, Lucy A; Lelliott, Patrick M; Parish, Christopher R

    2017-12-28

    Extracellular histones have been shown to play an important pathogenic role in many diseases, primarily through their cytotoxicity toward nucleated cells and their ability to promote platelet activation with resultant thrombosis and thrombocytopenia. In contrast, little is known about the effect of extracellular histones on erythrocyte function. We demonstrate in this study that histones promote erythrocyte aggregation, sedimentation, and using a novel in vitro shear stress model, we show that histones induce erythrocyte fragility and lysis in a concentration-dependent manner. Furthermore, histones impair erythrocyte deformability based on reduced passage of erythrocytes through an artificial spleen. These in vitro results were mirrored in vivo with the injection of histones inducing anemia within minutes of administration, with a concomitant increase in splenic hemoglobin content. Thrombocytopenia and leukopenia were also observed. These findings suggest that histones binding to erythrocytes may contribute to the elevated erythrocyte sedimentation rates observed in inflammatory conditions. Furthermore, histone-induced increases in red blood cell lysis and splenic clearance may be a significant factor in the unexplained anemias seen in critically ill patients. © 2017 by The American Society of Hematology.

  20. The International Society for Extracellular Vesicles launches the first massive open online course on extracellular vesicles.

    Science.gov (United States)

    Lässer, Cecilia; Théry, Clotilde; Buzás, Edit I; Mathivanan, Suresh; Zhao, Weian; Gho, Yong Song; Lötvall, Jan

    2016-01-01

    The International Society for Extracellular Vesicles (ISEV) has organised its first educational online course for students and beginners in the field of extracellular vesicles (EVs). This course, "Basics of Extracellular Vesicles," uses recorded lectures from experts in the field and will be open for an unlimited number of participants. The course is divided into 5 modules and can be accessed at www.coursera.org/learn/extracellular-vesicles. The first module is an introduction to the field covering the nomenclature and history of EVs. Module 2 focuses on the biogenesis and uptake mechanisms of EVs, as well as their RNA, protein and lipid cargo. Module 3 covers the collection and processing of cell culture media and body fluids such as blood, breast milk, cerebrospinal fluid and urine prior to isolation of EVs. Modules 4 and 5 present different isolation methods and characterisation techniques utilised in the EV field. Here, differential ultracentrifugation, size-exclusion chromatography, density gradient centrifugation, kit-based precipitation, electron microscopy, cryo-electron microscopy, flow cytometry, atomic-force microscopy and nanoparticle-tracking analysis are covered. This first massive open online course (MOOC) on EVs was launched on 15 August 2016 at the platform "Coursera" and is free of charge.

  1. Extracellular matrix in canine mammary tumors with special focus on versican, a versatile extracellular proteoglycan

    NARCIS (Netherlands)

    Erdélyi, Ildikó

    2006-01-01

    The extracellular matrix (ECM) research has become fundamental to understand cancer. This thesis focuses on the exploration of ECM composition and organization in canine mammary tumors, with a special interest in the large chondroitin-sulfate proteoglycan (PG), versican. Chapter 1 gives an

  2. The International Society for Extracellular Vesicles launches the first massive open online course on extracellular vesicles

    Directory of Open Access Journals (Sweden)

    Cecilia Lässer

    2016-12-01

    Full Text Available The International Society for Extracellular Vesicles (ISEV has organised its first educational online course for students and beginners in the field of extracellular vesicles (EVs. This course, “Basics of Extracellular Vesicles,” uses recorded lectures from experts in the field and will be open for an unlimited number of participants. The course is divided into 5 modules and can be accessed at www.coursera.org/learn/extracellular-vesicles. The first module is an introduction to the field covering the nomenclature and history of EVs. Module 2 focuses on the biogenesis and uptake mechanisms of EVs, as well as their RNA, protein and lipid cargo. Module 3 covers the collection and processing of cell culture media and body fluids such as blood, breast milk, cerebrospinal fluid and urine prior to isolation of EVs. Modules 4 and 5 present different isolation methods and characterisation techniques utilised in the EV field. Here, differential ultracentrifugation, size-exclusion chromatography, density gradient centrifugation, kit-based precipitation, electron microscopy, cryo-electron microscopy, flow cytometry, atomic-force microscopy and nanoparticle-tracking analysis are covered. This first massive open online course (MOOC on EVs was launched on 15 August 2016 at the platform “Coursera” and is free of charge.

  3. The Extracellular Matrix of Candida albicans Biofilms Impairs Formation of Neutrophil Extracellular Traps.

    Science.gov (United States)

    Johnson, Chad J; Cabezas-Olcoz, Jonathan; Kernien, John F; Wang, Steven X; Beebe, David J; Huttenlocher, Anna; Ansari, Hamayail; Nett, Jeniel E

    2016-09-01

    Neutrophils release extracellular traps (NETs) in response to planktonic C. albicans. These complexes composed of DNA, histones, and proteins inhibit Candida growth and dissemination. Considering the resilience of Candida biofilms to host defenses, we examined the neutrophil response to C. albicans during biofilm growth. In contrast to planktonic C. albicans, biofilms triggered negligible release of NETs. Time lapse imaging confirmed the impairment in NET release and revealed neutrophils adhering to hyphae and migrating on the biofilm. NET inhibition depended on an intact extracellular biofilm matrix as physical or genetic disruption of this component resulted in NET release. Biofilm inhibition of NETosis could not be overcome by protein kinase C activation via phorbol myristate acetate (PMA) and was associated with suppression of neutrophil reactive oxygen species (ROS) production. The degree of impaired NET release correlated with resistance to neutrophil attack. The clinical relevance of the role for extracellular matrix in diminishing NET production was corroborated in vivo using a rat catheter model. The C. albicans pmr1Δ/Δ, defective in production of matrix mannan, appeared to elicit a greater abundance of NETs by scanning electron microscopy imaging, which correlated with a decreased fungal burden. Together, these findings show that C. albicans biofilms impair neutrophil response through an inhibitory pathway induced by the extracellular matrix.

  4. Multiple Roles of the Extracellular Vestibule Amino Acid Residues in the Function of the Rat P2X4 Receptor

    Czech Academy of Sciences Publication Activity Database

    Rokic, Milos Boro; Stojilkovic, S. S.; Vávra, Vojtěch; Kuzyk, Pavlo; Tvrdoňová, Vendula; Zemková, Hana

    2013-01-01

    Roč. 8, č. 3 (2013), e59411 E-ISSN 1932-6203 R&D Projects: GA AV ČR(CZ) IAA500110910; GA ČR(CZ) GBP304/12/G069 Institutional research plan: CEZ:AV0Z50110509 Institutional support: RVO:67985823 Keywords : ATP * purinergic P2X receptor channels * transmembrane domain * extracellular vestibule * gating * ivermectin Subject RIV: ED - Physiology Impact factor: 3.534, year: 2013

  5. Supersymmetric domain walls

    NARCIS (Netherlands)

    Bergshoeff, Eric A.; Kleinschmidt, Axel; Riccioni, Fabio

    2012-01-01

    We classify the half-supersymmetric "domain walls," i.e., branes of codimension one, in toroidally compactified IIA/IIB string theory and show to which gauged supergravity theory each of these domain walls belong. We use as input the requirement of supersymmetric Wess-Zumino terms, the properties of

  6. Extracellular RNAs: development as biomarkers of human disease

    Directory of Open Access Journals (Sweden)

    Joseph F. Quinn

    2015-08-01

    Full Text Available Ten ongoing studies designed to test the possibility that extracellular RNAs may serve as biomarkers in human disease are described. These studies, funded by the NIH Common Fund Extracellular RNA Communication Program, examine diverse extracellular body fluids, including plasma, serum, urine and cerebrospinal fluid. The disorders studied include hepatic and gastric cancer, cardiovascular disease, chronic kidney disease, neurodegenerative disease, brain tumours, intracranial haemorrhage, multiple sclerosis and placental disorders. Progress to date and the plans for future studies are outlined.

  7. Proteolytic processing of lysyl oxidase-like-2 in the extracellular matrix is required for crosslinking of basement membrane collagen IV.

    Science.gov (United States)

    López-Jiménez, Alberto J; Basak, Trayambak; Vanacore, Roberto M

    2017-10-13

    Lysyl oxidase-like-2 (LOXL2) is an enzyme secreted into the extracellular matrix that crosslinks collagens by mediating oxidative deamination of lysine residues. Our previous work demonstrated that this enzyme crosslinks the 7S domain, a structural domain that stabilizes collagen IV scaffolds in the basement membrane. Despite its relevant role in extracellular matrix biosynthesis, little is known about the structural requirements of LOXL2 that enable collagen IV crosslinking. In this study, we demonstrate that LOXL2 is processed extracellularly by serine proteases, generating a 65-kDa form lacking the first two scavenger receptor cysteine-rich domains. Site-specific mutagenesis to prevent proteolytic processing generated a full-length enzyme that is active in vitro toward a soluble substrate, but fails to crosslink insoluble collagen IV within the extracellular matrix. In contrast, the processed form of LOXL2 binds to collagen IV and crosslinks the 7S domain. Together, our data demonstrate that proteolytic processing is an important event that allows LOXL2-mediated crosslinking of basement membrane collagen IV. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  8. A Novel Molecular Diagnostic of Glioblastomas: Detection of an Extracellular Fragment of Protein Tyrosine Phosphatase μ

    Directory of Open Access Journals (Sweden)

    Susan M. Burden-Gulley

    2010-04-01

    Full Text Available We recently found that normal human brain and low-grade astrocytomas express the receptor protein tyrosine phosphatase mu (PTPμ and that the more invasive astrocytomas, glioblastoma multiforme (GBM, downregulate full-length PTPμ expression. Loss of PTPμ expression in GBMs is due to proteolytic cleavage that generates an intracellular and potentially a cleaved and released extracellular fragment of PTPμ. Here, we identify that a cleaved extracellular fragment containing the domains required for PTPμ-mediated adhesion remains associated with GBM tumor tissue. We hypothesized that detection of this fragment would make an excellent diagnostic tool for the localization of tumor tissue within the brain. To this end, we generated a series of fluorescently tagged peptide probes that bind the PTPμ fragment. The peptide probes specifically recognize GBM cells in tissue sections of surgically resected human tumors. To test whether the peptide probes are able to detect GBM tumors in vivo, the PTPμ peptide probes were tested in both mouse flank and intracranial xenograft human glioblastoma tumor model systems. The glial tumors were molecularly labeled with the PTPμ peptide probes within minutes of tail vein injection using the Maestro FLEX In Vivo Imaging System. The label was stable for at least 3 hours. Together, these results indicate that peptide recognition of the PTPμ extracellular fragment provides a novel molecular diagnostic tool for detection of human glioblastomas. Such a tool has clear translational applications and may lead to improved surgical resections and prognosis for patients with this devastating disease.

  9. Collagen VI disorders: Insights on form and function in the extracellular matrix and beyond.

    Science.gov (United States)

    Lamandé, Shireen R; Bateman, John F

    2017-12-22

    Mutations in the three canonical collagen VI genes, COL6A1, COL6A2 and COL6A3, cause a spectrum of muscle disease from Bethlem myopathy at the mild end to the severe Ullrich congenital muscular dystrophy. Mutations can be either dominant or recessive and the resulting clinical severity is influenced by the way mutations impact the complex collagen VI assembly process. Most mutations are found towards the N-terminus of the triple helical collagenous domain and compromise extracellular microfibril assembly. Outside the triple helix collagen VI is highly polymorphic and discriminating mutations from rare benign changes remains a major diagnostic challenge. Collagen VI deficiency alters extracellular matrix structure and biomechanical properties and leads to increased apoptosis and oxidative stress, decreased autophagy, and impaired muscle regeneration. Therapies that target these downstream consequences have been tested in a collagen VI null mouse and also in small human trials where they show modest clinical efficacy. An important role for collagen VI in obesity, cancer and diabetes is emerging. A major barrier to developing effective therapies is the paucity of information about how collagen VI deficiency in the extracellular matrix signals the final downstream consequences - the receptors involved and the intracellular messengers await further characterization. Copyright © 2017 International Society of Matrix Biology. Published by Elsevier B.V. All rights reserved.

  10. The change of cerebral blood flow after heart transplantation in congestive heart failure: a voxel-based and automatic VOl analysis of Tc-99m ECD SPECT

    Energy Technology Data Exchange (ETDEWEB)

    Hong, I. K.; Kim, J. J.; Lee, C. H.; Lim, K. C.; Moon, D. H.; Rhu, J. S.; Kim, J. S. [Asan Medical Center, Seoul (Korea, Republic of)

    2007-07-01

    To investigate the change of global and regional cerebral blood flow after heart transplantation (HT) in congestive heart failure (CHF) patients. Twenty-one patients with CHF who underwent HT (45{+-}12 yrs, M/F=19/2) and 10 healthy volunteers (39{+-}13 yrs, M/F = 7/3) were prospectively included. All patients underwent echocardiography and radionuclide angiography including brain and aorta with brain SPECT which was performed after iv bolus injection of Tc-99m ECD (740MBq) before (175{+-}253 days) and after (129{+-}82 days) HT. Patients were divided into two groups according to the interval between HT and postoperative SPECT [early follow-up (f/u): <6 mo, n=14; late f/u: >6 mo, n=7]. Global CBF (gCBF) of bilateral hemispheres were calculated by Patlak graphical analysis. Absolute rCBF map was obtained from brain SPECT by Lassen's correction algorithm. Age-corrected voxel-based analysis using SPM2 and automatic VOl analysis were performed to assess the rCBF change. Cardiac ejection fraction of all patients improved after HT (20.8%{yields}64.0%). gCBF was reduced compared to normal before HT (35.7{+-}3.9 vs. 49.1{+-}3.0 ml/100g/min; p<0.001) and improved postoperatively (46.6{+-}5.4, p<0.001). The preoperative gCBFs of early and late f/u group were not different (34.6{+-}3.2 vs. 38.0{+-}4.4, p=0.149) but postoperative gCBF (43.9{+-}3.7) of late f/u group was higher than those (52.0{+-}4.0) of early f/u group (p<0.001). On voxel-based analysis, preoperative rCBF was reduced in entire brain but most severely in bilateral superior and inferior frontal cortex, supplementary motor area, precuneus and anterior cingulum, compared to normals (uncorrected p<0.001). After HT, rCBF of these areas improved more significantly in late f/u group than in early f/u group but still lower than normals. Global CBF was significantly reduced in CHF patients and improved after HT. rCBFs of the frontal cortex, precuneus and cingulum were most severely reduced and slowly improved after

  11. Regional cerebral blood flow in children with autism spectrum disorders: a quantitative 99mTc-ECD brain SPECT study with statistical parametric mapping evaluation

    Institute of Scientific and Technical Information of China (English)

    YANG Wen-han; JING Jin; XIU Li-juan; CHENG Mu-hua; WANG Xin; BAO Peng; WANG Qing-xiong

    2011-01-01

    Background Autism spectrum disorders (ASD), which include autism, asperger syndrome (AS) and pervasive developmental disorder-not otherwise specified (PDD-NOS), are devastating neurodevelopmental disorders of childhood resulting in deficits in social interaction, repetitive patterns of behaviors, and restricted interests and activities. Single photon emission computed tomography (SPECT) is a common technique used to measure regional cerebral blood flow (rCBF). Several studies have measured rCBF in children with ASD using SPECT, however, findings are discordant. In addition, the majority of subjects used in these studies were autistic. In this study, we aimed to investigate changes in rCBF in children with ASD using SPECT.Methods A Technetium-99m-ethyl cysteinate dimmer (99mTc-ECD) brain SPECT study was performed on an ASD group consisting of 23 children (3 girls and 20 boys; mean age (7.2±3.0) years) who were diagnosed according to Diagnostic and Statistical Manual of Mental Disorders, 4th edition (DSM-Ⅳ) criteria and an age-matched control group with 8children (1 girl and 7 boys, mean age (5.5±2.4) years). Image data were evaluated with Statistical Parametric Mapping,5th version (SPM5). A Student's t test for unpaired data was used to compare rCBF and asymmetry in the autism and corresponding control group. The covariance analysis, taking age as covariance, was performed between the ASD and control group.Results There was a significant reduction in rCBF in the bilateral frontal lobe (frontal poles, arcula frontal gyrus) and the bilateral basal ganglia in the autism group, and a reduction in the bilateral frontal, temporal, parietal, legumina nucleus and cerebellum in the AS group compared to the control. In addition, asymmetry of hemispheric hypoperfusion in the ASD group was observed. Inner-group comparison analysis revealed that rCBF decreased significantly in the bilateral frontal lobe (42.7%), basal nucleus (24.9%) and temporal lobe (22.8%) in the autism

  12. Changes in regional cerebral blood flow correlate with symptoms of depression, severity of fatigue, and sleep disorders in 99mTc-ECD SPECT study in 45 workers

    International Nuclear Information System (INIS)

    Koyama, Fumihiko; Matsuura, Naoyuki; Kageyama, Junichi; Otsuki, Kenro

    2010-01-01

    We performed single photon emission computed tomography ( 99m Tc-ECD SPECT) of the brain in 25 workers suffering from major depression (22 males and 3 females at the average age of 47.5±7.7 standard deviation (S.D.) years and all patients were right handed) and identified frontal lobe hypoperfusion in 18 cases using the easy Z-score Imaging System (eZIS). During the remission period, which was within ten months of therapy, we observed normalization of the frontal cerebral perfusion in 75% of the patients. In further evaluation of the cerebral blood flow after the addition of 20 healthy volunteer workers (18 males and 2 females at the average age 47.1±9.8 S.D. years and all were right handed) as a control group, reduced cerebral blood flow in the dorsal area, which is supposed to be correlated to the severity of the feeling of fatigue estimated based on the self-rating depression scale (SDS), was demonstrated by statistical parametric mapping (SPM). The severity of the fatigue of all workers was estimated using a self-rating checklist disseminated by the Japan Ministry of Health, Labor, and Welfare, which pertains to the accumulation of fatigue due to labor. In the SPM analysis, limbic hypoperfusion, which is correlated to the total severity of fatigue in all worker cases, was estimated based on the reduction in the cerebral blood flow in the lower part of the temporal lobe. We also estimated the severity of insomnia in 21 cases (8 depressive, the 8 remitted and 5 controls), referred to as the Insomnia Score in this study, using the Structured Interview Guide for the Hamilton Depression Rating Scale (SIGH-D). The severity of the insomnia statistically correlates with frontoparietal hypoperfusion, and this region is situated very close to the region correlated to the feeling of fatigue expressed by the patients in the SDS. Accordingly, prolonged sleep disturbance due to, for instance, overwork may result in easy fatigability and a pre-depressive state which are

  13. A perspective on extracellular vesicles proteomics

    Science.gov (United States)

    Rosa-Fernandes, Livia; Rocha, Victória Bombarda; Carregari, Victor Corasolla; Urbani, Andrea; Palmisano, Giuseppe

    2017-11-01

    Increasing attention has been given to secreted extracellular vesicles (EVs) in the past decades, especially in the portrayal of their molecular cargo and role as messengers in both homeostasis and pathophysiological conditions. This review presents the state-of-the-art proteomic technologies to identify and quantify EVs proteins along with their PTMs, interacting partners and structural details. The rapid growth of mass spectrometry-based analytical strategies for protein sequencing, PTMs and structural characterization has improved the level of molecular details that can be achieve from limited amount of EVs isolated from different biological sources. Here we will provide a perspective view on the achievements and challenges on EVs proteome characterization using mass spectrometry. A detailed bioinformatics approach will help us to picture the molecular fingerprint of EVs and understand better their pathophysiological function.

  14. Why regenerative medicine needs an extracellular matrix.

    Science.gov (United States)

    Prestwich, Glenn D; Healy, Kevin E

    2015-01-01

    Regenerative medicine is now coming of age. Many attempts at cell therapy have failed to show significant efficacy, and the umbrella term 'stem cell therapy' is perceived in some quarters as hype or just expensive and unnecessary medical tourism. Here we present a short editorial in three parts. First, we examine the importance of using a semisynthetic extracellular matrix (ECM) mimetic, or sECM, to deliver and retain therapeutic cells at the site of administration. Second, we describe one approach in which biophysical and biochemical properties are tailored to each tissue type, which we call "design for optimal functionality." Third, we describe an alternative approach to sECM design and implementation, called "design for simplicity," in which a deconstructed, minimalist sECM is employed and biology is allowed to perform the customization in situ. We opine that an sECM, whether minimal or instructive, is an essential contributor to improve the outcomes of cell-based therapies.

  15. Extracellular matrix fluctuations during early embryogenesis

    International Nuclear Information System (INIS)

    Szabó, A; Rupp, P A; Rongish, B J; Little, C D; Czirók, A

    2011-01-01

    Extracellular matrix (ECM) movements and rearrangements were studied in avian embryos during early stages of development. We show that the ECM moves as a composite material, whereby distinct molecular components as well as spatially separated layers exhibit similar displacements. Using scanning wide field and confocal microscopy we show that the velocity field of ECM displacement is smooth in space and that ECM movements are correlated even at locations separated by several hundred micrometers. Velocity vectors, however, strongly fluctuate in time. The autocorrelation time of the velocity fluctuations is less than a minute. Suppression of the fluctuations yields a persistent movement pattern that is shared among embryos at equivalent stages of development. The high resolution of the velocity fields allows a detailed spatio-temporal characterization of important morphogenetic processes, especially tissue dynamics surrounding the embryonic organizer (Hensen's node)

  16. Extracellular Matrix Molecules Facilitating Vascular Biointegration

    Directory of Open Access Journals (Sweden)

    Martin K.C. Ng

    2012-08-01

    Full Text Available All vascular implants, including stents, heart valves and graft materials exhibit suboptimal biocompatibility that significantly reduces their clinical efficacy. A range of biomolecules in the subendothelial space have been shown to play critical roles in local regulation of thrombosis, endothelial growth and smooth muscle cell proliferation, making these attractive candidates for modulation of vascular device biointegration. However, classically used biomaterial coatings, such as fibronectin and laminin, modulate only one of these components; enhancing endothelial cell attachment, but also activating platelets and triggering thrombosis. This review examines a subset of extracellular matrix molecules that have demonstrated multi-faceted vascular compatibility and accordingly are promising candidates to improve the biointegration of vascular biomaterials.

  17. Identification of a receptor for extracellular renalase.

    Directory of Open Access Journals (Sweden)

    Ling Wang

    Full Text Available An increased risk for developing essential hypertension, stroke and diabetes is associated with single nucleotide gene polymorphisms in renalase, a newly described secreted flavoprotein with oxidoreductase activity. Gene deletion causes hypertension, and aggravates acute ischemic kidney (AKI and cardiac injury. Independent of its intrinsic enzymatic activities, extracellular renalase activates MAPK signaling and prevents acute kidney injury (AKI in wild type (WT mice. Therefore, we sought to identity the receptor for extracellular renalase.RP-220 is a previously identified, 20 amino acids long renalase peptide that is devoid of any intrinsic enzymatic activity, but it is equally effective as full-length recombinant renalase at protecting against toxic and ischemic injury. Using biotin transfer studies with RP-220 in the human proximal tubular cell line HK-2 and protein identification by mass spectrometry, we identified PMCA4b as a renalase binding protein. This previously characterized plasma membrane ATPase is involved in cell signaling and cardiac hypertrophy. Co-immunoprecipitation and co-immunolocalization confirmed protein-protein interaction between endogenous renalase and PMCA4b. Down-regulation of endogenous PMCA4b expression by siRNA transfection, or inhibition of its enzymatic activity by the specific peptide inhibitor caloxin1b each abrogated RP-220 dependent MAPK signaling and cytoprotection. In control studies, these maneuvers had no effect on epidermal growth factor mediated signaling, confirming specificity of the interaction between PMCA4b and renalase.PMCA4b functions as a renalase receptor, and a key mediator of renalase dependent MAPK signaling.

  18. Incorporation of Tenascin-C into the Extracellular Matrix by Periostin Underlies an Extracellular Meshwork Architecture*

    OpenAIRE

    Kii, Isao; Nishiyama, Takashi; Li, Minqi; Matsumoto, Ken-ichi; Saito, Mitsuru; Amizuka, Norio; Kudo, Akira

    2009-01-01

    Extracellular matrix (ECM) underlies a complicated multicellular architecture that is subjected to significant forces from mechanical environment. Although various components of the ECM have been enumerated, mechanisms that evolve the sophisticated ECM architecture remain to be addressed. Here we show that periostin, a matricellular protein, promotes incorporation of tenascin-C into the ECM and organizes a meshwork architecture of the ECM. We found that both periostin null mice and tenascin-C...

  19. Spatial and frequency domain ring source models for the single muscle fiber action potential

    DEFF Research Database (Denmark)

    Henneberg, Kaj-åge; R., Plonsey

    1994-01-01

    In the paper, single-fibre models for the extracellular action potential are developed that will allow the potential to the evaluated at an arbitrary field point in the extracellular space. Fourier-domain models are restricted in that they evaluate potentials at equidistant points along a line...... parallel to the fibre axis. Consequently, they cannot easily evaluate the potential at the boundary nodes of a boundary-element electrode model. The Fourier-domain models employ axial-symmetric ring source models, and thereby provide higher accuracy that the line source model, where the source is lumped...... including anisotropy show that the spatial models require extreme care in the integration procedure owing to the singularity in the weighting functions. With adequate sampling, the spatial models can evaluate extracellular potentials with high accuracy....

  20. The crystal structures of EAP domains from Staphylococcus aureus reveal an unexpected homology to bacterial superantigens.

    Science.gov (United States)

    Geisbrecht, Brian V; Hamaoka, Brent Y; Perman, Benjamin; Zemla, Adam; Leahy, Daniel J

    2005-04-29

    The Eap (extracellular adherence protein) of Staphylococcus aureus functions as a secreted virulence factor by mediating interactions between the bacterial cell surface and several extracellular host proteins. Eap proteins from different Staphylococcal strains consist of four to six tandem repeats of a structurally uncharacterized domain (EAP domain). We have determined the three-dimensional structures of three different EAP domains to 1.8, 2.2, and 1.35 A resolution, respectively. These structures reveal a core fold that is comprised of an alpha-helix lying diagonally across a five-stranded, mixed beta-sheet. Comparison of EAP domains with known structures reveals an unexpected homology with the C-terminal domain of bacterial superantigens. Examination of the structure of the superantigen SEC2 bound to the beta-chain of a T-cell receptor suggests a possible ligand-binding site within the EAP domain (Fields, B. A., Malchiodi, E. L., Li, H., Ysern, X., Stauffacher, C. V., Schlievert, P. M., Karjalainen, K., and Mariuzza, R. (1996) Nature 384, 188-192). These results provide the first structural characterization of EAP domains, relate EAP domains to a large class of bacterial toxins, and will guide the design of future experiments to analyze EAP domain structure/function relationships.

  1. Conserved Domain Database (CDD)

    Data.gov (United States)

    U.S. Department of Health & Human Services — CDD is a protein annotation resource that consists of a collection of well-annotated multiple sequence alignment models for ancient domains and full-length proteins.

  2. The extracellular proteome of Rhizobium etli CE3 in exponential and stationary growth phase

    Directory of Open Access Journals (Sweden)

    Mendoza-Hernández Guillermo

    2010-10-01

    Full Text Available Abstract Background The extracellular proteome or secretome of symbiotic bacteria like Rhizobium etli is presumed to be a key element of their infection strategy and survival. Rhizobia infect the roots of leguminous plants and establish a mutually beneficial symbiosis. To find out the possible role of secreted proteins we analyzed the extracellular proteome of R. etli CE3 in the exponential and stationary growth phases in minimal medium, supplemented with succinate-ammonium. Results The extracellular proteins were obtained by phenol extraction and identified by LC-ESI MS/MS. We identified 192 and 191 proteins for the exponential and stationary phases respectively. Using the software Signal P, we predicted signal peptides for 12.95% and 35.60% of the proteins identified in the exponential and stationary phases, respectively, which could therefore be secreted by the Sec pathway. For the exponential growth phase, we found in abundance proteins like the ribosomal proteins, toxins and proteins belonging to the group "defence mechanisms". For the stationary growth phase, we found that the most abundant proteins were those with unknown function, and in many of these we identified characteristic domains of proteases and peptidases. Conclusions Our study provided the first dataset of the secretome of R. etli and its modifications, which may lead to novel insights into the adaptive response of different stages of growth. In addition, we found a high number of proteins with unknown function; these proteins could be analyzed in future research to elucidate their role in the extracellular proteome of R. etli.

  3. Neutrophil extracellular traps - the dark side of neutrophils

    DEFF Research Database (Denmark)

    Sørensen, Ole E.; Borregaard, Niels

    2016-01-01

    Neutrophil extracellular traps (NETs) were discovered as extracellular strands of decondensed DNA in complex with histones and granule proteins, which were expelled from dying neutrophils to ensnare and kill microbes. NETs are formed during infection in vivo by mechanisms different from those ori...

  4. Production of extracellular laccase from the newly isolated Bacillus ...

    African Journals Online (AJOL)

    This study was carried out with aim of screening for extracellular thermostable laccase producing bacteria. Twenty-two (22) laccase positive strains were isolated from the selected environmental samples while extracellular laccase activity was detected only in six strains namely TM1, TMT1, PK4, PS1, TMS1 and ASP3.

  5. Molecular Logic of Neuronal Self-Recognition through Protocadherin Domain Interactions

    DEFF Research Database (Denmark)

    Rubinstein, Rotem; Thu, Chan Aye; Goodman, Kerry Marie

    2015-01-01

    Self-avoidance, a process preventing interactions of axons and dendrites from the same neuron during development, is mediated in vertebrates through the stochastic single-neuron expression of clustered protocadherin protein isoforms. Extracellular cadherin (EC) domains mediate isoform-specific ho...

  6. The ligand-binding domain of the cell surface receptor for urokinase-type plasminogen activator

    DEFF Research Database (Denmark)

    Behrendt, N; Ploug, M; Patthy, L

    1991-01-01

    with the internal repeats of u-PAR constitute the extracellular part of Ly-6 antigens and of the squid glycoprotein Sgp-2. Like u-PAR, these proteins are attached to the membrane by a glycosyl-phosphatidylinositol anchor. The hydrophilic, ligand-binding u-PAR domain identified in the present study has potential...

  7. The subunit structure of the extracellular hemoglobin of Biomphalaria glabrata

    International Nuclear Information System (INIS)

    Arndt, Marcio H.L.; Naves, Cristiani F.; Xavier, Luciana P.; Santoro, Marcelo M.

    1997-01-01

    Full text. The hemoglobin of Biomphalaria glabrata was purified to homogeneity by a two step purification protocol using a gel filtration column (Superose 6 HR/Pharmacia ) followed by an anion exchange chromatography (MONO-Q Sepharose/Pharmacia). The dissociation products were analysed by a 5 - 15 % Polyacrylamide gel electrophoresis containing Sodium Dodecyl Sulfate (SDS-PAGE) giving a band of 270 K Daltons and a band of 180 K Daltons after reduction with β-mercaptoethanol. The same profile was obtained in a 3.5 % Agarose gel electrophoresis containing SDS (SDS-AGE) showing additional bands of higher molecular weight. These bands were proposed to be monomers, dimers and trimers and, after reduction in a Bidimensional SDS-AGE, the proposed monomers and dimers were decomposed in two and four bands that were interpreted as 1 - 4 chains. The hemoglobin was digested by four different proteases ( Thrombin, Trypsin, Chymotrypsin and Subtilisin ) showing several equivalent fragments with molecular weights multiples of its minimum molecular weight ( 17.7 K Daltons). The circular dichroism spectrum of the protein showed a characteristic high α-helix content. We proposed that this hemoglobin is a pentamer of approx. 360 K Daltons subunits each formed by two 180 K Daltons chains linked in pairs by disulfide bridges and each of these chains comprises ten Heme binding domains. These data were compared to other Planorbidae extracellular hemoglobins. Up to now, the quaternary structure of this hemoglobin (shape and disposition of the subunits) is unknown. It is intended to elucidate its structure by Small Angle X-Ray Scattering in Brazilian National Laboratory of Synchrotron Light (LNLS). (author)

  8. Structure and function of ameloblastin as an extracellular matrix protein: adhesion, calcium binding, and CD63 interaction in human and mouse.

    Science.gov (United States)

    Zhang, Xu; Diekwisch, Thomas G H; Luan, Xianghong

    2011-12-01

    The functional significance of extracellular matrix proteins in the life of vertebrates is underscored by a high level of sequence variability in tandem with a substantial degree of conservation in terms of cell-cell and cell-matrix adhesion interactions. Many extracellular matrix proteins feature multiple adhesion domains for successful attachment to substrates, such as integrin, CD63, and heparin. Here we have used homology and ab initio modeling algorithms to compare mouse ameloblastin (mAMBN) and human ameloblastin (hABMN) isoforms and to analyze their potential for cell adhesion and interaction with other matrix molecules as well as calcium binding. Sequence comparison between mAMBN and hAMBN revealed a 26-amino-acid deletion in mAMBN, corresponding to a helix-loop-helix frameshift. The human AMBN domain (174Q-201G), homologous to the mAMBN 157E-178I helix-loop-helix region, formed a helix-loop motif with an extended loop, suggesting a higher degree of flexibility of hAMBN compared with mAMBN, as confirmed by molecular dynamics simulation. Heparin-binding domains, CD63-interaction domains, and calcium-binding sites in both hAMBN and mAMBN support the concept of AMBN as an extracellular matrix protein. The high level of conservation between AMBN functional domains related to adhesion and differentiation was remarkable when compared with only 61% amino acid sequence homology. © 2011 Eur J Oral Sci.

  9. Evaluation and investigation of regional cerebral blood flow by 1 point arterial blood collection method using 99mTc-ECD. Intravenous injection for 4 minutes with constant speed

    International Nuclear Information System (INIS)

    Itoh, Takeo; Shibata, Kazuhiro; Sudoh, Hideaki; Tanaka, Masato; Itoh, Kenjiro; Ueno, Yasushi

    1998-01-01

    Regional cerebral blood flow (rCBF) was measured using a 99m Tc-ECD through the 4-min constant intravenous infusion/one point arterial blood sampling method, proposed by Nakagawara et al. of Nakamura Memorial Hospital, and 133Xenon ( 133 Xe)-SPECT was performed on the same subjects to investigate the reproducibility of this method. We also determined whether cerebral blood flow (CBF) could be measured on the day of blood sampling through dilution of the obtained blood because it was difficult to measure the radioactivity in the blood on the day of blood sampling by this method. More, we investigated fixation of an octanol extraction rate and the substitution of venous blood for arterial blood in this method. The results revealed that CBF measured by this method with a 99m Tc-ECD were closely correlated to those measured by 133 Xe-SPECT, indicating the reliability as a method of measuring CBF. rCBF could be measured on the day of blood sampling through appropriate dilution of the obtained arterial blood. Octanol extraction rates were almost constant, indicating possible omission of cumbersome extraction procedure by fixation. However, the substitution of venous blood for arterial blood showed no correlation under the study system examined. (author)

  10. Studies on asymptomatic cerebral ischemia in coronary heart disease with special reference to evaluation of postural changes in cerebral blood flow with {sup 99m}Tc-ECD brain SPECT

    Energy Technology Data Exchange (ETDEWEB)

    Sinozaki, Hideko [Teikyo Univ., Tokyo (Japan). Faculty of Medicine

    2001-03-01

    Postural changes in cerebral blood flow in patients with coronary heart disease (CHD) were evaluated with SPECT to detect asymptomaic cerebral ischemia (ACI). {sup 99m}Tc-ECD was used as a tracer. We developed a new analysis system for the processing of multiple images, making it possible to avoid the spatial shift in ROIs in different positions. The severity of ACI was classified into 3 groups based on SPECT findings in the supine position: group 1 without any abnormalities in cerebral perfusion, group 2 with a single perfusion defect, and group 3 with a number of perfusion defects. No cerebral siteobserved in any group showed a significant difference between the supine and upright positions in cerebral perfusion. Each group in each position, however, revealed a consistent perfusion pattern characterized by a significant decrease in perfusion in the occipital, temporal and frontal lobes. Moreover, the decrease in the latter two sites was significantly greater than in the former site. Concerning clinical profiles, hypertension and the thickness of the intima and media complex (TIMC) of the common carotid artery significantly correlated with the severity of ACI. Furthermore, in multiple regression analysis, only TIMC was identified as a significant determinant of ACI. In conclusion, cerebral blood flow determined with {sup 99m}Tc-ECD SPECT could accurately detect ACI in patients with CHD. (author)

  11. The anchorless adhesin Eap (extracellular adherence protein) from Staphylococcus aureus selectively recognizes extracellular matrix aggregates but binds promiscuously to monomeric matrix macromolecules.

    Science.gov (United States)

    Hansen, Uwe; Hussain, Muzaffar; Villone, Daniela; Herrmann, Mathias; Robenek, Horst; Peters, Georg; Sinha, Bhanu; Bruckner, Peter

    2006-05-01

    Besides a number of cell wall-anchored adhesins, the majority of Staphylococcus aureus strains produce anchorless, cell wall-associated proteins, such as Eap (extracellular adherence protein). Eap contains four to six tandem repeat (EAP)-domains. Eap mediates diverse biological functions, including adherence and immunomodulation, thus contributing to S. aureus pathogenesis. Eap binding to host macromolecules is unusually promiscuous and includes matrix or matricellular proteins as well as plasma proteins. The structural basis of this promiscuity is poorly understood. Here, we show that in spite of the preferential location of the binding epitopes within triple helical regions in some collagens there is a striking specificity of Eap binding to different collagen types. Collagen I, but not collagen II, is a binding substrate in monomolecular form. However, collagen I is virtually unrecognized by Eap when incorporated into banded fibrils. By contrast, microfibrils containing collagen VI as well as basement membrane-associated networks containing collagen IV, or aggregates containing fibronectin bound Eap as effectively as the monomeric proteins. Therefore, Eap-binding to extracellular matrix ligands is promiscuous at the molecular level but not indiscriminate with respect to supramolecular structures containing the same macromolecules. In addition, Eap bound to banded fibrils after their partial disintegration by matrix-degrading proteinases, including matrix metalloproteinase 1. Therefore, adherence to matrix suprastructures by S. aureus can be supported by inflammatory reactions.

  12. Extracellular matrix dynamics during vertebrate axis formation.

    Science.gov (United States)

    Czirók, András; Rongish, Brenda J; Little, Charles D

    2004-04-01

    The first evidence for the dynamics of in vivo extracellular matrix (ECM) pattern formation during embryogenesis is presented below. Fibrillin 2 filaments were tracked for 12 h throughout the avian intraembryonic mesoderm using automated light microscopy and algorithms of our design. The data show that these ECM filaments have a reproducible morphogenic destiny that is characterized by directed transport. Fibrillin 2 particles initially deposited in the segmental plate mesoderm are translocated along an unexpected trajectory where they eventually polymerize into an intricate scaffold of cables parallel to the anterior-posterior axis. The cables coalesce near the midline before the appearance of the next-formed somite. Moreover, the ECM filaments define global tissue movements with high precision because the filaments act as passive motion tracers. Quantification of individual and collective filament "behaviors" establish fate maps, trajectories, and velocities. These data reveal a caudally propagating traveling wave pattern in the morphogenetic movements of early axis formation. We conjecture that within vertebrate embryos, long-range mechanical tension fields are coupled to both large-scale patterning and local organization of the ECM. Thus, physical forces or stress fields are essential requirements for executing an emergent developmental pattern-in this case, paraxial fibrillin cable assembly.

  13. Extracellular ice phase transitions in insects.

    Science.gov (United States)

    Hawes, T C

    2014-01-01

    At temperatures below their temperature of crystallization (Tc), the extracellular body fluids of insects undergo a phase transition from liquid to solid. Insects that survive the transition to equilibrium (complete freezing of the body fluids) are designated as freeze tolerant. Although this phenomenon has been reported and described in many Insecta, current nomenclature and theory does not clearly delineate between the process of transition (freezing) and the final solid phase itself (the frozen state). Thus freeze tolerant insects are currently, by convention, described in terms of the temperature at which the crystallization of their body fluids is initiated, Tc. In fact, the correct descriptor for insects that tolerate freezing is the temperature of equilibrium freezing, Tef. The process of freezing is itself a separate physical event with unique physiological stresses that are associated with ice growth. Correspondingly there are a number of insects whose physiological cryo-limits are very specifically delineated by this transitional envelope. The distinction also has considerable significance for our understanding of insect cryobiology: firstly, because the ability to manage endogenous ice growth is a fundamental segregator of cryotype; and secondly, because our understanding of internal ice management is still largely nascent.

  14. Towards traceable size determination of extracellular vesicles

    Directory of Open Access Journals (Sweden)

    Zoltán Varga

    2014-02-01

    Full Text Available Background: Extracellular vesicles (EVs have clinical importance due to their roles in a wide range of biological processes. The detection and characterization of EVs are challenging because of their small size, low refractive index, and heterogeneity. Methods: In this manuscript, the size distribution of an erythrocyte-derived EV sample is determined using state-of-the-art techniques such as nanoparticle tracking analysis, resistive pulse sensing, and electron microscopy, and novel techniques in the field, such as small-angle X-ray scattering (SAXS and size exclusion chromatography coupled with dynamic light scattering detection. Results: The mode values of the size distributions of the studied erythrocyte EVs reported by the different methods show only small deviations around 130 nm, but there are differences in the widths of the size distributions. Conclusion: SAXS is a promising technique with respect to traceability, as this technique was already applied for traceable size determination of solid nanoparticles in suspension. To reach the traceable measurement of EVs, monodisperse and highly concentrated samples are required.

  15. Tumorigenic Potential of Extracellular Matrix Metalloproteinase Inducer

    Science.gov (United States)

    Zucker, Stanley; Hymowitz, Michelle; Rollo, Ellen E.; Mann, Richard; Conner, Cathleen E.; Cao, Jian; Foda, Hussein D.; Tompkins, David C.; Toole, Bryan P.

    2001-01-01

    Extracellular matrix metalloproteinase inducer (EMMPRIN), a glycoprotein present on the cancer cell plasma membrane, enhances fibroblast synthesis of matrix metalloproteinases (MMPs). The demonstration that peritumoral fibroblasts synthesize most of the MMPs in human tumors rather than the cancer cells themselves has ignited interest in the role of EMMPRIN in tumor dissemination. In this report we have demonstrated a role for EMMPRIN in cancer progression. Human MDA-MB-436 breast cancer cells, which are tumorigenic but slow growing in vivo, were transfected with EMMPRIN cDNA and injected orthotopically into mammary tissue of female NCr nu/nu mice. Green fluorescent protein was used to visualize metastases. In three experiments, breast cancer cell clones transfected with EMMPRIN cDNA were considerably more tumorigenic and invasive than plasmid-transfected cancer cells. Increased gelatinase A and gelatinase B expression (demonstrated by in situ hybridization and gelatin substrate zymography) was demonstrated in EMMPRIN-enhanced tumors. In contrast to de novo breast cancers in humans, human tumors transplanted into mice elicited minimal stromal or inflammatory cell reactions. Based on these experimental studies and our previous demonstration that EMMPRIN is prominently displayed in human cancer tissue, we propose that EMMPRIN plays an important role in cancer progression by increasing synthesis of MMPs. PMID:11395366

  16. New extracellular resistance mechanism for cisplatin.

    Science.gov (United States)

    Centerwall, Corey R; Kerwood, Deborah J; Goodisman, Jerry; Toms, Bonnie B; Dabrowiak, James C

    2008-01-01

    The HSQC NMR spectrum of 15N-cisplatin in cell growth media shows resonances corresponding to the monocarbonato complex, cis-[Pt(NH3)2(CO3)Cl](-), 4, and the dicarbonato complex, cis-[Pt(NH3)2(CO3)2](-2), 5, in addition to cisplatin itself, cis-[Pt(NH3)2Cl2], 1. The presence of Jurkat cells reduces the amount of detectable carbonato species by (2.8+/-0.7) fmol per cell and has little effect on species 1. Jurkat cells made resistant to cisplatin reduce the amount of detectable carbonato species by (7.9+/-5.6) fmol per cell and also reduce the amount of 1 by (3.4+/-0.9) fmol per cell. The amount of detectable carbonato species is also reduced by addition of the drug to medium that has previously been in contact with normal Jurkat cells (cells removed); the reduction is greater when drug is added to medium previously in contact with resistant Jurkat cells (cells removed). This shows that the platinum species are modified by a cell-produced substance that is released to the medium. Since the modified species have been shown not to enter or bind to cells, and since resistant cells modify more than non-resistant cells, the modification constitutes a new extracellular mechanism for cisplatin resistance which merits further attention.

  17. Relevance of extracellular DNA in rhizosphere

    Science.gov (United States)

    Pietramellara, Giacomo; Ascher, Judith; Baraniya, Divyashri; Arfaioli, Paola; Ceccherini, Maria Teresa; Hawes, Martha

    2013-04-01

    One of the most promising areas for future development is the manipulation of the rhizosphere to produce sustainable and efficient agriculture production systems. Using Omics approaches, to define the distinctive features of eDNA systems and structures, will facilitate progress in rhizo-enforcement and biocontrol studies. The relevance of these studies results clear when we consider the plethora of ecological functions in which eDNA is involved. This fraction can be actively extruded by living cells or discharged during cellular lysis and may exert a key role in the stability and variability of the soil bacterial genome, resulting also a source of nitrogen and phosphorus for plants due to the root's capacity to directly uptake short DNA fragments. The adhesive properties of the DNA molecule confer to eDNA the capacity to inhibit or kill pathogenic bacteria by cation limitation induction, and to facilitate formation of biofilm and extracellular traps (ETs), that may protect microorganisms inhabiting biofilm and plant roots against pathogens and allelopathic substances. The ETs are actively extruded by root border cells when they are dispersed in the rhizosphere, conferring to plants the capacity to extend an endogenous pathogen defence system outside the organism. Moreover, eDNA could be involved in rhizoremediation in heavy metal polluted soil acting as a bioflotation reagent.

  18. Functional transferred DNA within extracellular vesicles

    International Nuclear Information System (INIS)

    Cai, Jin; Wu, Gengze; Jose, Pedro A.; Zeng, Chunyu

    2016-01-01

    Extracellular vesicles (EVs) are small membrane vesicles including exosomes and shedding vesicles that mediated a cell-to-cell communication. EVs are released from almost all cell types under both physiological and pathological conditions and incorporate nuclear and cytoplasmic molecules for intercellular delivery. Besides protein, mRNA, and microRNA of these molecules, as recent studies show, specific DNA are prominently packaged into EVs. It appears likely that some of exosomes or shedding vesicles, bearing nuclear molecules are released upon bubble-like blebs. Specific interaction of EVs with susceptible recipients performs the uptake of EVs into the target cells, discharging their cargo including nuclear and cytoplasmic macromolecules into the cytosol. These findings expand the nucleic acid content of EVs to include increased levels of specific DNA. Thus, EVs contain a repertoire of genetic information available for horizontal gene transfer and potential use as blood biomarkers for cancer and atherosclerosis. In this review, the focus is on the characteristics, biological functions, and roles in diseases of DNA within EVs. - Highlights: • This review is focused on the DNA within EVs including its characteristics, biological functions, and roles in diseases. • It is clear that DNA within EVs might have important physiological and pathological roles in various diseases. • Knowledge in this area may provides us alternative methods for disease diagnosis or therapy in the future.

  19. Extracellular small RNAs: what, where, why?

    Science.gov (United States)

    Hoy, Anna M.; Buck, Amy H.

    2012-01-01

    miRNAs (microRNAs) are a class of small RNA that regulate gene expression by binding to mRNAs and modulating the precise amount of proteins that get expressed in a cell at a given time. This form of gene regulation plays an important role in developmental systems and is critical for the proper function of numerous biological pathways. Although miRNAs exert their functions inside the cell, these and other classes of RNA are found in body fluids in a cell-free form that is resistant to degradation by RNases. A broad range of cell types have also been shown to secrete miRNAs in association with components of the RISC (RNA-induced silencing complex) and/or encapsulation within vesicles, which can be taken up by other cells. In the present paper, we provide an overview of the properties of extracellular miRNAs in relation to their capacity as biomarkers, stability against degradation and mediators of cell–cell communication. PMID:22817753

  20. Peroxidase enzymes regulate collagen extracellular matrix biosynthesis.

    Science.gov (United States)

    DeNichilo, Mark O; Panagopoulos, Vasilios; Rayner, Timothy E; Borowicz, Romana A; Greenwood, John E; Evdokiou, Andreas

    2015-05-01

    Myeloperoxidase and eosinophil peroxidase are heme-containing enzymes often physically associated with fibrotic tissue and cancer in various organs, without any direct involvement in promoting fibroblast recruitment and extracellular matrix (ECM) biosynthesis at these sites. We report herein novel findings that show peroxidase enzymes possess a well-conserved profibrogenic capacity to stimulate the migration of fibroblastic cells and promote their ability to secrete collagenous proteins to generate a functional ECM both in vitro and in vivo. Mechanistic studies conducted using cultured fibroblasts show that these cells are capable of rapidly binding and internalizing both myeloperoxidase and eosinophil peroxidase. Peroxidase enzymes stimulate collagen biosynthesis at a post-translational level in a prolyl 4-hydroxylase-dependent manner that does not require ascorbic acid. This response was blocked by the irreversible myeloperoxidase inhibitor 4-amino-benzoic acid hydrazide, indicating peroxidase catalytic activity is essential for collagen biosynthesis. These results suggest that peroxidase enzymes, such as myeloperoxidase and eosinophil peroxidase, may play a fundamental role in regulating the recruitment of fibroblast and the biosynthesis of collagen ECM at sites of normal tissue repair and fibrosis, with enormous implications for many disease states where infiltrating inflammatory cells deposit peroxidases. Copyright © 2015 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.

  1. Functional transferred DNA within extracellular vesicles

    Energy Technology Data Exchange (ETDEWEB)

    Cai, Jin [Department of Cardiology, Daping Hospital, The Third Military Medical University, Chongqing 400042 (China); Department of Neurology, Jinling Hospital, Nanjing University School of Medicine, Jiangsu Province (China); Wu, Gengze [Department of Cardiology, Daping Hospital, The Third Military Medical University, Chongqing 400042 (China); Jose, Pedro A. [Division of Nephrology, Department of Medicine and Physiology, University of Maryland, School of Medicine, Baltimore, MD 21201 (United States); Zeng, Chunyu, E-mail: Chunyuzeng01@163.com [Department of Cardiology, Daping Hospital, The Third Military Medical University, Chongqing 400042 (China)

    2016-11-15

    Extracellular vesicles (EVs) are small membrane vesicles including exosomes and shedding vesicles that mediated a cell-to-cell communication. EVs are released from almost all cell types under both physiological and pathological conditions and incorporate nuclear and cytoplasmic molecules for intercellular delivery. Besides protein, mRNA, and microRNA of these molecules, as recent studies show, specific DNA are prominently packaged into EVs. It appears likely that some of exosomes or shedding vesicles, bearing nuclear molecules are released upon bubble-like blebs. Specific interaction of EVs with susceptible recipients performs the uptake of EVs into the target cells, discharging their cargo including nuclear and cytoplasmic macromolecules into the cytosol. These findings expand the nucleic acid content of EVs to include increased levels of specific DNA. Thus, EVs contain a repertoire of genetic information available for horizontal gene transfer and potential use as blood biomarkers for cancer and atherosclerosis. In this review, the focus is on the characteristics, biological functions, and roles in diseases of DNA within EVs. - Highlights: • This review is focused on the DNA within EVs including its characteristics, biological functions, and roles in diseases. • It is clear that DNA within EVs might have important physiological and pathological roles in various diseases. • Knowledge in this area may provides us alternative methods for disease diagnosis or therapy in the future.

  2. Towards integrating extracellular matrix and immunological pathways.

    Science.gov (United States)

    Boyd, David F; Thomas, Paul G

    2017-10-01

    The extracellular matrix (ECM) is a complex and dynamic structure made up of an estimated 300 different proteins. The ECM is also a rich source of cytokines and growth factors in addition to numerous bioactive ECM degradation products that influence cell migration, proliferation, and differentiation. The ECM is constantly being remodeled during homeostasis and in a wide range of pathological contexts. Changes in the ECM modulate immune responses, which in turn regulate repair and regeneration of tissues. Here, we review the many components of the ECM, enzymes involved in ECM remodeling, and the signals that feed into immunological pathways in the context of a dynamic ECM. We highlight studies that have taken an integrative approach to studying immune responses in the context of the ECM and studies that use novel proteomic strategies. Finally, we discuss research challenges relevant to the integration of immune and ECM networks and propose experimental and translational approaches to resolve these issues. Copyright © 2017 Elsevier Ltd. All rights reserved.

  3. Force spectroscopy of hepatocytic extracellular matrix components

    Energy Technology Data Exchange (ETDEWEB)

    Yongsunthon, R., E-mail: YongsuntR@Corning.com [Corning Incorporated, SP-FR-01, R1S32D, Corning, NY 14831 (United States); Baker, W.A.; Bryhan, M.D.; Baker, D.E.; Chang, T.; Petzold, O.N.; Walczak, W.J.; Liu, J.; Faris, R.A.; Senaratne, W.; Seeley, L.A.; Youngman, R.E. [Corning Incorporated, SP-FR-01, R1S32D, Corning, NY 14831 (United States)

    2009-07-15

    We present atomic force microscopy and force spectroscopy data of live hepatocytes (HEPG2/C3A liver cell line) grown in Eagle's Minimum Essential Medium, a complex solution of salts and amino acids commonly used for cell culture. Contact-mode imaging and force spectroscopy of this system allowed correlation of cell morphology and extracellular matrix (ECM) properties with substrate properties. Force spectroscopy analysis of cellular 'footprints' indicated that the cells secrete large polymers (e.g., 3.5 {mu}m contour length and estimated MW 1000 kDa) onto their substrate surface. Although definitive identification of the polymers has not yet been achieved, fluorescent-labeled antibody staining has specified the presence of ECM proteins such as collagen and laminin in the cellular footprints. The stretched polymers appear to be much larger than single molecules of known ECM components, such as collagen and heparan sulfate proteoglycan, thus suggesting that the cells create larger entangled, macromolecular structures from smaller components. There is strong evidence which suggests that the composition of the ECM is greatly influenced by the hydrophobicity of the substrate surface, with preferential production and/or adsorption of larger macromolecules on hydrophobic surfaces.

  4. Routes and mechanisms of extracellular vesicle uptake

    Directory of Open Access Journals (Sweden)

    Laura Ann Mulcahy

    2014-08-01

    Full Text Available Extracellular vesicles (EVs are small vesicles released by donor cells that can be taken up by recipient cells. Despite their discovery decades ago, it has only recently become apparent that EVs play an important role in cell-to-cell communication. EVs can carry a range of nucleic acids and proteins which can have a significant impact on the phenotype of the recipient. For this phenotypic effect to occur, EVs need to fuse with target cell membranes, either directly with the plasma membrane or with the endosomal membrane after endocytic uptake. EVs are of therapeutic interest because they are deregulated in diseases such as cancer and they could be harnessed to deliver drugs to target cells. It is therefore important to understand the molecular mechanisms by which EVs are taken up into cells. This comprehensive review summarizes current knowledge of EV uptake mechanisms. Cells appear to take up EVs by a variety of endocytic pathways, including clathrin-dependent endocytosis, and clathrin-independent pathways such as caveolin-mediated uptake, macropinocytosis, phagocytosis, and lipid raft–mediated internalization. Indeed, it seems likely that a heterogeneous population of EVs may gain entry into a cell via more than one route. The uptake mechanism used by a given EV may depend on proteins and glycoproteins found on the surface of both the vesicle and the target cell. Further research is needed to understand the precise rules that underpin EV entry into cells.

  5. Protein Dynamics in the Plant Extracellular Space

    Directory of Open Access Journals (Sweden)

    Leonor Guerra-Guimarães

    2016-07-01

    Full Text Available The extracellular space (ECS or apoplast is the plant cell compartment external to the plasma membrane, which includes the cell walls, the intercellular space and the apoplastic fluid (APF. The present review is focused on APF proteomics papers and intends to draw information on the metabolic processes occurring in the ECS under abiotic and biotic stresses, as well as under non-challenged conditions. The large majority of the proteins detected are involved in “cell wall organization and biogenesis”, “response to stimulus” and “protein metabolism”. It becomes apparent that some proteins are always detected, irrespective of the experimental conditions, although with different relative contribution. This fact suggests that non-challenged plants have intrinsic constitutive metabolic processes of stress/defense in the ECS. In addition to the multiple functions ascribed to the ECS proteins, should be considered the interactions established between themselves and with the plasma membrane and its components. These interactions are crucial in connecting exterior and interior of the cell, and even simple protein actions in the ECS can have profound effects on plant performance. The proteins of the ECS are permanently contributing to the high dynamic nature of this plant compartment, which seems fundamental to plant development and adaptation to the environmental conditions.

  6. Bacterial Extracellular Polysaccharides Involved in Biofilm Formation

    Directory of Open Access Journals (Sweden)

    Elena P. Ivanova

    2009-07-01

    Full Text Available Extracellular polymeric substances (EPS produced by microorganisms are a complex mixture of biopolymers primarily consisting of polysaccharides, as well as proteins, nucleic acids, lipids and humic substances. EPS make up the intercellular space of microbial aggregates and form the structure and architecture of the biofilm matrix. The key functions of EPS comprise the mediation of the initial attachment of cells to different substrata and protection against environmental stress and dehydration. The aim of this review is to present a summary of the current status of the research into the role of EPS in bacterial attachment followed by biofilm formation. The latter has a profound impact on an array of biomedical, biotechnology and industrial fields including pharmaceutical and surgical applications, food engineering, bioremediation and biohydrometallurgy. The diverse structural variations of EPS produced by bacteria of different taxonomic lineages, together with examples of biotechnological applications, are discussed. Finally, a range of novel techniques that can be used in studies involving biofilm-specific polysaccharides is discussed.

  7. Transcription activator-like effector-mediated regulation of gene expression based on the inducible packaging and delivery via designed extracellular vesicles

    International Nuclear Information System (INIS)

    Lainšček, Duško; Lebar, Tina; Jerala, Roman

    2017-01-01

    Transcription activator-like effector (TALE) proteins present a powerful tool for genome editing and engineering, enabling introduction of site-specific mutations, gene knockouts or regulation of the transcription levels of selected genes. TALE nucleases or TALE-based transcription regulators are introduced into mammalian cells mainly via delivery of the coding genes. Here we report an extracellular vesicle-mediated delivery of TALE transcription regulators and their ability to upregulate the reporter gene in target cells. Designed transcriptional activator TALE-VP16 fused to the appropriate dimerization domain was enriched as a cargo protein within extracellular vesicles produced by mammalian HEK293 cells stimulated by Ca-ionophore and using blue light- or rapamycin-inducible dimerization systems. Blue light illumination or rapamycin increased the amount of the TALE-VP16 activator in extracellular vesicles and their addition to the target cells resulted in an increased expression of the reporter gene upon addition of extracellular vesicles to the target cells. This technology therefore represents an efficient delivery for the TALE-based transcriptional regulators. - Highlights: • Inducible dimerization enriched cargo proteins within extracellular vesicles (EV). • Farnesylation surpassed LAMP-1 fusion proteins for the EV packing. • Extracellular vesicles were able to deliver TALE regulators to mammalian cells. • TALE mediated transcriptional activation was achieved by designed EV.

  8. Activation of retinal glial (Müller cells by extracellular ATP induces pronounced increases in extracellular H+ flux.

    Directory of Open Access Journals (Sweden)

    Boriana K Tchernookova

    Full Text Available Small alterations in extracellular acidity are potentially important modulators of neuronal signaling within the vertebrate retina. Here we report a novel extracellular acidification mechanism mediated by glial cells in the retina. Using self-referencing H+-selective microelectrodes to measure extracellular H+ fluxes, we show that activation of retinal Müller (glial cells of the tiger salamander by micromolar concentrations of extracellular ATP induces a pronounced extracellular H+ flux independent of bicarbonate transport. ADP, UTP and the non-hydrolyzable analog ATPγs at micromolar concentrations were also potent stimulators of extracellular H+ fluxes, but adenosine was not. The extracellular H+ fluxes induced by ATP were mimicked by the P2Y1 agonist MRS 2365 and were significantly reduced by the P2 receptor blockers suramin and PPADS, suggesting activation of P2Y receptors. Bath-applied ATP induced an intracellular rise in calcium in Müller cells; both the calcium rise and the extracellular H+ fluxes were significantly attenuated when calcium re-loading into the endoplasmic reticulum was inhibited by thapsigargin and when the PLC-IP3 signaling pathway was disrupted with 2-APB and U73122. The anion transport inhibitor DIDS also markedly reduced the ATP-induced increase in H+ flux while SITS had no effect. ATP-induced H+ fluxes were also observed from Müller cells isolated from human, rat, monkey, skate and lamprey retinae, suggesting a highly evolutionarily conserved mechanism of potential general importance. Extracellular ATP also induced significant increases in extracellular H+ flux at the level of both the outer and inner plexiform layers in retinal slices of tiger salamander which was significantly reduced by suramin and PPADS. We suggest that the novel H+ flux mediated by ATP-activation of Müller cells and of other glia as well may be a key mechanism modulating neuronal signaling in the vertebrate retina and throughout the brain.

  9. Extracellular electron transfer mechanisms between microorganisms and minerals

    Energy Technology Data Exchange (ETDEWEB)

    Shi, Liang; Dong, Hailiang; Reguera, Gemma; Beyenal, Haluk; Lu, Anhuai; Liu, Juan; Yu, Han-Qing; Fredrickson, James K.

    2016-08-30

    Electrons can be transferred from microorganisms to multivalent metal ions that are associated with minerals and vice versa. As the microbial cell envelope is neither physically permeable to minerals nor electrically conductive, microorganisms have evolved strategies to exchange electrons with extracellular minerals. In this Review, we discuss the molecular mechanisms that underlie the ability of microorganisms to exchange electrons, such as c-type cytochromes and microbial nanowires, with extracellular minerals and with microorganisms of the same or different species. Microorganisms that have extracellular electron transfer capability can be used for biotechnological applications, including bioremediation, biomining and the production of biofuels and nanomaterials.

  10. Surface glycosylation profiles of urine extracellular vesicles.

    Directory of Open Access Journals (Sweden)

    Jared Q Gerlach

    Full Text Available Urinary extracellular vesicles (uEVs are released by cells throughout the nephron and contain biomolecules from their cells of origin. Although uEV-associated proteins and RNA have been studied in detail, little information exists regarding uEV glycosylation characteristics. Surface glycosylation profiling by flow cytometry and lectin microarray was applied to uEVs enriched from urine of healthy adults by ultracentrifugation and centrifugal filtration. The carbohydrate specificity of lectin microarray profiles was confirmed by competitive sugar inhibition and carbohydrate-specific enzyme hydrolysis. Glycosylation profiles of uEVs and purified Tamm Horsfall protein were compared. In both flow cytometry and lectin microarray assays, uEVs demonstrated surface binding, at low to moderate intensities, of a broad range of lectins whether prepared by ultracentrifugation or centrifugal filtration. In general, ultracentrifugation-prepared uEVs demonstrated higher lectin binding intensities than centrifugal filtration-prepared uEVs consistent with lesser amounts of co-purified non-vesicular proteins. The surface glycosylation profiles of uEVs showed little inter-individual variation and were distinct from those of Tamm Horsfall protein, which bound a limited number of lectins. In a pilot study, lectin microarray was used to compare uEVs from individuals with autosomal dominant polycystic kidney disease to those of age-matched controls. The lectin microarray profiles of polycystic kidney disease and healthy uEVs showed differences in binding intensity of 6/43 lectins. Our results reveal a complex surface glycosylation profile of uEVs that is accessible to lectin-based analysis following multiple uEV enrichment techniques, is distinct from co-purified Tamm Horsfall protein and may demonstrate disease-specific modifications.

  11. Modular Extracellular Matrices: Solutions for the Puzzle

    Science.gov (United States)

    Serban, Monica A.; Prestwich, Glenn D.

    2008-01-01

    The common technique of growing cells in two-dimensions (2-D) is gradually being replaced by culturing cells on matrices with more appropriate composition and stiffness, or by encapsulation of cells in three-dimensions (3-D). The universal acceptance of the new 3-D paradigm has been constrained by the absence of a commercially available, biocompatible material that offers ease of use, experimental flexibility, and a seamless transition from in vitro to in vivo applications. The challenge – the puzzle that needs a solution – is to replicate the complexity of the native extracellular matrix (ECM) environment with the minimum number of components necessary to allow cells to rebuild and replicate a given tissue. For use in drug discovery, toxicology, cell banking, and ultimately in reparative medicine, the ideal matrix would therefore need to be highly reproducible, manufacturable, approvable, and affordable. Herein we describe the development of a set of modular components that can be assembled into biomimetic materials that meet these requirements. These semi-synthetic ECMs, or sECMs, are based on hyaluronan derivatives that form covalently crosslinked, biodegradable hydrogels suitable for 3-D culture of primary and stem cells in vitro, and for tissue formation in vivo. The sECMs can be engineered to provide appropriate biological cues needed to recapitulate the complexity of a given ECM environment. Specific applications for different sECM compositions include stem cell expansion with control of differentiation, scar-free wound healing, growth factor delivery, cell delivery for osteochondral defect and liver repair, and development of vascularized tumor xenografts for personalized chemotherapy. PMID:18442709

  12. Neutrophil extracellular trap formation in supragingival biofilms.

    Science.gov (United States)

    Hirschfeld, Josefine; Dommisch, Henrik; Skora, Philipp; Horvath, Gabor; Latz, Eicke; Hoerauf, Achim; Waller, Tobias; Kawai, Toshihisa; Jepsen, Søren; Deschner, James; Bekeredjian-Ding, Isabelle

    2015-01-01

    Oral biofilms are the causative agents of the highly prevalent oral diseases periodontitis and caries. Additionally, the host immune response is thought to play a critical role in disease onset. Neutrophils are known to be a key host response factor to bacterial challenge on host surfaces. Release of neutrophil extracellular traps (NETs) as a novel antimicrobial defense strategy has gained increasing attention in the past years. Here, we investigated the influx of neutrophils into the dental plaque and the ability of oral bacteria to trigger intra-biofilm release of NETs and intracellular proteins. Supragingival biofilms and whole saliva were sampled from systemically healthy subjects participating in an experimental gingivitis study. Biofilms were analysed by immunofluorescence followed by confocal and fluorescence microscopy. Moreover, concentrations of cytokines and immune-associated proteins in biofilm suspensions and saliva were assessed by ELISA. Neutrophils obtained from blood were stimulated with twelve bacterial species isolated from cultured biofilms or with lipopolysaccharide to monitor NET formation. Neutrophils, NETs, neutrophil-associated proteins (myeloperoxidase, elastase-2, cathepsin G, cathelicidin LL-37), interleukin-8, interleukin-1β and tumor necrosis factor were detected within plaque samples and saliva. All tested bacterial species as well as the polymicrobial samples isolated from the plaque of each donor induced release of NETs and interleukin-8. The degree of NET formation varied among different subjects and did not correlate with plaque scores or clinical signs of local inflammation. Our findings indicate that neutrophils are attracted towards dental biofilms, in which they become incorporated and where they are stimulated by microbes to release NETs and immunostimulatory proteins. Thus, neutrophils and NETs may be involved in host biofilm control, although their specific role needs to be further elucidated. Moreover, inter

  13. Extracellular gadolinium contrast agents: Differences in stability

    International Nuclear Information System (INIS)

    Morcos, S.K.

    2008-01-01

    Extracellular gadolinium contrast agents (Gd-CA) are either linear or macrocyclic chelates available as ionic or non-ionic preparations. The molecular structure whether cyclic or linear and ionicity determines the stability of Gd-CA. Linear chelates are flexible open chains which do not offer a strong binding to Gd 3+ . In contrast, the macrocyclic chelates offer a strong binding to Gd 3+ by the virtue of being preorganized rigid rings of almost optimal size to cage the gadolinium atom. Non-ionic preparations are also less stable in comparison to the ionic ones as the binding between Gd 3+ with the negatively charged carboxyl groups is stronger in comparison to that with amides or alcohol in the non-ionic preparations. According to stability constants and kinetic measurements, the most stable Gd-CM is the ionic-macrocyclic chelate Gd-DOTA and the least stable agents are the non-ionic linear chelates gadodiamide and gadoversetamide. In vivo data confirmed the low stability of non-ionic linear chelates but no significant difference was observed amongst the macrocyclic agents whether ionic (Gd-DOTA) or non-ionic such as Gd-HP-DO3A and Gd-BT-DO3A. The stability of Gd-CA seems to be an important factor in the pathogenesis of the serious complication of nephrogenic systemic fibrosis. Gd-CA of low stability are likely to undergo transmetallation and release free Gd ions that deposit in tissue and attract circulating fibrocytes to initiate the process of fibrosis. No cases of NSF have been observed so far after the exclusive use of the stable macrocyclic Gd-CA

  14. Creating leptin-like biofunctions by active immunization against chicken leptin receptor in growing chickens.

    Science.gov (United States)

    Lei, M M; Wu, S Q; Shao, X B; Li, X W; Chen, Z; Ying, S J; Shi, Z D

    2015-01-01

    In this study, immunization against chicken leptin receptor (cLEPR) extracellular domain (ECD) was applied to investigate leptin regulation and LEPR biofunction in growing chicken pullets. A recombinant protein (cLEPR ECD) based on the cLEPR complemenary DNA sequence corresponding to the 582nd to 796th amino acid residues of cLEPR mature peptide was prepared and used as antigen. Immunization against cLEPR ECD in growing chickens increased anti-cLEPR ECD antibody titers in blood, enhanced proportions of phosphorylated janus kinase 2 (JAK2) and served as signal transducer and activator of transcription 3 (STAT3) protein in liver tissue. Chicken live weight gain and abdominal fat mass were significantly decreased (P chickens. Copyright © 2015 Elsevier Inc. All rights reserved.

  15. Domain: Labour market

    NARCIS (Netherlands)

    Oude Mulders, J.; Wadensjö, E.; Hasselhorn, H.M.; Apt, W.

    This domain chapter is dedicated to summarize research on the effects of labour market contextual factors on labour market participation of older workers (aged 50+) and identify research gaps. While employment participation and the timing of (early) retirement is often modelled as an individual

  16. Cellulose binding domain proteins

    Science.gov (United States)

    Shoseyov, Oded; Shpiegl, Itai; Goldstein, Marc; Doi, Roy

    1998-01-01

    A cellulose binding domain (CBD) having a high affinity for crystalline cellulose and chitin is disclosed, along with methods for the molecular cloning and recombinant production thereof. Fusion products comprising the CBD and a second protein are likewise described. A wide range of applications are contemplated for both the CBD and the fusion products, including drug delivery, affinity separations, and diagnostic techniques.

  17. Domain-Specific Multimodeling

    DEFF Research Database (Denmark)

    Hessellund, Anders

    the overall level of abstraction. It does, however, also introduce a new problem of coordinating multiple different languages in a single system. We call this problem the coordination problem. In this thesis, we present the coordination method for domain-specific multimodeling that explicitly targets...

  18. GlycoDomainViewer

    DEFF Research Database (Denmark)

    Joshi, Hiren J; Jørgensen, Anja; Schjoldager, Katrine T

    2018-01-01

    features, which enhances visibility and accessibility of the wealth of glycoproteomic data being generated. The GlycoDomainViewer enables visual exploration of glycoproteomic data, incorporating information from recent N- and O-glycoproteome studies on human and animal cell lines and some organs and body...

  19. Molecular Mechanisms of SH2- and PTB-Domain-Containing Proteins in Receptor Tyrosine Kinase Signaling

    Science.gov (United States)

    Wagner, Melany J.; Stacey, Melissa M.; Liu, Bernard A.; Pawson, Tony

    2013-01-01

    Intracellular signaling is mediated by reversible posttranslational modifications (PTMs) that include phosphorylation, ubiquitination, and acetylation, among others. In response to extracellular stimuli such as growth factors, receptor tyrosine kinases (RTKs) typically dimerize and initiate signaling through phosphorylation of their cytoplasmic tails and downstream scaffolds. Signaling effectors are recruited to these phosphotyrosine (pTyr) sites primarily through Src homology 2 (SH2) domains and pTyr-binding (PTB) domains. This review describes how these conserved domains specifically recognize pTyr residues and play a major role in mediating precise downstream signaling events. PMID:24296166

  20. Molecular mechanisms of SH2- and PTB-domain-containing proteins in receptor tyrosine kinase signaling.

    Science.gov (United States)

    Wagner, Melany J; Stacey, Melissa M; Liu, Bernard A; Pawson, Tony

    2013-12-01

    Intracellular signaling is mediated by reversible posttranslational modifications (PTMs) that include phosphorylation, ubiquitination, and acetylation, among others. In response to extracellular stimuli such as growth factors, receptor tyrosine kinases (RTKs) typically dimerize and initiate signaling through phosphorylation of their cytoplasmic tails and downstream scaffolds. Signaling effectors are recruited to these phosphotyrosine (pTyr) sites primarily through Src homology 2 (SH2) domains and pTyr-binding (PTB) domains. This review describes how these conserved domains specifically recognize pTyr residues and play a major role in mediating precise downstream signaling events.

  1. Neutrophil Extracellular Trap-Related Extracellular Histones Cause Vascular Necrosis in Severe GN.

    Science.gov (United States)

    Kumar, Santhosh V R; Kulkarni, Onkar P; Mulay, Shrikant R; Darisipudi, Murthy N; Romoli, Simone; Thomasova, Dana; Scherbaum, Christina R; Hohenstein, Bernd; Hugo, Christian; Müller, Susanna; Liapis, Helen; Anders, Hans-Joachim

    2015-10-01

    Severe GN involves local neutrophil extracellular trap (NET) formation. We hypothesized a local cytotoxic effect of NET-related histone release in necrotizing GN. In vitro, histones from calf thymus or histones released by neutrophils undergoing NETosis killed glomerular endothelial cells, podocytes, and parietal epithelial cells in a dose-dependent manner. Histone-neutralizing agents such as antihistone IgG, activated protein C, or heparin prevented this effect. Histone toxicity on glomeruli ex vivo was Toll-like receptor 2/4 dependent, and lack of TLR2/4 attenuated histone-induced renal thrombotic microangiopathy and glomerular necrosis in mice. Anti-glomerular basement membrane GN involved NET formation and vascular necrosis, whereas blocking NET formation by peptidylarginine inhibition or preemptive anti-histone IgG injection significantly reduced all aspects of GN (i.e., vascular necrosis, podocyte loss, albuminuria, cytokine induction, recruitment or activation of glomerular leukocytes, and glomerular crescent formation). To evaluate histones as a therapeutic target, mice with established GN were treated with three different histone-neutralizing agents. Anti-histone IgG, recombinant activated protein C, and heparin were equally effective in abrogating severe GN, whereas combination therapy had no additive effects. Together, these results indicate that NET-related histone release during GN elicits cytotoxic and immunostimulatory effects. Furthermore, neutralizing extracellular histones is still therapeutic when initiated in established GN. Copyright © 2015 by the American Society of Nephrology.

  2. The framing of scientific domains

    DEFF Research Database (Denmark)

    Dam Christensen, Hans

    2014-01-01

    domains, and UNISIST helps understanding this navigation. Design/methodology/approach The UNISIST models are tentatively applied to the domain of art history at three stages, respectively two modern, partially overlapping domains, as well as an outline of an art historical domain anno c1820...

  3. Functional Elements on SIRPα IgV domain Mediate Cell Surface Binding to CD47

    Science.gov (United States)

    Liu, Yuan; Tong, Qiao; Zhou, Yubin; Lee, Hsiau-Wei; Yang, Jenny J.; Bühring, Hans-Jörg; Chen, Yi-Tien; Ha, Binh; Chen, Celia X-J.; Zen, Ke

    2007-01-01

    Summary SIRPα and SIRPβ1, the two major isoforms of the signal regulatory protein (SIRP) family, are co-expressed in human leukocytes but mediate distinct extracellular binding interactions and divergent cell signaling responses. Previous studies have demonstrated that binding of SIRPα with CD47, another important cell surface molecule, through the extracellular IgV domain regulates important leukocyte functions including macrophage recognition, leukocyte adhesion and transmigration. Although SIRPβ1 shares highly homologous extracellular IgV structure with SIRPα, it does not bind to CD47. In this study, we defined key amino acid residues exclusively expressing in the IgV domain of SIRPα, but not SIRPβ1, which determine the extracellular binding interaction of SIRPα to CD47. These key residues include Gln67, a small hydrophobic amino acid (Ala or Val) at the 57th position and Met102. We found that Gln67 and Ala/Val57 are critical. Mutation of either of these residues abates SIRPα directly binding to CD47. Functional cell adhesion and leukocyte transmigration assays further demonstrated central roles of Gln67 and Ala/Val57 in SIRPα extracellular binding mediated cell interactions and cell migration. Another SIRPα-specific residue, Met102, appears to assist SIRPα IgV binding through Gln67 and Ala/Val57. An essential role of these amino acids in SIRPα binding to CD47 was further confirmed by introducing these residues into the SIRPβ1 IgV domain, which dramatically converts SIRPβ1 into a CD47-binding molecule. Our results thus revealed the molecular basis by which SIRPα selectively binds to CD47 and shed new light into the structural mechanisms of SIRP isoform mediated distinctive extracellular interactions and cellular responses. PMID:17070842

  4. Functional elements on SIRPalpha IgV domain mediate cell surface binding to CD47.

    Science.gov (United States)

    Liu, Yuan; Tong, Qiao; Zhou, Yubin; Lee, Hsiau-Wei; Yang, Jenny J; Bühring, Hans-Jörg; Chen, Yi-Tien; Ha, Binh; Chen, Celia X-J; Yang, Yang; Zen, Ke

    2007-01-19

    SIRPalpha and SIRPbeta1, the two major isoforms of the signal regulatory protein (SIRP) family, are co-expressed in human leukocytes but mediate distinct extracellular binding interactions and divergent cell signaling responses. Previous studies have demonstrated that binding of SIRPalpha with CD47, another important cell surface molecule, through the extracellular IgV domain regulates important leukocyte functions including macrophage recognition, leukocyte adhesion and transmigration. Although SIRPbeta1 shares highly homologous extracellular IgV structure with SIRPalpha, it does not bind to CD47. Here, we defined key amino acid residues exclusively expressing in the IgV domain of SIRPalpha, but not SIRPbeta1, which determine the extracellular binding interaction of SIRPalpha to CD47. These key residues include Gln67, a small hydrophobic amino acid (Ala or Val) at the 57th position and Met102. We found that Gln67 and Ala/Val57 are critical. Mutation of either of these residues abates SIRPalpha directly binding to CD47. Functional cell adhesion and leukocyte transmigration assays further demonstrated central roles of Gln67 and Ala/Val57 in SIRPalpha extracellular binding mediated cell interactions and cell migration. Another SIRPalpha-specific residue, Met102, appears to assist SIRPalpha IgV binding through Gln67 and Ala/Val57. An essential role of these amino acid residues in SIRPalpha binding to CD47 was further confirmed by introducing these residues into the SIRPbeta1 IgV domain, which dramatically converts SIRPbeta1 into a CD47-binding molecule. Our results thus revealed the molecular basis by which SIRPalpha binds to CD47 and shed new light into the structural mechanisms of SIRP isoform mediated distinctive extracellular interactions and cellular responses.

  5. Improved Methods of Producing and Administering Extracellular Vesicles | Poster

    Science.gov (United States)

    An efficient method of producing purified extracellular vesicles (EVs), in conjunction with a method that blocks liver macrophages from clearing EVs from the body, has produced promising results for the use of EVs in cancer therapy.

  6. EVpedia : a community web portal for extracellular vesicles research

    NARCIS (Netherlands)

    Kim, Dae-Kyum; Lee, Jaewook; Kim, Sae Rom; Choi, Dong-Sic; Yoon, Yae Jin; Kim, Ji Hyun; Go, Gyeongyun; Nhung, Dinh; Hong, Kahye; Jang, Su Chul; Kim, Si-Hyun; Park, Kyong-Su; Kim, Oh Youn; Park, Hyun Taek; Seo, Ji Hye; Aikawa, Elena; Baj-Krzyworzeka, Monika; van Balkom, Bas W M; Belting, Mattias; Blanc, Lionel; Bond, Vincent; Bongiovanni, Antonella; Borràs, Francesc E; Buée, Luc; Buzás, Edit I; Cheng, Lesley; Clayton, Aled; Cocucci, Emanuele; Dela Cruz, Charles S; Desiderio, Dominic M; Di Vizio, Dolores; Ekström, Karin; Falcon-Perez, Juan M; Gardiner, Chris; Giebel, Bernd; Greening, David W; Gross, Julia Christina; Gupta, Dwijendra; Hendrix, An; Hill, Andrew F; Hill, Michelle M; Nolte-'t Hoen, Esther; Hwang, Do Won; Inal, Jameel; Jagannadham, Medicharla V; Jayachandran, Muthuvel; Jee, Young-Koo; Jørgensen, Malene; Kim, Kwang Pyo; Kim, Yoon-Keun; Kislinger, Thomas; Lässer, Cecilia; Lee, Dong Soo; Lee, Hakmo; van Leeuwen, Johannes; Lener, Thomas; Liu, Ming-Lin; Lötvall, Jan; Marcilla, Antonio; Mathivanan, Suresh; Möller, Andreas; Morhayim, Jess; Mullier, François; Nazarenko, Irina; Nieuwland, Rienk; Nunes, Diana N; Pang, Ken; Park, Jaesung; Patel, Tushar; Pocsfalvi, Gabriella; Del Portillo, Hernando; Putz, Ulrich; Ramirez, Marcel I; Rodrigues, Marcio L; Roh, Tae-Young; Royo, Felix; Sahoo, Susmita; Schiffelers, Raymond|info:eu-repo/dai/nl/212909509; Sharma, Shivani; Siljander, Pia; Simpson, Richard J; Soekmadji, Carolina; Stahl, Philip; Stensballe, Allan; Stępień, Ewa; Tahara, Hidetoshi; Trummer, Arne; Valadi, Hadi; Vella, Laura J; Wai, Sun Nyunt; Witwer, Kenneth; Yáñez-Mó, María; Youn, Hyewon; Zeidler, Reinhard; Gho, Yong Song; Nolte - t Hoen, Esther|info:eu-repo/dai/nl/261632175

    2014-01-01

    MOTIVATION: Extracellular vesicles (EVs) are spherical bilayered proteolipids, harboring various bioactive molecules. Due to the complexity of the vesicular nomenclatures and components, online searches for EV-related publications and vesicular components are currently challenging. RESULTS: We

  7. EVpedia: a community web portal for extracellular vesicles research

    NARCIS (Netherlands)

    Kim, Dae-Kyum; Lee, Jaewook; Kim, Sae Rom; Choi, Dong-Sic; Yoon, Yae Jin; Kim, Ji Hyun; Go, Gyeongyun; Nhung, Dinh; Hong, Kahye; Jang, Su Chul; Kim, Si-Hyun; Park, Kyong-Su; Kim, Oh Youn; Park, Hyun Taek; Seo, Ji Hye; Aikawa, Elena; Baj-Krzyworzeka, Monika; van Balkom, Bas W. M.; Belting, Mattias; Blanc, Lionel; Bond, Vincent; Bongiovanni, Antonella; Borràs, Francesc E.; Buée, Luc; Buzás, Edit I.; Cheng, Lesley; Clayton, Aled; Cocucci, Emanuele; Dela Cruz, Charles S.; Desiderio, Dominic M.; Di Vizio, Dolores; Ekström, Karin; Falcon-Perez, Juan M.; Gardiner, Chris; Giebel, Bernd; Greening, David W.; Gross, Julia Christina; Gupta, Dwijendra; Hendrix, An; Hill, Andrew F.; Hill, Michelle M.; Nolte-'t Hoen, Esther; Hwang, Do Won; Inal, Jameel; Jagannadham, Medicharla V.; Jayachandran, Muthuvel; Jee, Young-Koo; Jørgensen, Malene; Kim, Kwang Pyo; Kim, Yoon-Keun; Kislinger, Thomas; Lässer, Cecilia; Lee, Dong Soo; Lee, Hakmo; van Leeuwen, Johannes; Lener, Thomas; Liu, Ming-Lin; Lötvall, Jan; Marcilla, Antonio; Mathivanan, Suresh; Möller, Andreas; Morhayim, Jess; Mullier, François; Nazarenko, Irina; Nieuwland, Rienk; Nunes, Diana N.; Pang, Ken; Park, Jaesung; Patel, Tushar; Pocsfalvi, Gabriella; del Portillo, Hernando; Putz, Ulrich; Ramirez, Marcel I.; Rodrigues, Marcio L.; Roh, Tae-Young; Royo, Felix; Sahoo, Susmita; Schiffelers, Raymond; Sharma, Shivani; Siljander, Pia; Simpson, Richard J.; Soekmadji, Carolina; Stahl, Philip; Stensballe, Allan; Stępień, Ewa; Tahara, Hidetoshi; Trummer, Arne; Valadi, Hadi; Vella, Laura J.; Wai, Sun Nyunt; Witwer, Kenneth; Yáñez-Mó, María; Youn, Hyewon; Zeidler, Reinhard; Gho, Yong Song

    2015-01-01

    Extracellular vesicles (EVs) are spherical bilayered proteolipids, harboring various bioactive molecules. Due to the complexity of the vesicular nomenclatures and components, online searches for EV-related publications and vesicular components are currently challenging. We present an improved

  8. Biological properties of extracellular vesicles and their physiological functions

    NARCIS (Netherlands)

    Yáñez-Mó, María; Siljander, Pia R-M; Andreu, Zoraida; Zavec, Apolonija Bedina; Borràs, Francesc E; Buzas, Edit I; Buzas, Krisztina; Casal, Enriqueta; Cappello, Francesco; Carvalho, Joana; Colás, Eva; Cordeiro-da Silva, Anabela; Fais, Stefano; Falcon-Perez, Juan M; Ghobrial, Irene M; Giebel, Bernd; Gimona, Mario; Graner, Michael; Gursel, Ihsan; Gursel, Mayda; Heegaard, Niels H H; Hendrix, An; Kierulf, Peter; Kokubun, Katsutoshi; Kosanovic, Maja; Kralj-Iglic, Veronika; Krämer-Albers, Eva-Maria; Laitinen, Saara; Lässer, Cecilia; Lener, Thomas; Ligeti, Erzsébet; Linē, Aija; Lipps, Georg; Llorente, Alicia; Lötvall, Jan; Manček-Keber, Mateja; Marcilla, Antonio; Mittelbrunn, Maria; Nazarenko, Irina; Nolte-'t Hoen, Esther N M; Nyman, Tuula A; O'Driscoll, Lorraine; Olivan, Mireia; Oliveira, Carla; Pállinger, Éva; Del Portillo, Hernando A; Reventós, Jaume; Rigau, Marina; Rohde, Eva; Sammar, Marei; Sánchez-Madrid, Francisco; Santarém, N; Schallmoser, Katharina; Ostenfeld, Marie Stampe; Stoorvogel, Willem|info:eu-repo/dai/nl/074352385; Stukelj, Roman; Van der Grein, Susanne G|info:eu-repo/dai/nl/412755211; Vasconcelos, M Helena; Wauben, Marca H M|info:eu-repo/dai/nl/112675735; De Wever, Olivier

    2015-01-01

    In the past decade, extracellular vesicles (EVs) have been recognized as potent vehicles of intercellular communication, both in prokaryotes and eukaryotes. This is due to their capacity to transfer proteins, lipids and nucleic acids, thereby influencing various physiological and pathological

  9. Electrochemical roles of extracellular polymeric substances in biofilms

    DEFF Research Database (Denmark)

    Xiao, Yong; Zhao, Feng

    2017-01-01

    Most microbial cells in nature are surrounded by extracellular polymeric substances (EPS), which are fundamental components and determine the physiochemical properties of a biofilm. This review highlights the EPS properties of conductivity and redox ability from an electrochemical perspective, em...

  10. Extracellular matrix scaffolds for cartilage and bone regeneration

    NARCIS (Netherlands)

    Benders, K.E.M.; van Weeren, P.R.; Badylak, S.F.; Saris, Daniël B.F.; Dhert, W.J.A.; Malda, J.

    2013-01-01

    Regenerative medicine approaches based on decellularized extracellular matrix (ECM) scaffolds and tissues are rapidly expanding. The rationale for using ECM as a natural biomaterial is the presence of bioactive molecules that drive tissue homeostasis and regeneration. Moreover, appropriately

  11. Extracellular vesicles provide a means for tissue crosstalk during exercise

    DEFF Research Database (Denmark)

    Whitham, Martin; Parker, Benjamin L; Friedrichsen, Martin

    2018-01-01

    Exercise stimulates the release of molecules into the circulation, supporting the concept that inter-tissue signaling proteins are important mediators of adaptations to exercise. Recognizing that many circulating proteins are packaged in extracellular vesicles (EVs), we employed quantitative prot...

  12. EVpedia : A community web portal for extracellular vesicles research

    NARCIS (Netherlands)

    Kim, Dae Kyum; Lee, Jaewook; Kim, Sae Rom; Choi, Dong Sic; Yoon, Yae Jin; Kim, Ji Hyun; Go, Gyeongyun; Nhung, Dinh; Hong, Kahye; Jang, Su Chul; Kim, Si Hyun; Park, Kyong Su; Kim, Oh Youn; Park, Hyun Taek; Seo, Ji Hye; Aikawa, Elena; Baj-Krzyworzeka, Monika; Van Balkom, Bas W M; Belting, Mattias; Blanc, Lionel; Bond, Vincent; Bongiovanni, Antonella; Borràs, Francesc E.; Buée, Luc; Buzás, Edit I.; Cheng, Lesley; Clayton, Aled; Cocucci, Emanuele; Dela Cruz, Charles S.; Desiderio, Dominic M.; Di Vizio, Dolores; Ekström, Karin; Falcon-Perez, Juan M.; Gardiner, Chris; Giebel, Bernd; Greening, David W.; Christina Gross, Julia; Gupta, Dwijendra; Hendrix, An; Hill, Andrew F.; Hill, Michelle M.; Nolte-'T Hoen, Esther; Hwang, Do Won; Inal, Jameel; Jagannadham, Medicharla V.; Jayachandran, Muthuvel; Jee, Young Koo; Jørgensen, Malene; Kim, Kwang Pyo; Kim, Yoon Keun; Kislinger, Thomas; Lässer, Cecilia; Lee, Dong Soo; Lee, Hakmo; Van Leeuwen, Johannes; Lener, Thomas; Liu, Ming Lin; Lötvall, Jan; Marcilla, Antonio; Mathivanan, Suresh; Möller, Andreas; Morhayim, Jess; Mullier, Francois; Nazarenko, Irina; Nieuwland, Rienk; Nunes, Diana N.; Pang, Ken; Park, Jaesung; Patel, Tushar; Pocsfalvi, Gabriella; Del Portillo, Hernando; Putz, Ulrich; Ramirez, Marcel I.; Rodrigues, Marcio L.; Roh, Tae Young; Royo, Felix; Sahoo, Susmita; Schiffelers, Raymond; Sharma, Shivani; Siljander, Pia; Simpson, Richard J.; Soekmadji, Carolina; Stahl, Philip; Stensballe, Allan; Stepień, Ewa; Tahara, Hidetoshi; Trummer, Arne; Valadi, Hadi; Vella, Laura J.; Wai, Sun Nyunt; Witwer, Kenneth; Yánez-Mó, Maria; Youn, Hyewon; Zeidler, Reinhard; Gho, Yong Song

    2015-01-01

    Motivation: Extracellular vesicles (EVs) are spherical bilayered proteolipids, harboring various bioactive molecules. Due to the complexity of the vesicular nomenclatures and components, online searches for EV-related publications and vesicular components are currently challenging. Results: We

  13. The role of extracellular histones in haematological disorders.

    Science.gov (United States)

    Alhamdi, Yasir; Toh, Cheng-Hock

    2016-06-01

    Over the past decades, chromosomal alterations have been extensively investigated for their pathophysiological relevance in haematological malignancies. In particular, epigenetic modifications of intra-nuclear histones are now known as key regulators of healthy cell cycles that have also evolved into novel therapeutic targets for certain blood cancers. Thus, for most haematologists, histones are DNA-chained proteins that are buried deep within chromatin. However, the plot has deepened with recent revelations on the function of histones when unchained and released extracellularly upon cell death or from activated neutrophils as part of neutrophil extracellular traps (NETs). Extracellular histones and NETs are increasingly recognized for profound cytotoxicity and pro-coagulant effects. This article highlights the importance of recognizing this new paradigm of extracellular histones as a key player in host defence through its damage-associated molecular patterns, which could translate into novel diagnostic and therapeutic biomarkers in various haematological and critical disorders. © 2016 John Wiley & Sons Ltd.

  14. In vivo extracellular matrix protein expression by human periodontal ...

    African Journals Online (AJOL)

    ONOS

    2010-08-23

    Aug 23, 2010 ... Extracellular matrix proteins (ECM) are described as molecular regulators of these events. ..... zation and adhesive interaction of cells (Yamada, 1983). .... periodontal ligament fibroblasts after simulation of orthodontic force.

  15. Dopamine transporters govern diurnal variation in extracellular dopamine tone

    OpenAIRE

    Ferris, Mark J.; España, Rodrigo A.; Locke, Jason L.; Konstantopoulos, Joanne K.; Rose, Jamie H.; Chen, Rong; Jones, Sara R.

    2014-01-01

    The mechanism for diurnal (i.e., light/dark) oscillations in extracellular dopamine tone in mesolimbic and nigrostriatal systems is unknown. This is because, unlike other neurotransmitter systems, variation in dopamine tone does not correlate with variation in dopamine cell firing. The current research pinpoints the dopamine transporter as a critical governor of diurnal variation in both extracellular dopamine tone and the intracellular availability of releasable dopamine. These data describe...

  16. The extracellular matrix of plants: Molecular, cellular and developmental biology

    Energy Technology Data Exchange (ETDEWEB)

    NONE

    1996-12-31

    A symposium entitled ``The Extracellular Matrix of Plants: Molecular, Cellular and Developmental Biology was held in Tamarron, Colorado, March 15--21, 1996. The following topics were explored in addresses by 43 speakers: structure and biochemistry of cell walls; biochemistry, molecular biology and biosynthesis of lignin; secretory pathway and synthesis of glycoproteins; biosynthesis of matrix polysaccharides, callose and cellulose; role of the extracellular matrix in plant growth and development; plant cell walls in symbiosis and pathogenesis.

  17. Effects of ionizing radiation on extracellular matrix

    Energy Technology Data Exchange (ETDEWEB)

    Mohamed, F. [School of Physics, University of Exeter, Exeter EX44QL (United Kingdom)], E-mail: f.mohamed@ex.ac.uk; Bradley, D.A. [Department of Physics, University of Surrey, Guildford GU72XH (United Kingdom); Winlove, C.P. [School of Physics, University of Exeter, Exeter EX44QL (United Kingdom)

    2007-09-21

    The extracellular matrix is a ubiquitous and important component of tissues. We investigated the effects of ionizing radiation on the physical properties of its principal macromolecular components, pericardial collagen, ligament elastin and hyaluronan, a representative glycosaminoglycan. Samples were exposed to X-rays from an electron linear accelerator in the range of 10-100 Gy to cover the range of irradiation exposure during radiotherapy. A uniaxial mechanical testing protocol was used to characterize the fibrous proteins. For pericardial tissue the major change was an increase in the elastic modulus in the toe region of the curve ({<=}20% strain), from 23{+-}18 kPa for controls to 57{+-}22 kPa at a dose of 10 Gy (p=0.01, {alpha}=0.05). At larger strain ({>=}20% strain), the elastic modulus in the linear region decreased from 1.92{+-}0.70 MPa for control pericardium tissue to 1.31{+-}0.56 MPa (p=0.01, {alpha}=0.05) for 10 Gy X-irradiated sample. Similar observations have been made previously on tendon collagen at larger strains. For elastin, the stress-strain relationship was linear up to 30% strain, but the elastic modulus decreased significantly with irradiation (controls 626{+-}65 kPa, irradiated 474{+-}121 kPa (p=0.02, {alpha}=0.05), at 10 Gy X-irradiation). The results suggest that for collagen the primary effect of irradiation is generation of additional cross-links, while for elastin chain scissions are important. The viscosity of HA (at 1.25% w/v and 0.125% w/v) was measured by both cone and plate and capillary viscometry, the former providing measurement at uniform shear rate and the latter providing a more sensitive indication of changes at low viscosity. Both techniques revealed a dose-dependent reduction in viscosity (from 3400{+-}194 cP for controls to 1500{+-}88 cP at a shear rate of 2 s{sup -1} and dose of 75 Gy), again suggesting depolymerization.

  18. Effects of ionizing radiation on extracellular matrix

    International Nuclear Information System (INIS)

    Mohamed, F.; Bradley, D.A.; Winlove, C.P.

    2007-01-01

    The extracellular matrix is a ubiquitous and important component of tissues. We investigated the effects of ionizing radiation on the physical properties of its principal macromolecular components, pericardial collagen, ligament elastin and hyaluronan, a representative glycosaminoglycan. Samples were exposed to X-rays from an electron linear accelerator in the range of 10-100 Gy to cover the range of irradiation exposure during radiotherapy. A uniaxial mechanical testing protocol was used to characterize the fibrous proteins. For pericardial tissue the major change was an increase in the elastic modulus in the toe region of the curve (≤20% strain), from 23±18 kPa for controls to 57±22 kPa at a dose of 10 Gy (p=0.01, α=0.05). At larger strain (≥20% strain), the elastic modulus in the linear region decreased from 1.92±0.70 MPa for control pericardium tissue to 1.31±0.56 MPa (p=0.01, α=0.05) for 10 Gy X-irradiated sample. Similar observations have been made previously on tendon collagen at larger strains. For elastin, the stress-strain relationship was linear up to 30% strain, but the elastic modulus decreased significantly with irradiation (controls 626±65 kPa, irradiated 474±121 kPa (p=0.02, α=0.05), at 10 Gy X-irradiation). The results suggest that for collagen the primary effect of irradiation is generation of additional cross-links, while for elastin chain scissions are important. The viscosity of HA (at 1.25% w/v and 0.125% w/v) was measured by both cone and plate and capillary viscometry, the former providing measurement at uniform shear rate and the latter providing a more sensitive indication of changes at low viscosity. Both techniques revealed a dose-dependent reduction in viscosity (from 3400±194 cP for controls to 1500±88 cP at a shear rate of 2 s -1 and dose of 75 Gy), again suggesting depolymerization

  19. Glioblastoma extracellular vesicles: reservoirs of potential biomarkers

    Directory of Open Access Journals (Sweden)

    Redzic JS

    2014-02-01

    Full Text Available Jasmina S Redzic,1 Timothy H Ung,2 Michael W Graner2 1Skaggs School of Pharmacy and Pharmaceutical Sciences, 2Department of Neurosurgery, School of Medicine, University of Colorado Denver, Aurora, CO, USA Abstract: Glioblastoma multiforme (GBM is the most frequent and most devastating of the primary central nervous system tumors, with few patients living beyond 2 years postdiagnosis. The damage caused by the disease and our treatments for the patients often leave them physically and cognitively debilitated. Generally, GBMs appear after very short clinical histories and are discovered by imaging (using magnetic resonance imaging [MRI], and the diagnosis is validated by pathology, following surgical resection. The treatment response and diagnosis of tumor recurrence are also tracked by MRI, but there are numerous problems encountered with these monitoring modalities, such as ambiguous interpretation and forms of pseudoprogression. Diagnostic, prognostic, and predictive biomarkers would be an immense boon in following treatment schemes and in determining recurrence, which often requires an invasive intracranial biopsy to verify imaging data. Extracellular vesicles (EVs are stable, membrane-enclosed, virus-sized particles released from either the cell surface or from endosomal pathways that lead to the systemic release of EVs into accessible biofluids, such as serum/plasma, urine, cerebrospinal fluid, and saliva. EVs carry a wide variety of proteins, nucleic acids, lipids, and other metabolites, with many common features but with enough individuality to be able to identify the cell of origin of the vesicles. These components, if properly interrogated, could allow for the identification of tumor-derived EVs in biofluids, indicating tumor progression, relapse, or treatment failure. That knowledge would allow clinicians to continue with treatment regimens that were actually effective or to change course if the therapies were failing. Here, we review

  20. The Role of Extracellular Histones in Influenza Virus Pathogenesis.

    Science.gov (United States)

    Ashar, Harshini K; Mueller, Nathan C; Rudd, Jennifer M; Snider, Timothy A; Achanta, Mallika; Prasanthi, Maram; Pulavendran, Sivasami; Thomas, Paul G; Ramachandran, Akhilesh; Malayer, Jerry R; Ritchey, Jerry W; Rajasekhar, Rachakatla; Chow, Vincent T K; Esmon, Charles T; Teluguakula, Narasaraju

    2018-01-01

    Although exaggerated host immune responses have been implicated in influenza-induced lung pathogenesis, the etiologic factors that contribute to these events are not completely understood. We previously demonstrated that neutrophil extracellular traps exacerbate pulmonary injury during influenza pneumonia. Histones are the major protein components of neutrophil extracellular traps and are known to have cytotoxic effects. Here, we examined the role of extracellular histones in lung pathogenesis during influenza. Mice infected with influenza virus displayed high accumulation of extracellular histones, with widespread pulmonary microvascular thrombosis. Occluded pulmonary blood vessels with vascular thrombi often exhibited endothelial necrosis surrounded by hemorrhagic effusions and pulmonary edema. Histones released during influenza induced cytotoxicity and showed strong binding to platelets within thrombi in infected mouse lungs. Nasal wash samples from influenza-infected patients also showed increased accumulation of extracellular histones, suggesting a possible clinical relevance of elevated histones in pulmonary injury. Although histones inhibited influenza growth in vitro, in vivo treatment with histones did not yield antiviral effects and instead exacerbated lung pathology. Blocking with antihistone antibodies caused a marked decrease in lung pathology in lethal influenza-challenged mice and improved protection when administered in combination with the antiviral agent oseltamivir. These findings support the pathogenic effects of extracellular histones in that pulmonary injury during influenza was exacerbated. Targeting histones provides a novel therapeutic approach to influenza pneumonia. Copyright © 2018 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.

  1. TENCompetence Domain Model

    NARCIS (Netherlands)

    2006-01-01

    This is the version 1.1 of the TENCompetence Domain Model (version 1.0 released at 19-6-2006; version 1.1 at 9-11-2008). It contains several files: a) a pdf with the model description, b) three jpg files with class models (also in the pdf), c) a MagicDraw zip file with the model itself, d) a release

  2. Phosphorylation of carbonic anhydrase IX controls its ability to mediate extracellular acidification in hypoxic tumors.

    Science.gov (United States)

    Ditte, Peter; Dequiedt, Franck; Svastova, Eliska; Hulikova, Alzbeta; Ohradanova-Repic, Anna; Zatovicova, Miriam; Csaderova, Lucia; Kopacek, Juraj; Supuran, Claudiu T; Pastorekova, Silvia; Pastorek, Jaromir

    2011-12-15

    In the hypoxic regions of a tumor, carbonic anhydrase IX (CA IX) is an important transmembrane component of the pH regulatory machinery that participates in bicarbonate transport. Because tumor pH has implications for growth, invasion, and therapy, determining the basis for the contributions of CA IX to the hypoxic tumor microenvironment could lead to new fundamental and practical insights. Here, we report that Thr443 phosphorylation at the intracellular domain of CA IX by protein kinase A (PKA) is critical for its activation in hypoxic cells, with the fullest activity of CA IX also requiring dephosphorylation of Ser448. PKA is activated by cAMP, which is elevated by hypoxia, and we found that attenuating PKA in cells disrupted CA IX-mediated extracellular acidification. Moreover, following hypoxia induction, CA IX colocalized with the sodium-bicarbonate cotransporter and other PKA substrates in the leading edge membranes of migrating tumor cells, in support of the concept that bicarbonate metabolism is spatially regulated at cell surface sites with high local ion transport and pH control. Using chimeric CA IX proteins containing heterologous catalytic domains derived from related CA enzymes, we showed that CA IX activity was modulated chiefly by the intracellular domain where Thr443 is located. Our findings indicate that CA IX is a pivotal mediator of the hypoxia-cAMP-PKA axis, which regulates pH in the hypoxic tumor microenvironment.

  3. SH2 Domain Histochemistry.

    Science.gov (United States)

    Buhs, Sophia; Nollau, Peter

    2017-01-01

    Among posttranslational modifications, the phosphorylation of tyrosine residues is a key modification in cell signaling. Because of its biological importance, characterization of the cellular state of tyrosine phosphorylation is of great interest. Based on the unique properties of endogenously expressed SH2 domains recognizing tyrosine phosphorylated signaling proteins with high specificity we have developed an alternative approach, coined SH2 profiling, enabling us to decipher complex patterns of tyrosine phosphorylation in various normal and cancerous tissues. So far, SH2 profiling has largely been applied for the analysis of protein extracts with the limitation that information on spatial distribution and intensity of tyrosine phosphorylation within a tissue is lost. Here, we describe a novel SH2 domain based strategy for differential characterization of the state of tyrosine phosphorylation in formaldehyde-fixed and paraffin-embedded tissues. This approach demonstrates that SH2 domains may serve as very valuable tools for the analysis of the differential state of tyrosine phosphorylation in primary tissues fixed and processed under conditions frequently applied by routine pathology laboratories.

  4. Molecular Basis of the Extracellular Ligands Mediated Signaling by the Calcium Sensing Receptor

    Directory of Open Access Journals (Sweden)

    Chen Zhang

    2016-09-01

    Full Text Available Ca2+-sensing receptors (CaSRs play a central role in regulating extracellular calcium concentration ([Ca2+]o homeostasis and many (pathophysiological processes in multiple organs. This regulation is orchestrated by a cooperative response to extracellular stimuli such as small changes in Ca2+, Mg2+, amino acids and other ligands. In addition, CaSR is a pleiotropic receptor regulating several intracellular signaling pathways, including calcium mobilization and intracellular calcium oscillation. Nearly 200 mutations and polymorphisms have been found in CaSR in relation to a variety of human disorders associated with abnormal Ca2+ homeostasis. In this review, we summarize efforts directed at identifying binding sites for calcium and amino acids. Both homotropic cooperativity among multiple calcium binding sites and heterotropic cooperativity between calcium and amino acid were revealed using computational modeling, predictions, and site-directed mutagenesis coupled with functional assays. The hinge region of the bilobed Venus flytrap (VFT domain of CaSR plays a pivotal role in coordinating multiple extracellular stimuli, leading to cooperative responses from the receptor. We further highlight the extensive number of disease-associated mutations that have also been shown to affect CaSR’s cooperative action via several types of mechanisms. These results provide insights into the molecular bases of the structure and functional cooperativity of this receptor and other members of family C of the G protein-coupled receptors (cGPCRs in health and disease states, and may assist in the prospective development of novel receptor-based therapeutics.

  5. Perfusion MRI before and after acetazolamide administration for assessment of cerebrovascular reserve capacity in patients with symptomatic internal carotid artery (ICA) occlusion: comparison with {sup 99m}Tc-ECD SPECT

    Energy Technology Data Exchange (ETDEWEB)

    Ma, J.; Holtmannspoetter, M.; Brueckmann, H. [University of Munich, Department of Neuroradiology, Klinikum Grosshadern, Munich (Germany); Mehrkens, J.H.; Schmid-Elsaesser, R. [University of Munich, Department of Neurosurgery, Klinikum Grosshadern, Munich (Germany); Linke, R. [University of Munich, Department of Nuclear Medicine, Klinikum Grosshadern, Munich (Germany); Steiger, H.J. [Heinrich-Heine University, Department of Neurosurgery, Duesseldorf (Germany); Bruening, R. [University of Munich, Department of Neuroradiology, Klinikum Grosshadern, Munich (Germany); Asklepios Klinik Barmbek, Department of Radiology, Hamburg (Germany)

    2007-04-15

    Impaired cerebral vascular reserve (CVR) in patients with symptomatic internal carotid artery (ICA) occlusion is regarded as a possible indication for performing extra-/intracranial (EC/IC) bypass surgery. As perfusion MR imaging (MRI) can demonstrate cerebral haemodynamics at capillary level, our hypothesis was that perfusion MRI could be used in these patients for the evaluation of CVR following acetazolamide challenge in a similar way to single photon emission CT (SPECT) and might provide additional information. Enrolled in the study were 12 patients (mean age 61.3 years; 11 male, 1 female) with symptomatic unilateral ICA occlusion proven by angiography. Both perfusion MRI and 99m-technetium-ethyl-cysteinate dimer ({sup 99m}Tc-ECD) SPECT were performed before and after injection of acetazolamide (Diamox,1000 mg i.v.). CVR parameters including regional cerebral blood flow (rCBF) and volume (rCBV), and mean transit times (MTT) were measured by perfusion MRI. The patients with impaired CVR proven by SPECT (n = 9) had a negative mean rCBF increment (-46.52%), negative rCBV increment (-13.5%) and delayed MTT (mean +2.98 s), respectively, on the occluded side (Student's t-test all P < 0.05). The patients with sufficient CVR (n = 3) had a mean rCBF increment of 1.2%, a decrement of rCBV of 10.46%, and a mean MTT shortening of 0.27 s following the acetazolamide injection. Perfusion MRI before and after acetazolamide administration compares favourably with {sup 99m}Tc-ECD SPECT for the detection of impaired CVR. The impact that perfusion MRI studies (before and after acetazolamide administration) might have on the treatment decision in patients with ICA occlusion has yet to be determined by a prospective study. (orig.)

  6. Highly efficient gene targeting in Aspergillus oryzae industrial strains under ligD mutation introduced by genome editing: Strain-specific differences in the effects of deleting EcdR, the negative regulator of sclerotia formation.

    Science.gov (United States)

    Nakamura, Hidetoshi; Katayama, Takuya; Okabe, Tomoya; Iwashita, Kazuhiro; Fujii, Wataru; Kitamoto, Katsuhiko; Maruyama, Jun-Ichi

    2017-07-11

    Numerous strains of Aspergillus oryzae are industrially used for Japanese traditional fermentation and for the production of enzymes and heterologous proteins. In A. oryzae, deletion of the ku70 or ligD genes involved in non-homologous end joining (NHEJ) has allowed high gene targeting efficiency. However, this strategy has been mainly applied under the genetic background of the A. oryzae wild strain RIB40, and it would be laborious to delete the NHEJ genes in many A. oryzae industrial strains, probably due to their low gene targeting efficiency. In the present study, we generated ligD mutants from the A. oryzae industrial strains by employing the CRISPR/Cas9 system, which we previously developed as a genome editing method. Uridine/uracil auxotrophic strains were generated by deletion of the pyrG gene, which was subsequently used as a selective marker. We examined the gene targeting efficiency with the ecdR gene, of which deletion was reported to induce sclerotia formation under the genetic background of the strain RIB40. As expected, the deletion efficiencies were high, around 60~80%, in the ligD mutants of industrial strains. Intriguingly, the effects of the ecdR deletion on sclerotia formation varied depending on the strains, and we found sclerotia-like structures under the background of the industrial strains, which have never been reported to form sclerotia. The present study demonstrates that introducing ligD mutation by genome editing is an effective method allowing high gene targeting efficiency in A. oryzae industrial strains.

  7. [Inhibitory proteins of neuritic regeneration in the extracellular matrix: structure, molecular interactions and their functions. Mechanisms of extracellular balance].

    Science.gov (United States)

    Vargas, Javier; Uribe-Escamilla, Rebeca; Alfaro-Rodríguez, Alfonso

    2013-01-01

    After injury of the central nervous system (CNS) in higher vertebrates, neurons neither grow nor reconnect with their targets because their axons or dendrites cannot regenerate within the injured site. In the CNS, the signal from the environment regulating neurite regeneration is not exclusively generated by one molecular group. This signal is generated by the interaction of various types of molecules such as extracellular matrix proteins, soluble factors and surface membrane molecules; all these elements interact with one another generating the matrix's biological state: the extracellular balance. Proteins in the balanced extracellular matrix, support and promote cellular physiological states, including neuritic regeneration. We have reviewed three types of proteins of the extracellular matrix possessing an inhibitory effect and that are determinant of neuritic regeneration failure in the CNS: chondroitin sulfate proteoglycans, keratan sulfate proteoglycans and tenascin. We also review some of the mechanisms involved in the balance of extracellular proteins such as isomerization, epimerization, sulfation and glycosylation as well as the assemblage of the extracellular matrix, the interaction between the matrix and soluble factors and its proteolytic degradation. In the final section, we have presented some examples of the matrix's role in development and in tumor propagation.

  8. Extracellular vesicle associated long non-coding RNAs functionally enhance cell viability

    Directory of Open Access Journals (Sweden)

    Chris Hewson

    2016-10-01

    Full Text Available Cells communicate with one another to create microenvironments and share resources. One avenue by which cells communicate is through the action of exosomes. Exosomes are extracellular vesicles that are released by one cell and taken up by neighbouring cells. But how exosomes instigate communication between cells has remained largely unknown. We present evidence here that particular long non-coding RNA molecules are preferentially packaged into exosomes. We also find that a specific class of these exosome associated non-coding RNAs functionally modulate cell viability by direct interactions with l-lactate dehydrogenase B (LDHB, high-mobility group protein 17 (HMG-17, and CSF2RB, proteins involved in metabolism, nucleosomal architecture and cell signalling respectively. Knowledge of this endogenous cell to cell pathway, those proteins interacting with exosome associated non-coding transcripts and their interacting domains, could lead to a better understanding of not only cell to cell interactions but also the development of exosome targeted approaches in patient specific cell-based therapies. Keywords: Non-coding RNA, Extracellular RNA, Exosomes, Retroelement, Pseudogene

  9. Ligand-specific regulation of the extracellular surface of a G-protein-coupled receptor

    Energy Technology Data Exchange (ETDEWEB)

    Bokoch, Michael P.; Zou, Yaozhong; Rasmussen, Søren G.F.; Liu, Corey W.; Nygaard, Rie; Rosenbaum, Daniel M.; Fung, Juan José; Choi, Hee-Jung; Thian, Foon Sun; Kobilka, Tong Sun; Puglisi, Joseph D.; Weis, William I.; Pardo, Leonardo; Prosser, R. Scott; Mueller, Luciano; Kobilka, Brian K. (Stanford-MED); (Toronto); (BMS); (UAB, Spain)

    2010-01-14

    G-protein-coupled receptors (GPCRs) are seven-transmembrane proteins that mediate most cellular responses to hormones and neurotransmitters. They are the largest group of therapeutic targets for a broad spectrum of diseases. Recent crystal structures of GPCRs have revealed structural conservation extending from the orthosteric ligand-binding site in the transmembrane core to the cytoplasmic G-protein-coupling domains. In contrast, the extracellular surface (ECS) of GPCRs is remarkably diverse and is therefore an ideal target for the discovery of subtype-selective drugs. However, little is known about the functional role of the ECS in receptor activation, or about conformational coupling of this surface to the native ligand-binding pocket. Here we use NMR spectroscopy to investigate ligand-specific conformational changes around a central structural feature in the ECS of the {beta}{sub 2} adrenergic receptor: a salt bridge linking extracellular loops 2 and 3. Small-molecule drugs that bind within the transmembrane core and exhibit different efficacies towards G-protein activation (agonist, neutral antagonist and inverse agonist) also stabilize distinct conformations of the ECS. We thereby demonstrate conformational coupling between the ECS and the orthosteric binding site, showing that drugs targeting this diverse surface could function as allosteric modulators with high subtype selectivity. Moreover, these studies provide a new insight into the dynamic behaviour of GPCRs not addressable by static, inactive-state crystal structures.

  10. Chondrocyte secreted CRTAC1: a glycosylated extracellular matrix molecule of human articular cartilage.

    Science.gov (United States)

    Steck, Eric; Bräun, Jessica; Pelttari, Karoliina; Kadel, Stephanie; Kalbacher, Hubert; Richter, Wiltrud

    2007-01-01

    Cartilage acidic protein 1 (CRTAC1), a novel human marker which allowed discrimination of human chondrocytes from osteoblasts and mesenchymal stem cells in culture was so far studied only on the RNA-level. We here describe its genomic organisation and detect a new brain expressed (CRTAC1-B) isoform resulting from alternate last exon usage which is highly conserved in vertebrates. In humans, we identify an exon sharing process with the neighbouring tail-to-tail orientated gene leading to CRTAC1-A. This isoform is produced by cultured human chondrocytes, localized in the extracellular matrix of articular cartilage and its secretion can be stimulated by BMP4. Of five putative O-glycosylation motifs in the last exon of CRTAC1-A, the most C-terminal one is modified according to exposure of serial C-terminal deletion mutants to the O-glycosylation inhibitor Benzyl-alpha-GalNAc. Both isoforms contain four FG-GAP repeat domains and an RGD integrin binding motif, suggesting cell-cell or cell-matrix interaction potential. In summary, CRTAC1 acquired an alternate last exon from the tail-to-tail oriented neighbouring gene in humans resulting in the glycosylated isoform CRTAC1-A which represents a new extracellular matrix molecule of articular cartilage.

  11. Skeletal muscle expresses the extracellular cyclic AMP–adenosine pathway

    Science.gov (United States)

    Chiavegatti, T; Costa, V L; Araújo, M S; Godinho, R O

    2007-01-01

    Background and purpose: cAMP is a key intracellular signalling molecule that regulates multiple processes of the vertebrate skeletal muscle. We have shown that cAMP can be actively pumped out from the skeletal muscle cell. Since in other tissues, cAMP efflux had been associated with extracellular generation of adenosine, in the present study we have assessed the fate of interstitial cAMP and the existence of an extracellular cAMP-adenosine signalling pathway in skeletal muscle. Experimental approach: cAMP efflux and/or its extracellular degradation were analysed by incubating rat cultured skeletal muscle with exogenous cAMP, forskolin or isoprenaline. cAMP and its metabolites were quantified by radioassay or HPLC, respectively. Key results: Incubation of cells with exogenous cAMP was followed by interstitial accumulation of 5′-AMP and adenosine, a phenomenon inhibited by selective inhibitors of ecto-phosphodiesterase (DPSPX) and ecto-nucleotidase (AMPCP). Activation of adenylyl cyclase (AC) in cultured cells with forskolin or isoprenaline increased cAMP efflux and extracellular generation of 5′-AMP and adenosine. Extracellular cAMP-adenosine pathway was also observed after direct and receptor-dependent stimulation of AC in rat extensor muscle ex vivo. These events were attenuated by probenecid, an inhibitor of ATP binding cassette family transporters. Conclusions and implications: Our results show the existence of an extracellular biochemical cascade that converts cAMP into adenosine. The functional relevance of this extracellular signalling system may involve a feedback modulation of cellular response initiated by several G protein-coupled receptor ligands, amplifying cAMP influence to a paracrine mode, through its metabolite, adenosine. PMID:18157164

  12. Domain decomposition method for solving elliptic problems in unbounded domains

    International Nuclear Information System (INIS)

    Khoromskij, B.N.; Mazurkevich, G.E.; Zhidkov, E.P.

    1991-01-01

    Computational aspects of the box domain decomposition (DD) method for solving boundary value problems in an unbounded domain are discussed. A new variant of the DD-method for elliptic problems in unbounded domains is suggested. It is based on the partitioning of an unbounded domain adapted to the given asymptotic decay of an unknown function at infinity. The comparison of computational expenditures is given for boundary integral method and the suggested DD-algorithm. 29 refs.; 2 figs.; 2 tabs

  13. Interaction with Shc prevents aberrant Erk activation in the absence of extracellular stimuli

    KAUST Repository

    Suen, KinMan

    2013-05-01

    Control mechanisms that prevent aberrant signaling are necessary to maintain cellular homeostasis. We describe a new mechanism by which the adaptor protein Shc directly binds the MAP kinase Erk, thus preventing its activation in the absence of extracellular stimuli. The Shc-Erk complex restricts Erk nuclear translocation, restraining Erk-dependent transcription of genes, including those responsible for oncogenic growth. The complex forms through unique binding sites on both the Shc PTB domain and the N-terminal lobe of Erk. Upon receptor tyrosine kinase stimulation, a conformational change within Shc - induced through interaction with the phosphorylated receptor - releases Erk, allowing it to fulfill its role in signaling. Thus, in addition to its established role in promoting MAP kinase signaling in stimulated cells, Shc negatively regulates Erk activation in the absence of growth factors and thus could be considered a tumor suppressor in human cells. © 2013 Nature America, Inc. All rights reserved.

  14. Interaction with Shc prevents aberrant Erk activation in the absence of extracellular stimuli

    KAUST Repository

    Suen, KinMan; Lin, Chichuan; George, Roger R.; Melo, Fernando A.; Biggs, Eleanor R.; Ahmed, Zamal; Drake, Melanie N.; Arur, Swathi; Arold, Stefan T.; Ladbury, John E S D

    2013-01-01

    Control mechanisms that prevent aberrant signaling are necessary to maintain cellular homeostasis. We describe a new mechanism by which the adaptor protein Shc directly binds the MAP kinase Erk, thus preventing its activation in the absence of extracellular stimuli. The Shc-Erk complex restricts Erk nuclear translocation, restraining Erk-dependent transcription of genes, including those responsible for oncogenic growth. The complex forms through unique binding sites on both the Shc PTB domain and the N-terminal lobe of Erk. Upon receptor tyrosine kinase stimulation, a conformational change within Shc - induced through interaction with the phosphorylated receptor - releases Erk, allowing it to fulfill its role in signaling. Thus, in addition to its established role in promoting MAP kinase signaling in stimulated cells, Shc negatively regulates Erk activation in the absence of growth factors and thus could be considered a tumor suppressor in human cells. © 2013 Nature America, Inc. All rights reserved.

  15. Ligand-specific regulation of the extracellular surface of a G-protein-coupled receptor

    DEFF Research Database (Denmark)

    Bokoch, Michael P; Zou, Yaozhong; Rasmussen, Søren Gøgsig Faarup

    2010-01-01

    extending from the orthosteric ligand-binding site in the transmembrane core to the cytoplasmic G-protein-coupling domains. In contrast, the extracellular surface (ECS) of GPCRs is remarkably diverse and is therefore an ideal target for the discovery of subtype-selective drugs. However, little is known...... conformational coupling between the ECS and the orthosteric binding site, showing that drugs targeting this diverse surface could function as allosteric modulators with high subtype selectivity. Moreover, these studies provide a new insight into the dynamic behaviour of GPCRs not addressable by static, inactive...... about the functional role of the ECS in receptor activation, or about conformational coupling of this surface to the native ligand-binding pocket. Here we use NMR spectroscopy to investigate ligand-specific conformational changes around a central structural feature in the ECS of the beta(2) adrenergic...

  16. Mechanistic understanding of nanoparticles' interactions with extracellular matrix: the cell and immune system.

    Science.gov (United States)

    Engin, Ayse Basak; Nikitovic, Dragana; Neagu, Monica; Henrich-Noack, Petra; Docea, Anca Oana; Shtilman, Mikhail I; Golokhvast, Kirill; Tsatsakis, Aristidis M

    2017-06-24

    Extracellular matrix (ECM) is an extraordinarily complex and unique meshwork composed of structural proteins and glycosaminoglycans. The ECM provides essential physical scaffolding for the cellular constituents, as well as contributes to crucial biochemical signaling. Importantly, ECM is an indispensable part of all biological barriers and substantially modulates the interchange of the nanotechnology products through these barriers. The interactions of the ECM with nanoparticles (NPs) depend on the morphological characteristics of intercellular matrix and on the physical characteristics of the NPs and may be either deleterious or beneficial. Importantly, an altered expression of ECM molecules ultimately affects all biological processes including inflammation. This review critically discusses the specific behavior of NPs that are within the ECM domain, and passing through the biological barriers. Furthermore, regenerative and toxicological aspects of nanomaterials are debated in terms of the immune cells-NPs interactions.

  17. Neutrophil elastase processing of Gelatinase A is mediated by extracellular matrix

    Energy Technology Data Exchange (ETDEWEB)

    Rice, A.; Banda, M.J. [Univ. of California, San Franciso, CA (United States)

    1995-07-18

    Gelatinase A (72-kDa type IV collagenase) is a metalloproteinase that is expressed by many cells in culture and is overexpressed by some tumor cells. It has been suggested that the serine proteinase neutrophil elastase might play a role iii the posttranslational processing of gelatinase A and that noncatalytic interactions between gelatinase A and components of the extracellular matrix might alter potential processing pathways. These questions were addressed with the use of gelatin substrate zymography, gelatinolytic activity assays, and amino acid sequence analysis. We found that neutrophil elastase does proteolytically modify gelatinase A by cleaving at a number of sites within gelatinase A. Sequential treatment of gelatinase A with 4-aminophenylmercuric acetate (APMA) and neutrophil elastase yielded an active gelatinase with a 4-fold increase in gelatinolytic activity. The increased gelatinolytic activity correlated with that of a 40-kDa fragment of gelatinase A. Matrix components altered the proteolytic modifications in gelatinase A that were mediated by neutrophil elastase. In the absence of gelatin, neutrophil elastase destructively degraded gelatinase A by hydrolyzing at least two bonds within the fibronectin-like gelatin-binding domain of gelatinase A. In the presence of gelatin, these two inactivating cleavage sites were protected, and cleavage at a site within the hemopexin-like carboxyl-terminal domain resulted in a truncated yet active gelatinase. The results suggest a regulatory role for extracellular matrix molecules in stabilizing gelatinase A fragments and in altering the availability of sites susceptible to destructive proteolysis by neutrophil elastase. 32 refs., 10 figs.

  18. Akt1 binds focal adhesion kinase via the Akt1 kinase domain independently of the pleckstrin homology domain.

    Science.gov (United States)

    Basson, M D; Zeng, B; Wang, S

    2015-10-01

    Akt1 and focal adhesion kinase (FAK) are protein kinases that play key roles in normal cell signaling. Individually, aberrant expression of these kinases has been linked to a variety of cancers. Together, Akt1/FAK interactions facilitate cancer metastasis by increasing cell adhesion under conditions of increased extracellular pressure. Pathological and iatrogenic sources of pressure arise from tumor growth against constraining stroma or direct perioperative manipulation. We previously reported that 15 mmHg increased extracellular pressure causes Akt1 to both directly interact with FAK and to phosphorylate and activate it. We investigated the nature of the Akt1/FAK binding by creating truncations of recombinant FAK, conjugated to glutathione S-transferase (GST), to pull down full-length Akt1. Western blots probing for Akt1 showed that FAK/Akt1 binding persisted in FAK truncations consisting of only amino acids 1-126, FAK(NT1), which contains the F1 subdomain of its band 4.1, ezrin, radixin, and moesin (FERM) domain. Using FAK(NT1) as bait, we then pulled down truncated versions of recombinant Akt1 conjugated to HA (human influenza hemagglutinin). Probes for GST-FAK(NT1) showed Akt1-FAK binding to occur in the absence of the both the Akt1 (N)-terminal pleckstrin homology (PH) domain and its adjacent hinge region. The Akt1 (C)-terminal regulatory domain was equally unnecessary for Akt1/FAK co-immunoprecipitation. Truncations involving the Akt1 catalytic domain showed that the domain by itself was enough to pull down FAK. Additionally, a fragment spanning from the PH domain to half way through the catalytic domain demonstrated increased FAK binding compared to full length Akt1. These results begin to delineate the Akt1/FAK interaction and can be used to manipulate their force-activated signal interactions. Furthermore, the finding that the N-terminal half of the Akt1 catalytic domain binds so strongly to FAK when cleaved from the rest of the protein may suggest a means

  19. Multiple functions of the von Willebrand Factor A domain in matrilins: secretion, assembly, and proteolysis

    Directory of Open Access Journals (Sweden)

    Kanbe Katsuaki

    2008-06-01

    Full Text Available Abstract The von Willebrand Factor A (vWF A domain is one of the most widely distributed structural modules in cell-matrix adhesive molecules such as intergrins and extracellular matrix proteins. Mutations in the vWF A domain of matrilin-3 cause multiple epiphyseal dysplasia (MED, however the pathological mechanism remains to be determined. Previously we showed that the vWF A domain in matrilin-1 mediates formation of a filamentous matrix network through metal-ion dependent adhesion sites in the domain. Here we show two new functions of the vWF A domain in cartilage-specific matrilins (1 and 3. First, vWF A domain regulates oligomerization of matrilins. Insertion of a vWF A domain into matrilin-3 converts the formation of a mixture of matrilin-3 tetramer, trimer, and dimer into a tetramer only, while deletion of a vWF A domain from matrilin-1 converts the formation of the native matrilin-1 trimer into a mixture of trimer and dimer. Second, the vWF A domain protects matrilin-1 from proteolysis. We identified a latent proteolytic site next to the vWF A2 domain in matrilin-1, which is sensitive to the inhibitors of matrix proteases. Deletion of the abutting vWF A domain results in degradation of matrilin-1, presumably by exposing the adjacent proteolytic site. In addition, we also confirmed the vWF A domain is vital for the secretion of matrilin-3. Secretion of the mutant matrilin-3 harbouring a point mutation within the vWF A domain, as occurred in MED patients, is markedly reduced and delayed, resulting from intracellular retention of the mutant matrilin-3. Taken together, our data suggest that different mutations/deletions of the vWF A domain in matrilins may lead to distinct pathological mechanisms due to the multiple functions of the vWF A domain.

  20. Functional Domain Driven Design

    OpenAIRE

    Herrera Guzmán, Sergio

    2016-01-01

    Las tecnologías están en constante expansión y evolución, diseñando nuevas técnicas para cumplir con su fin. En el desarrollo de software, las herramientas y pautas para la elaboración de productos software constituyen una pieza en constante evolución, necesarias para la toma de decisiones sobre los proyectos a realizar. Uno de los arquetipos para el desarrollo de software es el denominado Domain Driven Design, donde es importante conocer ampliamente el negocio que se desea modelar en form...

  1. Feature-level domain adaptation

    DEFF Research Database (Denmark)

    Kouw, Wouter M.; Van Der Maaten, Laurens J P; Krijthe, Jesse H.

    2016-01-01

    -level domain adaptation (flda), that models the dependence between the two domains by means of a feature-level transfer model that is trained to describe the transfer from source to target domain. Subsequently, we train a domain-adapted classifier by minimizing the expected loss under the resulting transfer...... modeled via a dropout distribution, which allows the classiffier to adapt to differences in the marginal probability of features in the source and the target domain. Our experiments on several real-world problems show that flda performs on par with state-of-the-art domainadaptation techniques.......Domain adaptation is the supervised learning setting in which the training and test data are sampled from different distributions: training data is sampled from a source domain, whilst test data is sampled from a target domain. This paper proposes and studies an approach, called feature...

  2. Compensating for Incomplete Domain Knowledge

    National Research Council Canada - National Science Library

    Scott, Lynn M; Drezner, Steve; Rue, Rachel; Reyes, Jesse

    2007-01-01

    .... First, many senior leader positions require experience in more than one functional or operational domain, but it is difficult to develop a corps of senior leaders with all the required combinations of domain knowledge...

  3. Extracellular Metabolites from Industrial Microalgae and Their Biotechnological Potential

    Directory of Open Access Journals (Sweden)

    Lu Liu

    2016-10-01

    Full Text Available Industrial microalgae, as a big family of promising producers of renewable biomass feedstock, have been commercially exploited for functional food, living feed and feed additives, high-value chemicals in nutraceuticals, cosmeceuticals, and chemical reagents. Recently, microalgae have also been considered as a group that might play an important role in biofuel development and environmental protection. Almost all current products of industrial microalgae are derived from their biomass; however, large amounts of spent cell-free media are available from mass cultivation that is mostly unexploited. In this contribution we discuss that these media, which may contain a remarkable diversity of bioactive substances are worthy to be recovered for further use. Obviously, the extracellular metabolites from industrial microalgae have long been neglected in the development of production methods for valuable metabolites. With the advances in the last ten years, more and more structures and properties from extracellular metabolites have been identified, and the potential utilization over wide fields is attracting attention. Some of these extracellular metabolites can be potentially used as drugs, antioxidants, growth regulators or metal chelators. The purpose of this review is to provide an overview of the known extracellular metabolites from industrial microalgae which might be of commercial interest. The attention mainly focuses on the reports of extracellular bioactive metabolites and their potential application in biotechnology.

  4. In vitro Determination of Extracellular Proteins from Xylella fastidiosa.

    Science.gov (United States)

    Mendes, Juliano S; Santiago, André S; Toledo, Marcelo A S; Horta, Maria A C; de Souza, Alessandra A; Tasic, Ljubica; de Souza, Anete P

    2016-01-01

    The phytopathogen Xylella fastidiosa causes economic losses in important agricultural crops. Xylem vessel occlusion caused by biofilm formation is the major mechanism underlying the pathogenicity of distinct strains of X. fastidiosa . Here, we provide a detailed in vitro characterization of the extracellular proteins of X. fastidiosa . Based on the results, we performed a comparison with a strain J1a12, which cannot induce citrus variegated chlorosis symptoms when inoculated into citrus plants. We then extend this approach to analyze the extracellular proteins of X. fastidiosa in media supplemented with calcium. We verified increases in extracellular proteins concomitant with the days of growth and, consequently, biofilm development (3-30 days). Outer membrane vesicles carrying toxins were identified beginning at 10 days of growth in the 9a5c strain. In addition, a decrease in extracellular proteins in media supplemented with calcium was observed in both strains. Using mass spectrometry, 71 different proteins were identified during 30 days of X. fastidiosa biofilm development, including proteases, quorum-sensing proteins, biofilm formation proteins, hypothetical proteins, phage-related proteins, chaperones, toxins, antitoxins, and extracellular vesicle membrane components.

  5. Ligand binding by PDZ domains

    DEFF Research Database (Denmark)

    Chi, Celestine N.; Bach, Anders; Strømgaard, Kristian

    2012-01-01

    , for example, are particularly rich in these domains. The general function of PDZ domains is to bring proteins together within the appropriate cellular compartment, thereby facilitating scaffolding, signaling, and trafficking events. The many functions of PDZ domains under normal physiological as well...... as pathological conditions have been reviewed recently. In this review, we focus on the molecular details of how PDZ domains bind their protein ligands and their potential as drug targets in this context....

  6. Summarization by domain ontology navigation

    DEFF Research Database (Denmark)

    Andreasen, Troels; Bulskov, Henrik

    2013-01-01

    of the subject. In between these two extremes, conceptual summaries encompass selected concepts derived using background knowledge. We address in this paper an approach where conceptual summaries are provided through a conceptualization as given by an ontology. The ontology guiding the summarization can...... be a simple taxonomy or a generative domain ontology. A domain ontology can be provided by a preanalysis of a domain corpus and can be used to condense improved summaries that better reflects the conceptualization of a given domain....

  7. The conserved WW-domain binding sites in Dystroglycan C-terminus are essential but partially redundant for Dystroglycan function

    DEFF Research Database (Denmark)

    Yatsenko, A S; Kucherenko, M M; Pantoja, M

    2009-01-01

    BACKGROUND: Dystroglycan (Dg) is a transmembrane protein that is a part of the Dystrophin Glycoprotein Complex (DGC) which connects the extracellular matrix to the actin cytoskeleton. The C-terminal end of Dg contains a number of putative SH3, SH2 and WW domain binding sites. The most C...

  8. Imaging hydrated microbial extracellular polymers: Comparative analysis by electron microscopy

    Energy Technology Data Exchange (ETDEWEB)

    Dohnalkova, A.C.; Marshall, M. J.; Arey, B. W.; Williams, K. H.; Buck, E. C.; Fredrickson, J. K.

    2011-01-01

    Microbe-mineral and -metal interactions represent a major intersection between the biosphere and geosphere but require high-resolution imaging and analytical tools for investigating microscale associations. Electron microscopy has been used extensively for geomicrobial investigations and although used bona fide, the traditional methods of sample preparation do not preserve the native morphology of microbiological components, especially extracellular polymers. Herein, we present a direct comparative analysis of microbial interactions using conventional electron microscopy approaches of imaging at room temperature and a suite of cryogenic electron microscopy methods providing imaging in the close-to-natural hydrated state. In situ, we observed an irreversible transformation of the hydrated bacterial extracellular polymers during the traditional dehydration-based sample preparation that resulted in their collapse into filamentous structures. Dehydration-induced polymer collapse can lead to inaccurate spatial relationships and hence could subsequently affect conclusions regarding nature of interactions between microbial extracellular polymers and their environment.

  9. Extracellular matrix as a driver for lung regeneration.

    Science.gov (United States)

    Balestrini, Jenna L; Niklason, Laura E

    2015-03-01

    Extracellular matrix has manifold roles in tissue mechanics, guidance of cellular behavior, developmental biology, and regenerative medicine. Over the past several decades, various pre-clinical and clinical studies have shown that many connective tissues may be replaced and/or regenerated using suitable extracellular matrix scaffolds. More recently, decellularization of lung tissue has shown that gentle removal of cells can leave behind a "footprint" within the matrix that may guide cellular adhesion, differentiation and homing following cellular repopulation. Fundamental issues like understanding matrix composition and micro-mechanics remain difficult to tackle, largely because of a lack of available assays and tools for systematically characterizing intact matrix from tissues and organs. This review will critically examine the role of engineered and native extracellular matrix in tissue and lung regeneration, and provide insights into directions for future research and translation.

  10. ISEV position paper: extracellular vesicle RNA analysis and bioinformatics

    Directory of Open Access Journals (Sweden)

    Andrew F. Hill

    2013-12-01

    Full Text Available Extracellular vesicles (EVs are the collective term for the various vesicles that are released by cells into the extracellular space. Such vesicles include exosomes and microvesicles, which vary by their size and/or protein and genetic cargo. With the discovery that EVs contain genetic material in the form of RNA (evRNA has come the increased interest in these vesicles for their potential use as sources of disease biomarkers and potential therapeutic agents. Rapid developments in the availability of deep sequencing technologies have enabled the study of EV-related RNA in detail. In October 2012, the International Society for Extracellular Vesicles (ISEV held a workshop on “evRNA analysis and bioinformatics.” Here, we report the conclusions of one of the roundtable discussions where we discussed evRNA analysis technologies and provide some guidelines to researchers in the field to consider when performing such analysis.

  11. Multistability in a neuron model with extracellular potassium dynamics

    Science.gov (United States)

    Wu, Xing-Xing; Shuai, J. W.

    2012-06-01

    Experiments show a primary role of extracellular potassium concentrations in neuronal hyperexcitability and in the generation of epileptiform bursting and depolarization blocks without synaptic mechanisms. We adopt a physiologically relevant hippocampal CA1 neuron model in a zero-calcium condition to better understand the function of extracellular potassium in neuronal seizurelike activities. The model neuron is surrounded by interstitial space in which potassium ions are able to accumulate. Potassium currents, Na+-K+ pumps, glial buffering, and ion diffusion are regulatory mechanisms of extracellular potassium. We also consider a reduced model with a fixed potassium concentration. The bifurcation structure and spiking frequency of the two models are studied. We show that, besides hyperexcitability and bursting pattern modulation, the potassium dynamics can induce not only bistability but also tristability of different firing patterns. Our results reveal the emergence of the complex behavior of multistability due to the dynamical [K+]o modulation on neuronal activities.

  12. Extracellular DNA as matrix component in microbial biofilms

    DEFF Research Database (Denmark)

    Chiang, Wen-Chi; Tolker-Nielsen, Tim

    2010-01-01

    Bacteria in nature primarily live in surface-associated communities commonly known as biofilms. Because bacteria in biofilms, in many cases, display tolerance to host immune systems, antibiotics, and biocides, they are often difficult or impossible to eradicate. Biofilm formation, therefore, leads...... to various persistent infections in humans and animals, and to a variety of complications in industry, where solid–water interfaces occur. Knowledge about the molecular mechanisms involved in biofilm formation is necessary for creating strategies to control biofilms. Recent studies have shown...... that extracellular DNA is an important component of the extracellular matrix of microbial biofilms. The present chapter is focussed on extracellular DNA as matrix component in biofilms formed by Pseudomonas aeruginosa as an example from the Gram-negative bacteria, and Streptococcus and Staphylococcus as examples...

  13. Gap junction modulation by extracellular signaling molecules: the thymus model

    Directory of Open Access Journals (Sweden)

    Alves L.A.

    2000-01-01

    Full Text Available Gap junctions are intercellular channels which connect adjacent cells and allow direct exchange of molecules of low molecular weight between them. Such a communication has been described as fundamental in many systems due to its importance in coordination, proliferation and differentiation. Recently, it has been shown that gap junctional intercellular communication (GJIC can be modulated by several extracellular soluble factors such as classical hormones, neurotransmitters, interleukins, growth factors and some paracrine substances. Herein, we discuss some aspects of the general modulation of GJIC by extracellular messenger molecules and more particularly the regulation of such communication in the thymus gland. Additionally, we discuss recent data concerning the study of different neuropeptides and hormones in the modulation of GJIC in thymic epithelial cells. We also suggest that the thymus may be viewed as a model to study the modulation of gap junction communication by different extracellular messengers involved in non-classical circuits, since this organ is under bidirectional neuroimmunoendocrine control.

  14. Extracellular Protease Activity of Enteropathogenic Escherechia coli on Mucin Substrate

    Directory of Open Access Journals (Sweden)

    SRI BUDIARTI

    2007-03-01

    Full Text Available Enteropathogenic Escherichia coli (EPEC causes gastrointestinal infections in human. EPEC invasion was initiated by attachment and aggressive colonization on intestinal surface. Attachment of EPEC alter the intestine mucosal cells. Despite this, the pathogenic mechanism of EPEC infectior has not been fully understood. This research hypothesizes that extracellular proteolytic enzymes is necessary for EPEC colonization. The enzyme is secreted into gastrointestinal milieu and presumably destroy mucus layer cover the gastrointestinal tract. The objective of this study was to assay EPEC extracellular protease enzyme by using mucin substrate. The activity of EPEC extracellular proteolytic enzyme on 1% mucin substrate was investigated. Non-pathogenic E. coli was used as a negative control. Positive and tentative controls were Yersinia enterocolitica and Salmonella. Ten EPEC strains were assayed, seven of them were able to degrade mucin, and the highest activity was produced by K1.1 strain. Both positive and tentative controls also showed the ability to digest 0.20% mucin.

  15. Selenoprotein P: an extracellular protein with unique physical characteristics and a role in selenium homeostasis.

    Science.gov (United States)

    Burk, Raymond F; Hill, Kristina E

    2005-01-01

    Selenoprotein P is an abundant extracellular glycoprotein that is rich in selenocysteine. It has two domains with respect to selenium content. The N-terminal domain of the rat protein contains one selenocysteine residue in a UxxC redox motif. This domain also has a pH-sensitive heparin-binding site and two histidine-rich amino acid stretches. The smaller C-terminal domain contains nine selenocysteine and ten cysteine residues. Four isoforms of selenoprotein P are present in rat plasma. They share the same N terminus and amino acid sequence. One isoform is full length and the three others terminate at the positions of the second, third, and seventh selenocysteine residues. Selenoprotein P turns over rapidly in rat plasma with the consequence that approximately 25% of the amount of whole-body selenium passes through it each day. Evidence supports functions of the protein in selenium homeostasis and oxidant defense. Selenoprotein P knockout mice have very low selenium concentrations in the brain, the testis, and the fetus, with severe pathophysiological consequences in each tissue. In addition, those mice waste moderate amounts of selenium in the urine. Selenoprotein P binds to endothelial cells in the rat, and plasma levels of the protein correlate with prevention of diquat-induced lipid peroxidation and hepatic endothelial cell injury. The mechanisms of these apparent functions remain speculative and much work on the mechanism of selenoprotein P function lies ahead. Measurement of selenoprotein P in human plasma has shown that it is depressed by selenium deficiency and by cirrhosis. Selenium supplementation of selenium-deficient human subjects showed that glutathione peroxidase activity was optimized before selenoprotein P concentration was optimized, indicating that plasma selenoprotein P is the better index of human selenium nutritional status.

  16. The Staphylococcus aureus extracellular adherence protein (Eap) adopts an elongated but structured conformation in solution.

    Science.gov (United States)

    Hammel, Michal; Nemecek, Daniel; Keightley, J Andrew; Thomas, George J; Geisbrecht, Brian V

    2007-12-01

    The extracellular adherence protein (Eap) of Staphylococcus aureus participates in a wide range of protein-protein interactions that facilitate the initiation and dissemination of Staphylococcal disease. In this report, we describe the use of a multidisciplinary approach to characterize the solution structure of full-length Eap. In contrast to previous reports suggesting that a six-domain isoform of Eap undergoes multimerization, sedimentation equilibrium analytical ultracentrifugation data revealed that a four-domain isoform of Eap is a monomer in solution. In vitro proteolysis and solution small angle X-ray scattering studies both indicate that Eap adopts an extended conformation in solution, where the linkers connecting sequential EAP modules are solvent exposed. Construction of a low-resolution model of full-length Eap using a combination of ab initio deconvolution of the SAXS data and rigid body modeling of the EAP domain crystal structure suggests that full-length Eap may present several unique concave surfaces capable of participating in ligand binding. These results also raise the possibility that such surfaces may be held together by additional interactions between adjacent EAP modules. This hypothesis is supported by a comparative Raman spectroscopic analysis of full-length Eap and a stoichiometric solution of the individual EAP modules, which indicates the presence of additional secondary structure and a greater extent of hydrogen/deuterium exchange protection in full-length Eap. Our results provide the first insight into the solution structure of full-length Eap and an experimental basis for interpreting the EAP domain crystal structures within the context of the full-length molecule. They also lay a foundation for future studies into the structural and molecular bases of Eap-mediated protein-protein interactions with its many ligands.

  17. Expansion of protein domain repeats.

    Directory of Open Access Journals (Sweden)

    Asa K Björklund

    2006-08-01

    Full Text Available Many proteins, especially in eukaryotes, contain tandem repeats of several domains from the same family. These repeats have a variety of binding properties and are involved in protein-protein interactions as well as binding to other ligands such as DNA and RNA. The rapid expansion of protein domain repeats is assumed to have evolved through internal tandem duplications. However, the exact mechanisms behind these tandem duplications are not well-understood. Here, we have studied the evolution, function, protein structure, gene structure, and phylogenetic distribution of domain repeats. For this purpose we have assigned Pfam-A domain families to 24 proteomes with more sensitive domain assignments in the repeat regions. These assignments confirmed previous findings that eukaryotes, and in particular vertebrates, contain a much higher fraction of proteins with repeats compared with prokaryotes. The internal sequence similarity in each protein revealed that the domain repeats are often expanded through duplications of several domains at a time, while the duplication of one domain is less common. Many of the repeats appear to have been duplicated in the middle of the repeat region. This is in strong contrast to the evolution of other proteins that mainly works through additions of single domains at either terminus. Further, we found that some domain families show distinct duplication patterns, e.g., nebulin domains have mainly been expanded with a unit of seven domains at a time, while duplications of other domain families involve varying numbers of domains. Finally, no common mechanism for the expansion of all repeats could be detected. We found that the duplication patterns show no dependence on the size of the domains. Further, repeat expansion in some families can possibly be explained by shuffling of exons. However, exon shuffling could not have created all repeats.

  18. Neutrophil extracellular traps promote deep vein thrombosis in mice

    Science.gov (United States)

    Brill, A.; Fuchs, T.A.; Savchenko, A.S.; Thomas, G.M.; Martinod, K.; De Meyer, S.F.; Bhandari, A.A.; Wagner, D.D.

    2011-01-01

    Summary Background Upon activation, neutrophils can release nuclear material known as neutrophil extracellular traps (NETs), which were initially described as a part of antimicrobial defense. Extracellular chromatin was recently reported to be pro-thrombotic in vitro and to accumulate in plasma and thrombi of baboons with experimental deep vein thrombosis (DVT). Objective To explore the source and role of extracellular chromatin in DVT. Methods We used an established murine model of DVT induced by flow restriction (stenosis) in the inferior vena cava (IVC). Results We demonstrate that the levels of extracellular DNA increase in plasma after 6 h IVC stenosis, compared to sham-operated mice. Immunohistochemical staining revealed the presence of Gr-1-positive neutrophils in both red (RBC-rich) and white (platelet-rich) parts of thrombi. Citrullinated histone H3 (CitH3), an element of NETs’ structure, was present only in the red part of thrombi and was frequently associated with the Gr-1 antigen. Immunofluorescent staining of thrombi showed proximity of extracellular CitH3 and von Willebrand factor (VWF), a platelet adhesion molecule crucial for thrombus development in this model. Infusion of Deoxyribonuclease 1 (DNase 1) protected mice from DVT after 6 h and also 48 h IVC stenosis. Infusion of an unfractionated mixture of calf thymus histones increased plasma VWF and promoted DVT early after stenosis application. Conclusions Extracellular chromatin, likely originating from neutrophils, is a structural part of a venous thrombus and both the DNA scaffold and histones appear to contribute to the pathogenesis of DVT in mice. NETs may provide new targets for DVT drug development. PMID:22044575

  19. Brain Extracellular Space: The Final Frontier of Neuroscience.

    Science.gov (United States)

    Nicholson, Charles; Hrabětová, Sabina

    2017-11-21

    Brain extracellular space is the narrow microenvironment that surrounds every cell of the central nervous system. It contains a solution that closely resembles cerebrospinal fluid with the addition of extracellular matrix molecules. The space provides a reservoir for ions essential to the electrical activity of neurons and forms an intercellular chemical communication channel. Attempts to reveal the size and structure of the extracellular space using electron microscopy have had limited success; however, a biophysical approach based on diffusion of selected probe molecules has proved useful. A point-source paradigm, realized in the real-time iontophoresis method using tetramethylammonium, as well as earlier radiotracer methods, have shown that the extracellular space occupies ∼20% of brain tissue and small molecules have an effective diffusion coefficient that is two-fifths that in a free solution. Monte Carlo modeling indicates that geometrical constraints, including dead-space microdomains, contribute to the hindrance to diffusion. Imaging the spread of macromolecules shows them increasingly hindered as a function of size and suggests that the gaps between cells are predominantly ∼40 nm with wider local expansions that may represent dead-spaces. Diffusion measurements also characterize interactions of ions and proteins with the chondroitin and heparan sulfate components of the extracellular matrix; however, the many roles of the matrix are only starting to become apparent. The existence and magnitude of bulk flow and the so-called glymphatic system are topics of current interest and controversy. The extracellular space is an exciting area for research that will be propelled by emerging technologies. Copyright © 2017 Biophysical Society. Published by Elsevier Inc. All rights reserved.

  20. Focus on Extracellular Vesicles: Physiological Role and Signalling Properties of Extracellular Membrane Vesicles

    Directory of Open Access Journals (Sweden)

    Nunzio Iraci

    2016-02-01

    Full Text Available Extracellular vesicles (EVs are a heterogeneous population of secreted membrane vesicles, with distinct biogenesis routes, biophysical properties and different functions both in physiological conditions and in disease. The release of EVs is a widespread biological process, which is conserved across species. In recent years, numerous studies have demonstrated that several bioactive molecules are trafficked with(in EVs, such as microRNAs, mRNAs, proteins and lipids. The understanding of their final impact on the biology of specific target cells remains matter of intense debate in the field. Also, EVs have attracted great interest as potential novel cell-free therapeutics. Here we describe the proposed physiological and pathological functions of EVs, with a particular focus on their molecular content. Also, we discuss the advances in the knowledge of the mechanisms regulating the secretion of EV-associated molecules and the specific pathways activated upon interaction with the target cell, highlighting the role of EVs in the context of the immune system and as mediators of the intercellular signalling in the brain.

  1. Extracellular DNA and lipoteichoic acids interact with exopolysaccharides in the extracellular matrix of Streptococcus mutans biofilms

    Science.gov (United States)

    Castillo Pedraza, Midian C.; Novais, Tatiana F.; Faustoferri, Roberta C.; Quivey, Robert G.; Terekhov, Anton; Hamaker, Bruce R.; Klein, Marlise I.

    2018-01-01

    Streptococcus mutans -derived exopolysaccharides are virulence determinants in the matrix of biofilms that cause caries. Extracellular DNA (eDNA) and lipoteichoic acid (LTA) are found in cariogenic biofilms, but their functions are unclear. Therefore, strains of S. mutans carrying single deletions that would modulate matrix components were used: eDNA – ΔlytS and ΔlytT; LTA – ΔdltA and ΔdltD; and insoluble exopolysaccharide – ΔgtfB. Single-species (parental strain S. mutans UA159 or individual mutant strains) and mixed-species (UA159 or mutant strain, Actinomyces naeslundii and Streptococcus gordonii) biofilms were evaluated. Distinct amounts of matrix components were detected, depending on the inactivated gene. eDNA was found to be cooperative with exopolysaccharide in early phases, while LTA played a larger role in the later phases of biofilm development. The architecture of mutant strains biofilms was distinct (vs UA159), demonstrating that eDNA and LTA influence exopolysaccharide distribution and microcolony organization. Thus, eDNA and LTA may shape exopolysaccharide structure, affecting strategies for controlling pathogenic biofilms. PMID:28946780

  2. Extracellular DNA and lipoteichoic acids interact with exopolysaccharides in the extracellular matrix of Streptococcus mutans biofilms.

    Science.gov (United States)

    Castillo Pedraza, Midian C; Novais, Tatiana F; Faustoferri, Roberta C; Quivey, Robert G; Terekhov, Anton; Hamaker, Bruce R; Klein, Marlise I

    2017-10-01

    Streptococcus mutans-derived exopolysaccharides are virulence determinants in the matrix of biofilms that cause caries. Extracellular DNA (eDNA) and lipoteichoic acid (LTA) are found in cariogenic biofilms, but their functions are unclear. Therefore, strains of S. mutans carrying single deletions that would modulate matrix components were used: eDNA - ∆lytS and ∆lytT; LTA - ∆dltA and ∆dltD; and insoluble exopolysaccharide - ΔgtfB. Single-species (parental strain S. mutans UA159 or individual mutant strains) and mixed-species (UA159 or mutant strain, Actinomyces naeslundii and Streptococcus gordonii) biofilms were evaluated. Distinct amounts of matrix components were detected, depending on the inactivated gene. eDNA was found to be cooperative with exopolysaccharide in early phases, while LTA played a larger role in the later phases of biofilm development. The architecture of mutant strains biofilms was distinct (vs UA159), demonstrating that eDNA and LTA influence exopolysaccharide distribution and microcolony organization. Thus, eDNA and LTA may shape exopolysaccharide structure, affecting strategies for controlling pathogenic biofilms.

  3. Extracellular Electron Uptake: Among Autotrophs and Mediated by Surfaces

    DEFF Research Database (Denmark)

    Tremblay, Pier-Luc; Angenent, Largus T.; Zhang, Tian

    2017-01-01

    Autotrophic microbes can acquire electrons from solid donors such as steel, other microbial cells, or electrodes. Based on this feature, bioprocesses are being developed for the microbial electrosynthesis (MES) of useful products from the greenhouse gas CO2. Extracellular electron-transfer mechan......Autotrophic microbes can acquire electrons from solid donors such as steel, other microbial cells, or electrodes. Based on this feature, bioprocesses are being developed for the microbial electrosynthesis (MES) of useful products from the greenhouse gas CO2. Extracellular electron......; or (iii) mediator-generating enzymes detached from cells. This review explores the interactions of autotrophs with solid electron donors and their importance in nature and for biosustainable technologies....

  4. Syndecans as receptors and organizers of the extracellular matrix

    DEFF Research Database (Denmark)

    Xian, Xiaojie; Gopal, Sandeep; Couchman, John

    2009-01-01

    , the collagens and glycoproteins of the extracellular matrix are prominent. Frequently, they do so in conjunction with other receptors, most notably the integrins. For this reason, they are often referred to as "co-receptors". However, just as with integrins, syndecans can interact with actin-associated proteins...... and signalling molecules, such as protein kinases. Some aspects of syndecan signalling are understood but much remains to be learned. The functions of syndecans in regulating cell adhesion and extracellular matrix assembly are described here. Evidence from null mice suggests that syndecans have roles...

  5. Production of extracellular fatty acid using engineered Escherichia coli

    Directory of Open Access Journals (Sweden)

    Liu Hui

    2012-04-01

    Full Text Available Abstract Background As an alternative for economic biodiesel production, the microbial production of extracellular fatty acid from renewable resources is receiving more concerns recently, since the separation of fatty acid from microorganism cells is normally involved in a series of energy-intensive steps. Many attempts have been made to construct fatty acid producing strains by targeting genes in the fatty acid biosynthetic pathway, while few studies focused on the cultivation process and the mass transfer kinetics. Results In this study, both strain improvements and cultivation process strategies were applied to increase extracellular fatty acid production by engineered Escherichia coli. Our results showed overexpressing ‘TesA and the deletion of fadL in E. coli BL21 (DE3 improved extracellular fatty acid production, while deletion of fadD didn’t strengthen the extracellular fatty acid production for an undetermined mechanism. Moreover, the cultivation process controls contributed greatly to extracellular fatty acid production with respect to titer, cell growth and productivity by adjusting the temperature, adding ampicillin and employing on-line extraction. Under optimal conditions, the E. coli strain (pACY-‘tesA-ΔfadL produced 4.8 g L−1 extracellular fatty acid, with the specific productivity of 0.02 g h−1 g−1dry cell mass, and the yield of 4.4% on glucose, while the ratios of cell-associated fatty acid versus extracellular fatty acid were kept below 0.5 after 15 h of cultivation. The fatty acids included C12:1, C12:0, C14:1, C14:0, C16:1, C16:0, C18:1, C18:0. The composition was dominated by C14 and C16 saturated and unsaturated fatty acids. Using the strain pACY-‘tesA, similar results appeared under the same culture conditions and the titer was also much higher than that ever reported previously, which suggested that the supposedly superior strain did not necessarily perform best for the efficient production of desired

  6. Physical, Spatial, and Molecular Aspects of Extracellular Matrix of In Vivo Niches and Artificial Scaffolds Relevant to Stem Cells Research

    Directory of Open Access Journals (Sweden)

    Maria Akhmanova

    2015-01-01

    Full Text Available Extracellular matrix can influence stem cell choices, such as self-renewal, quiescence, migration, proliferation, phenotype maintenance, differentiation, or apoptosis. Three aspects of extracellular matrix were extensively studied during the last decade: physical properties, spatial presentation of adhesive epitopes, and molecular complexity. Over 15 different parameters have been shown to influence stem cell choices. Physical aspects include stiffness (or elasticity, viscoelasticity, pore size, porosity, amplitude and frequency of static and dynamic deformations applied to the matrix. Spatial aspects include scaffold dimensionality (2D or 3D and thickness; cell polarity; area, shape, and microscale topography of cell adhesion surface; epitope concentration, epitope clustering characteristics (number of epitopes per cluster, spacing between epitopes within cluster, spacing between separate clusters, cluster patterns, and level of disorder in epitope arrangement, and nanotopography. Biochemical characteristics of natural extracellular matrix molecules regard diversity and structural complexity of matrix molecules, affinity and specificity of epitope interaction with cell receptors, role of non-affinity domains, complexity of supramolecular organization, and co-signaling by growth factors or matrix epitopes. Synergy between several matrix aspects enables stem cells to retain their function in vivo and may be a key to generation of long-term, robust, and effective in vitro stem cell culture systems.

  7. Time Domain Induced Polarization

    DEFF Research Database (Denmark)

    Fiandaca, Gianluca; Auken, Esben; Christiansen, Anders Vest

    2012-01-01

    Time-domain-induced polarization has significantly broadened its field of reference during the last decade, from mineral exploration to environmental geophysics, e.g., for clay and peat identification and landfill characterization. Though, insufficient modeling tools have hitherto limited the use...... of time-domaininduced polarization for wider purposes. For these reasons, a new forward code and inversion algorithm have been developed using the full-time decay of the induced polarization response, together with an accurate description of the transmitter waveform and of the receiver transfer function......, to reconstruct the distribution of the Cole-Cole parameters of the earth. The accurate modeling of the transmitter waveform had a strong influence on the forward response, and we showed that the difference between a solution using a step response and a solution using the accurate modeling often is above 100...

  8. Domain architecture conservation in orthologs

    Science.gov (United States)

    2011-01-01

    Background As orthologous proteins are expected to retain function more often than other homologs, they are often used for functional annotation transfer between species. However, ortholog identification methods do not take into account changes in domain architecture, which are likely to modify a protein's function. By domain architecture we refer to the sequential arrangement of domains along a protein sequence. To assess the level of domain architecture conservation among orthologs, we carried out a large-scale study of such events between human and 40 other species spanning the entire evolutionary range. We designed a score to measure domain architecture similarity and used it to analyze differences in domain architecture conservation between orthologs and paralogs relative to the conservation of primary sequence. We also statistically characterized the extents of different types of domain swapping events across pairs of orthologs and paralogs. Results The analysis shows that orthologs exhibit greater domain architecture conservation than paralogous homologs, even when differences in average sequence divergence are compensated for, for homologs that have diverged beyond a certain threshold. We interpret this as an indication of a stronger selective pressure on orthologs than paralogs to retain the domain architecture required for the proteins to perform a specific function. In general, orthologs as well as the closest paralogous homologs have very similar domain architectures, even at large evolutionary separation. The most common domain architecture changes observed in both ortholog and paralog pairs involved insertion/deletion of new domains, while domain shuffling and segment duplication/deletion were very infrequent. Conclusions On the whole, our results support the hypothesis that function conservation between orthologs demands higher domain architecture conservation than other types of homologs, relative to primary sequence conservation. This supports the

  9. Protein domain organisation: adding order

    Directory of Open Access Journals (Sweden)

    Kummerfeld Sarah K

    2009-01-01

    Full Text Available Abstract Background Domains are the building blocks of proteins. During evolution, they have been duplicated, fused and recombined, to produce proteins with novel structures and functions. Structural and genome-scale studies have shown that pairs or groups of domains observed together in a protein are almost always found in only one N to C terminal order and are the result of a single recombination event that has been propagated by duplication of the multi-domain unit. Previous studies of domain organisation have used graph theory to represent the co-occurrence of domains within proteins. We build on this approach by adding directionality to the graphs and connecting nodes based on their relative order in the protein. Most of the time, the linear order of domains is conserved. However, using the directed graph representation we have identified non-linear features of domain organization that are over-represented in genomes. Recognising these patterns and unravelling how they have arisen may allow us to understand the functional relationships between domains and understand how the protein repertoire has evolved. Results We identify groups of domains that are not linearly conserved, but instead have been shuffled during evolution so that they occur in multiple different orders. We consider 192 genomes across all three kingdoms of life and use domain and protein annotation to understand their functional significance. To identify these features and assess their statistical significance, we represent the linear order of domains in proteins as a directed graph and apply graph theoretical methods. We describe two higher-order patterns of domain organisation: clusters and bi-directionally associated domain pairs and explore their functional importance and phylogenetic conservation. Conclusion Taking into account the order of domains, we have derived a novel picture of global protein organization. We found that all genomes have a higher than expected

  10. Protein domain organisation: adding order.

    Science.gov (United States)

    Kummerfeld, Sarah K; Teichmann, Sarah A

    2009-01-29

    Domains are the building blocks of proteins. During evolution, they have been duplicated, fused and recombined, to produce proteins with novel structures and functions. Structural and genome-scale studies have shown that pairs or groups of domains observed together in a protein are almost always found in only one N to C terminal order and are the result of a single recombination event that has been propagated by duplication of the multi-domain unit. Previous studies of domain organisation have used graph theory to represent the co-occurrence of domains within proteins. We build on this approach by adding directionality to the graphs and connecting nodes based on their relative order in the protein. Most of the time, the linear order of domains is conserved. However, using the directed graph representation we have identified non-linear features of domain organization that are over-represented in genomes. Recognising these patterns and unravelling how they have arisen may allow us to understand the functional relationships between domains and understand how the protein repertoire has evolved. We identify groups of domains that are not linearly conserved, but instead have been shuffled during evolution so that they occur in multiple different orders. We consider 192 genomes across all three kingdoms of life and use domain and protein annotation to understand their functional significance. To identify these features and assess their statistical significance, we represent the linear order of domains in proteins as a directed graph and apply graph theoretical methods. We describe two higher-order patterns of domain organisation: clusters and bi-directionally associated domain pairs and explore their functional importance and phylogenetic conservation. Taking into account the order of domains, we have derived a novel picture of global protein organization. We found that all genomes have a higher than expected degree of clustering and more domain pairs in forward and

  11. Prediction Reweighting for Domain Adaptation.

    Science.gov (United States)

    Shuang Li; Shiji Song; Gao Huang

    2017-07-01

    There are plenty of classification methods that perform well when training and testing data are drawn from the same distribution. However, in real applications, this condition may be violated, which causes degradation of classification accuracy. Domain adaptation is an effective approach to address this problem. In this paper, we propose a general domain adaptation framework from the perspective of prediction reweighting, from which a novel approach is derived. Different from the major domain adaptation methods, our idea is to reweight predictions of the training classifier on testing data according to their signed distance to the domain separator, which is a classifier that distinguishes training data (from source domain) and testing data (from target domain). We then propagate the labels of target instances with larger weights to ones with smaller weights by introducing a manifold regularization method. It can be proved that our reweighting scheme effectively brings the source and target domains closer to each other in an appropriate sense, such that classification in target domain becomes easier. The proposed method can be implemented efficiently by a simple two-stage algorithm, and the target classifier has a closed-form solution. The effectiveness of our approach is verified by the experiments on artificial datasets and two standard benchmarks, a visual object recognition task and a cross-domain sentiment analysis of text. Experimental results demonstrate that our method is competitive with the state-of-the-art domain adaptation algorithms.

  12. Structure of the C-type lectin carbohydrate recognition domain of human Tetranectin

    DEFF Research Database (Denmark)

    Kastrup, Jette Sandholm Jensen; Nielsen, Bettina Bryde; Rasmussen, Hanne B.

    1998-01-01

    Tetranectin (TN) is a C-type lectin involved in fibrinolysis, being the only endogenous ligand known to bind specifically to the kringle 4 domain of plasminogen. TN was originally isolated from plasma, but shows a wide tissue distribution. Furthermore, TN has been found in the extracellular matri...... molecules. One sulfate ion has been located at the surface of TN3, forming contacts to Glu120, Lys148, Asn106 of a symmetry-related molecule, and to an ethanol molecule....

  13. Crystallization and preliminary crystallographic analysis of Gibberella zeae extracellular lipase

    International Nuclear Information System (INIS)

    Sun, Yuna; Li, Ming; Zhang, Yan; Liu, Lifang; Liu, Ye; Liu, Zheng; Li, Xumei; Lou, Zhiyong

    2008-01-01

    G. zeae extracellular lipase has been overexpressed, purified and crystallized. Diffraction data were collected to 2.8 Å resolution. Fusarium head blight, one of the most destructive crop diseases, is mainly caused by Fusarium graminearum (known in its sexual stage as Gibberella zeae). F. graminearum secretes various extracellular enzymes that have been hypothesized to be involved in host infection. One of the extracellular enzymes secreted by this organism is the G. zeae extracellular lipase (GZEL), which is encoded by the FGL1 gene. In order to solve the crystal structure of GZEL and to gain a better understanding of the biological functions of the protein and of possible inhibitory mechanisms of lipase inhibitors, recombinant GZEL was crystallized at 291 K using PEG 3350 as a precipitant. A data set was collected to 2.8 Å resolution from a single flash-cooled crystal (100 K). The crystal belonged to space group P2 1 2 1 2 1 , with unit-cell parameters a = 78.4, b = 91.0, c = 195.8 Å, α = β = γ = 90°. The presence of four molecules was assumed per asymmetric unit, which gave a Matthews coefficient of 2.6 Å 3 Da −1

  14. Filtration recovery of extracellular DNA from environmental water samples

    Science.gov (United States)

    qPCR methods are able to analyze DNA from microbes within hours of collecting water samples, providing the promptest notification and public awareness possible when unsafe pathogenic levels are reached. Health risk, however, may be overestimated by the presence of extracellular ...

  15. Glycosylation of extracellular vesicles : current knowledge, tools and clinical perspectives

    NARCIS (Netherlands)

    Williams, Charles; Royo, Felix; Aizpurua-Olaizola, Oier; Pazos, Raquel; Boons, Geert-Jan; Reichardt, Niels-Christian; Falcon-Perez, Juan M

    2018-01-01

    It is now acknowledged that extracellular vesicles (EVs) are important effectors in a vast number of biological processes through intercellular transfer of biomolecules. Increasing research efforts in the EV field have yielded an appreciation for the potential role of glycans in EV function. Indeed,

  16. Extracellular protease produced by Bacillus subtilis isolated from ...

    African Journals Online (AJOL)

    In a study to evaluate the microbiological safety of some paracetamol oral solutions sold in some Nigerian drug stores, 40.0% of the samples examined was contaminated with protease-producing Bacillus subtilis. The production of extracellular protease was induced by casein in the minimal medium and was found to be the ...

  17. Extracellular Membrane Vesicles and Phytopathogenicity of Acholeplasma laidlawii PG8

    Directory of Open Access Journals (Sweden)

    Vladislav M. Chernov

    2012-01-01

    Full Text Available For the first time, the phytopathogenicity of extracellular vesicles of Acholeplasma laidlawii PG8 (a ubiquitous mycoplasma that is one of the five common species of cell culture contaminants and is a causative agent for phytomycoplasmoses in Oryza sativa L. plants was studied. Data on the ability of extracellular vesicles of Acholeplasma laidlawii PG8 to penetrate from the nutrient medium into overground parts of Oryza sativa L. through the root system and to cause alterations in ultrastructural organization of the plants were presented. As a result of the analysis of ultrathin leaf sections of plants grown in medium with A. laidlawii PG8 vesicles, we detected significant changes in tissue ultrastructure characteristic to oxidative stress in plants as well as their cultivation along with bacterial cells. The presence of nucleotide sequences of some mycoplasma genes within extracellular vesicles of Acholeplasma laidlawii PG8 allowed a possibility to use PCR (with the following sequencing to perform differential detection of cells and bacterial vesicles in samples under study. The obtained data may suggest the ability of extracellular vesicles of the mycoplasma to display in plants the features of infection from the viewpoint of virulence criteria—invasivity, infectivity—and toxigenicity—and to favor to bacterial phytopathogenicity.

  18. Optimization of culture media for extracellular expression of ...

    African Journals Online (AJOL)

    Purpose: To investigate the enhancement of streptokinase extracellular expression in Escherichia coli by adjusting culture media. Methods: Screening of 10 chemical factors (EDTA, peptone, glycine, triton X-100, glycerol, K2HPO4,. KH2PO4, Ca2+ (calcium chloride), yeast and NaCl) in order to increase the secretion of ...

  19. Immunological and biochemical characterization of extracellular polysaccharides of mucoralean moulds

    NARCIS (Netherlands)

    Ruiter, de G.A.

    1993-01-01

    In this thesis the characterization is described of the antigenic determinants (epitopes) of the extracellular polysaccharides (EPSs) from moulds belonging to the order of Mucorales. Detailed knowledge of the structure of these epitopes allows for further development of a new generation of

  20. Brain washing : Transport of cerebral extracellular fluids and solutes

    NARCIS (Netherlands)

    Bedussi, B.

    2017-01-01

    Regulation of extracellular volume and fluid composition provides a robust microenvironment for brain cells. In peripheral tissue, fluid surplus and solutes are removed from the interstitium via drainage into lymphatic channels. Since the central nervous system lacks a proper lymphatic vasculature,

  1. Extracellular matrix components direct porcine muscle stem cell behavior

    International Nuclear Information System (INIS)

    Wilschut, Karlijn J.; Haagsman, Henk P.; Roelen, Bernard A.J.

    2010-01-01

    In muscle tissue, extracellular matrix proteins, together with the vasculature system, muscle-residence cells and muscle fibers, create the niche for muscle stem cells. The niche is important in controlling proliferation and directing differentiation of muscle stem cells to sustain muscle tissue. Mimicking the extracellular muscle environment improves tools exploring the behavior of primary muscle cells. Optimizing cell culture conditions to maintain muscle commitment is important in stem cell-based studies concerning toxicology screening, ex vivo skeletal muscle tissue engineering and in the enhancement of clinical efficiency. We used the muscle extracellular matrix proteins collagen type I, fibronectin, laminin, and also gelatin and Matrigel as surface coatings of tissue culture plastic to resemble the muscle extracellular matrix. Several important factors that determine myogenic commitment of the primary muscle cells were characterized by quantitative real-time RT-PCR and immunofluorescence. Adhesion of high PAX7 expressing satellite cells was improved if the cells were cultured on fibronectin or laminin coatings. Cells cultured on Matrigel and laminin coatings showed dominant integrin expression levels and exhibited an activated Wnt pathway. Under these conditions both stem cell proliferation and myogenic differentiation capacity were superior if compared to cells cultured on collagen type I, fibronectin and gelatin. In conclusion, Matrigel and laminin are the preferred coatings to sustain the proliferation and myogenic differentiation capacity of the primary porcine muscle stem cells, when cells are removed from their natural environment for in vitro culture.

  2. Integration of concepts: cardiac extracellular matrix remodeling after myocardial infarction

    NARCIS (Netherlands)

    Cleutjens, Jack P. M.; Creemers, Esther E. J. M.

    2002-01-01

    The cardiac extracellular matrix consists of a three-dimensional structural network of interstitial collagens to which other matrix components are attached. The main physiological functions of this network are to retain tissue integrity and cardiac pump function. Collagen deposition is controlled

  3. Extracellular matrix proteins: a positive feedback loop in lung fibrosis?

    NARCIS (Netherlands)

    Blaauboer, M.E.; van Boeijen, F.R.; Emson, C.L.; Turner, S.M.; Zandieh-Doulabi, B.; Hanemaaijer, R.; Smit, T.H.; Stoop, R.; Everts, V.

    2014-01-01

    Lung fibrosis is characterized by excessive deposition of extracellular matrix. This not only affects tissue architecture and function, but it also influences fibroblast behavior and thus disease progression. Here we describe the expression of elastin, type V collagen and tenascin C during the

  4. Extracellular matrix proteins: A positive feedback loop in lung fibrosis?

    NARCIS (Netherlands)

    Blaauboer, M.E.; Boeijen, F.R.; Emson, C.L.; Turner, S.M.; Zandieh-Doulabi, B.; Hanemaaijer, R.; Smit, T.H.; Stoop, R.; Everts, V.

    2014-01-01

    Lung fibrosis is characterized by excessive deposition of extracellular matrix. This not only affects tissue architecture and function, but it also influences fibroblast behavior and thus disease progression. Here we describe the expression of elastin, type V collagen and tenascin C during the

  5. Extracellular matrix components direct porcine muscle stem cell behavior

    Energy Technology Data Exchange (ETDEWEB)

    Wilschut, Karlijn J. [Department of Farm Animal Health, Faculty of Veterinary Medicine, Utrecht University, Yalelaan 104, 3584 CM, Utrecht (Netherlands); Haagsman, Henk P. [Department of Infectious Diseases and Immunology, Faculty of Veterinary Medicine, Utrecht University, Yalelaan 1, 3584 CL, Utrecht (Netherlands); Roelen, Bernard A.J., E-mail: b.a.j.roelen@uu.nl [Department of Farm Animal Health, Faculty of Veterinary Medicine, Utrecht University, Yalelaan 104, 3584 CM, Utrecht (Netherlands)

    2010-02-01

    In muscle tissue, extracellular matrix proteins, together with the vasculature system, muscle-residence cells and muscle fibers, create the niche for muscle stem cells. The niche is important in controlling proliferation and directing differentiation of muscle stem cells to sustain muscle tissue. Mimicking the extracellular muscle environment improves tools exploring the behavior of primary muscle cells. Optimizing cell culture conditions to maintain muscle commitment is important in stem cell-based studies concerning toxicology screening, ex vivo skeletal muscle tissue engineering and in the enhancement of clinical efficiency. We used the muscle extracellular matrix proteins collagen type I, fibronectin, laminin, and also gelatin and Matrigel as surface coatings of tissue culture plastic to resemble the muscle extracellular matrix. Several important factors that determine myogenic commitment of the primary muscle cells were characterized by quantitative real-time RT-PCR and immunofluorescence. Adhesion of high PAX7 expressing satellite cells was improved if the cells were cultured on fibronectin or laminin coatings. Cells cultured on Matrigel and laminin coatings showed dominant integrin expression levels and exhibited an activated Wnt pathway. Under these conditions both stem cell proliferation and myogenic differentiation capacity were superior if compared to cells cultured on collagen type I, fibronectin and gelatin. In conclusion, Matrigel and laminin are the preferred coatings to sustain the proliferation and myogenic differentiation capacity of the primary porcine muscle stem cells, when cells are removed from their natural environment for in vitro culture.

  6. Astrocytes and extracellular matrix in extrasynaptic volume transmission

    Czech Academy of Sciences Publication Activity Database

    Vargová, Lýdia; Syková, Eva

    2014-01-01

    Roč. 369, č. 1654 (2014) ISSN 0962-8436 R&D Projects: GA ČR GA13-11867S; GA ČR(CZ) GBP304/12/G069 Institutional support: RVO:68378041 Keywords : extracellular space * diffusion * astrocytes Subject RIV: FH - Neurology Impact factor: 7.055, year: 2014

  7. Sortilin mediates vascular calcification via its recruitment into extracellular vesicles

    DEFF Research Database (Denmark)

    Goettsch, Claudia; Hutscheson, JD; Aikawa, M

    2016-01-01

    obscure. Here, we have demonstrated that sortilin is a key regulator of smooth muscle cell (SMC) calcification via its recruitment to extracellular vesicles. Sortilin localized to calcifying vessels in human and mouse atheromata and participated in formation of microcalcifications in SMC culture. Sortilin...

  8. The effect of nutrients on extracellular polymeric substance ...

    African Journals Online (AJOL)

    The effect of nutrients on extracellular polymeric substance (EPS) production and its impact on sludge properties and removal efficiencies were investigated in an in-depth field survey of wastewater treatment plants. Thereafter, laboratory studies were performed to evaluate the effect of a combination of nutrients - nitrogen ...

  9. Inflammation leads to distinct populations of extracellular vesicles from microglia

    DEFF Research Database (Denmark)

    Yang, Yiyi; Boza-Serrano, Antonio; Dunning, Christopher J.R.

    2018-01-01

    Background: Activated microglia play an essential role in inflammatory responses elicited in the central nervous system (CNS). Microglia-derived extracellular vesicles (EVs) are suggested to be involved in propagation of inflammatory signals and in the modulation of cell-to-cell communication...

  10. Extracellular products of photosynthesis in a tropical environment

    Digital Repository Service at National Institute of Oceanography (India)

    Gomes, H.R.; Pant, A.

    Extracellular products in the Arabian Sea averaged 0.42 g/cm2/day and represented 35% of the total carbon fixed by phytoplankton. No seasonal changes are observed during the two seasons i.e. premonsoon (Jan-Feb) and onset of southwest monsoon (April...

  11. Nucleases from Prevotella intermedia can degrade neutrophil extracellular traps.

    Science.gov (United States)

    Doke, M; Fukamachi, H; Morisaki, H; Arimoto, T; Kataoka, H; Kuwata, H

    2017-08-01

    Periodontitis is an inflammatory disease caused by periodontal bacteria in subgingival plaque. These bacteria are able to colonize the periodontal region by evading the host immune response. Neutrophils, the host's first line of defense against infection, use various strategies to kill invading pathogens, including neutrophil extracellular traps (NETs). These are extracellular net-like fibers comprising DNA and antimicrobial components such as histones, LL-37, defensins, myeloperoxidase, and neutrophil elastase from neutrophils that disarm and kill bacteria extracellularly. Bacterial nuclease degrades the NETs to escape NET killing. It has now been shown that extracellular nucleases enable bacteria to evade this host antimicrobial mechanism, leading to increased pathogenicity. Here, we compared the DNA degradation activity of major Gram-negative periodontopathogenic bacteria, Porphyromonas gingivalis, Prevotella intermedia, Fusobacterium nucleatum, and Aggregatibacter actinomycetemcomitans. We found that Pr. intermedia showed the highest DNA degradation activity. A genome search of Pr. intermedia revealed the presence of two genes, nucA and nucD, putatively encoding secreted nucleases, although their enzymatic and biological activities are unknown. We cloned nucA- and nucD-encoding nucleases from Pr. intermedia ATCC 25611 and characterized their gene products. Recombinant NucA and NucD digested DNA and RNA, which required both Mg 2+ and Ca 2+ for optimal activity. In addition, NucA and NucD were able to degrade the DNA matrix comprising NETs. © 2016 The Authors Molecular Oral Microbiology Published by John Wiley & Sons Ltd.

  12. Neutrophil extracellular traps in patients with pulmonary tuberculosis

    NARCIS (Netherlands)

    van der Meer, Anne Jan; Zeerleder, Sacha; Blok, Dana C.; Kager, Liesbeth M.; Lede, Ivar O.; Rahman, Wahid; Afroz, Rumana; Ghose, Aniruddha; Visser, Caroline E.; Zahed, Abu Shahed Md; Husain, Md Anwar; Alam, Khan Mashrequl; Barua, Pravat Chandra; Hassan, Mahtabuddin; Tayab, Md Abu; Dondorp, Arjen M.; van der Poll, Tom

    2017-01-01

    Tuberculosis is a devastating infectious disease causing many deaths worldwide. Recent investigations have implicated neutrophil extracellular traps (NETs) in the host response to tuberculosis. The aim of the current study was to obtain evidence for NETs release in the circulation during human

  13. Extracellular β-D-fructofuranosidase from Aspergillus parasiticus ...

    African Journals Online (AJOL)

    The β-D-fructofuranosidases are enzymes with biotechnological potential that can be used in different industrial sectors as food and beverage. In this context, microorganisms are important producers of these biomolecules, especially filamentous fungi. The production of extracellular β-Dfructofuranosidase from Aspergillus ...

  14. Extracellular clusterin promotes neuronal network complexity in vitro

    DEFF Research Database (Denmark)

    Wicher, Grzegorz; Velsecchi, Isabel; Charnay, Yves

    2008-01-01

    Clusterin (apolipoprotein J), a highly conserved amphiphatic glycoprotein and chaperone, has been implicated in a wide range of physiological and pathological processes. As a secreted protein, clusterin has been shown to act extracellularly where it is involved in lipid transportation and clearan...

  15. In vivo extracellular matrix protein expression by human periodontal ...

    African Journals Online (AJOL)

    It is well known that the orthodontic force applied to teeth generates a series of events that remodel the periodontal ligament (PDL). Extracellular matrix proteins (ECM) are described as molecular regulators of these events. However, the exact contribution of these proteins in human PDL modeling by orthodontic force ...

  16. Focus on Extracellular Vesicles: Therapeutic Potential of Stem Cell-Derived Extracellular Vesicles

    Directory of Open Access Journals (Sweden)

    Bin Zhang

    2016-02-01

    Full Text Available The intense research focus on stem and progenitor cells could be attributed to their differentiation potential to generate new cells to replace diseased or lost cells in many highly intractable degenerative diseases, such as Alzheimer disease, multiple sclerosis, and heart diseases. However, experimental and clinical studies have increasingly attributed the therapeutic efficacy of these cells to their secretion. While stem and progenitor cells secreted many therapeutic molecules, none of these molecules singly or in combination could recapitulate the functional effects of stem cell transplantations. Recently, it was reported that extracellular vesicles (EVs could recapitulate the therapeutic effects of stem cell transplantation. Based on the observations reported thus far, the prevailing hypothesis is that stem cell EVs exert their therapeutic effects by transferring biologically active molecules such as proteins, lipids, mRNA, and microRNA from the stem cells to injured or diseased cells. In this respect, stem cell EVs are similar to EVs from other cell types. They are both primarily vehicles for intercellular communication. Therefore, the differentiating factor is likely due to the composition of their cargo. The cargo of EVs from different cell types are known to include a common set of proteins and also proteins that reflect the cell source of the EVs and the physiological or pathological state of the cell source. Hence, elucidation of the stem cell EV cargo would provide an insight into the multiple physiological or biochemical changes necessary to affect the many reported stem cell-based therapeutic outcomes in a variety of experimental models and clinical trials.

  17. Soluble extracellular matrix metalloproteinase inducer (EMMPRIN, EMN) regulates cancer-related cellular functions by homotypic interactions with surface CD147.

    Science.gov (United States)

    Knutti, Nadine; Kuepper, Michael; Friedrich, Karlheinz

    2015-11-01

    EMMPRIN (extracellular matrix metalloproteinase inducer) is a widely expressed glycoprotein and a member of the immunoglobulin superfamily which exists in both a membrane-spanning and a soluble form. Homotypic interactions of EMMPRIN underlie its multiple roles in normal development and pathological situations such as viral infections, Alzheimer's disease and cancer. This study employed a recombinant soluble, fully glycosylated EMMPRIN domain (rhsEMN) as a tool to characterize the structural basis of EMMPRIN-EMMPRIN receptor (EMNR) contacts and their functional effects on MCF-7 breast carcinoma cells. rhsEMN did not form dimers in solution but bound to surface EMMPRIN (EMN) on MCF-7 cells with high affinity and was readily internalized. The interaction interface for the homotypic contact was localized to the N-terminal Ig domain. rhsEMN exerted a stimulatory effect on proliferation of MCF-7 cells whereas it reduced cell migration in a dose-dependent manner. These effects were accompanied by an upregulation of endogenous EMMPRIN as well as of matrix metalloproteinase-14 (MMP-14), a membrane-bound protease involved in the extracellular release of soluble EMMPRIN, indicating a regulatory feedback mechanism. The proliferation-promoting activity of rhsEMN was mimicked by a novel functional antibody directed to EMMPRIN, underscoring that crosslinking of cell surface EMMPRIN (EMNR) is crucial for eliciting intracellular signalling. Addressing malignancy-related signal transduction in HEK-293 cells, we could show that rhsEMN triggers the oncogenic Wnt pathway. © 2015 FEBS.

  18. CD81 Receptor Regions outside the Large Extracellular Loop Determine Hepatitis C Virus Entry into Hepatoma Cells

    Directory of Open Access Journals (Sweden)

    Pia Banse

    2018-04-01

    Full Text Available Hepatitis C virus (HCV enters human hepatocytes using four essential entry factors, one of which is human CD81 (hCD81. The tetraspanin hCD81 contains a large extracellular loop (LEL, which interacts with the E2 glycoprotein of HCV. The role of the non-LEL regions of hCD81 (intracellular tails, four transmembrane domains, small extracellular loop and intracellular loop is poorly understood. Here, we studied the contribution of these domains to HCV susceptibility of hepatoma cells by generating chimeras of related tetraspanins with the hCD81 LEL. Our results show that non-LEL regions in addition to the LEL determine susceptibility of cells to HCV. While closely related tetraspanins (X. tropicalis CD81 and D. rerio CD81 functionally complement hCD81 non-LEL regions, distantly related tetraspanins (C. elegans TSP9 amd D. melanogaster TSP96F do not and tetraspanins with intermediate homology (hCD9 show an intermediate phenotype. Tetraspanin homology and susceptibility to HCV correlate positively. For some chimeras, infectivity correlates with surface expression. In contrast, the hCD9 chimera is fully surface expressed, binds HCV E2 glycoprotein but is impaired in HCV receptor function. We demonstrate that a cholesterol-coordinating glutamate residue in CD81, which hCD9 lacks, promotes HCV infection. This work highlights the hCD81 non-LEL regions as additional HCV susceptibility-determining factors.

  19. Extended and structurally supported insights into extracellular hormone binding, signal transduction and organization of the thyrotropin receptor.

    Directory of Open Access Journals (Sweden)

    Gerd Krause

    Full Text Available The hormone thyrotropin (TSH and its receptor (TSHR are crucial for the growth and function of the thyroid gland. The TSHR is evolutionary linked with the receptors of follitropin (FSHR and lutropin/choriogonadotropin (LHR and their sequences and structures are similar. The extracellular region of TSHR contains more than 350 amino acids and binds hormone and antibodies. Several important questions related to functions and mechanisms of TSHR are still not comprehensively understood. One major reason for these open questions is the lack of any structural information about the extracellular segment of TSHR that connects the N-terminal leucine-rich repeat domain (LRRD with the transmembrane helix (TMH 1, the hinge region. It has been shown experimentally that this segment is important for fine tuning of signaling and ligand interactions. A new crystal structure containing most of the extracellular hFSHR region in complex with hFSH has recently been published. Now, we have applied these new structural insights to the homologous TSHR and have generated a structural model of the TSHR LRRD/hinge-region/TSH complex. This structural model is combined and evaluated with experimental data including hormone binding (bTSH, hTSH, thyrostimulin, super-agonistic effects, antibody interactions and signaling regulation. These studies and consideration of significant and non-significant amino acids have led to a new description of mechanisms at the TSHR, including ligand-induced displacements of specific hinge region fragments. This event triggers conformational changes at a convergent center of the LRRD and the hinge region, activating an "intramolecular agonistic unit" close to the transmembrane domain.

  20. Extended and structurally supported insights into extracellular hormone binding, signal transduction and organization of the thyrotropin receptor.

    Science.gov (United States)

    Krause, Gerd; Kreuchwig, Annika; Kleinau, Gunnar

    2012-01-01

    The hormone thyrotropin (TSH) and its receptor (TSHR) are crucial for the growth and function of the thyroid gland. The TSHR is evolutionary linked with the receptors of follitropin (FSHR) and lutropin/choriogonadotropin (LHR) and their sequences and structures are similar. The extracellular region of TSHR contains more than 350 amino acids and binds hormone and antibodies. Several important questions related to functions and mechanisms of TSHR are still not comprehensively understood. One major reason for these open questions is the lack of any structural information about the extracellular segment of TSHR that connects the N-terminal leucine-rich repeat domain (LRRD) with the transmembrane helix (TMH) 1, the hinge region. It has been shown experimentally that this segment is important for fine tuning of signaling and ligand interactions. A new crystal structure containing most of the extracellular hFSHR region in complex with hFSH has recently been published. Now, we have applied these new structural insights to the homologous TSHR and have generated a structural model of the TSHR LRRD/hinge-region/TSH complex. This structural model is combined and evaluated with experimental data including hormone binding (bTSH, hTSH, thyrostimulin), super-agonistic effects, antibody interactions and signaling regulation. These studies and consideration of significant and non-significant amino acids have led to a new description of mechanisms at the TSHR, including ligand-induced displacements of specific hinge region fragments. This event triggers conformational changes at a convergent center of the LRRD and the hinge region, activating an "intramolecular agonistic unit" close to the transmembrane domain.

  1. Multifunctionalities driven by ferroic domains

    Science.gov (United States)

    Yang, J. C.; Huang, Y. L.; He, Q.; Chu, Y. H.

    2014-08-01

    Considerable attention has been paid to ferroic systems in pursuit of advanced applications in past decades. Most recently, the emergence and development of multiferroics, which exhibit the coexistence of different ferroic natures, has offered a new route to create functionalities in the system. In this manuscript, we step from domain engineering to explore a roadmap for discovering intriguing phenomena and multifunctionalities driven by periodic domain patters. As-grown periodic domains, offering exotic order parameters, periodic local perturbations and the capability of tailoring local spin, charge, orbital and lattice degrees of freedom, are introduced as modeling templates for fundamental studies and novel applications. We discuss related significant findings on ferroic domain, nanoscopic domain walls, and conjunct heterostructures based on the well-organized domain patterns, and end with future prospects and challenges in the field.

  2. Extracellular small heat shock proteins: exosomal biogenesis and function.

    Science.gov (United States)

    Reddy, V Sudhakar; Madala, Satish K; Trinath, Jamma; Reddy, G Bhanuprakash

    2018-05-01

    Small heat shock proteins (sHsps) belong to the family of heat shock proteins (Hsps): some are induced in response to multiple stressful events to protect the cells while others are constitutively expressed. Until now, it was believed that Hsps, including sHsps, are present inside the cells and perform intracellular functions. Interestingly, several groups recently reported the extracellular presence of Hsps, and sHsps have also been detected in sera/cerebrospinal fluids in various pathological conditions. Secretion into the extracellular milieu during many pathological conditions suggests additional or novel functions of sHsps in addition to their intracellular properties. Extracellular sHsps are implicated in cell-cell communication, activation of immune cells, and promoting anti-inflammatory and anti-platelet responses. Interestingly, exogenous administration of sHsps showed therapeutic effects in multiple disease models implying that extracellular sHsps are beneficial in pathological conditions. sHsps do not possess signal sequence and, hence, are not exported through the classical Endoplasmic reticulum-Golgi complex (ER-Golgi) secretory pathway. Further, export of sHsps is not inhibited by ER-Golgi secretory pathway inhibitors implying the involvement of a nonclassical secretory pathway in sHsp export. In lieu, lysoendosomal and exosomal pathways have been proposed for the export of sHsps. Heat shock protein 27 (Hsp27), αB-crystallin (αBC), and Hsp20 are shown to be exported by exosomes. Exosomes packaged with sHsps have beneficial effects in in vivo disease models. However, secretion mechanisms and therapeutic use of sHsps have not been elucidated in detail. Therefore, this review aimed at highlighting the current understanding of sHsps (Hsp27, αBC, and Hsp20) in the extracellular medium.

  3. Mapping the Moral Domain

    Science.gov (United States)

    Graham, Jesse; Nosek, Brian A.; Haidt, Jonathan; Iyer, Ravi; Koleva, Spassena; Ditto, Peter H.

    2010-01-01

    The moral domain is broader than the empathy and justice concerns assessed by existing measures of moral competence, and it is not just a subset of the values assessed by value inventories. To fill the need for reliable and theoretically-grounded measurement of the full range of moral concerns, we developed the Moral Foundations Questionnaire (MFQ) based on a theoretical model of five universally available (but variably developed) sets of moral intuitions: Harm/care, Fairness/reciprocity, Ingroup/loyalty, Authority/respect, and Purity/sanctity. We present evidence for the internal and external validity of the scale and the model, and in doing so present new findings about morality: 1. Comparative model fitting of confirmatory factor analyses provides empirical justification for a five-factor structure of moral concerns. 2. Convergent/discriminant validity evidence suggests that moral concerns predict personality features and social group attitudes not previously considered morally relevant. 3. We establish pragmatic validity of the measure in providing new knowledge and research opportunities concerning demographic and cultural differences in moral intuitions. These analyses provide evidence for the usefulness of Moral Foundations Theory in simultaneously increasing the scope and sharpening the resolution of psychological views of morality. PMID:21244182

  4. Domain wall networks on solitons

    International Nuclear Information System (INIS)

    Sutcliffe, Paul

    2003-01-01

    Domain wall networks on the surface of a soliton are studied in a simple theory. It consists of two complex scalar fields, in 3+1 dimensions, with a global U(1)xZ n symmetry, where n>2. Solutions are computed numerically in which one of the fields forms a Q ball and the other field forms a network of domain walls localized on the surface of the Q ball. Examples are presented in which the domain walls lie along the edges of a spherical polyhedron, forming junctions at its vertices. It is explained why only a small restricted class of polyhedra can arise as domain wall networks

  5. Topological domain walls in helimagnets

    Science.gov (United States)

    Schoenherr, P.; Müller, J.; Köhle