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Sample records for spleen trypsin inhibitor

  1. Aggregation of trypsin and trypsin inhibitor by Al cation.

    Science.gov (United States)

    Chanphai, P; Kreplak, L; Tajmir-Riahi, H A

    2017-04-01

    Al cation may trigger protein structural changes such as aggregation and fibrillation, causing neurodegenerative diseases. We report the effect of Al cation on the solution structures of trypsin (try) and trypsin inhibitor (tryi), using thermodynamic analysis, UV-Visible, Fourier transform infrared (FTIR) spectroscopic methods and atomic force microscopy (AFM). Thermodynamic parameters showed Al-protein bindings occur via H-bonding and van der Waals contacts for trypsin and trypsin inhibitor. AFM showed that Al cations are able to force trypsin into larger or more robust aggregates than trypsin inhibitor, with trypsin 5±1 SE (n=52) proteins per aggregate and for trypsin inhibitor 8.3±0.7 SE (n=118). Thioflavin T test showed no major protein fibrillation in the presence of Al cation. Al complexation induced more alterations of trypsin inhibitor conformation than trypsin. Copyright © 2017 Elsevier B.V. All rights reserved.

  2. Laura: Soybean variety lacking Kunitz trypsin inhibitor

    Directory of Open Access Journals (Sweden)

    Srebrić Mirjana

    2010-01-01

    Full Text Available Grain of conventional soybean varieties requires heat processing to break down trypsin inhibitor's activity before using as food or animal feed. At the same time, protein denaturation and other qualitative changes occur in soybean grain, especially if the temperature of heating is not controlled. Two types of trypsin inhibitor were found in soybean grain the Kunitz trypsin inhibitor and the Bowman-Birk inhibitor. Mature grain of soybean Laura is lacking Kunitz trypsin inhibitor. Grain yield of variety Laura is equal to high yielding varieties from the maturity group I, where it belongs. Lacking of Kunitz-trypsin inhibitor makes soybean grain suitable for direct feeding in adult non ruminant animals without previous thermal processing. Grain of variety Laura can be processed for a shorter period of time than conventional soybeans. This way we save energy, and preserve valuable nutritional composition of soybean grain, which is of interest in industrial processing.

  3. Inga laurina trypsin inhibitor (ILTI) obstructs Spodoptera frugiperda trypsins expressed during adaptive mechanisms against plant protease inhibitors.

    Science.gov (United States)

    Machado, Suzy Wider; de Oliveira, Caio Fernando Ramalho; Zério, Neide Graciano; Parra, José Roberto Postali; Macedo, Maria Lígia Rodrigues

    2017-08-01

    Plant protease inhibitors (PIs) are elements of a common plant defense mechanism induced in response to herbivores. The fall armyworm, Spodoptera frugiperda, a highly polyphagous lepidopteran pest, responds to various PIs in its diet by expressing genes encoding trypsins. This raises the question of whether the PI-induced trypsins are also inhibited by other PIs, which we posed as the hypothesis that Inga laurina trypsin inhibitor (ILTI) inhibits PI-induced trypsins in S. frugiperda. In the process of testing our hypothesis, we compared its properties with those of selected PIs, soybean Kunitz trypsin inhibitor (SKTI), Inga vera trypsin inhibitor (IVTI), Adenanthera pavonina trypsin inhibitor (ApTI), and Entada acaciifolia trypsin inhibitor (EATI). We report that ILTI is more effective in inhibiting the induced S. frugiperda trypsins than SKTI and the other PIs, which supports our hypothesis. ILTI may be more appropriate than SKTI for studies regarding adaptive mechanisms to dietary PIs. © 2017 Wiley Periodicals, Inc.

  4. The kinetics of interaction of porcine - alpha-, and porcine - beta -trypsin with intact and modified soybean trypsin inhibitor (kunitz)

    International Nuclear Information System (INIS)

    Hamid, M.A.

    1994-01-01

    The association of porcine trypsin with soybean trypsin inhibitor (Kunitz) resulted in characteristic changes in absorption spectrum, indicating an alteration of the micro environments of the enzyme chromophores as a consequence of the interaction. The rates of formation of the stable trypsin - inhibitor complexes from porcine - alpha - trypsin and soybean trypsin inhibitor and from porcine - beta - trypsin and either intact or modified soybean trypsin inhibitor were measured by mixing the equimolar concentration of the reactants in a Stopped - Flow apparatus at pH (4.5 to 10.0). The reaction of trypsin with soybean trypsin inhibitor was of first order with respect to the concentration of the reactants used. The rates of dissociation of the stable complexes, alpha - trypsin - soybean trypsin inhibitor, beta -trypsin - soybean trypsin inhibitor and beta -trypsin modified soybean trypsin inhibitor were also measured at pH (1.92 to 3.58). The values of first order rate constant, k/sub D/ obtained for the dissociation of all the three complexes were identical with one another. The kinetics results obtained for the porcine trypsin were compared with those of bovine trypsin system and it was suggested that the reaction mechanisms in both these systems were identical. (author)

  5. Cytogenetic study of Ascaris trypsin inhibitor in cultured human ...

    Indian Academy of Sciences (India)

    2009-04-01

    Apr 1, 2009 ... Although the physical and chemical properties of Ascaris trypsin inhibitors ... male of Ascaris suum according to the method of Pudles and. Rola (1967). ..... inhibitor isolated from Ascaris resulted in the appearance of dominant ...

  6. Hydrogen exchange kinetics changes upon formation of the soybean trypsin inhibitor: trypsin complex

    International Nuclear Information System (INIS)

    Woodward, C.K.; Ellis, L.M.

    1975-01-01

    The hydrogen exchange kinetics of the complex of trypsin--soybean trypsin inhibitor (Kunitz) have been compared to the calculated sum of the exchange kinetics for the inhibitor and trypsin measured separately. The exchange rates observed for the complex are substantially less than the sum of the exchange rates in the two individual proteins. These results cannot be accounted for by changes in intermolecular or intramolecular hydrogen bonding. The decrease in exchange rates in the complex are ascribed to changes in solvent accessibility in the component proteins. (U.S.)

  7. Bovine Pancreatic Trypsin Inhibitor-Trypsin Complex as a Detection System for Recombinant Proteins

    Science.gov (United States)

    Borjigin, Jimo; Nathans, Jeremy

    1993-01-01

    Bovine pancreatic trypsin inhibitor (BPTI) binds to trypsin and anhydrotrypsin (an enzymatically inactive derivative of trypsin) with affinities of 6 x 10-14 and 1.1 x 10-13 M, respectively. We have taken advantage of the high affinity and specificity of this binding reaction to develop a protein tagging system in which biotinylated trypsin or biotinylated anhydrotrypsin is used as the reagent to detect recombinant fusion proteins into which BPTI has been inserted. Two proteins, opsin and growth hormone, were used as targets for insertional mutagenesis with BPTI. In each case, both domains of the fusion protein appear to be correctly folded. The fusion proteins can be specifically and efficiently detected by biotinylated trypsin or biotinylated anhydrotrypsin, as demonstrated by staining of transfected cells, protein blotting, affinity purification, and a mobility shift assay in SDS/polyacrylamide gels.

  8. ELISA analysis of soybean trypsin inhibitors in processed foods.

    Science.gov (United States)

    Brandon, D L; Bates, A H; Friedman, M

    1991-01-01

    Soybean proteins are widely used in human foods in a variety of forms, including infant formulas, flour, protein concentrates, protein isolates, soy sauces, textured soy fibers, and tofu. The presence of inhibitors of digestive enzymes in soy proteins impairs the nutritional quality and possibly the safety of soybeans and other legumes. Processing, based on the use of heat or fractionation of protein isolates, does not completely inactivate or remove these inhibitors, so that residual amounts of inhibitors are consumed by animals and humans. New monoclonal antibody-based immunoassays can measure low levels of the soybean Kunitz trypsin inhibitor (KTI) and the Bowman-Birk trypsin and chymotrypsin inhibitor (BBI) and the Bowman-Birk foods. The enzyme-linked immunosorbent assay (ELISA) was used to measure the inhibitor content of soy concentrates, isolates, and flours, both heated and unheated; a commercial soy infant formula; KTI and BBI with rearranged disulfide bonds; browning products derived from heat-treatment of KTI with glucose and starch; and KTI exposed to high pH. The results indicate that even low inhibitor isolates contain significant amounts of specific inhibitors. Thus, infants on soy formula consume about 10 mg of KTI plus BBI per day. The immunoassays complement the established enzymatic assays of trypsin and chymotrypsin inhibitors, and have advantages in (a) measuring low levels of inhibitors in processed foods; and (b) differentiating between the Kunitz and Bowman-Birk inhibitors. The significance of our findings for food safety are discussed.

  9. Study on transformation of cowpea trypsin inhibitor gene into ...

    African Journals Online (AJOL)

    Cowpea Trypsin Inhibitor (CpTI) gene was transferred into cauliflower by agrobacterium-mediated transformation method, and 14 transgenic cauliflower plants were obtained. Cotyledons and hypocotyls were used as explants. The putative transformants were assayed by PCR and Southern blotting analysis. The results ...

  10. Low molecular weight squash trypsin inhibitors from Sechium edule seeds.

    Science.gov (United States)

    Laure, Hélen J; Faça, Vítor M; Izumi, Clarice; Padovan, Júlio C; Greene, Lewis J

    2006-02-01

    Nine chromatographic components containing trypsin inhibitor activity were isolated from Sechium edule seeds by acetone fractionation, gel filtration, affinity chromatography and RP-HPLC in an overall yield of 46% of activity and 0.05% of protein. The components obtained with highest yield of total activity and highest specific activity were sequenced by Edman degradation and their molecular masses determined by mass spectrometry. The inhibitors contained 31, 32 and 27 residues per molecule and their sequences were: SETI-IIa, EDRKCPKILMRCKRDSDCLAKCTCQESGYCG; SETI-IIb, EEDRKCPKILMRCKRDSDCLAKCTCQESGYCG and SETI-V, CPRILMKCKLDTDCFPTCTCRPSGFCG. SETI-IIa and SETI-IIb, which differed by an amino-terminal E in the IIb form, were not separable under the conditions employed. The sequences are consistent with consensus sequences obtained from 37 other inhibitors: CPriI1meCk_DSDCla_C_C_G_CG, where capital letters are invariant amino acid residues and lower case letters are the most preserved in this position. SETI-II and SETI-V form complexes with trypsin with a 1:1 stoichiometry and have dissociation constants of 5.4x10(-11)M and 1.1x10(-9)M, respectively.

  11. Urinary trypsin inhibitor - an experimental and clinical study

    International Nuclear Information System (INIS)

    Berling, B.M.

    1991-01-01

    The urinary trypsin inhibitor (UTI) is an acid stable proteinase inhibitor present in blood and urine. It was purified from urine using affinity chromatography, ion exchange chromatography and gel filtration. Two forms of UTI were present in urine, A and B. A radioimmunoassay for measurement of UTI in urine and plasma was performed. The normal level of UTI in plasma and serum was about 2 mg/l. The normal excretion in urine was about 8 mg per 24 hours. The plasma and urine levels of UTI were studied in patients with acute pancreatitis and in patients undergoing cholecystectomy. Uremic patients had a marked increase of UTI in plasma compatible with decreased glomerular filtration. In samples from healthy persons as well as from patients only inhibitor A was found. Inhibitor B has recently been renamed bikunin because of its two Kunitz-type inhibiting domains. Inhibitor A might be called tetrakunin. Radioactively labeled UTI (inhibitor A) was injected intravenously in three male volunteers. The plasma half-life of 125 I UTI was 2 hours. Free biologically active inhibitor was found in the urine during the first four hours after injection. The organ distribution of intravenously injected 125 I UTI was studied in rats. Fifteen minutes after injection the major part of the radioactivity was found in the kidneys, suggesting that the kidneys are the primary site of UTI metabolism. Using immunohistochemical techniques UTI was found in the proximal tubules of the normal human kidney further indicating the tubular reabsorption and methabolisms of UTI

  12. Carbohydrate as covalent crosslink in human inter-alpha-trypsin inhibitor

    DEFF Research Database (Denmark)

    Jessen, T E; Faarvang, K L; Ploug, M

    1988-01-01

    The primary structure of inter-alpha-trypsin inhibitor is partially elucidated, but controversy about the construction of the polypeptide backbone still exists. We present evidence suggesting that inter-alpha-trypsin inhibitor represents a novel plasma protein structure with two separate polypept...... polypeptide chains covalently crosslinked only by carbohydrate (chondroitin sulphate)....

  13. Gamma rays induced mutation for low phytic acid and trypsin inhibitor content in soybean

    International Nuclear Information System (INIS)

    Gupta, S.K.; Manjaya, J.G.

    2017-01-01

    Soybean (Glycine max (L.) Merrill) is an important source of vegetable protein and is used as a food, feed and health supplement. However, consumption of soybean as food is limited because of the presence of many anti-nutritional factors. Trypsin inhibitors and phytic acid are two major anti-nutritional factors present in soybean that need to be removed for increasing the soybean consumption as food. Trypsin inhibitor is known to inhibit the trypsin/chymotrpsin activity and phytic acid reduces the bioavailability of essential micronutrients in digestive tract, resulting in adverse effect on health. Therefore, developing soybean cultivars having low trypsin inhibitors and phytic acid content is highly desirable. Soybean cultivar JS 93-05 was irradiated with 250 Gy gamma rays to induce mutation for various morphological and biochemical characters. A large number of mutants with altered morphological characters were identified. Ninety true breeding mutant lines in M6 generation were screened for trypsin inhibitor and phytic acid content. The phytic acid content was estimated using modified colorimetric method and trypsin inhibitor concentration was estimated using BAPNA as substrate in colorimetric method. The phytic acid content in the mutants varied from 7.59 to 24.14 mg g -1 . Two mutants lines TSG - 62 (7.59 mg g -1 ) and TSG - 66 (9.62 mg g -1 ) showed significant low phytic acid content as compared to the parent JS 93-05 (20.19 mg g -1 ). The trypsin inhibitor concentration in the mutants varied from 19.92 to 53.64 TIU mg -1 and one mutant line (TSG -14) was found with the lowest trypsin inhibitor concentration of 19.92 TIU mg -1 compared to parent JS 93-05 (50.90 TIU mg -1 ). The mutant lines identified in this study will serve as important genetic resources for developing low phytic acid and low trypsin inhibitor cultivars in soybean. (author)

  14. Inactivation Methods of Trypsin Inhibitor in Legumes: A Review.

    Science.gov (United States)

    Avilés-Gaxiola, Sara; Chuck-Hernández, Cristina; Serna Saldívar, Sergio O

    2018-01-01

    Seed legumes have played a major role as a crop worldwide, being cultivated on about 12% to 15% of Earth's arable land; nevertheless, their use is limited by, among other things, the presence of several antinutritional factors (ANFs - naturally occurring metabolites that the plant produces to protect itself from pest attacks.) Trypsin inhibitors (TIs) are one of the most relevant ANFs because they reduce digestion and absorption of dietary proteins. Several methods have been developed in order to inactivate TIs, and of these, thermal treatments are the most commonly used. They cause loss of nutrients, affect functional properties, and require high amounts of energy. Given the above, new processes have emerged to improve the nutritional quality of legumes while trying to solve the problems caused by the use of thermal treatments. This review examines and discusses the methods developed by researchers to inactivate TI present in legumes and their effects over nutritional and functional properties. © 2017 Institute of Food Technologists®.

  15. Influence of Different Genotypes on Trypsin Inhibitor Levels and Activity in Soybeans

    Directory of Open Access Journals (Sweden)

    Viktor A. Nedovic

    2007-01-01

    Full Text Available This study describes the relationship between the two major trypsin inhibitors (TI in soybean, i.e., the Kunitz (KTI and Bowman-Birk (BBI trypsin inhibitors, as well as between them and the corresponding trypsin inhibitor activity (TIA. Twelve investigated soybean genotypes showed significant differences in TI levels and TIA. A very strong positive correlation was found between the levels of KTI and total BBI (r = 0.94, P < 0.05. No relationship was found between KTI, BBI or total TI and TIA. Based on this data, it appears that the levels of major TI in soybean are related. Understanding the relationship between trypsin inhibitors and their activities could be useful for further improvement of the health impacts of soy proteins.

  16. A new protein inhibitor of trypsin and activated Hageman factor from pumpkin (Cucurbita maxima) seeds.

    Science.gov (United States)

    Krishnamoorthi, R; Gong, Y X; Richardson, M

    1990-10-29

    A protein inhibitor (CMTI-V; Mr 7106) of trypsin and activated Hageman factor (Factor XIIa), a serine protease involved in blood coagulation, has been isolated for the first time from pumpkin (Cucurbita maxima) seeds by means of trypsin-affinity chromatography and reverse phase high performance liquid chromatography (HPLC). The dissociation constants of the inhibitor complexes with trypsin and Factor XIIa have been determined to be 1.6 x 10(-8) and 4.1 x 10(-8) M, respectively. The primary structure of CMTI-V is reported. The protein has 68 amino acid residues and one disulfide bridge and shows a high level of sequence homology to the Potato I inhibitor family. Furthermore, its amino terminus consists of an N-acetylates Ser. The reactive site has been established to be the peptide bond between Lys44-Asp45. The modified inhibitor which has the reactive site peptide bond hydrolyzed inhibits trypsin but not the Hageman factor.

  17. Purification and characterization of a trypsin inhibitor from the seeds of Artocarpus heterophyllus Lam.

    Science.gov (United States)

    Lyu, Junchen; Liu, Yuan; An, Tianchen; Liu, Yujun; Wang, Manchuriga; Song, Yanting; Zheng, Feifei; Wu, Dan; Zhang, Yingxia; Deng, Shiming

    2015-05-01

    A proteinaceous inhibitor against trypsin was isolated from the seeds of Artocarpus heterophyllus Lam. by successive ammonium sulfate precipitation, ion-exchange, and gel-filtration chromatography. The trypsin inhibitor, named as AHLTI (A. heterophyllus Lam. trypsin inhibitor), consisted of a single polypeptide chain with a molecular weight of 28.5 kDa, which was confirmed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and gel-filtration chromatography. The N-terminal sequence of AHLTI was DEPPSELDAS, which showed no similarity to other known trypsin inhibitor sequence. AHLTI completely inhibited bovine trypsin at a molar ratio of 1:2 (AHLTI:trypsin) analyzed by native polyacrylamide gel electrophoresis, inhibition activity assay, and gel-filtration chromatography. Moreover, kinetic enzymatic studies were carried out to understand the inhibition mechanism of AHLTI against trypsin. Results showed that AHLTI was a competitive inhibitor with an equilibrium dissociation constant (Ki) of 3.7 × 10(-8) M. However, AHLTI showed weak inhibitory activity toward chymotrypsin and elastase. AHLTI was stable over a broad range of pH 4-8 and temperature 20-80°C. The reduction agent, dithiothreitol, had no obvious effect on AHLTI. The trypsin inhibition assays of AHLTI toward digestive enzymes from insect pest guts in vitro demonstrated that AHLTI was effective against enzymes from Locusta migratoria manilensis (Meyen). These results suggested that AHLTI might be a novel trypsin inhibitor from A. heterophyllus Lam. belonging to Kunitz family, and play an important role in protecting from insect pest. © The Author 2015. Published by ABBS Editorial Office in association with Oxford University Press on behalf of the Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences.

  18. Silica-supported Macroporous Chitosan Bead for Affinity Purification of Trypsin Inhibitor

    Institute of Scientific and Technical Information of China (English)

    Feng Na XI; Jian Min WU; Ming Ming LUAN

    2005-01-01

    Macroporous cross-linking chitosan layer coated on silica gel (CTS-SiO2) was prepared by phase inversion and polyethylene glycol (PEG) molecular imprinting methods. Formation of macroporous surface was investigated by scanning electron microscopy (SEM) and BET analysis.The prepared bead was activated by reacting with 1,2-ethylene diglycidyl ether for introducing epoxy groups, and trypsin could be efficiently immobilized on the bead as a biospecific ligand.The bead bearing trypsin was employed to purify trypsin inhibitor (TIs) from egg white as affinity adsorbent.

  19. Buckwheat trypsin inhibitor with helical hairpin structure belongs to a new family of plant defence peptides.

    Science.gov (United States)

    Oparin, Peter B; Mineev, Konstantin S; Dunaevsky, Yakov E; Arseniev, Alexander S; Belozersky, Mikhail A; Grishin, Eugene V; Egorov, Tsezi A; Vassilevski, Alexander A

    2012-08-15

    A new peptide trypsin inhibitor named BWI-2c was obtained from buckwheat (Fagopyrum esculentum) seeds by sequential affinity, ion exchange and reversed-phase chromatography. The peptide was sequenced and found to contain 41 amino acid residues, with four cysteine residues involved in two intramolecular disulfide bonds. Recombinant BWI-2c identical to the natural peptide was produced in Escherichia coli in a form of a cleavable fusion with thioredoxin. The 3D (three-dimensional) structure of the peptide in solution was determined by NMR spectroscopy, revealing two antiparallel α-helices stapled by disulfide bonds. Together with VhTI, a trypsin inhibitor from veronica (Veronica hederifolia), BWI-2c represents a new family of protease inhibitors with an unusual α-helical hairpin fold. The linker sequence between the helices represents the so-called trypsin inhibitory loop responsible for direct binding to the active site of the enzyme that cleaves BWI-2c at the functionally important residue Arg(19). The inhibition constant was determined for BWI-2c against trypsin (1.7×10(-1)0 M), and the peptide was tested on other enzymes, including those from various insect digestive systems, revealing high selectivity to trypsin-like proteases. Structural similarity shared by BWI-2c, VhTI and several other plant defence peptides leads to the acknowledgement of a new widespread family of plant peptides termed α-hairpinins.

  20. Ecological costs and benefits correlated with trypsin protease inhibitor production in Nicotiana attenuata

    NARCIS (Netherlands)

    Glawe, G.A.; Zavala, J.A.; Kessler, A.; Van Dam, N.M.; Baldwin, I.T.

    2003-01-01

    Genotypes of the wild tobacco Nicotiana attenuata from different geographic regions in North America vary considerably in the level of constitutive and inducible trypsin protease inhibitors (TrypPIs), a potent direct defense, as well as in the production of herbivore-induced volatiles that function

  1. Frequency of inhibitors of daphnid trypsin in the widely distributed cyanobacterial genus Planktothrix

    DEFF Research Database (Denmark)

    Rohrlack, T.; Christoffersen, K.; Friberg-Jensen, U.

    2005-01-01

    on the frequency of such compounds in the widely distributed cyanobacterial genus Planktothrix. Of the 89 Planktothrix strains analysed, about 70% produced inhibitors of daphnid trypsin. The strains tested positive represented three common Planktothrix species and were isolated from diverse localities...

  2. Design, chemical synthesis and kinetic studies of trypsin chromogenic substrates based on the proteinase binding loop of Cucurbita maxima trypsin inhibitor (CMTI-III).

    Science.gov (United States)

    Lesner, A; Brzozowski, K; Kupryszewski, G; Rolka, K

    2000-03-05

    A series of trypsin chromogenic substrates with formula: Y-Ala-X-Abu-Pro-Lys-pNA, where X = Gly, Ala, Abu, Val, Leu, Phe, Ser, Glu and Y = Ac, H; pNA = p-nitroanilide was synthesized. The Cucurbita maxima trypsin inhibitor CMTI-III molecule was used as a vehicle to design the trypsin substrates. To evaluate the influence of position P(4) on the substrate-enzyme interaction, kinetic parameters of newly synthesized substrates with bovine beta-trypsin were determined. The increasing hydrophobicity of the amino acid residue (Gly, Ala, Abu, Val) introduced in position P(4) significantly enhanced the substrate specificity (k(cat)/K(m)) which was over 8 times higher for the last residue than that for the first one. The introduction of residues with more hydrophilic side chain (Glu, Ser) in this position reduced the value of this parameter. These results correspond well with those obtained using molecular dynamics of bovine beta-trypsin with monosubstituted CMTI-I analogues, indicating that in both trypsin substrate and inhibitor position 4 plays an important role in the interaction with the enzyme. Copyright 2000 Academic Press.

  3. A colostrum trypsin inhibitor gene expressed in the Cape fur seal mammary gland during lactation.

    Science.gov (United States)

    Pharo, Elizabeth A; Cane, Kylie N; McCoey, Julia; Buckle, Ashley M; Oosthuizen, W H; Guinet, Christophe; Arnould, John P Y

    2016-03-01

    The colostrum trypsin inhibitor (CTI) gene and transcript were cloned from the Cape fur seal mammary gland and CTI identified by in silico analysis of the Pacific walrus and polar bear genomes (Order Carnivora), and in marine and terrestrial mammals of the Orders Cetartiodactyla (yak, whales, camel) and Perissodactyla (white rhinoceros). Unexpectedly, Weddell seal CTI was predicted to be a pseudogene. Cape fur seal CTI was expressed in the mammary gland of a pregnant multiparous seal, but not in a seal in its first pregnancy. While bovine CTI is expressed for 24-48 h postpartum (pp) and secreted in colostrum only, Cape fur seal CTI was detected for at least 2-3 months pp while the mother was suckling its young on-shore. Furthermore, CTI was expressed in the mammary gland of only one of the lactating seals that was foraging at-sea. The expression of β-casein (CSN2) and β-lactoglobulin II (LGB2), but not CTI in the second lactating seal foraging at-sea suggested that CTI may be intermittently expressed during lactation. Cape fur seal and walrus CTI encode putative small, secreted, N-glycosylated proteins with a single Kunitz/bovine pancreatic trypsin inhibitor (BPTI) domain indicative of serine protease inhibition. Mature Cape fur seal CTI shares 92% sequence identity with Pacific walrus CTI, but only 35% identity with BPTI. Structural homology modelling of Cape fur seal CTI and Pacific walrus trypsin based on the model of the second Kunitz domain of human tissue factor pathway inhibitor (TFPI) and porcine trypsin (Protein Data Bank: 1TFX) confirmed that CTI inhibits trypsin in a canonical fashion. Therefore, pinniped CTI may be critical for preventing the proteolytic degradation of immunoglobulins that are passively transferred from mother to young via colostrum and milk. Copyright © 2015 Elsevier B.V. All rights reserved.

  4. Electronic structure of trypsin inhibitor from squash seeds in aqueous solution

    Science.gov (United States)

    Zheng, Haoping

    2000-10-01

    The electronic structure of the trypsin inhibitor from seeds of the squash Cucurbita maxima (CMTI-I) in aqueous solution is obtained by ab initio, all-electron, full-potential calculations using the self-consistent cluster-embedding (SCCE) method. The reactive site of the inhibitor is explained theoretically, which is in agreement with the experimental results. It is shown that the coordinates of oxygen atoms in the inhibitor, determined by nuclear magnetic resonance and combination of distance geometry and dynamical simulated annealing, are systematically less accurate than that of other kinds of heavy atoms.

  5. Bioinsecticidal activity of a novel Kunitz trypsin inhibitor from Catanduva (Piptadenia moniliformis) seeds.

    Science.gov (United States)

    Cruz, Ana C B; Massena, Fábio S; Migliolo, Ludovico; Macedo, Leonardo L P; Monteiro, Norberto K V; Oliveira, Adeliana S; Macedo, Francisco P; Uchoa, Adriana F; Grossi de Sá, Maria F; Vasconcelos, Ilka M; Murad, Andre M; Franco, Octavio L; Santos, Elizeu A

    2013-09-01

    The present study aims to provide new in vitro and in vivo biochemical information about a novel Kunitz trypsin inhibitor purified from Piptadenia moniliformis seeds. The purification process was performed using TCA precipitation, Trypsin-Sepharose and reversed-phase C18 HPLC chromatography. The inhibitor, named PmTKI, showed an apparent molecular mass of around 19 kDa, visualized by SDS-PAGE, which was confirmed by mass spectrometry MALDI-ToF demonstrating a monoisotopic mass of 19.296 Da. The inhibitor was in vitro active against trypsin, chymotrypsin and papain. Moreover, kinetic enzymatic studies were performed aiming to understand the inhibition mode of PmTKI, which competitively inhibits the target enzyme, presenting Ki values of 1.5 × 10(-8) and 3.0 × 10(-1) M against trypsin and chymotrypsin, respectively. Also, the inhibitory activity was assayed at different pH ranges, temperatures and reduction environments (DTT). The inhibitor was stable in all conditions maintaining an 80% residual activity. N-terminal sequence was obtained by Edman degradation and the primary sequence presented identity with members of Kunitz-type inhibitors from the same subfamily. Finally after biochemical characterization the inhibitory effect was evaluated in vitro on insect digestive enzymes from different orders, PmTKI demonstrated remarkable activity against enzymes from Anthonomus grandis (90%), Plodia interpuncptella (60%), and Ceratitis capitata (70%). Furthermore, in vivo bioinsecticidal assays of C. capitata larvae were also performed and the concentration of PmTKI (w/w) in an artificial diet required to LD50 and ED50 larvae were 0.37 and 0.3% respectively. In summary, data reported here shown the biotechnological potential of PmTKI for insect pest control. Copyright © 2013 Elsevier Masson SAS. All rights reserved.

  6. Spleen

    International Nuclear Information System (INIS)

    Mittelstaedt, C.A.; McCartney, W.H.; Mauro, M.A.; Vincent, L.M.; Peterson, N.P.; Staab, E.V.

    1984-01-01

    Imaging modalities currently available for the evaluation of splenic disorders include /sup 99m/Tc-sulfur colloid (TcSC) radionuclide splenic scintigraphy, ultrasound, and computed tomography (CT). These techniques produce images of the spleen on the basis of reticuloendothelial function, echogenic properties, and x-ray attenuation, respectively. No one technique is clearly superior to another for all clinical situations or disease processes. Each modality has its strengths and limitations that relate to the properties being measured (function, echogenicity, attenuation) and the disease processes involving the spleen. This chapter offers a combined approach to splenic imaging in the evaluation of focal disease, diffuse disease, traumatic disorders, normal variations, congenital anomalies, and perisplenic disorders

  7. Bauhinia variegata var. variegata trypsin inhibitor: From isolation to potential medicinal applications

    Energy Technology Data Exchange (ETDEWEB)

    Fang, Evandro Fei; Wong, Jack Ho [School of Biomedical Sciences, Faculty of Medicine, The Chinese University of Hong Kong, Shatin, Hong Kong SAR (China); Bah, Clara Shui Fern [Department of Food Science, Division of Sciences, University of Otago (New Zealand); Lin, Peng [School of Biomedical Sciences, Faculty of Medicine, The Chinese University of Hong Kong, Shatin, Hong Kong SAR (China); Tsao, Sai Wah [Department of Anatomy, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Sassoon Road, Pokfulam, Hong Kong SAR (China); Ng, Tzi Bun, E-mail: b021770@mailserv.cuhk.edu.hk [School of Biomedical Sciences, Faculty of Medicine, The Chinese University of Hong Kong, Shatin, Hong Kong SAR (China)

    2010-06-11

    Here we report for the first time of a new Kunitz-type trypsin inhibitor (termed BvvTI) from seeds of the Camel's foot tree, Bauhinia variegata var. variegata. BvvTI shares the same reactive site residues (Arg, Ser) and exhibits a homology of N-terminal amino acid sequence to other Bauhinia protease inhibitors. The trypsin inhibitory activity (K{sub i}, 0.1 x 10{sup -9} M) of BvvTI ranks the highest among them. Besides anti-HIV-1 reverse transcriptase activity, BvvTI could significantly inhibit the proliferation of nasopharyngeal cancer CNE-1 cells in a selective way. This may partially be contributed by its induction of cytokines and apoptotic bodies. These results unveil potential medicinal applications of BvvTI.

  8. Bauhinia variegata var. variegata trypsin inhibitor: from isolation to potential medicinal applications.

    Science.gov (United States)

    Fang, Evandro Fei; Wong, Jack Ho; Bah, Clara Shui Fern; Lin, Peng; Tsao, Sai Wah; Ng, Tzi Bun

    2010-06-11

    Here we report for the first time of a new Kunitz-type trypsin inhibitor (termed BvvTI) from seeds of the Camel's foot tree, Bauhinia variegata var. variegata. BvvTI shares the same reactive site residues (Arg, Ser) and exhibits a homology of N-terminal amino acid sequence to other Bauhinia protease inhibitors. The trypsin inhibitory activity (K(i), 0.1 x 10(-9)M) of BvvTI ranks the highest among them. Besides anti-HIV-1 reverse transcriptase activity, BvvTI could significantly inhibit the proliferation of nasopharyngeal cancer CNE-1 cells in a selective way. This may partially be contributed by its induction of cytokines and apoptotic bodies. These results unveil potential medicinal applications of BvvTI. (c) 2010 Elsevier Inc. All rights reserved.

  9. Bauhinia variegata var. variegata trypsin inhibitor: From isolation to potential medicinal applications

    International Nuclear Information System (INIS)

    Fang, Evandro Fei; Wong, Jack Ho; Bah, Clara Shui Fern; Lin, Peng; Tsao, Sai Wah; Ng, Tzi Bun

    2010-01-01

    Here we report for the first time of a new Kunitz-type trypsin inhibitor (termed BvvTI) from seeds of the Camel's foot tree, Bauhinia variegata var. variegata. BvvTI shares the same reactive site residues (Arg, Ser) and exhibits a homology of N-terminal amino acid sequence to other Bauhinia protease inhibitors. The trypsin inhibitory activity (K i , 0.1 x 10 -9 M) of BvvTI ranks the highest among them. Besides anti-HIV-1 reverse transcriptase activity, BvvTI could significantly inhibit the proliferation of nasopharyngeal cancer CNE-1 cells in a selective way. This may partially be contributed by its induction of cytokines and apoptotic bodies. These results unveil potential medicinal applications of BvvTI.

  10. Crystallization and preliminary X-ray diffraction studies of Murraya koenigii trypsin inhibitor

    Energy Technology Data Exchange (ETDEWEB)

    Shee, Chandan [Department of Biotechnology, Indian Institute of Technology Roorkee, Roorkee 247 667 (India); Singh, Tej P. [Department of Biophysics, All India Institute of Medical Sciences, New Delhi 100 029 (India); Kumar, Pravindra, E-mail: kumarfbs@iitr.ernet.in; Sharma, Ashwani K., E-mail: kumarfbs@iitr.ernet.in [Department of Biotechnology, Indian Institute of Technology Roorkee, Roorkee 247 667 (India)

    2007-04-01

    A Kunitz-type trypsin inhibitor purified from the seeds of Murraya koenigii has been crystallized by the sitting-drop vapour-diffusion method using PEG 8000 as the precipitating agent. A Kunitz-type trypsin inhibitor purified from the seeds of Murraya koenigii has been crystallized by the sitting-drop vapour-diffusion method using PEG 8000 as the precipitating agent. The crystals belong to the tetragonal space group P4{sub 3}2{sub 1}2, with unit-cell parameters a = b = 75.8, c = 150.9 Å. The crystals contain two molecules in the asymmetric unit with a V{sub M} value of 2.5 Å{sup 3} Da{sup −1}. Diffraction was observed to 2.65 Å resolution and a complete data set was collected to 2.9 Å resolution.

  11. Effects of soybean Kunitz trypsin inhibitor on the cotton boll weevil (Anthonomus grandis).

    Science.gov (United States)

    Franco, Octávio L; Dias, Simoni C; Magalhães, Claudio P; Monteiro, Ana C S; Bloch, Carlos; Melo, Francislete R; Oliveira-Neto, Osmundo B; Monnerat, Rose G; Grossi-de-Sá, Maria Fátima

    2004-01-01

    The cotton boll weevil, Anthonomus grandis, is an economically important pest of cotton in tropical and subtropical areas of several countries in the Americas, causing severe losses due to their damage in cotton floral buds. Enzymatic assays using gut extracts from larval and adult boll weevil have demonstrated the presence of digestive serine proteinase-like activities. Furthermore, in vitro assays showed that soybean Kunitz trypsin inhibitor (SKTI) was able to inhibit these enzymes. Previously, in vivo effects of black-eyed pea trypsin chymotrypsin inhibitor (BTCI) have been demonstrated towards the boll weevil pest. Here, when neonate larvae were reared on an artificial diet containing SKTI at three different concentrations, a reduction of larval weight of up to 64% was observed for highest SKTI concentration 500 microM. The presence of SKTI caused an increase in mortality and severe deformities of larvae, pupae and adult insects. This work therefore represents the first observation of a Kunitz trypsin inhibitor active in vivo and in vitro against A. grandis. Bioassays suggested that SKTI could be used as a tool in engineering crop plants, which might exhibit increased resistance against cotton boll weevil.

  12. Proteolytic and Trypsin Inhibitor Activity in Germinating Jojoba Seeds (Simmondsia chinensis) 1

    Science.gov (United States)

    Samac, Deborah; Storey, Richard

    1981-01-01

    Changes in proteolytic activity (aminopeptidase, carboxypeptidase, endopeptidase) were followed during germination (imbibition through seedling development) in extracts from cotyledons of jojoba seeds (Simmondsia chinensis). After imbibition, the cotyledons contained high levels of sulfhydryl aminopeptidase activity (APA) but low levels of serine carboxypeptidase activity (CPA). CPA increased with germination through the apparent loss of a CPA inhibitor substance in the seed. Curves showing changes in endopeptidase activity (EPA) assayed at pH 4, 5, 6, 7, and 8 during germination were distinctly different. EPA at pH 4, 5, 6, and 7 showed characteristics of sulfhydryl enzymes while activity at pH 8 was probably due to a serine type enzyme. EPA at pH 6 was inhibited early in germination by one or more substances in the seed. Activities at pH 5 and later at pH 6 were the highest of all EPA throughout germination and increases in these activities were associated with a rapid loss of protein from the cotyledons of the developing seedling. Jojoba cotyledonary extracts were found to inhibit the enzymic activity of trypsin, chymotrypsin, and pepsin but not the protease from Aspergillus saotoi. The heat-labile trypsin inhibitor substance(s) was found in commercially processed jojoba seed meal and the albumin fraction of seed proteins. Trypsin inhibitor activity decreased with germination. PMID:16662104

  13. Proteolytic and Trypsin Inhibitor Activity in Germinating Jojoba Seeds (Simmondsia chinensis).

    Science.gov (United States)

    Samac, D; Storey, R

    1981-12-01

    Changes in proteolytic activity (aminopeptidase, carboxypeptidase, endopeptidase) were followed during germination (imbibition through seedling development) in extracts from cotyledons of jojoba seeds (Simmondsia chinensis). After imbibition, the cotyledons contained high levels of sulfhydryl aminopeptidase activity (APA) but low levels of serine carboxypeptidase activity (CPA). CPA increased with germination through the apparent loss of a CPA inhibitor substance in the seed. Curves showing changes in endopeptidase activity (EPA) assayed at pH 4, 5, 6, 7, and 8 during germination were distinctly different. EPA at pH 4, 5, 6, and 7 showed characteristics of sulfhydryl enzymes while activity at pH 8 was probably due to a serine type enzyme. EPA at pH 6 was inhibited early in germination by one or more substances in the seed. Activities at pH 5 and later at pH 6 were the highest of all EPA throughout germination and increases in these activities were associated with a rapid loss of protein from the cotyledons of the developing seedling.Jojoba cotyledonary extracts were found to inhibit the enzymic activity of trypsin, chymotrypsin, and pepsin but not the protease from Aspergillus saotoi. The heat-labile trypsin inhibitor substance(s) was found in commercially processed jojoba seed meal and the albumin fraction of seed proteins. Trypsin inhibitor activity decreased with germination.

  14. Overlapping binding sites for trypsin and papain on a Kunitz-type proteinase inhibitor from Prosopis juliflora.

    Science.gov (United States)

    Franco, Octávio L; Grossi de Sá, Maria F; Sales, Maurício P; Mello, Luciane V; Oliveira, Adeliana S; Rigden, Daniel J

    2002-11-15

    Proteinase inhibitors are among the most promising candidates for expression by transgenic plants and consequent protection against insect predation. However, some insects can respond to the threat of the proteinase inhibitor by the production of enzymes insensitive to inhibition. Inhibitors combining more than one favorable activity are therefore strongly favored. Recently, a known small Kunitz trypsin inhibitor from Prosopis juliflora (PTPKI) has been shown to possess unexpected potent cysteine proteinase inhibitory activity. Here we show, by enzyme assay and gel filtration, that, unlike other Kunitz inhibitors with dual activities, this inhibitor is incapable of simultaneous inhibition of trypsin and papain. These data are most readily interpreted by proposing overlapping binding sites for the two enzymes. Molecular modeling and docking experiments favor an interaction mode in which the same inhibitor loop that interacts in a canonical fashion with trypsin can also bind into the papain catalytic site cleft. Unusual residue substitutions at the proposed interface can explain the relative rarity of twin trypsin/papain inhibition. Other changes seem responsible for the relative low affinity of PTPKI for trypsin. The predicted coincidence of trypsin and papain binding sites, once confirmed, would facilitate the search, by phage display for example, for mutants highly active against both proteinases. Copyright 2002 Wiley-Liss, Inc.

  15. Amino Acid Composition, Urease Activity and Trypsin Inhibitor Activity after Toasting of Soybean in Thick and Thin Layer

    OpenAIRE

    Krička, Tajana; Jurišić, Vanja; Voća, Neven; Ćurić, Duška; Brlek Savić, Tea; Matin, Ana

    2009-01-01

    The objective of this study was to determine amino acid content, urease activity and trypsin inhibitor activity in soybean grain for polygastric animals’ feed aft er toasting with the aim to introduce thick layer in toasting technology. Hence, soybean was toasted both in thick and thin layer at 130 oC during 10 minutes. In order to properly monitor the technological process of soybean thermal processing, it was necessary to study crude protein content, urease activity, trypsin inhibitor activ...

  16. Screening and purification of a novel trypsin inhibitor from Prosopis juliflora seeds with activity toward pest digestive enzymes.

    Science.gov (United States)

    Sivakumar, S; Franco, O L; Tagliari, P D; Bloch, C; Mohan, M; Thayumanavan, B

    2005-08-01

    Several pests are capable of decreasing crop production causing severe economical and social losses. Aiming to find novel molecules that could impede the digestion process of different pests, a screening of alpha-amylase and trypsin-like proteinase inhibitors was carried out in Prosopis juliflora, showing the presence of both in dry seeds. Furthermore, a novel trypsin inhibitor, with molecular mass of 13,292 Da, was purified showing remarkable in vitro activity against T. castaneum and C. maculatus.

  17. Modifications outside the proteinase binding loop in Cucurbita maxima trypsin inhibitor III (CMTI-III) analogues change the binding energy with bovine beta-trypsin.

    Science.gov (United States)

    Jaśkiewicz, A; Lis, K; Rózycki, J; Kupryszewski, G; Rolka, K; Ragnarsson, U; Zbyryt, T; Wilusz, T

    1998-10-02

    Five 26-peptide analogues of the trypsin inhibitor [Pro18]CMTI-III containing Leu or Tyr in position 7 and Val or Tyr in position 27: 1 (Leu7, Tyr27), 2 (Tyr7, Val27), 3 (Tyr7, Tyr27), 4 (Leu7, Val27) and 5 (Leu7, Ala18, Tyr27) were synthesized by the solid-phase method. Analogues 1-4 displayed Ka with bovine beta-trypsin of the same order of magnitude as the wild CMTI-III inhibitor, whereas for analogue 5, this value was lower by about 3 orders of magnitude. This indicated that for the analogues with Pro (but not with Ala) in position 18, the side-chain interactions between positions 7 and 27 did not play a critical role for the stabilization of the active structure. In addition, these results also suggest that Tyr7 is involved in an additional aromatic interaction with position 41 of the enzyme.

  18. Changing the inhibitory specificity and function of Cucurbita maxima trypsin inhibitor-V by site-directed mutagenesis.

    Science.gov (United States)

    Wen, L; Lee, I; Chen, G; Huang, J K; Gong, Y; Krishnamoorthi, R

    1995-02-27

    Cucurbita maxima trypsin inhibitor-V (CMTI-V) is also a specific inhibitor of human blood coagulation factor beta-factor XIIa. A recombinant version of CMTI-V has allowed probing of roles of individual amino acid residues including the reactive site residue, lysine (P1), by site-directed mutagenesis. The K44R showed at least a 5-fold increase in inhibitory activity toward human beta-factor XIIa, while there was no change toward bovine trypsin. This result demonstrates that beta-factor-XIIa prefers an arginine residue over lysine residue, while trypsin is non-specific to lysine or arginine in its binding pocket. On the other hand, the specificity of CMTI-V could be changed from trypsin to chymotrypsin inhibition by mutation of the P1 residue to either leucine or methionine (K44L or K44M).

  19. BstXI RFLP in the human inter-alpha-trypsin inhibitor light chain gene

    Energy Technology Data Exchange (ETDEWEB)

    Leveillard, T; Bourguignon, J; Sesbouee, R; Hanauer, A; Salier, J P; Diarra-Mehrpour, M; Martin, J P

    1988-03-25

    The 1.2 kb EcoRI/SmaI fragment of lambdaHuLITI2 was used as probe. lambdaHuLITI2 is a full length cDNA clone coding for human inter-alpha-trypsin inhibitor light chain isolated from immunochemical screening of a lambdagt11 library. Its sequence coding for HI-30 and alpha-1-microglobulin is in agreement. BstXI identifies five invariant bands at 5.0 kb, 2.3 kb, 1.5 kb, 1.1 kb, and 0.7 kb and a diallelic polymorphism with DNA fragments at 2.0 kb or 1.7 kb.

  20. New analogues of Cucurbita maxima trypsin inhibitor III (CMTI III) with simplified structure.

    Science.gov (United States)

    Rolka, K; Kupryszewski, G; Rózycki, J; Ragnarsson, U; Zbyryt, T; Otlewski, J

    1992-10-01

    Seven new analogues of trypsin inhibitor CMTI III were obtained by solid-phase peptide synthesis. Three analogues contained only two, instead of three, disulfide bridges, whereas the molecules of the next four analogues were shortened at the N- and/or C-terminus. The elimination of one disulfide bridge in CMTI III induces a decrease in the association equilibrium constants by 6-7 orders of magnitude, whereas the removal of one, two or three amino-acid residues at the N- and/or C-terminus does not significantly affect the activity.

  1. Toxicity to cotton boll weevil Anthonomus grandis of a trypsin inhibitor from chickpea seeds.

    Science.gov (United States)

    de P G Gomes, Angélica; Dias, Simoni C; Bloch, Carlos; Melo, Francislete R; Furtado, José R; Monnerat, Rose G; Grossi-de-Sá, Maria F; Franco, Octávio L

    2005-02-01

    Cotton (Gossypium hirsutum L.) is an important agricultural commodity, which is attacked by several pests such as the cotton boll weevil Anthonomus grandis. Adult A. grandis feed on fruits and leaf petioles, reducing drastically the crop production. The predominance of boll weevil digestive serine proteinases has motivated inhibitor screenings in order to discover new ones with the capability to reduce the digestion process. The present study describes a novel proteinase inhibitor from chickpea seeds (Cicer arietinum L.) and its effects against A. grandis. This inhibitor, named CaTI, was purified by using affinity Red-Sepharose Cl-6B chromatography, followed by reversed-phase HPLC (Vydac C18-TP). SDS-PAGE and MALDI-TOF analyses, showed a unique monomeric protein with a mass of 12,877 Da. Purified CaTI showed significant inhibitory activity against larval cotton boll weevil serine proteinases (78%) and against bovine pancreatic trypsin (73%), when analyzed by fluorimetric assays. Although the molecular mass of CaTI corresponded to alpha-amylase/trypsin bifunctional inhibitors masses, no inhibitory activity against insect and mammalian alpha-amylases was observed. In order to observe CaTI in vivo effects, an inhibitor rich fraction was added to an artificial diet at different concentrations. At 1.5% (w/w), CaTI caused severe development delay, several deformities and a mortality rate of approximately 45%. These results suggested that CaTI could be useful in the production of transgenic cotton plants with enhanced resistance toward cotton boll weevil.

  2. Effect of radiation and soaking on trypsin inhibitor and protein content of chickpea (Cicer arietinum L.)

    International Nuclear Information System (INIS)

    Sattar, A.; Atta, S.; Akhtar, M.A.

    1990-01-01

    Composition of Ascochyta-blight resistant chickpea for proximate components, vitamins, energy and trypsin inhibitor, was determined. The influence of irradiation and soaking at ambient temperatures (25-35deg C) on trypsin inhibitor activity (TIA) and protein content of chickpea, was investigated. A significant linear relation (r = -0.960 to -0.987) was found between the loss of TIA and soaking time of irradiated and unirradiated seeds (p < 0.05) and the rate of loss increased with increasing radiation dose (0.25-1.00 kGy). However, effect of radiation alone was negligible. Maximum decrease (30.7%) in TIA (from 330.0 to 228.6 TiU/g) occurred during soaking for 12 h of 1.00 kGy sample. The protein contents increased from an initial value of 21.7% to 23.4% and 22.7% as a result of soaking for 12 h in tap and distilled waters, respectively. Radiation treatment exhibited little or no effect. (author)

  3. Allium sativum Protease Inhibitor: A Novel Kunitz Trypsin Inhibitor from Garlic Is a New Comrade of the Serpin Family.

    Science.gov (United States)

    Shamsi, Tooba Naz; Parveen, Romana; Amir, Mohd; Baig, Mohd Affan; Qureshi, M Irfan; Ali, Sher; Fatima, Sadaf

    2016-01-01

    This study was aimed to purify and characterize the Protease inhibitor (PI) from a plant Allium sativum (garlic) with strong medicinal properties and to explore its phytodrug potentials. Allium sativum Protease Inhibitor (ASPI) was purified using ammonium sulphate fractionation and Fast Protein Liquid Chromatography on anion exchanger Hi-Trap DEAE column. The purified protein was analyzed for its purity and molecular weight by SDS PAGE. The confirmation of presence of trypsin inhibiting PI was performed by MALDI TOF-TOF and analyzed by MASCOT database. The ASPI was further investigated for its kinetic properties and stability under extreme conditions of pH, temperature and chemical denaturants. Secondary structure was determined by Circular Dichorism (CD) spectroscopy. ASPI of ~15 kDa inhibited trypsin and matched "truncated kunitz Trypsin Inhibitor (Glycine max)" in MASCOT database. The purified ASPI showed 30376.1371 U/mg specific activity with a fold purity of 159.92 and yield ~93%. ASPI was quite stable in the range of pH 2-12 showing a decline in the activity around pH 4-5 suggesting that the pI value of the protein as ASPI aggregates in this range. ASPI showed stability to a broad range of temperature (10-80°C) but declined beyond 80°C. Further, detergents, oxidizing agents and reducing agents demonstrated change in ASPI activity under varying concentrations. The kinetic analysis revealed sigmoidal relationship of velocity with substrate concentration with Vmax 240.8 (μM/min) and Km value of 0.12 μM. ASPI showed uncompetitive inhibition with a Ki of 0.08±0.01 nM). The Far UV CD depicted 2.0% α -helices and 51% β -sheets at native pH. To conclude, purified ~15 kDa ASPI exhibited fair stability in wide range of pH and temperature Overall, there was an increase in purification fold with remarkable yield. Chemical modification studies suggested the presence of lysine and tryptophan residues as lead amino acids present in the reactive sites. Therefore, ASPI

  4. Allium sativum Protease Inhibitor: A Novel Kunitz Trypsin Inhibitor from Garlic Is a New Comrade of the Serpin Family.

    Directory of Open Access Journals (Sweden)

    Tooba Naz Shamsi

    Full Text Available This study was aimed to purify and characterize the Protease inhibitor (PI from a plant Allium sativum (garlic with strong medicinal properties and to explore its phytodrug potentials.Allium sativum Protease Inhibitor (ASPI was purified using ammonium sulphate fractionation and Fast Protein Liquid Chromatography on anion exchanger Hi-Trap DEAE column. The purified protein was analyzed for its purity and molecular weight by SDS PAGE. The confirmation of presence of trypsin inhibiting PI was performed by MALDI TOF-TOF and analyzed by MASCOT database. The ASPI was further investigated for its kinetic properties and stability under extreme conditions of pH, temperature and chemical denaturants. Secondary structure was determined by Circular Dichorism (CD spectroscopy.ASPI of ~15 kDa inhibited trypsin and matched "truncated kunitz Trypsin Inhibitor (Glycine max" in MASCOT database. The purified ASPI showed 30376.1371 U/mg specific activity with a fold purity of 159.92 and yield ~93%. ASPI was quite stable in the range of pH 2-12 showing a decline in the activity around pH 4-5 suggesting that the pI value of the protein as ASPI aggregates in this range. ASPI showed stability to a broad range of temperature (10-80°C but declined beyond 80°C. Further, detergents, oxidizing agents and reducing agents demonstrated change in ASPI activity under varying concentrations. The kinetic analysis revealed sigmoidal relationship of velocity with substrate concentration with Vmax 240.8 (μM/min and Km value of 0.12 μM. ASPI showed uncompetitive inhibition with a Ki of 0.08±0.01 nM. The Far UV CD depicted 2.0% α -helices and 51% β -sheets at native pH.To conclude, purified ~15 kDa ASPI exhibited fair stability in wide range of pH and temperature Overall, there was an increase in purification fold with remarkable yield. Chemical modification studies suggested the presence of lysine and tryptophan residues as lead amino acids present in the reactive sites

  5. A trypsin inhibitor from rambutan seeds with antitumor, anti-HIV-1 reverse transcriptase, and nitric oxide-inducing properties.

    Science.gov (United States)

    Fang, Evandro Fei; Ng, Tzi Bun

    2015-04-01

    Nephelium lappaceum L., commonly known as "rambutan," is a typical tropical tree and is well known for its juicy and sweet fruit which has an exotic flavor. Chemical studies on rambutan have led to the identification of various components such as monoterpene lactones and volatile compounds. Here, a 22.5-kDa trypsin inhibitor (N . lappaceum trypsin inhibitor (NLTI)) was isolated from fresh rambutan seeds using liquid chromatographical techniques. NLTI reduced the proteolytic activities of both trypsin and α-chymotrypsin. Dithiothreitol reduced the trypsin inhibitory activity of NLTI at a concentration of 1 mM, indicating that an intact disulfide bond is essential to the activity. NLTI inhibited HIV-1 reverse transcriptase with an IC50 of 0.73 μM. In addition, NLTI manifested a time- and dose-dependent inhibitory effect on growth in many tumor cells. NLTI is one of the few trypsin inhibitors with nitric oxide-inducing activity and may find application in tumor therapy.

  6. New active analogues of Cucurbita maxima trypsin inhibitor III (CMTI-III) modified in the non-contact region.

    Science.gov (United States)

    Rózycki, J; Kupryszewski, G; Rolka, K; Ragnarsson, U; Zbyryt, T; Krokoszyńska, I; Wilusz, T

    1994-01-01

    Four new analogues of trypsin inhibitor CMTI-III(3-28) = [desArg1,desVal2,desGly29]CMTI-III which was recently shown to be fully active, were synthesized by the solid-phase method. The introduction of glycine in position 9 (peptide 1) and Gly-Pro-Gly (peptide 2) and Gly-Pro-Asn (peptide 3) in the regions 17-19 and 23-25, respectively, did not change the antitrypsin activity of all modified peptides. All of these substitutions are presumed to be outside the trypsin-binding loop as judged from the X-ray structure of the complex between beta-trypsin and the related inhibitor CMTI-I. Also the fourth analogue which was substituted in all the positions mentioned, exhibited the full activity.

  7. Effect of irradiation and germination on trypsin inhibitor and protein content of chickpea (Cicer arietinum L.)

    International Nuclear Information System (INIS)

    Sattar, A.; Atta, S.; Akhtar, M.A.

    1990-01-01

    Effect of irradiation (0.005-0.20 kGy) and subsequent germination on trypsin inhibitor activity (TIA) and protein content of chickpea, was studied. The results revealed a significant linear relation (r = -0.981 to -0.992) between the loss of TIA and germination time, and the rate of TIA destruction increased with irradiation dose (p < 0.05). Maximum destruction (43.8%) of the TIA occurred on germination for 120 hr of 0.20 kGy sample (from 330.0 to 185.3 TIU/g). Initially protein content was 21.7% and the value significantly increased to maximum levels of 27.5% and 27.9% in distilled and tap water respectively during germination for 120 hr of 0.10 kGy sample (p < 0.05). Protein contents were not affected by irradiation

  8. Biochemical response of Anticarsia gemmatalis fed with soybean plants pulverized with the synthetic trypsin inhibitor benzamidine

    International Nuclear Information System (INIS)

    Oliveira, M.G.A.; Pilon, A.M.; Pilon, F.M.; Ribeiro, F.R.; Silva, F.C.; Ribon, A.O.B.; Reis, A.P.; Visotto, L.E.; Guedes, R.N.C.; Oliveira, J.A.

    2008-01-01

    Full text: Insects are responsible for severe crop losses. New alternatives for pest control other than agrochemicals have been investigated. Protease inhibitors are one of the prime candidates effective against insect pests. In this work we studied the effect of the synthetic trypsin inhibitor benzamidine on the development of Anticarsia gemmatalis, an important pest of the soybean culture. Larvae were reared on soybean plants containing 0.00, 0.15, 0.30, 0.45, 0.60 and 0.75% (w/w) of benzamidine. After 6, 12, 24 and 48 h of feeding midgut extracts were prepared and assayed for enzymatic activity (proteolytic, amidasic and stearic). Benzamidine altered the activity patterns but was not able to totally abolish enzyme activity. The proteolytic, amidasic and stearic activity showed the higher time of inhibition in 48 h in concentration of 0,75%, the inhibition was the around 93%, 63.1% and 36.6%, respectively. We suggest that the presence of inhibitor has made insects to adapt and produce proteases which are insensitive to the action of benzamidine. (author)

  9. Biochemical response of Anticarsia gemmatalis fed with soybean plants pulverized with the synthetic trypsin inhibitor benzamidine

    Energy Technology Data Exchange (ETDEWEB)

    Oliveira, M.G.A.; Pilon, A.M.; Pilon, F.M.; Ribeiro, F.R.; Silva, F.C.; Ribon, A.O.B.; Reis, A.P.; Visotto, L.E. [Universidade Federal de Vicosa (UFV), Belo Horizonte, MG (Brazil). Dept. de Bioquimica e Biologia Molecular; Guedes, R.N.C. [Universidade Federal de Vicosa (UFV), Belo Horizonte, MG (Brazil). Dept. de Biologia Animal; Oliveira, J.A. [Universidade Federal de Vicosa (UFV), Belo Horizonte, MG (Brazil). Dept. de Quimica

    2008-07-01

    Full text: Insects are responsible for severe crop losses. New alternatives for pest control other than agrochemicals have been investigated. Protease inhibitors are one of the prime candidates effective against insect pests. In this work we studied the effect of the synthetic trypsin inhibitor benzamidine on the development of Anticarsia gemmatalis, an important pest of the soybean culture. Larvae were reared on soybean plants containing 0.00, 0.15, 0.30, 0.45, 0.60 and 0.75% (w/w) of benzamidine. After 6, 12, 24 and 48 h of feeding midgut extracts were prepared and assayed for enzymatic activity (proteolytic, amidasic and stearic). Benzamidine altered the activity patterns but was not able to totally abolish enzyme activity. The proteolytic, amidasic and stearic activity showed the higher time of inhibition in 48 h in concentration of 0,75%, the inhibition was the around 93%, 63.1% and 36.6%, respectively. We suggest that the presence of inhibitor has made insects to adapt and produce proteases which are insensitive to the action of benzamidine. (author)

  10. Amino-acid sequences of trypsin inhibitors from watermelon (Citrullus vulgaris) and red bryony (Bryonia dioica) seeds.

    Science.gov (United States)

    Otlewski, J; Whatley, H; Polanowski, A; Wilusz, T

    1987-11-01

    The amino-acid sequences of two trypsin inhibitors isolated from red bryony (Bryonia dioica) and watermelon (Citrullus vulgaris) seeds are reported. Both species represent different genera of the Cucurbitaceae family, which have not been previously investigated as a source of proteinase inhibitors. The sequences are unique but are very similar to those of other proteinase inhibitors which have been isolated from squash seeds. Based on structural homology we assume that the Arg5-Ile6 peptide bond represents the reactive site bond of both inhibitors.

  11. An unusual helix-turn-helix protease inhibitory motif in a novel trypsin inhibitor from seeds of Veronica (Veronica hederifolia L.).

    Science.gov (United States)

    Conners, Rebecca; Konarev, Alexander V; Forsyth, Jane; Lovegrove, Alison; Marsh, Justin; Joseph-Horne, Timothy; Shewry, Peter; Brady, R Leo

    2007-09-21

    The storage tissues of many plants contain protease inhibitors that are believed to play an important role in defending the plant from invasion by pests and pathogens. These proteinaceous inhibitor molecules belong to a number of structurally distinct families. We describe here the isolation, purification, initial inhibitory properties, and three-dimensional structure of a novel trypsin inhibitor from seeds of Veronica hederifolia (VhTI). The VhTI peptide inhibits trypsin with a submicromolar apparent K(i) and is expected to be specific for trypsin-like serine proteases. VhTI differs dramatically in structure from all previously described families of trypsin inhibitors, consisting of a helix-turn-helix motif, with the two alpha helices tightly associated by two disulfide bonds. Unusually, the crystallized complex is in the form of a stabilized acyl-enzyme intermediate with the scissile bond of the VhTI inhibitor cleaved and the resulting N-terminal portion of the inhibitor remaining attached to the trypsin catalytic serine 195 by an ester bond. A synthetic, truncated version of the VhTI peptide has also been produced and co-crystallized with trypsin but, surprisingly, is seen to be uncleaved and consequently forms a noncovalent complex with trypsin. The VhTI peptide shows that effective enzyme inhibitors can be constructed from simple helical motifs and provides a new scaffold on which to base the design of novel serine protease inhibitors.

  12. Three-dimensional solution structure of Cucurbita maxima trypsin inhibitor-V determined by NMR spectroscopy.

    Science.gov (United States)

    Cai, M; Gong, Y; Kao, J L; Krishnamoorthi, R

    1995-04-18

    The solution structure of Cucurbita maxima trypsin inhibitor-V (CMTI-V), which is also a specific inhibitor of the blood coagulation protein, factor XIIa, was determined by 1H NMR spectroscopy in combination with a distance-geometry and simulated annealing algorithm. Sequence-specific resonance assignments were made for all the main-chain and most of the side-chain hydrogens. Stereospecific assignments were also made for some of the beta-, gamma-, delta-, and epsilon-hydrogens and valine methyl hydrogens. The ring conformations of all six prolines in the inhibitor were determined on the basis of 1H-1H vicinal coupling constant patterns; most of the proline ring hydrogens were stereospecifically assigned on the basis of vicinal coupling constant and intraresidue nuclear Overhauser effect (NOE) patterns. Distance constraints were determined on the basis of NOEs between pairs of hydrogens. Dihedral angle constraints were determined from estimates of scalar coupling constants and intraresidue NOEs. On the basis of 727 interproton distance and 111 torsion angle constraints, which included backbone phi angles and side-chain chi 1, chi 2, chi 3, and chi 4 angles, 22 structures were calculated by a distance geometry algorithm and refined by energy minimization and simulated annealing methods. Both main-chain and side-chain atoms are well-defined, except for a loop region, two terminal residues, and some side-chain atoms located on the molecular surface. The average root mean squared deviation in the position for equivalent atoms between the 22 individual structures and the mean structure obtained by averaging their coordinates is 0.58 +/- 0.06 A for the main-chain atoms and 1.01 +/- 0.07 A for all the non-hydrogen atoms of residues 3-40 and 49-67. These structures were compared to the X-ray crystallographic structure of another protein of the same inhibitor family-chymotrypsin inhibitor-2 from barley seeds [CI-2; McPhalen, C. A., & James, M. N. G. (1987) Biochemistry 26

  13. Nuclear magnetic resonance studies of trypsin inhibitors isolated from seeds of Cucurbitaceae plants

    International Nuclear Information System (INIS)

    Sobczyk, K.; Markley, J.L.

    1986-01-01

    Two-dimensional NMR techniques were used to assign individual protons in two trypsin inhibitors isolated from squash seeds (Cucurbita maxima and C. pepo). The assignments of 1 H NMR signals from these unusually small proteins (29 amino acid residues) were based entirely on the sequences of the proteins plus the analysis of data from two-dimensional 1 H{ 1 H} chemical shift correlation spectroscopy (COSY), relayed COSY, and nuclear Overhauser effect spectroscopy (NOESY). The pH dependence (pH range 1 to 12) of NMR chemical shifts has been analyzed in detail. The complex pH titration profiles exhibited by the 1 H peaks of His-25 and Tyr-27 may be interpreted in terms of side-chain interactions between these residues. The protons of Gly-26 demonstrate a well-resolved AB spin system. These results suggest that this region of the protein has restricted conformational freedom. The kinetics of amide hydrogen/deuterium exchange were measured as a function of pH. The exchange rate for all amide hydrogens is governed by acid/base catalysis

  14. Serum inter-alpha-trypsin inhibitor and matrix hyaluronan promote angiogenesis in fibrotic lung injury.

    Science.gov (United States)

    Garantziotis, Stavros; Zudaire, Enrique; Trempus, Carol S; Hollingsworth, John W; Jiang, Dianhua; Lancaster, Lisa H; Richardson, Elizabeth; Zhuo, Lisheng; Cuttitta, Frank; Brown, Kevin K; Noble, Paul W; Kimata, Koji; Schwartz, David A

    2008-11-01

    The etiology and pathogenesis of angiogenesis in idiopathic pulmonary fibrosis (IPF) is poorly understood. Inter-alpha-trypsin inhibitor (IaI) is a serum protein that can bind to hyaluronan (HA) and may contribute to the angiogenic response to tissue injury. To determine whether IaI promotes HA-mediated angiogenesis in tissue injury. An examination was undertaken of angiogenesis in IaI-sufficient and -deficient mice in the bleomycin model of pulmonary fibrosis and in angiogenesis assays in vivo and in vitro. IaI and HA in patients with IPF were examined. IaI significantly enhances the angiogenic response to short-fragment HA in vivo and in vitro. lal deficiency Ieads to decreased angiogenesis in the matrigel model, and decreases lung angiogenesis after bleomycin exposure in mice. IaI is found in fibroblastic foci in IPF, where it colocalizes with HA. The colocalization is particularly strong in vascular areas around fibroblastic foci. Serum levels of IaI and HA are significantly elevated in patients with IPF compared with control subjects. High serum IaI and HA levels are associated with decreased lung diffusing capacity, but not FVC. Our findings indicate that serum IaI interacts with HA, and promotes angiogenesis in lung injury. IaI appears to contribute to the vascular response to lung injury and may lead to aberrant angiogenesis. Clinical trial registered with www.clinicaltrials.gov (NCT00016627).

  15. Serum Inter–α-Trypsin Inhibitor and Matrix Hyaluronan Promote Angiogenesis in Fibrotic Lung Injury

    Science.gov (United States)

    Garantziotis, Stavros; Zudaire, Enrique; Trempus, Carol S.; Hollingsworth, John W.; Jiang, Dianhua; Lancaster, Lisa H.; Richardson, Elizabeth; Zhuo, Lisheng; Cuttitta, Frank; Brown, Kevin K.; Noble, Paul W.; Kimata, Koji; Schwartz, David A.

    2008-01-01

    Rationale: The etiology and pathogenesis of angiogenesis in idiopathic pulmonary fibrosis (IPF) is poorly understood. Inter-α-trypsin inhibitor (IaI) is a serum protein that can bind to hyaluronan (HA) and may contribute to the angiogenic response to tissue injury. Objectives: To determine whether IaI promotes HA-mediated angiogenesis in tissue injury. Methods: An examination was undertaken of angiogenesis in IaI-sufficient and -deficient mice in the bleomycin model of pulmonary fibrosis and in angiogenesis assays in vivo and in vitro. IaI and HA in patients with IPF were examined. Measurements and Main Results: IaI significantly enhances the angiogenic response to short-fragment HA in vivo and in vitro. lal deficiency Ieads to decreased angiogenesis in the matrigel model, and decreases lung angiogenesis after bleomycin exposure in mice. IaI is found in fibroblastic foci in IPF, where it colocalizes with HA. The colocalization is particularly strong in vascular areas around fibroblastic foci. Serum levels of IaI and HA are significantly elevated in patients with IPF compared with control subjects. High serum IaI and HA levels are associated with decreased lung diffusing capacity, but not FVC. Conclusions: Our findings indicate that serum IaI interacts with HA, and promotes angiogenesis in lung injury. IaI appears to contribute to the vascular response to lung injury and may lead to aberrant angiogenesis. Clinical trial registered with www.clinicaltrials.gov (NCT00016627). PMID:18703791

  16. Kunitz trypsin inhibitor in addition to Bowman-Birk inhibitor influence stability of lunasin against pepsin-pancreatin hydrolysis.

    Science.gov (United States)

    Price, Samuel J; Pangloli, Philipus; Krishnan, Hari B; Dia, Vermont P

    2016-12-01

    Soybean contains several biologically active components and one of this belongs to the bioactive peptide group. The objectives of this study were to produce different lunasin-enriched preparations (LEP) and determine the effect of Bowman-Birk inhibitor (BBI) and Kunitz trypsin inhibitor (KTI) concentrations on the stability of lunasin against pepsin-pancreatin hydrolysis (PPH). In addition, the effect of KTI mutation on lunasin stability against PPH was determined. LEP were produced by calcium and pH precipitation methods of 30% aqueous ethanol extract from defatted soybean flour. LEP, lunasin-enriched commercially available products and KTI control and mutant flours underwent PPH and samples were taken after pepsin and pepsin-pancreatin hydrolysis. The concentrations of BBI, KTI, and lunasin all decreased after hydrolysis, but they had varying results. BBI concentration ranged from 167.5 to 655.8μg/g pre-hydrolysis and 171.5 to 250.1μg/g after hydrolysis. KTI concentrations ranged from 0.3 to 122.3μg/g pre-hydrolysis and 9.0 to 18.7μg/g after hydrolysis. Lunasin concentrations ranged from 8.5 to 71.0μg/g pre-hydrolysis and 4.0 to 13.2μg/g after hydrolysis. In all products tested, lunasin concentration after PPH significantly correlated with BBI and KTI concentrations. Mutation in two KTI isoforms led to a lower concentration of lunasin after PPH. This is the first report on the potential role of KTI in lunasin stability against PPH and must be considered in designing lunasin-enriched products that could potentially survive digestion after oral ingestion. Copyright © 2016 Elsevier Ltd. All rights reserved.

  17. {sup 13}C-NMR studies on disulfide bond isomerization in bovine pancreatic trypsin inhibitor (BPTI)

    Energy Technology Data Exchange (ETDEWEB)

    Takeda, Mitsuhiro [Kumamoto University, Department of Structural BioImaging, Faculty of Life Sciences (Japan); Miyanoiri, Yohei [Nagoya University, Structural Biology Research Center, Graduate School of Science (Japan); Terauchi, Tsutomu [Tokyo Metropolitan University, Graduate School of Science and Engineering (Japan); Kainosho, Masatsune, E-mail: kainosho@tmu.ac.jp [Nagoya University, Structural Biology Research Center, Graduate School of Science (Japan)

    2016-09-15

    Conformational isomerization of disulfide bonds is associated with the dynamics and thus the functional aspects of proteins. However, our understanding of the isomerization is limited by experimental difficulties in probing it. We explored the disulfide conformational isomerization of the Cys14–Cys38 disulfide bond in bovine pancreatic trypsin inhibitor (BPTI), by performing an NMR line-shape analysis of its Cys carbon peaks. In this approach, 1D {sup 13}C spectra were recorded at small temperature intervals for BPTI samples selectively labeled with site-specifically {sup 13}C-enriched Cys, and the recorded peaks were displayed in the order of the temperature after the spectral scales were normalized to a carbon peak. Over the profile of the line-shape, exchange broadening that altered with temperature was manifested for the carbon peaks of Cys14 and Cys38. The Cys14–Cys38 disulfide bond reportedly exists in equilibrium between a high-populated (M) and two low-populated states (m{sub c14} and m{sub c38}). Consistent with the three-site exchange model, biphasic exchange broadening arising from the two processes was observed for the peak of the Cys14 α-carbon. As the exchange broadening is maximized when the exchange rate equals the chemical shift difference in Hz between equilibrating sites, semi-quantitative information that was useful for establishing conditions for {sup 13}C relaxation dispersion experiments was obtained through the carbon line-shape profile. With respect to the m{sub c38} isomerization, the {sup 1}H-{sup 13}C signals at the β-position of the minor state were resolved from the major peaks and detected by exchange experiments at a low temperature.

  18. Effects of black-eyed pea trypsin/chymotrypsin inhibitor on proteolytic activity and on development of Anthonomus grandis.

    Science.gov (United States)

    Franco, Octávio L; dos Santos, Roseane C; Batista, João A N; Mendes, Ana Cristina M; de Araújo, Marcus Aurélio M; Monnerat, Rose G; Grossi-de-Sá, Maria Fátima; de Freitas, Sonia M

    2003-06-01

    The cotton boll weevil Anthonomus grandis (Boheman) is one of the major pests of cotton (Gossypium hirsutum L.) in tropical and sub-tropical areas of the New World. This feeds on cotton floral fruits and buds causing severe crop losses. Digestion in the boll weevil is facilitated by high levels of serine proteinases, which are responsible for the almost all proteolytic activity. Aiming to reduce the proteolytic activity, the inhibitory effects of black-eyed pea trypsin/chymotrypsin inhibitor (BTCI), towards trypsin and chymotrypsin from bovine pancreas and from midguts of A. grandis larvae and adult insects were analyzed. BTCI, purified from Vigna unguiculata (L.) seeds, was highly active against different trypsin-like proteinases studied and moderately active against the digestive chymotrypsin of adult insects. Nevertheless, no inhibitory activity was observed against chymotrypsin from A. grandis larval guts. To test the BTCI efficiency in vivo, neonate larvae were reared on artificial diet containing BTCI at 10, 50 and 100 microM. A reduction of larval weight of up to approximately 54% at the highest BTCI concentration was observed. At this concentration, the insect mortality was 65%. This work constitutes the first observation of a Bowman-Birk type inhibitor active in vitro and in vivo toward the cotton boll weevil A. grandis. The results of bioassays strongly suggest that BTCI may have potential as a transgene protein for use in engineered crop plants modified for heightened resistance to the cotton boll weevil.

  19. [Comparative analysis of sensitivity of proteases (chymotrypsin and trypsin) and cholinesterases of different origin to some organophosphorus inhibitors].

    Science.gov (United States)

    Rozengart, E V

    2009-01-01

    The antichymotrypsin, antitrypsin, and anticholinesterase efficiencies of four homologous series of organophosphorus inhibitors are compared: O-ethyl-S-(n-alkyl)methylthiophosphonates, O-(n-alkyl)-S-(n-butyl)methylthiophosphonates, O-(n-alkyl)-S-beta-(ethylmercaptoethylene)methylthiophosphonates, and their methylsulfomethylates. As sources of a-chymotrypsin and trypsin, commercial compounds of Worthington Biochemical Corporation and Leningrad Myasokombinat were tested. Bimolecular constant of the reaction rate was used as the measure of antienzyme efficiency. In all cases, the antichymotrypsin efficiency was lower, while the antitrypsin--essentially higher than the anticholinesterase activity of the studied inhibitors. These differences were found to much depend both on the inhibitor structure and on nature of the cholinesterase compounds.

  20. Understanding binding affinity : A combined isothermal titration calorimetry/molecular dynamics study of the binding of a series of hydrophobically modified benzamidinium chloride inhibitors to trypsin

    NARCIS (Netherlands)

    Talhout, Reinskje; Villa, Alessandra; Mark, AE; Engberts, JBFN

    2003-01-01

    The binding of a series of p-alkylbenzamidinium chloride inhibitors to the serine proteinase trypsin over a range of temperatures has been studied using isothermal titration (micro)calorimetry and molecular dynamics simulation techniques. The inhibitors have small structural variations at the para

  1. Bovine pancreatic trypsin inhibitor immobilized onto sepharose as a new strategy to purify a thermostable alkaline peptidase from cobia (Rachycentron canadum) processing waste.

    Science.gov (United States)

    França, Renata Cristina da Penha; Assis, Caio Rodrigo Dias; Santos, Juliana Ferreira; Torquato, Ricardo José Soares; Tanaka, Aparecida Sadae; Hirata, Izaura Yoshico; Assis, Diego Magno; Juliano, Maria Aparecida; Cavalli, Ronaldo Olivera; Carvalho, Luiz Bezerra de; Bezerra, Ranilson Souza

    2016-10-15

    A thermostable alkaline peptidase was purified from the processing waste of cobia (Rachycentron canadum) using bovine pancreatic trypsin inhibitor (BPTI) immobilized onto Sepharose. The purified enzyme had an apparent molecular mass of 24kDa by both sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and mass spectrometry. Its optimal temperature and pH were 50°C and 8.5, respectively. The enzyme was thermostable until 55°C and its activity was strongly inhibited by the classic trypsin inhibitors N-ρ-tosyl-l-lysine chloromethyl ketone (TLCK) and benzamidine. BPTI column allowed at least 15 assays without loss of efficacy. The purified enzyme was identified as a trypsin and the N-terminal amino acid sequence of this trypsin was IVGGYECTPHSQAHQVSLNSGYHFC, which was highly homologous to trypsin from cold water fish species. Using Nα-benzoyl-dl-arginine ρ-nitroanilide hydrochloride (BApNA) as substrate, the apparent km value of the purified trypsin was 0.38mM, kcat value was 3.14s(-1), and kcat/km was 8.26s(-1)mM(-1). The catalytic proficiency of the purified enzyme was 2.75×10(12)M(-1) showing higher affinity for the substrate at the transition state than other fish trypsin. The activation energy (AE) of the BApNA hydrolysis catalyzed by this enzyme was estimated to be 11.93kcalmol(-1) while the resulting rate enhancement of this reaction was found to be approximately in a range from 10(9) to 10(10)-fold evidencing its efficiency in comparison to other trypsin. This new purification strategy showed to be appropriate to obtain an alkaline peptidase from cobia processing waste with high purification degree. According with N-terminal homology and kinetic parameters, R. canadum trypsin may gathers desirable properties of psychrophilic and thermostable enzymes. Copyright © 2016 Elsevier B.V. All rights reserved.

  2. JcTI-I: a novel trypsin inhibitor from Jatropha curcas seed cake with potential for bacterial infection treatment.

    Science.gov (United States)

    Costa, Helen P S; Oliveira, Jose T A; Sousa, Daniele O B; Morais, Janne K S; Moreno, Frederico B; Monteiro-Moreira, Ana Cristina O; Viegas, Ricardo A; Vasconcelos, Ilka M

    2014-01-01

    Jatropha curcas seed cake is a low-value by-product resulting from biodiesel production. The seed cake is highly toxic, but it has great potential for biotechnology applications as it is a repository of biomolecules that could be important in agriculture, medicine, and industry. To explore this potential, a novel trypsin inhibitor called JcTI-I was purified by fractionation of the crude extract with trichloroacetic acid (2.5%, v/v) followed by affinity chromatography (Trypsin-Sepharose 4B) and molecular exclusion (Sephacryl S-200). Non-reducing sodium dodecyl sulfate-polyacrylamide gel electrophoresis and gel filtration showed that JcTI-I has approximately 20.0~kDa. Mass spectrometry analysis revealed that the intact molecular mass of JcTI-I is 10.252~kDa. Moreover, JcTI-I is a glycoprotein with 6.4% (m/m) carbohydrates, pI of 6.6, N-terminal sequence similarity around 60% to plant albumins and high stability to heat, pH, and salinity. JcTI-I presented antibacterial activity against the human pathogenic bacteria Salmonella enterica subspecies enterica serovar choleraesuis and Staphylococcus aureus, with minimum inhibitory concentration less than 5~μg/mL. Furthermore, JcTI-I did have inhibitory activity against the serine proteases from the tested bacteria. Otherwise, no hemolytic activity of human erythrocytes and signs of acute toxicity to mice were observed for JcTI-I. The results demonstrate the benefits of J. curcas seed cake as a source of trypsin inhibitor with potential for biotechnological application as a new antimicrobial agent against human pathogenic bacteria.

  3. JcTI-I, a novel trypsin inhibitor from Jatropha curcas seed cake with potential for bacterial infection treatment

    Directory of Open Access Journals (Sweden)

    Helen Paula S Costa

    2014-01-01

    Full Text Available Jatropha curcas seed cake is a low-value by-product resulting from biodiesel production. The seed cake is highly toxic, but it has great potential for biotechnology applications as it is a repository of biomolecules that could be important in agriculture, medicine and industry. To explore this potential, a novel trypsin inhibitor called JcTI-I was purified by fractionation of the crude extract with trichloroacetic acid (2.5%, v/v followed by affinity chromatography (Trypsin-Sepharose 4B and molecular exclusion (Sephacryl S-200. Non-reducing SDS-PAGE and gel filtration showed that JcTI-I has approximately 20.0 kDa. Mass spectrometry analysis revealed that the intact molecular mass of JcTI-I is 10.252 kDa. Moreover, JcTI-I is a glycoprotein with 6.4% (m/m carbohydrates, pI of 6.6, N-terminal sequence similarity around 60% to plant albumins and high stability to heat, pH and salinity. JcTI-I presented antibacterial activity against the human pathogenic bacteria Salmonella enterica subspecies enterica serovar choleraesuis and Staphylococcus aureus, with minimum inhibitory concentration (MIC less than 5 µg/mL. Furthermore, JcTI-I did have inhibitory activity against the serine proteases from the tested bacteria. Otherwise, no hemolytic activity of human erythrocytes and signs of acute toxicity to mice were observed for JcTI-I. The results demonstrate the benefits of J. curcas seed cake as a source of trypsin inhibitor with potential for biotechnological application as a new antimicrobial agent against human pathogenic bacteria.

  4. Disulfide bond effects on protein stability: designed variants of Cucurbita maxima trypsin inhibitor-V.

    Science.gov (United States)

    Zavodszky, M; Chen, C W; Huang, J K; Zolkiewski, M; Wen, L; Krishnamoorthi, R

    2001-01-01

    Attempts to increase protein stability by insertion of novel disulfide bonds have not always been successful. According to the two current models, cross-links enhance stability mainly through denatured state effects. We have investigated the effects of removal and addition of disulfide cross-links, protein flexibility in the vicinity of a cross-link, and disulfide loop size on the stability of Cucurbita maxima trypsin inhibitor-V (CMTI-V; 7 kD) by differential scanning calorimetry. CMTI-V offers the advantage of a large, flexible, and solvent-exposed loop not involved in extensive intra-molecular interactions. We have uncovered a negative correlation between retention time in hydrophobic column chromatography, a measure of protein hydrophobicity, and melting temperature (T(m)), an indicator of native state stabilization, for CMTI-V and its variants. In conjunction with the complete set of thermodynamic parameters of denaturation, this has led to the following deductions: (1) In the less stable, disulfide-removed C3S/C48S (Delta Delta G(d)(50 degrees C) = -4 kcal/mole; Delta T(m) = -22 degrees C), the native state is destabilized more than the denatured state; this also applies to the less-stable CMTI-V* (Delta Delta G(d)(50 degrees C) = -3 kcal/mole; Delta T(m) = -11 degrees C), in which the disulfide-containing loop is opened by specific hydrolysis of the Lys(44)-Asp(45) peptide bond; (2) In the less stable, disulfide-inserted E38C/W54C (Delta Delta G(d)(50 degrees C) = -1 kcal/mole; Delta T(m) = +2 degrees C), the denatured state is more stabilized than the native state; and (3) In the more stable, disulfide-engineered V42C/R52C (Delta Delta G(d)(50 degrees C) = +1 kcal/mole; Delta T(m) = +17 degrees C), the native state is more stabilized than the denatured state. These results show that a cross-link stabilizes both native and denatured states, and differential stabilization of the two states causes either loss or gain in protein stability. Removal of hydrogen

  5. Two-dimensional NMR studies of squash family inhibitors. Sequence-specific proton assignments and secondary structure of reactive-site hydrolyzed Cucurbita maxima trypsin inhibitor III.

    Science.gov (United States)

    Krishnamoorthi, R; Gong, Y X; Lin, C L; VanderVelde, D

    1992-01-28

    The solution structure of reactive-site hydrolyzed Cucurbita maxima trypsin inhibitor III (CMTI-III*) was investigated by two-dimensional proton nuclear magnetic resonance (2D NMR) spectroscopy. CMTI-III*, prepared by reacting CMTI-III with trypsin which cleaved the Arg5-Ile6 peptide bond, had the two fragments held together by a disulfide linkage. Sequence-specific 1H NMR resonance assignments were made for all the 29 amino acid residues of the protein. The secondary structure of CMTI-III*, as deduced from NOESY cross peaks and identification of slowly exchanging hydrogens, contains two turns (residues 8-12 and 24-27), a 3(10)-helix (residues 13-16), and a triple-stranded beta-sheet (residues 8-10, 29-27, and 21-25). This secondary structure is similar to that of CMTI-I [Holak, T. A., Gondol, D., Otlewski, J., & Wilusz, T. (1989) J. Mol. Biol. 210, 635-648], which has a Glu instead of a Lys at position 9. Sequential proton assignments were also made for the virgin inhibitor, CMTI-III, at pH 4.71, 30 degrees C. Comparison of backbone hydrogen chemical shifts of CMTI-III and CMTI-III* revealed significant changes for residues located far away from the reactive-site region as well as for those located near it, indicating tertiary structural changes that are transmitted through most of the 29 residues of the inhibitor protein. Many of these residues are functionally important in that they make contact with atoms of the enzyme in the trypsin-inhibitor complex, as revealed by X-ray crystallography [Bode, W., Greyling, H. J., Huber, R., Otlewski, J., & Wilusz, T. (1989) FEBS Lett. 242, 285-292].(ABSTRACT TRUNCATED AT 250 WORDS)

  6. Reactive-site hydrolyzed Cucurbita maxima trypsin inhibitor-V: function, thermodynamic stability, and NMR solution structure.

    Science.gov (United States)

    Cai, M; Gong, Y; Prakash, O; Krishnamoorthi, R

    1995-09-26

    Reactive-site (Lys44-Asp45 peptide bond) hydrolyzed Cucurbita maxima trypsin inhibitor-V (CMTI-V*) was prepared and characterized: In comparison to the intact form, CMTI-V* exhibited markedly reduced inhibitory properties and binding affinities toward trypsin and human blood coagulation factor XIIa. The equilibrium constant of trypsin-catalyzed hydrolysis, Khyd, defined as [CMTI-V*]/[CMTI-V], was measured to be approximately 9.4 at 25 degrees C (delta G degrees = -1.3 kcal.mol-1). From the temperature dependence of delta G degrees, the following thermodynamic parameters were estimated: delta H degrees = 1.6 kcal.mol-1 and delta S degrees = 9.8 eu. In order to understand the functional and thermodynamic differences between the two forms, the three-dimensional solution structure of CMTI-V* was determined by a combined approach of NMR, distance geometry, and simulated annealing methods. Thus, following sequence-specific and stereospecific resonance assignments, including those of beta-, gamma-, delta-, and epsilon-hydrogens and valine methyl hydrogens, 809 interhydrogen distances and 123 dihedral angle constraints were determined, resulting in the computation and energy-minimization of 20 structures for CMTI-V*. The average root mean squared deviation in position for equivalent atoms between the 20 individual structures and the mean structure obtained by averaging their coordinates is 0.67 +/- 0.15 A for the main chain atoms and 1.19 +/- 0.23 A for all the non-hydrogen atoms of residues 5-40 and residues 48-67.(ABSTRACT TRUNCATED AT 250 WORDS)

  7. Effects of gamma radiation on total phenolics, trypsin and tannin inhibitors in soybean grains

    Science.gov (United States)

    de Toledo, T. C. F.; Canniatti-Brazaca, S. G.; Arthur, V.; Piedade, S. M. S.

    2007-10-01

    The objective was determining possible radiation-induced alterations (with doses of 2, 4 and 8 kGy) in raw or cooked grains from five soybean cultivars through the analysis of some antinutrient. Total phenolic ranged from 2.46 to 10.83 mg/g, the trypsin inhibited from 18.19 to 71.64 UTI/g and tannins from 0.01 to 0.39 mg/g. All the antinutrient studied underwent reduction with increases in the doses and cooking process was effective too.

  8. Effects of gamma radiation on total phenolics, trypsin and tannin inhibitors in soybean grains

    International Nuclear Information System (INIS)

    de Toledo, T.C.F; Canniatti-Brazaca, S.G.; Arthur, V.; Piedade, S.M.S.

    2007-01-01

    The objective was determining possible radiation-induced alterations (with doses of 2, 4 and 8 kGy) in raw or cooked grains from five soybean cultivars through the analysis of some antinutrient. Total phenolic ranged from 2.46 to 10.83 mg/g, the trypsin inhibited from 18.19 to 71.64 UTI/g and tannins from 0.01 to 0.39 mg/g. All the antinutrient studied underwent reduction with increases in the doses and cooking process was effective too

  9. Effects of gamma radiation on total phenolics, trypsin and tannin inhibitors in soybean grains

    Energy Technology Data Exchange (ETDEWEB)

    de Toledo, T.C.F [Department of Agroindustry, Food and Nutrition Escola Superior de Agricultura ' Luiz de Queiroz' , Universidade de Sao Paulo (ESALQ/USP), Piracicaba, SP (Brazil)]. E-mail: tcftoled@esalq.usp.br; Canniatti-Brazaca, S.G. [Department of Agroindustry, Food and Nutrition, Escola Superior de Agricultura ' Luiz de Queiroz' , Universidade de Sao Paulo (ESALQ/USP), Av. Padua Dias, 11 Box 9, CEP 13418-900, Piracicaba, SP (Brazil)]. E-mail: sgcbraza@esalq.usp.br; Arthur, V. [Food Irradiation and Entomology Laboratory, Center for Nuclear Agriculture-CENA/USP, Av. Centenario 303, Caixa Postal 96, 13400-970, Piracicaba, SP (Brazil)]. E-mail: arthur@cena.usp.br; Piedade, S.M.S. [Department of Mathematic Sciences, Escola Superior de Agricultura ' Luiz de Queiroz' , Universidade de Sao Paulo (ESALQ/USP), Piracicaba, SP (Brazil)]. E-mail: soniamsp@esalq.usp.br

    2007-10-15

    The objective was determining possible radiation-induced alterations (with doses of 2, 4 and 8 kGy) in raw or cooked grains from five soybean cultivars through the analysis of some antinutrient. Total phenolic ranged from 2.46 to 10.83 mg/g, the trypsin inhibited from 18.19 to 71.64 UTI/g and tannins from 0.01 to 0.39 mg/g. All the antinutrient studied underwent reduction with increases in the doses and cooking process was effective too.

  10. Purification, crystallization and X-ray characterization of a Kunitz-type trypsin inhibitor protein from the seeds of chickpea (Cicer arietinum)

    International Nuclear Information System (INIS)

    Sharma, Urvashi; Suresh, C. G.

    2011-01-01

    The purification, characterization and crystallization of a trypsin inhibitor protein isolated from chickpea seeds are reported. A Kunitz-type trypsin inhibitor protein (CPTI) purified from chickpea seeds was estimated to have a molecular mass of 18 kDa on SDS–PAGE. The IC 50 value of CPTI was determined to be 2.5 µg against trypsin. The inhibitory activity of CPTI is 114 TIU (trypsin inhibitory units) per milligram of protein, which is high compared with those of other known Kunitz-type trypsin inhibitors from legumes. CPTI crystallized in three different orthorhombic crystal forms: P2 1 2 1 2 form A, P2 1 2 1 2 form B and P2 1 2 1 2 1 . The crystals of P2 1 2 1 2 form A, with unit-cell parameters a = 37.2, b = 41.2, c = 104.6 Å, diffracted to 2.0 Å resolution at the home source and to 1.4 Å on beamline BM14 at the ESRF. Data were also collected from crystals grown in the presence of iodine. The Matthews coefficient for these crystals was calculated to be 2.37 Å 3 Da −1 , corresponding to a solvent content of 42%. The other two crystal forms (P2 1 2 1 2 form B and P2 1 2 1 2 1 ) diffracted comparatively poorly

  11. Two-dimensional NMR studies of squash family inhibitors. Sequence-specific proton assignments and secondary structure of reactive-site hydrolyzed Cucurbita maxima trypsin inhibitor III

    Energy Technology Data Exchange (ETDEWEB)

    Krisnamoorthi, R.; Yuxi Gong; Chanlan Sun Lin (Kansas State Univ., Manhattan (United States)); VanderVelde, D. (Univ. of Kansas, Lawrence (United States))

    1992-01-28

    The solution structure of reactive-site hydrolyzed Cucurbita maxima trypsin inhibitor III (CMTI-III*) was investigated by two-dimensional proton nuclear magnetic resonance (2D NMR) spectroscopy. CMTI-III*, prepared by reacting CMTI-III with trypsin which cleaved the Arg5-Ile6 peptide bond, had the two fragments held together by a disulfide linkage. Sequence-specific {sup 1}H NMR resonance assignments were made for all the 29 amino acid residues of the protein. The secondary structure of CMTI-III*, as deduced from NOESY cross peaks and identification of slowly exchanging hydrogens, contains two turns, a 3{sub 10}-helix, and a triple-stranded {beta}-sheet. Sequential proton assignments were also made for the virgin inhibitor, CMTI-III, at pH 4.71, 30C. Comparison of backbone hydrogen chemical shifts of CMTI-III and CMTI-III* revealed significant changes for residues located far away from the reactive-site region as well as for those located near it, indicating tertiary structural changes that are transmitted through most of the 29 residues of the inhibitor protein. These chemical shift changes were relatively small compared to changes that occurred upon hydrolysis of the reactive-site peptide bond between Arg 5 and Ile6 in CMTI-III.

  12. Dioscorin, the major tuber storage protein of yam (Dioscorea batatas decne) with carbonic anhydrase and trypsin inhibitor activities.

    Science.gov (United States)

    Hou, W C; Liu, J S; Chen, H J; Chen, T E; Chang, C F; Lin, Y H

    1999-05-01

    Dioscorin, the tuber storage protein of yam (Dioscorea batatas Decne), was purified successively by ammonium sulfate fractionation, DE-52 ion exchange chromatography, and Sephadex G-75 column. Two protein bands (82 and 28 kDa) were found under nonreducing conditions after SDS-PAGE; but only one band (32 kDa) was detected under reducing conditions. The first 21 amino acids in the N-terminal region of the 28 kDa form were VEDEFSYIEGNPNGPENWGNL, which was highly homologous to deductive sequence of dioscorin from cDNA of another yam species (Dioscoreacayenensis Lam) reported by Conlan et al. (Plant Mol. Biol. 1995, 28, 369-380). Hewett-Emmett and Tashian (Mol. Phylogenet. Evol. 1996, 5, 50 -77) mentioned that, according to DNA alignments, dioscorin from yam (D. cayenensis) was alpha-carbonic anhydrase (alpha-CA) related. In this report, we found that the purified dioscorin showed both CA dehydration activity using sodium bicarbonate as a substrate and CA activity staining after SDS-PAGE. A polyclonal antibody, which was raised against trypsin inhibitor (TI), a storage protein of sweet potato (Ipomoea batatas [L.] Lam var. Tainong 57), cross-reacted with dioscorin, which also showed TI activity determined by both activity staining after SDS-PAGE and trypsin inhibition determination.

  13. Proton NMR studies of Cucurbita maxima trypsin inhibitors: Evidence for pH-dependent conformational change and his25 - try27 interaction

    Energy Technology Data Exchange (ETDEWEB)

    Krishnamoorthi, R.; Chanlan Sun Lin; Yuxi Gong (Kansas State Univ., Manhattan (United States)); VanderVelde, D. (Univ. of Kansas, Lawrence (United States)); Hahn, K. (Univ. of Colorado, Denver (United States))

    1992-01-28

    A pH-dependent His25-Tyr27 interaction was demonstrated in the case of Cucurbita maxima trypsin inhibitors (CMTI-I and CMTI-III) by means of nuclear magnetic resonance (NMR) spectroscopy. pH titration, line widths, peak shapes, deuterium exchange kinetics, and two-dimensional nuclear Overhauser effect spectroscopy (NOESY) were employed to characterize a conformational change involving Tyr27, which was shown to be triggered by deprotonation of His25 around pH 6. A hydrogen bond is proposed to be formed between N{sub {epsilon}} of His25 and OH of Tyr27, as a distance between the atoms, His25 N{epsilon} and Tyr25 OH, of 3.02 {angstrom} is consistent with a model built with NOE-derived distance constraints. The presently characterized relative orientations of His25 and Tyr27 are of functional significance, as these residues make contact with the enzyme in the enzyme-inhibitor complex. Furthermore, trypsin assay and inhibitor-binding studies showed that conformations of trypsin and the squash inhibitor complex. Furthermore, trypsin assay and inhibitor-binding studies showed that conformations of trypsin and the squash inhibitor were functionally relevant only in the pH range 6-8. The pK{sub a} of His25 was determined and found to be influenced by Glu9/Lys substitution and by the hydrolysis of the reactive-site peptide bond between Arg5 and Ile6. As these sites are located far (>10 {angstrom}) from His25, the results point out conformational changes that are propagated to a distant site in the protein molecule.

  14. Effect of traditional processing methods on the β-carotene, ascorbic acid and trypsin inhibitor content of orange-fleshed sweet potato for production of amala in Nigeria.

    Science.gov (United States)

    Yusuf, Abbas Bazata; Fuchs, Richard; Nicolaides, Linda

    2016-05-01

    The aim of the work was to study the effect of traditional processing methods on the β-carotene, ascorbic acid and trypsin inhibitor contents of orange-fleshed sweet potato amala. The most common sweet potato in Nigeria is white or yellow fleshed, which is very low in provitamin A. However, efforts are underway to promote orange-fleshed sweet potato to improve provitamin A intake. This paper describes how orange-fleshed sweet potato slices were traditionally processed into amala, which is increasingly consumed in Nigeria. The study revealed that both the cold and hot fermentation methods resulted in increased vitamin A levels and lower vitamin C levels in orange-fleshed sweet potato. Further processing to make amala resulted in a fall in both vitamin A and C content. The study found an increase in trypsin inhibitor activity following the cold-water fermentation and a decrease following the hot-water fermentation compared to raw orange-fleshed sweet potato. Trypsin inhibitor activity in amala produced using both the cold and hot methods was below detectable levels. The results indicate that amala produced from traditionally fermented orange-fleshed sweet potato could be a good source of vitamins A and C for the rural poor and that the processing removes any potential negative effects of trypsin inhibitors. © 2015 Society of Chemical Industry. © 2015 Society of Chemical Industry.

  15. Carboxamide Spleen Tyrosine Kinase (Syk) Inhibitors: Leveraging Ground State Interactions To Accelerate Optimization

    Energy Technology Data Exchange (ETDEWEB)

    Ellis, J. Michael; Altman, Michael D.; Cash, Brandon; Haidle, Andrew M.; Kubiak, Rachel L.; Maddess, Matthew L.; Yan, Youwei; Northrup, Alan B. (Merck)

    2016-12-08

    Optimization of a series of highly potent and kinome selective carbon-linked carboxamide spleen tyrosine kinase (Syk) inhibitors with favorable drug-like properties is described. A pervasive Ames liability in an analogous nitrogen-linked carboxamide series was obviated by replacement with a carbon-linked moiety. Initial efforts lacked on-target potency, likely due to strain induced between the hinge binding amide and solvent front heterocycle. Consideration of ground state and bound state energetics allowed rapid realization of improved solvent front substituents affording subnanomolar Syk potency and high kinome selectivity. These molecules were also devoid of mutagenicity risk as assessed via the Ames test using the TA97a Salmonella strain.

  16. Glycine-rich analogues of Cucurbita maxima trypsin inhibitor (CMTI-III) substituted by valine in position 27 display relatively low antitrypsin activity.

    Science.gov (United States)

    Rózycki, J; Kupryszewski, G; Rolka, K; Ragnarsson, U; Zbytryt, T; Krokoszyńska, I; Otlewski, J

    1993-09-01

    Five new analogues of the trypsin inhibitor CMTI-III were synthesized by the solid-phase method. All analogues containing a valine residue in position 27 and glycine residues in some or all of the positions 9, 11, 14, 17, 19, 29 as well as in two cases a norleucine residue in position 8 displayed association equilibrium constants by 6-7 orders of magnitude lower than the native CMTI-III inhibitor.

  17. The refined 2.0 A X-ray crystal structure of the complex formed between bovine beta-trypsin and CMTI-I, a trypsin inhibitor from squash seeds (Cucurbita maxima). Topological similarity of the squash seed inhibitors with the carboxypeptidase A inhibitor from potatoes.

    Science.gov (United States)

    Bode, W; Greyling, H J; Huber, R; Otlewski, J; Wilusz, T

    1989-01-02

    The stoichiometric complex formed between bovine beta-trypsin and the Cucurbita maxima trypsin inhibitor I (CMTI-I) was crystallized and its X-ray crystal structure determined using Patterson search techniques. Its structure has been crystallographically refined to a final R value of 0.152 (6.0-2.0 A). CMTI-I is of ellipsoidal shape; it lacks helices or beta-sheets, but consists of turns and connecting short polypeptide stretches. The disulfide pairing is CYS-3I-20I, Cys-10I-22I and Cys-16I-28I. According to the polypeptide fold and disulfide connectivity its structure resembles that of the carboxypeptidase A inhibitor from potatoes. Thirteen of the 29 inhibitor residues are in direct contact with trypsin; most of them are in the primary binding segment Val-2I (P4)-Glu-9I (P4') which contains the reactive site bond Arg-5I-Ile-6I and is in a conformation observed also for other serine proteinase inhibitors.

  18. Purification, crystallization and X-ray characterization of a Kunitz-type trypsin inhibitor protein from the seeds of chickpea (Cicer arietinum).

    Science.gov (United States)

    Sharma, Urvashi; Suresh, C G

    2011-06-01

    A Kunitz-type trypsin inhibitor protein (CPTI) purified from chickpea seeds was estimated to have a molecular mass of 18 kDa on SDS-PAGE. The IC(50) value of CPTI was determined to be 2.5 µg against trypsin. The inhibitory activity of CPTI is 114 TIU (trypsin inhibitory units) per milligram of protein, which is high compared with those of other known Kunitz-type trypsin inhibitors from legumes. CPTI crystallized in three different orthorhombic crystal forms: P2(1)2(1)2 form A, P2(1)2(1)2 form B and P2(1)2(1)2(1). The crystals of P2(1)2(1)2 form A, with unit-cell parameters a = 37.2, b = 41.2, c = 104.6 Å, diffracted to 2.0 Å resolution at the home source and to 1.4 Å on beamline BM14 at the ESRF. Data were also collected from crystals grown in the presence of iodine. The Matthews coefficient for these crystals was calculated to be 2.37 Å(3) Da(-1), corresponding to a solvent content of 42%. The other two crystal forms (P2(1)2(1)2 form B and P2(1)2(1)2(1)) diffracted comparatively poorly.

  19. Levels of Urinary Trypsin Inhibitor and Structure of Its Chondroitin Sulphate Moiety in Type 1 and Type 2 Diabetes

    Science.gov (United States)

    Ucciferri, Nadia; Idini, Michela; De Muro, Pierina

    2018-01-01

    Background Diabetes mellitus is a global health problem representing the fifth leading cause of mortality and a major risk factor for cardiovascular diseases. In the last years, we reported an association among urinary trypsin inhibitor (UTI), a small proteoglycan that plays pleiotropic roles in many inflammatory processes, and both type 1 and 2 diabetes and developed a method for its direct quantitation and structural characterization. Methods Urine from 39 patients affected by type 1 diabetes, 32 patients with type 2 diabetes, and 52 controls were analysed. UTI was separated from the main glycosaminoglycans physiologically present in urine by anion exchange chromatography, treated for chondroitin sulphate (CS) chain complete depolymerisation, and analysed for both UTI content and CS structure. UTI identification was performed by nano-LC-MS/MS analysis. Results We evidenced increased UTI levels, as well as reduced sulphation of its CS moiety in association with diabetes, regardless of both age and medium-term glycaemic control. Furthermore, no association between UTI and albumin excretion rate was found. Conclusions Evidences suggest that UTI levels are not directly correlated with renal function or, otherwise, that they may increase before the onset of renal impairment in diabetes, representing a potential marker for the underlying inflammatory condition. PMID:29541644

  20. Application and bioactive properties of CaTI, a trypsin inhibitor from Capsicum annuum seeds: membrane permeabilization, oxidative stress and intracellular target in phytopathogenic fungi cells.

    Science.gov (United States)

    Silva, Marciele S; Ribeiro, Suzanna Ff; Taveira, Gabriel B; Rodrigues, Rosana; Fernandes, Katia Vs; Carvalho, André O; Vasconcelos, Ilka Maria; Mello, Erica Oliveira; Gomes, Valdirene M

    2017-08-01

    During the last few years, a growing number of antimicrobial peptides have been isolated from plants and particularly from seeds. Recent results from our laboratory have shown the purification of a new trypsin inhibitor, named CaTI, from chilli pepper (Capsicum annuum L.) seeds. This study aims to evaluate the antifungal activity and mechanism of action of CaTI on phytopathogenic fungi and detect the presence of protease inhibitors in other species of this genus. Our results show that CaTI can inhibit the growth of the phytopathogenic fungi Colletotrichum gloeosporioides and C. lindemuthianum. CaTI can also permeabilize the membrane of all tested fungi. When testing the inhibitor on its ability to induce reactive oxygen species, an induction of reactive oxygen species (ROS) and nitric oxide (NO) particularly in Fusarium species was observed. Using CaTI coupled to fluorescein isothiocyanate (FITC), it was possible to determine the presence of the inhibitor inside the hyphae of the Fusarium oxysporum fungus. The search for protease inhibitors in other Capsicum species revealed their presence in all tested species. This paper shows the antifungal activity of protease inhibitors such as CaTI against phytopathogenic fungi. Antimicrobial peptides, among which the trypsin protease inhibitor family stands out, are present in different species of the genus Capsicum and are part of the chemical arsenal that plants use to defend themselves against pathogens. © 2017 Society of Chemical Industry. © 2017 Society of Chemical Industry.

  1. Immunotoxicity assessment for the novel Spleen tyrosine kinase inhibitor R406

    International Nuclear Information System (INIS)

    Zhu Yanhong; Herlaar, Ellen; Masuda, Esteban S.; Burleson, Gary R.; Nelson, Andrew J.; Grossbard, Elliott B.; Clemens, George R.

    2007-01-01

    Spleen tyrosine kinase (Syk) is a novel pharmaceutical target for treatment of allergic, autoimmune, and neoplastic disorders. Previous studies have indicated that Syk signaling plays critical roles in regulating the lymphohematopoietic system. These observations prompted us to investigate whether inhibition of Syk would promote immunotoxicity. In a series of studies, rats were treated orally with R406, at dose levels up to and including 100 mg/kg/day (or its prodrug R788 at dose levels up to and including 100 mg/kg/day, reduced to 50 mg/kg/day for females as MTD was exceeded), a potent Syk inhibitor, twice daily for 28 days. In addition to standard toxicological assessments, immunophenotyping by flow cytometric analysis, and a study of humoral immune response measuring anti-KLH IgM and IgG levels, were undertaken. Other immunotoxicity studies included three host resistance models in female Balb/c mice to further ascertain effects of R406 on innate and acquired immunity. Following R406 treatment, expected immunomodulating effects (e.g., decreased thymic and spleen weight, hypocellularity of bone marrow, and reduced lymphocyte counts, including T and B cells) were observed in the rat studies. These changes essentially resolved during a 14-day treatment-free recovery period. A KLH challenge in rats demonstrated no adverse effects on IgG or IgM response. R788/406, administered orally at dose levels up to and including 80 mg/kg/day for 28 days, did not affect bacterial or viral clearance in the Listeria, Streptococcal, or Influenza host resistance mouse models, respectively. This correlated with previous in vitro macrophage and neutrophil function assays (assessing migration, phagocytosis, oxidative burst and microbicidal activity), which revealed that R406 did not adversely affect macrophage or neutrophil function in innate immune responses. Collectively, these results demonstrate that R406 has minimal functional immunotoxicity notwithstanding its lymphocytopenic

  2. Supplementation with a new trypsin inhibitor from peanut is associated with reduced fasting glucose, weight control, and increased plasma CCK secretion in an animal model.

    Science.gov (United States)

    Serquiz, Alexandre C; Machado, Richele J A; Serquiz, Raphael P; Lima, Vanessa C O; de Carvalho, Fabiana Maria C; Carneiro, Marcella A A; Maciel, Bruna L L; Uchôa, Adriana F; Santos, Elizeu A; Morais, Ana H A

    2016-12-01

    Ingestion of peanuts may have a beneficial effect on weight control, possibly due to the satietogenic action of trypsin inhibitors. The aim of this study was to isolate a new trypsin inhibitor in a typical Brazilian peanut sweet (paçoca) and evaluate its effect in biochemical parameters, weight gain and food intake in male Wistar rats. The trypsin inhibitor in peanut paçoca (AHTI) was isolated. Experimental diets were prepared with AIN-93G supplemented with AHTI. Animals had their weight and food intake monitored. Animals were anesthetized, euthanized, and their bloods collected by cardiac puncture for dosage of cholecystokinin (CCK) and other biochemical parameters. Supplementation with AHTI significantly decreased fasting glucose, body weight gain, and food intake. These effects may be attributed to increased satiety, once supplemented animals showed no evidence of impaired nutritional status and also because AHTI increased CCK production. Thus, our results indicate that AHTI, besides reducing fasting glucose, can reduce weight gain via food intake reduction.

  3. A Trypsin Inhibitor from Tamarind Reduces Food Intake and Improves Inflammatory Status in Rats with Metabolic Syndrome Regardless of Weight Loss

    Directory of Open Access Journals (Sweden)

    Fabiana M. C. Carvalho

    2016-09-01

    Full Text Available Trypsin inhibitors are studied in a variety of models for their anti-obesity and anti-inflammatory bioactive properties. Our group has previously demonstrated the satietogenic effect of tamarind seed trypsin inhibitors (TTI in eutrophic mouse models and anti-inflammatory effects of other trypsin inhibitors. In this study, we evaluated TTI effect upon satiety, biochemical and inflammatory parameters in an experimental model of metabolic syndrome (MetS. Three groups of n = 5 male Wistar rats with obesity-based MetS received for 10 days one of the following: (1 Cafeteria diet; (2 Cafeteria diet + TTI (25 mg/kg; and (3 Standard diet. TTI reduced food intake in animals with MetS. Nevertheless, weight gain was not different between studied groups. Dyslipidemia parameters were not different with the use of TTI, only the group receiving standard diet showed lower very low density lipoprotein (VLDL and triglycerides (TG (Kruskal–Wallis, p < 0.05. Interleukin-6 (IL-6 production did not differ between groups. Interestingly, tumor necrosis factor-alpha (TNF-α was lower in animals receiving TTI. Our results corroborate the satietogenic effect of TTI in a MetS model. Furthermore, we showed that TTI added to a cafeteria diet may decrease inflammation regardless of weight loss. This puts TTI as a candidate for studies to test its effectiveness as an adjuvant in MetS treatment.

  4. Increased serum levels of tumour-associated trypsin inhibitor independently predict a poor prognosis in colorectal cancer patients

    Directory of Open Access Journals (Sweden)

    Gaber Alexander

    2010-09-01

    Full Text Available Abstract Background There is an insufficient number of reliable prognostic and response predictive biomarkers in colorectal cancer (CRC management. In a previous study, we found that high tumour tissue expression of tumour-associated trypsin inhibitor (TATI correlated with liver metastasis and an impaired prognosis in CRC. The aim of this study was to investigate the prognostic validity of serum TATI (s-TATI in CRC. We further assessed the prognostic value of carcino-embryonic antigen in serum (s-CEA and the interrelationship between s-TATI and TATI in tissue (t-TATI. Methods Using an immunofluorometric assay, s-TATI levels were analysed in 334 preoperatively collected serum samples from patients with CRC. Spearman's Rho and Chi-square test were used for analysis of correlations between s-TATI and clinicopathological parameters, s-CEA and t-TATI. Kaplan-Meier analysis and Cox uni- and multivariate regression analysis were used to estimate disease free survival (DFS and overall survival (OS according to quartiles of s-TATI and cut-offs derived from ROC-analysis of s-TATI and s-CEA. Results Increased levels of s-TATI were associated with a reduced DFS (HR = 2.00; 95% CI 1.40-2.84, P P P = 0.034 for DFS and HR = 1.78; 95% CI 1.25-2.53, P = 0.001 for OS. There was no significant association between s-TATI and t-TATI. The prognostic value of s-CEA was also evident, but somewhat weaker than for s-TATI. Conclusions High preoperative s-TATI levels predict a poor prognosis in patients with CRC, and the prognostic value is independent of established prognostic parameters and t-TATI expression. These data suggest that s-TATI might be a useful marker for prognostic stratification in CRC.

  5. Protective effect of urinary trypsin inhibitor on the development of radiation-induced lung fibrosis in mice

    International Nuclear Information System (INIS)

    Katoh, Hiroyuki; Ishikawa, Hitoshi; Suzuki, Yoshiyuki; Ohno, Tatsuya; Takahashi, Takeo; Nakano, Takashi; Hasegawa, Masatoshi; Yoshida, Yukari

    2010-01-01

    This study aimed to analyze whether Ulinastatin, a urinary trypsin inhibitor (UTI), inhibits the transforming growth factor (TGF)-β signaling pathway and lung fibrosis induced by thoracic irradiation in a lung injury mouse model. The thoraces of 9-week-old female fibrosis-sensitive C57BL/6 mice were irradiated with a single X-ray dose of 12 Gy or 24 Gy. UTI was administrated intraperitoneally at a dose of 200,000 units/kg concurrently with radiation (concurrent UTI) or daily during the post-irradiation period for 8-14 days (post-RT UTI). Mice were sacrificed at 16 weeks after irradiation to assess the histological grade of lung fibrosis and immunohistochemical TGF-β expression. Survival rates of mice given 24 Gy to the whole lung ±UTI were also compared. Post-RT UTI reduced the score of lung fibrosis in mice, but concurrent UTI had no beneficial effects in irradiated mice. The fibrosis score in post-RT UTI mice was 3.2±1.0, which was significantly smaller than that of irradiated mice without UTI treatment (RT alone; 6.0±1.3; p 2 =0.26, p<0.01). The survival rate at 30 weeks for post-RT UTI mice was significantly better than that of RT alone mice (33% vs. 10%, p<0.05). The administration of post-RT UTI suppressed TGF-β expression and radiation-induced lung fibrosis, which resulted in significant survival prolongation of the irradiated mice. (author)

  6. Proton NMR studies of Cucurbita maxima trypsin inhibitors: evidence for pH-dependent conformational change and His25-Tyr27 interaction.

    Science.gov (United States)

    Krishnamoorthi, R; Lin, C L; Gong, Y X; VanderVelde, D; Hahn, K

    1992-01-28

    A pH-dependent His25-Tyr27 interaction was demonstrated in the case of Cucurbita maxima trypsin inhibitors (CMTI-I and CMTI-III) by means of nuclear magnetic resonance (NMR) spectroscopy. pH titration, line widths, peak shapes, deuterium exchange kinetics, and two-dimensional nuclear Overhauser effect spectroscopy (NOESY) were employed to characterize a conformational change involving Tyr27, which was shown to be triggered by deprotonation of His25 around pH 6. A hydrogen bond is proposed to be formed between N epsilon of His25 and OH of Tyr27, as a distance between the atoms, His25 N epsilon and Tyr27 OH, of 3.02 A is consistent with a model built with NOE-derived distance constraints. Both the X-ray [Bode, W., Greyling, J.H., Huber, R., Otlewski, J., & Wilusz, T. (1989) FEBS Lett. 242, 282-292] and NMR [Holak, T.A., Gondol, D., Otlewski, J., & Wilusz, T. (1989) J. Mol. Biol. 210, 635-648] structures of CMTI-I at low pH (4.7-5.3) rule out such an interaction between the two aromatic rings, as the ring planes are oriented about 10 A away from each other. The presently characterized relative orientations of His25 and Tyr27 are of functional significance, as these residues make contact with the enzyme in the enzyme-inhibitor complex. Furthermore, trypsin assay and inhibitor-binding studies showed that conformations of trypsin and the squash inhibitor were functionally relevant only in the pH range 6-8. The pKa of His25 was determined and found to be influenced by Glu9/Lys substitution and by the hydrolysis of the reactive-site peptide bond between Arg5 and Ile6.(ABSTRACT TRUNCATED AT 250 WORDS)

  7. Solution structure and backbone dynamics of recombinant Cucurbita maxima trypsin inhibitor-V determined by NMR spectroscopy.

    Science.gov (United States)

    Liu, J; Prakash, O; Cai, M; Gong, Y; Huang, Y; Wen, L; Wen, J J; Huang, J K; Krishnamoorthi, R

    1996-02-06

    The solution structure of recombinant Cucurbita maxima trypsin inhibitor-V (rCMTI-V), whose N-terminal is unacetylated and carries an extra glycine residue, was determined by means of two-dimensional (2D) homo and 3D hetero NMR experiments in combination with a distance geometry and simulated annealing algorithm. A total of 927 interproton distances and 123 torsion angle constraints were utilized to generate 18 structures. The root mean squared deviation (RMSD) of the mean structure is 0.53 A for main-chain atoms and 0.95 A for all the non-hydrogen atoms of residues 3-40 and 49-67. The average structure of rCMTI-V is found to be almost the same as that of the native protein [Cai, M., Gong, Y., Kao, J.-L., & Krishnamoorthi, R. (1995) Biochemistry 34, 5201-5211]. The backbone dynamics of uniformly 15N-labeled rCMTI-V were characterized by 2D 1H-15N NMR methods. 15N spin-lattice and spin-spin relaxation rate constants (R1 and R2, respectively) and [1H]-15N steady-state heteronuclear Overhauser effect enhancements were measured for the peptide NH units and, using the model-free formalism [Lipari, G., & Szabo, A. (1982) J. Am. Chem. Soc. 104, 4546-4559, 4559-4570], the following parameters were determined: overall tumbling correlation time for the protein molecule (tau m), generalized order parameters for the individual N-H vectors (S2), effective correlation times for their internal motions (tau e), and terms to account for motions on a slower time scale (second) due to chemical exchange and/or conformational averaging (R(ex)). Most of the backbone NH groups of rCMTI-V are found to be highly constrained ((S2) = 0.83) with the exception of those in the binding loop (residues 41-48, (S2) = 0.71) and the N-terminal region ((S2) = 0.73). Main-chain atoms in these regions show large RMSD values in the average NMR structure. Residues involved in turns also appear to have more mobility ((S2) = 0.80). Dynamical properties of rCMTI-V were compared with those of two other

  8. Effect of trypsin inhibitor activity in soya bean on growth performance, protein digestibility and incidence of sub-clinical necrotic enteritis in broiler chicken flocks.

    Science.gov (United States)

    Palliyeguru, M W C D; Rose, S P; Mackenzie, A M

    2011-06-01

    1. The effect of three different levels of dietary trypsin inhibitor activity (achieved by varying the amount of non-toasted full fat soya bean in replacement for toasted full fat soya bean) on the incidence of spontaneously-occurring sub-clinical necrotic enteritis (NE) in broiler chickens was compared. A fourth dietary treatment compared the effect of a diet that used potato protein concentrate as the major protein source. The determined trypsin inhibitor activity increased with the increasing content of non-toasted soya bean: 1·90, 6·21, 8·46 and 3·72 mg/g for the three soya bean diets (0, 100 and 200 g of non-toasted soya bean/kg) and the potato protein diet respectively. 2. Although increasing amounts of the non-toasted full-fat soya bean increased the feed intakes of the birds, there was a marked reduction in protein digestibility, weight gain and feed conversion efficiency. 3. There was a linear increase in sub-clinical NE lesions in the duodenum, jejunum, mid small intestine and ileum with increasing non-toasted soya bean. Caecal Clostridium perfringens counts increased with the increasing dietary content of non-toasted soya bean. Serum α-toxin antibodies were higher in the birds fed the 200 g non-toasted soya bean/kg diet compared with the other diets. 4. The results demonstrated that variation in the amount of non-toasted dietary soya bean not only affects growth performance of broilers but also affects the incidence of sub-clinical necrotic enteritis in the flock. Ensuring the lowest possible trypsin-inhibitor activity in soya bean samples is a valuable tool to improve the health and welfare of birds and in reducing the financial losses from this disease.

  9. Trypsin radioimmunoassay

    International Nuclear Information System (INIS)

    Scheithauer, R.; Wolf, F.; Tympner, F.

    1981-01-01

    In 29 patients with suspicion of pancreatic disease standard secretin-pancreozymin-test was performed parallel to trypsin determination by radioimmunoassay before and after stimulation with secretin. Mean serum trypsin in normal subjects was 175 ng BIT/ml (range 90-250), the maximum after stimulation being 20 minutes after secreting injection (range 110-550). Preliminary normal values are 270 ng BIT/ml for basal concentration and 650 ng BIT/ml for 20 min after secretin stimulation. In the group of normals there was no case of misinterpretation. In patients with several pathological parameters (n = 10) basal trypsin concentration was increased in 7 cases, the stimulated value was concordant with the definite diagnosis in every case. Significant advantage for diagnostics was derived in patients having had pancreatic diseases before, however being actually normal with respect to standard diagnostic parameters. All these patients revealed increased trypsin concentrations after stimulation and 50% of them showed increased basal values. Equivocal results were seen in patients with endstage pancreatitis as well as in case of obstruction during reduced secretion. (orig.) [de

  10. Effect of gamma irradiation on lipoxygenases, trypsin inhibitor, raffinose family oligosaccharides and nutritional factors of different seed coat colored soybean (Glycine max L.)

    Energy Technology Data Exchange (ETDEWEB)

    Kumar Dixit, Amit [School of Biochemistry, Devi Ahilya University, Khandwa Road, Indore-452017 (India); Kumar, Vineet; Rani, Anita [National Research Centre for Soybean, Khandwa Road, Indore-452017 (India); Manjaya, J.G. [Board of Research in Nuclear Sciences, Bhabha Atomic Research Centre, Trombay, Mumbai (India); Bhatnagar, Deepak, E-mail: bhatnagarbio@yahoo.co [School of Biochemistry, Devi Ahilya University, Khandwa Road, Indore-452017 (India)

    2011-04-15

    Three soybean genotypes Kalitur, Hara soya and NRC37 with black, green and yellow seed coat color, respectively, were gamma irradiated at 0.5, 2.0 and 5.0 kGy and tested for antinutritional and nutritional factors. Gamma irradiation at all doses reduced the level of lipoxygenase isomers, trypsin inhibitor (TI) and ascorbic acid in all the 3 soybean genotypes as compared to the unirradiated control. However, irradiation dose of 5.0 kGy increased the sucrose content of the soybean genotypes. No significant change was observed in oil, protein fatty acids and total tocopherol content of the 3 genotypes at any irradiation dose. It is suggested that inhibition of lipoxygenase, reduction in TI and ascorbic acid may be due to the breakage or oxidation of protein structure by the gamma irradiation. Similarly, gamma irradiation at higher doses may break glycosidic linkages in oligosaccharides to produce more sucrose and decrease the content of flatulence causing oligosaccharides.

  11. Synthesis of a wild-type and three mutant Cucurbita maxima trypsin inhibitor-encoding genes by a single-strand approach.

    Science.gov (United States)

    Botes, D P; Qobose, M D; Corfield, V A

    1991-09-15

    A single-strand approach to gene assembly, based on a modification of an in vitro complementary oligodeoxyribonucleotide template-directed ligation of the desired sequence to a linearized vector [Chen et al., Nucleic Acids Res. 18 (1990) 871-878], is described. The gene coding for the wild-type Cucurbita maxima trypsin inhibitor of 29 amino acid residues [Bode et al., FEBS Lett. 242 (1989) 285-292], as well as three mutant forms of the gene, in which two of the three disulfide bonds have been replaced singly or as a pair, have been synthesized in a single synthesis run with minimal manual intervention. Subsequent to ligation to pUC9 and in vivo gapped duplex repair by Escherichia coli, their sequences have been verified.

  12. Effect of gamma irradiation on lipoxygenases, trypsin inhibitor, raffinose family oligosaccharides and nutritional factors of different seed coat colored soybean (Glycine max L.)

    International Nuclear Information System (INIS)

    Kumar Dixit, Amit; Kumar, Vineet; Rani, Anita; Manjaya, J.G.; Bhatnagar, Deepak

    2011-01-01

    Three soybean genotypes Kalitur, Hara soya and NRC37 with black, green and yellow seed coat color, respectively, were gamma irradiated at 0.5, 2.0 and 5.0 kGy and tested for antinutritional and nutritional factors. Gamma irradiation at all doses reduced the level of lipoxygenase isomers, trypsin inhibitor (TI) and ascorbic acid in all the 3 soybean genotypes as compared to the unirradiated control. However, irradiation dose of 5.0 kGy increased the sucrose content of the soybean genotypes. No significant change was observed in oil, protein fatty acids and total tocopherol content of the 3 genotypes at any irradiation dose. It is suggested that inhibition of lipoxygenase, reduction in TI and ascorbic acid may be due to the breakage or oxidation of protein structure by the gamma irradiation. Similarly, gamma irradiation at higher doses may break glycosidic linkages in oligosaccharides to produce more sucrose and decrease the content of flatulence causing oligosaccharides.

  13. The Effect of Gamma radiation, microwave radiation, their interaction and storage on chemical composition, antinutritional factors and the activities of trypsin inhibitor and lipoxygenase of soybean seeds

    Energy Technology Data Exchange (ETDEWEB)

    Abdel-Rahim, E A; Abdel-Fatah, O M [Dept. of Biochem., Faculty of Agric., Cairo University. (Egypt); El-Adawy, M; Badea, M Y [Food Technol. Dept., National Center for Research and Radiation Technol., Atomic Energy Authority (Egypt)

    2000-07-01

    The effect of gamma radiation, microwave radiation, interaction between them; and storage of radiated soybean seeds were investigated to find out the best treatment which had to the maximum reduction of antinutrional factors (Trypsin inhibitor and lipoxygenase activities) without significant effect on the chemical constituents. The gamma rays was used at three doses of 2.5, 5.0 and 8.0 kGy, microwave radiation was at 70 level power for 2 and 4 min; and the storage of seeds was at temperature, R.H. 50-55% for six months. The data revealed that, effects of interaction treatments were more effective than the treatment with microwave or gamma radiation alone.

  14. The Effect of Gamma radiation, microwave radiation, their interaction and storage on chemical composition, antinutritional factors and the activities of trypsin inhibitor and lipoxygenase of soybean seeds

    International Nuclear Information System (INIS)

    Abdel-Rahim, E.A.; Abdel-Fatah, O.M.; El-Adawy, M.; Badea, M.Y.

    2000-01-01

    The effect of gamma radiation, microwave radiation, interaction between them; and storage of radiated soybean seeds were investigated to find out the best treatment which had to the maximum reduction of antinutrional factors (Trypsin inhibitor and lipoxygenase activities) without significant effect on the chemical constituents. The gamma rays was used at three doses of 2.5, 5.0 and 8.0 kGy, microwave radiation was at 70 level power for 2 and 4 min; and the storage of seeds was at temperature, R.H. 50-55% for six months. The data revealed that, effects of interaction treatments were more effective than the treatment with microwave or gamma radiation alone

  15. Kinetic intermediates en route to the final serpin-protease complex: studies of complexes of α1-protease inhibitor with trypsin.

    Science.gov (United States)

    Maddur, Ashoka A; Swanson, Richard; Izaguirre, Gonzalo; Gettins, Peter G W; Olson, Steven T

    2013-11-01

    Serpin protein protease inhibitors inactivate their target proteases through a unique mechanism in which a major serpin conformational change, resulting in a 70-Å translocation of the protease from its initial reactive center loop docking site to the opposite pole of the serpin, kinetically traps the acyl-intermediate complex. Although the initial Michaelis and final trapped acyl-intermediate complexes have been well characterized structurally, the intermediate stages involved in this remarkable transformation are not well understood. To better characterize such intermediate steps, we undertook rapid kinetic studies of the FRET and fluorescence perturbation changes of site-specific fluorophore-labeled derivatives of the serpin, α1-protease inhibitor (α1PI), which report the serpin and protease conformational changes involved in transforming the Michaelis complex to the trapped acyl-intermediate complex in reactions with trypsin. Two kinetically resolvable conformational changes were observed in the reactions, ascribable to (i) serpin reactive center loop insertion into sheet A with full protease translocation but incomplete protease distortion followed by, (ii) full conformational distortion and movement of the protease and coupled serpin conformational changes involving the F helix-sheet A interface. Kinetic studies of calcium effects on the labeled α1PI-trypsin reactions demonstrated both inactive and low activity states of the distorted protease in the final complex that were distinct from the intermediate distorted state. These studies provide new insights into the nature of the serpin and protease conformational changes involved in trapping the acyl-intermediate complex in serpin-protease reactions and support a previously proposed role for helix F in the trapping mechanism.

  16. A trypsin inhibitor from Tecoma stans leaves inhibits growth and promotes ATP depletion and lipid peroxidation in Candida albicans and Candida krusei

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    Leydianne Leite de Siqueira Patriota

    2016-04-01

    Full Text Available Tecoma stans (yellow elder has shown medicinal properties and antimicrobial activity. Previous reports on antifungal activity of T. stans preparations and presence of trypsin inhibitor activity from T. stans leaves stimulated the investigation reported here. In this work, we proceeded to the purification and characterization of a trypsin inhibitor (TesTI, which was investigated for anti-Candida activity. Finally, in order to determine the potential of TesTI as a new natural chemotherapeutic product, its cytotoxicity to human peripheral blood mononuclear cells (PBMCs was evaluated. TesTI was isolated from saline extract by ammonium sulphate fractionation followed by ion exchange and gel filtration chromatographies. Antifungal activity was evaluated by determining the minimal inhibitory (MIC and fungicide (MFC concentrations using fungal cultures containing only yeast form or both yeast and hyphal forms. Candida cells treated with TesTI were evaluated for intracellular ATP levels and lipid peroxidation. Cytotoxicity of TesTI to PBMCs was evaluated by MTT assay. TesTI (39.8 kDa, pI 3.41, Ki 43 nM inhibited similarly the growth of both C. albicans and C. krusei culture types at MIC of 100 µg/mL. The MFCs were 200 µg/mL for C. albicans and C. krusei. Time-response curves revealed that TesTI (at MIC was more effective at inhibiting the replication of C. albicans cells. At MIC, TesTI promoted reduction of ATP levels and lipid peroxidation in the Candida cells, being not cytotoxic to PBMCs. In conclusion, TesTI is an antifungal agent against C. albicans and C. krusei, without toxicity to human cells.

  17. Perspectives of digestive pest control with proteinase inhibitors that mainly affect the trypsin-like activity of Anticarsia gemmatalis Hübner (Lepidoptera: Noctuidae

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    M.E. Pereira

    2005-11-01

    Full Text Available The present study describes the main characteristics of the proteolytic activities of the velvetbean caterpillar, Anticarsia gemmatalis Hübner, and their sensitivity to proteinase inhibitors and activators. Midguts of last instar larvae reared on an artificial diet were homogenized in 0.15 M NaCl and centrifuged at 14,000 g for 10 min at 4ºC and the supernatants were used in enzymatic assays at 30ºC, pH 10.0. Basal total proteolytic activity (azocasein hydrolysis was 1.14 ± 0.15 absorbance variation min-1 mg protein-1, at 420 nm; basal trypsin-like activity (N-benzoyl-L-arginine-p-nitroanilide, BApNA, hydrolysis was 0.217 ± 0.02 mmol p-nitroaniline min-1 mg protein-1. The maximum proteolytic activities were observed at pH 10.5 using azocasein and at pH 10.0 using BApNA, this pH being identical to the midgut pH of 10.0. The maximum trypsin-like activity occurred at 50ºC, a temperature that reduces enzyme stability to 80 and 60% of the original, when pre-incubated for 5 and 30 min, respectively. Phenylmethylsulfonyl fluoride inhibited the proteolytic activities with an IC50 of 0.39 mM for azocasein hydrolysis and of 1.35 mM for BApNA hydrolysis. Benzamidine inhibited the hydrolysis with an IC50 of 0.69 and 0.076 mM for azocasein and BApNA, respectively. The absence of cysteine-proteinases is indicated by the fact that 2-mercaptoethanol and L-cysteine did not increase the rate of azocasein hydrolysis. These results demonstrate the presence of serine-proteinases and the predominance of trypsin-like activity in the midgut of Lepidoptera insects, now also detected in A. gemmatalis, and suggest this enzyme as a major target for pest control based on disruption of protein metabolism using proteinase inhibitors.

  18. Differential modulation of binding loop flexibility and stability by Arg50 and Arg52 in Cucurbita maxima trypsin inhibitor-V deduced by trypsin-catalyzed hydrolysis and NMR spectroscopy.

    Science.gov (United States)

    Cai, M; Huang, Y; Prakash, O; Wen, L; Dunkelbarger, S P; Huang, J K; Liu, J; Krishnamoorthi, R

    1996-04-16

    The side chains of Arg50 and Arg52 iin Cucurbita maxima trypsin inhibitor-V (CMTI-V) anchor the binding loop to the scaffold region [Cai, M., Gong, Y., Kao, J.L-F., & Krishnamoorthi, R. (1995) Biochemistry 34, 5201-5211]. The consequences of these hydrogen-bonding and electrostatic interactions on the conformational flexibility and stability of the binding loop were evaluated by trypsin-catalyzed hydrolysis of CMTI-V mutants, in which each of the arginines was individually replaced with Ala, Lys, or Gln by genetic engineering methods. All mutants exhibited significantly increased vulnerability to the protease attack at many sites, including the reactive-site (Lys44-Asp45 peptide bond), with the R50 mutants showing much more pronounced effects than the R52 counterparts. For CmTI-V and the mutants studied, a qualitative correlation was inferred between binding loop flexibility and retention time on a reverse-phase high-pressure liquid chromatography C-18 column. The R50 mutants were found to be more flexible than the corresponding R52 versions. These results demonstrate that Arg50 contributes more to the stability and function of CMTI-V. The differing strengths of the hydrogen bonds made by Arg50 and Arg52 were characterized by determining the internal dynamics of their side chains at pH 5.0 and 2.5: 15N NMR longitudinal and transverse relaxation rates and 15N-1H nuclear Overhauser effect (NOE) enhancements were measured for the main-chain and side-chain NH groups in 15N-labeled recombinant CMTI-V (rCMTI-V) and the model-free parameters [Lipari, G., & Szabo, A.(1982) J. Am. Chem. Soc. 104, 4546-59; 4559-4570] were calculated. At both pH 5.0 and 2.5, the arginines at positions 26, 47, 58 and 66 are found to be highly mobile, as the caluculated general order parameters, S2 values, of their NepsilonH groups fall in the range 0.03-0.18. The corresponding values for Arg50 amd Arg52 are 0.73 and 0.63, respectively, at pH 5.0, thus confirming that the two arginines are

  19. Structural consequences of the natural substitution, E9K, on reactive-site-hydrolyzed squash (Cucurbita maxima) trypsin inhibitor (CMTI), as studied by two-dimensional NMR.

    Science.gov (United States)

    Krishnamoorthi, R; Lin, C L; VanderVelde, D

    1992-06-02

    Sequence-specific hydrogen-1 NMR assignments were made to all of the 29 amino acid residues of reactive-site-hydrolyzed Cucurbita maxima trypsin inhibitor I (CMTI-I*) by the application of two-dimensional NMR (2D NMR) techniques, and its secondary structural elements (two tight turns, a 3(10)-helix, and a triple-stranded beta-sheet) were identified on the basis of short-range NOESY cross peaks and deuterium-exchange kinetics. These secondary structural elements are present in the intact inhibitor [Holak, T. A., Gondol, D., Otlewski, J., & Wilusz, T. (1989) J. Mol. Biol. 210, 635-648] and are unaffected by the hydrolysis of the reactive-site peptide bond between Arg5 and Ile6, in accordance with the earlier conclusion reached for CMTI-III* [Krishnamoorthi, R., Gong, Y.-X., Lin, C. S., & VanderVelde, D. (1992) Biochemistry 31, 898-904]. Chemical shifts of backbone hydrogen atoms, peptide NH's, and C alpha H's, of CMTI-I* were compared with those of the intact inhibitor, CMTI-I, and of the reactive-site-hydrolyzed, natural, E9K variant, CMTI-III*. Cleavage of the Arg5-Ile6 peptide bond resulted in changes of chemical shifts of most of the backbone atoms of CMTI-I, in agreement with the earlier results obtained for CMTI-III. Comparison of chemical shifts of backbone hydrogen atoms of CMTI-I* and CMTI-III* revealed no changes, except for residues Glu9 and His25. However, the intact forms of the same two proteins, CMTI-I and CMTI-III, showed small but significant perturbations of chemical shifts of residues that made up the secondary structural elements of the inhibitors.(ABSTRACT TRUNCATED AT 250 WORDS)

  20. Diastereotopic covalent binding of the natural inhibitor leupeptin to trypsin: Detection of two interconverting hemiacetals by solution and solid-state NMR spectroscopy

    International Nuclear Information System (INIS)

    Ortiz, C.; Tellier, C.; Williams, H.; Stolowich, N.J.; Scott, A.I.

    1991-01-01

    The naturally occurring peptidyl protease inhibitor leupeptin (N-acetyl-L-leucyl-L-leucyl-L-argininal) has been prepared labeled with 13 C at the argininal carbonyl. 13 C chemical shift data for the trypsin-leupeptin inhibitor complex in the pH range 3.0-7.6 reveal the presence of two pH-dependent covalent complexes, suggestive of two interconverting diastereomers at the new asymmetric tetrahedral center created by covalent addition of Ser195 to either side of the 13 C-enriched aldehyde of the inhibitor. At pH 7 two signals are observable at δ 98.8 and δ 97.2 (84:16 ratio), while at pH 3.0 the latter signal predominates. In the selective proton 13 C-edited NOE spectrum of the major diastereomer at pH 7.4, a strong NOE is observed between the hemiacetal proton of the inhibitor and the C2 proton of His57 of the enzyme, thus defining the stereochemistry of the high pH complex to the S configuration in which the hemiacetal oxygen resides in the oxyanion hole. pH titration studies further indicate that the 13 C chemical shift of the S diastereomer follows a titration curve with a pK a of 4.69, the magnitude of which is consistent with direct titration of the hemiacetal oxygen. Similar pH-dependent chemical shifts were obtained by using CPMAS 13 C NMR, providing evidence for the existence of the same diastereomeric equilibrium in the solid state

  1. Natural abundance 15N NMR assignments delineate structural differences between intact and reactive-site hydrolyzed Cucurbita maxima trypsin inhibitor III.

    Science.gov (United States)

    Krishnamoorthi, R; Nemmers, S; Tobias, B

    1992-06-15

    15N NMR assignments were made to the backbone amide nitrogen atoms at natural isotopic abundance of intact and reactive-site (Arg5-Ile6) hydrolyzed Cucurbita maxima trypsin inhibitor III (CMTI-III and CMTI-III*, respectively) by means of 2D proton-detected heteronuclear single bond chemical shift correlation (HSBC) spectroscopy, utilizing the previously made sequence-specific 1H NMR assignments (Krishnamoorthi et al. (1992) Biochemistry 31, 898-904). Comparison of the 15N chemical shifts of the two forms of the inhibitor molecule revealed significant changes not only for residues located near the reactive-site region, but also for those distantly located. Residues Cys3, Arg5, Leu7, Met8, Cys10, Cys16, Glu19, His25, Tyr27, Cys28 and Gly29 showed significant chemical shift changes ranging from 0.3 to 6.1 ppm, thus indicating structural perturbations that were transmitted throughout the molecule. These findings confirm the earlier conclusions based on 1H NMR investigations.

  2. Does the oxidation of methionine residue precede the inactivation of the trypsin inhibitor (LUTI in germinating seeds of common flax (Linum usitatissimum?

    Directory of Open Access Journals (Sweden)

    Irena Lorenc-Kubis

    2011-01-01

    Full Text Available Antitrypsin activity in germinating common seeds of flax (Linum usitatissimum was investigated. At the early stage of germination an increase in antitrypsin activity was observed, followed by its decrease during the development of the seedlings. From 6-day-old seedlings a trypsin inhibitor (gerLUTI was purified. The purification procedure involved fractionation of proteins from seedling homogenate with alcohol and successive chromatography on CM-Sephadex C-25 on immobilised methylchymotrypsin in the presence of 5 M NaCl, and finally on a C18 column in RP-HPLC. The gerLUTI migrated in SDS PAGE as a single band, but in mass spectroscopy analysis it exhibited the presence of at least three forms with molecular masses of 7654 ± 3 Da, 7668/7670 ± 3 Da, and 7687 ± 3 Da. The preparation of LUTI isolated from resting seeds contained only one form, with a molecular mass of 7655 ± 3 Da. LUTI and gerLUTI differed also in methionine contents. LUTI contained two methionine residues, whereas in gerLUTI only a trace of methionine was detected. The obtained results might suggest that during flax seeds germination the inhibitor molecules undergo selective modification, e.g. oxidation at methionine residues, before being degraded by proteolytic enzymes.

  3. Loss of second and sixth conserved cysteine residues from trypsin inhibitor-like cysteine-rich domain-type protease inhibitors in Bombyx mori may induce activity against microbial proteases.

    Science.gov (United States)

    Li, Youshan; Liu, Huawei; Zhu, Rui; Xia, Qingyou; Zhao, Ping

    2016-12-01

    Previous studies have indicated that most trypsin inhibitor-like cysteine-rich domain (TIL)-type protease inhibitors, which contain a single TIL domain with ten conserved cysteines, inhibit cathepsin, trypsin, chymotrypsin, or elastase. Our recent findings suggest that Cys 2nd and Cys 6th were lost from the TIL domain of the fungal-resistance factors in Bombyx mori, BmSPI38 and BmSPI39, which inhibit microbial proteases and the germination of Beauveria bassiana conidia. To reveal the significance of these two missing cysteines in relation to the structure and function of TIL-type protease inhibitors in B. mori, cysteines were introduced at these two positions (D36 and L56 in BmSPI38, D38 and L58 in BmSPI39) by site-directed mutagenesis. The homology structure model of TIL domain of the wild-type and mutated form of BmSPI39 showed that two cysteine mutations may cause incorrect disulfide bond formation of B. mori TIL-type protease inhibitors. The results of Far-UV circular dichroism (CD) spectra indicated that both the wild-type and mutated form of BmSPI39 harbored predominantly random coil structures, and had slightly different secondary structure compositions. SDS-PAGE and Western blotting analysis showed that cysteine mutations affected the multimerization states and electrophoretic mobility of BmSPI38 and BmSPI39. Activity staining and protease inhibition assays showed that the introduction of cysteine mutations dramaticly reduced the activity of inhibitors against microbial proteases, such as subtilisin A from Bacillus licheniformis, protease K from Engyodontium album, protease from Aspergillus melleus. We also systematically analyzed the key residue sites, which may greatly influence the specificity and potency of TIL-type protease inhibitors. We found that the two missing cysteines in B. mori TIL-type protease inhibitors might be crucial for their inhibitory activities against microbial proteases. The genetic engineering of TIL-type protease inhibitors may be

  4. The belonging of gpMuc, a glycoprotein from Mucuna pruriens seeds, to the Kunitz-type trypsin inhibitor family explains its direct anti-snake venom activity.

    Science.gov (United States)

    Scirè, Andrea; Tanfani, Fabio; Bertoli, Enrico; Furlani, Emiliano; Nadozie, Hope-Onyekwere N; Cerutti, Helena; Cortelazzo, Alessio; Bini, Luca; Guerranti, Roberto

    2011-07-15

    In Nigeria, Mucuna pruriens seeds are locally prescribed as an oral prophylactic for snake bite and it is claimed that when two seeds are swallowed they protect the individual for a year against snake bites. In order to understand the Mucuna pruriens antisnake properties, the proteins from the acqueous extract of seeds were purified by three chromatographic steps: ConA affinity chromatography, tandem anionic-cationic exchange and gel filtration, obtaining a fraction conventionally called gpMucB. This purified fraction was analysed by SDS-PAGE obtaining 3 bands with apparent masses ranging from 20 to 24 kDa, and by MALDI-TOF which showed two main peaks of 21 and 23 kDa and another small peak of 19 kDa. On the other hand, gel filtration analysis of the native protein indicated a molecular mass of about 70 kDa suggesting that in its native form, gpMucB is most likely an oligomeric multiform protein. Infrared spectroscopy of gpMucB indicated that the protein is particularly thermostable both at neutral and acidic pHs and that it is an all beta protein. All data suggest that gpMucB belongs to the Kunitz-type trypsin inhibitor family explaining the direct anti-snake venom activity of Mucuna pruriens seeds. Copyright © 2011 Elsevier GmbH. All rights reserved.

  5. Spleen removal

    Science.gov (United States)

    ... spleen. Sickle cell anemia . Splenic artery aneurysm (rare). Trauma to the spleen. Risks Risks for anesthesia and surgery in general ... removal - series References Brandow AM, Camitta BM. Hyposplenism, splenic trauma, and splenectomy. In: Kliegman RM, Stanton BF, St. ...

  6. pH and temperature effects on the molecular conformation of the porcine pancreatic secretory trypsin inhibitor as detected by hydrogen-1 nuclear magnetic resonance

    International Nuclear Information System (INIS)

    De Marco, A.; Menegatti, E.; Guarneri, M.

    1982-01-01

    1 H NMR spectra of the porcine pancreatic secretory trypsin inhibitor (PSTI) have been recorded vs. pH and temperature. Of the two tyrosines, one titrates with a pK of 1.25, while the resonances from the other are pH insensitive in the investigated range 4.8 less than or equal to pH less than or equal to 12. This is consistent with PSTI having one Tyr solvent exposed (Try-20) and the other buried (Tyr-31). The resonances from the lysyl epsilon-CH 2 protons titrate with a pK of 10.95. The titration is accompanied by a pronounced line broadening, which starts near pH 8.5. Between pH 11.5 and pH 12 the epsilon-CH 2 resonances recover their low pH line width. Titration curves for the lysines and Tyr-20 reflect single proton ionization equilibria, suggesting that these residues do not interact among themselves. On the basis of double resonance experiments, combined with analysis of chemical shifts, spin-spin couplngs, and line widths, all methyl resonances are identified and followed as functions of pH and temperature. The γ-CH 3 doublet from the N-terminal Thr-1 is assigned by comparison between spectra of forms I and II of the inhibitor, the latter lacking the first four residues of form I. The β-CH 3 resonance from Ala-7 is also assigned. Proton resonance parameters of methyl groups are shown to afford useful NMR probes for the characterization of local nonbonded interactions, microenvironments, and mobilities

  7. Frequent expression loss of Inter-alpha-trypsin inhibitor heavy chain (ITIH) genes in multiple human solid tumors: A systematic expression analysis

    International Nuclear Information System (INIS)

    Hamm, Alexander; Knuechel, Ruth; Dahl, Edgar; Veeck, Juergen; Bektas, Nuran; Wild, Peter J; Hartmann, Arndt; Heindrichs, Uwe; Kristiansen, Glen; Werbowetski-Ogilvie, Tamra; Del Maestro, Rolando

    2008-01-01

    The inter-alpha-trypsin inhibitors (ITI) are a family of plasma protease inhibitors, assembled from a light chain – bikunin, encoded by AMBP – and five homologous heavy chains (encoded by ITIH1, ITIH2, ITIH3, ITIH4, and ITIH5), contributing to extracellular matrix stability by covalent linkage to hyaluronan. So far, ITIH molecules have been shown to play a particularly important role in inflammation and carcinogenesis. We systematically investigated differential gene expression of the ITIH gene family, as well as AMBP and the interacting partner TNFAIP6 in 13 different human tumor entities (of breast, endometrium, ovary, cervix, stomach, small intestine, colon, rectum, lung, thyroid, prostate, kidney, and pancreas) using cDNA dot blot analysis (Cancer Profiling Array, CPA), semiquantitative RT-PCR and immunohistochemistry. We found that ITIH genes are clearly downregulated in multiple human solid tumors, including breast, colon and lung cancer. Thus, ITIH genes may represent a family of putative tumor suppressor genes that should be analyzed in greater detail in the future. For an initial detailed analysis we chose ITIH2 expression in human breast cancer. Loss of ITIH2 expression in 70% of cases (n = 50, CPA) could be confirmed by real-time PCR in an additional set of breast cancers (n = 36). Next we studied ITIH2 expression on the protein level by analyzing a comprehensive tissue micro array including 185 invasive breast cancer specimens. We found a strong correlation (p < 0.001) between ITIH2 expression and estrogen receptor (ER) expression indicating that ER may be involved in the regulation of this ECM molecule. Altogether, this is the first systematic analysis on the differential expression of ITIH genes in human cancer, showing frequent downregulation that may be associated with initiation and/or progression of these malignancies

  8. Frequent expression loss of Inter-alpha-trypsin inhibitor heavy chain (ITIH genes in multiple human solid tumors: A systematic expression analysis

    Directory of Open Access Journals (Sweden)

    Werbowetski-Ogilvie Tamra

    2008-01-01

    Full Text Available Abstract Background The inter-alpha-trypsin inhibitors (ITI are a family of plasma protease inhibitors, assembled from a light chain – bikunin, encoded by AMBP – and five homologous heavy chains (encoded by ITIH1, ITIH2, ITIH3, ITIH4, and ITIH5, contributing to extracellular matrix stability by covalent linkage to hyaluronan. So far, ITIH molecules have been shown to play a particularly important role in inflammation and carcinogenesis. Methods We systematically investigated differential gene expression of the ITIH gene family, as well as AMBP and the interacting partner TNFAIP6 in 13 different human tumor entities (of breast, endometrium, ovary, cervix, stomach, small intestine, colon, rectum, lung, thyroid, prostate, kidney, and pancreas using cDNA dot blot analysis (Cancer Profiling Array, CPA, semiquantitative RT-PCR and immunohistochemistry. Results We found that ITIH genes are clearly downregulated in multiple human solid tumors, including breast, colon and lung cancer. Thus, ITIH genes may represent a family of putative tumor suppressor genes that should be analyzed in greater detail in the future. For an initial detailed analysis we chose ITIH2 expression in human breast cancer. Loss of ITIH2 expression in 70% of cases (n = 50, CPA could be confirmed by real-time PCR in an additional set of breast cancers (n = 36. Next we studied ITIH2 expression on the protein level by analyzing a comprehensive tissue micro array including 185 invasive breast cancer specimens. We found a strong correlation (p Conclusion Altogether, this is the first systematic analysis on the differential expression of ITIH genes in human cancer, showing frequent downregulation that may be associated with initiation and/or progression of these malignancies.

  9. Development of a method for urine bikunin/urinary trypsin inhibitor (UTI) quantitation and structural characterization: Application to type 1 and type 2 diabetes.

    Science.gov (United States)

    Lepedda, Antonio Junior; Nieddu, Gabriele; Rocchiccioli, Silvia; Fresu, Pietro; De Muro, Pierina; Formato, Marilena

    2013-12-01

    Bikunin is a plasma proteinase inhibitor often associated with inflammatory conditions. It has a half-life of few minutes and it is rapidly excreted into urine as urinary trypsin inhibitor (UTI). UTI levels are usually low in healthy individuals but they can increase up to tenfold in both acute and chronic inflammatory diseases. This article describes a sensitive method for both direct UTI quantitation and structural characterization. UTI purification was performed by anion exchange micro-chromatography followed by SDS-PAGE. A calibration curve for protein quantitation was set up by using a purified UTI fraction. UTI identification and structural characterization was performed by Nano-LC-MS/MS analysis. The method was applied on urine samples from 9 patients with type 1 diabetes, 11 patients with type 2 diabetes, and 28 healthy controls, matched for age and sex with patients, evidencing higher UTI levels in both groups of patients with respect to controls (p UTI levels and age in each group tested. Owing to the elevated sensitivity and specificity, the described method allows UTI quantitation from very low quantities of specimen. Furthermore, as UTI concentration is normalized for creatinine level, the analysis could be also performed on randomly collected urine samples. Finally, MS/MS analysis prospects the possibility of characterizing PTM sites potentially able to affect UTI localization, function, and pathophysiological activity. Preliminary results suggest that UTI levels could represent a useful marker of chronic inflammatory condition in type 1 and 2 diabetes. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  10. Wandering spleen

    Energy Technology Data Exchange (ETDEWEB)

    Park, Yong Tai; Lee, Sun Hwa; Lee, Dong Ho; Ko, Young Tae; Lim, Jae Hoon [Kyung Hee University Hospital, Seoul (Korea, Republic of)

    1987-12-15

    Wandering spleen is a rare entity, which is defined as the presence of the spleen in other than the left upper quadrant of the abdomen. The etiology of wandering spleen is unknown. Congenital factors such as incomplete fusion or laxity of the supporting structures of the spleen and acquired factors such as splenomegaly, trauma, abdominal laxity and hormonal effects of pregnancy may play a role. The clinical presentation is variable from asymptomatic to catastrophic if torsion of the splenic pedicle occurs. We present two cases of wandering spleen with a brief review of the literature.

  11. Wandering spleen

    International Nuclear Information System (INIS)

    Park, Yong Tai; Lee, Sun Hwa; Lee, Dong Ho; Ko, Young Tae; Lim, Jae Hoon

    1987-01-01

    Wandering spleen is a rare entity, which is defined as the presence of the spleen in other than the left upper quadrant of the abdomen. The etiology of wandering spleen is unknown. Congenital factors such as incomplete fusion or laxity of the supporting structures of the spleen and acquired factors such as splenomegaly, trauma, abdominal laxity and hormonal effects of pregnancy may play a role. The clinical presentation is variable from asymptomatic to catastrophic if torsion of the splenic pedicle occurs. We present two cases of wandering spleen with a brief review of the literature

  12. Two proteins for the price of one: Structural studies of the dual-destiny protein preproalbumin with sunflower trypsin inhibitor-1.

    Science.gov (United States)

    Franke, Bastian; James, Amy M; Mobli, Mehdi; Colgrave, Michelle L; Mylne, Joshua S; Rosengren, K Johan

    2017-07-28

    Seed storage proteins are both an important source of nutrition for humans and essential for seedling establishment. Interestingly, unusual napin-type 2S seed storage albumin precursors in sunflowers contain a sequence that is released as a macrocyclic peptide during post-translational processing. The mechanism by which such peptides emerge from linear precursor proteins has received increased attention; however, the structural characterization of intact precursor proteins has been limited. Here, we report the 3D NMR structure of the Helianthus annuus PawS1 ( p repro a lbumin w ith s unflower trypsin inhibitor- 1 ) and provide new insights into the processing of this remarkable dual-destiny protein. In seeds, PawS1 is matured by asparaginyl endopeptidases (AEPs) into the cyclic peptide SFTI-1 ( s un f lower t rypsin i nhibitor- 1 ) and a heterodimeric 2S albumin. The structure of PawS1 revealed that SFTI-1 and the albumin are independently folded into well-defined domains separated by a flexible linker. PawS1 was cleaved in vitro with recombinant sunflower HaAEP1 and in situ using a sunflower seed extract in a way that resembled the expected in vivo cleavages. Recombinant HaAEP1 cleaved PawS1 at multiple positions, and in situ , its flexible linker was removed, yielding fully mature heterodimeric albumin. Liberation and cyclization of SFTI-1, however, was inefficient, suggesting that specific seed conditions or components may be required for in vivo biosynthesis of SFTI-1. In summary, this study has revealed the 3D structure of a macrocyclic precursor protein and provided important mechanistic insights into the maturation of sunflower proalbumins into an albumin and a macrocyclic peptide. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  13. Effects of radiation therapy on tissue and serum concentrations of tumour associated trypsin inhibitor and their prognostic significance in rectal cancer patients

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    Stenman Ulf-Håkan

    2011-08-01

    Full Text Available Abstract Background We have previously demonstrated that elevated concentrations of tumour-associated trypsin inhibitor (TATI in both tumour tissue (t-TATI and in serum (s-TATI are associated with a poor prognosis in colorectal cancer patients. It was also found that s-TATI concentrations were lower in patients with rectal cancer compared to patients with colon cancer. In this study, we investigated the effects of neoadjuvant radiotherapy (RT on concentrations of t-TATI and s-TATI in patients with rectal cancer. Methods TATI was analysed in serum, normal mucosa and tumour tissue collected at various time points in 53 rectal cancer patients enrolled in a case-control study where 12 patients received surgery alone, 20 patients 5 × 5 Gy (short-term preoperative RT and 21 patients 25 × 2 Gy (long-term preoperative RT. T-TATI was analysed by immunohistochemistry and s-TATI was determined by an immunofluorometric assay. Mann-Whitney U test and Wilcoxon Z (Z test were used to assess t-TATI and s-TATI concentrations in relation to RT. Spearman's correlation (R test was used to explore the associations between t-TATI, s-TATI and clinicopathological parameters. Overall survival (OS according to high and low t-TATI and s-TATI concentrations was estimated by classification and regression tree analysis, Kaplan-Meier analysis and the log rank test. Results RT did not affect concentrations of t-TATI or s-TATI. In patients receiving short-term but not long-term RT, s-TATI concentrations were significantly higher 4 weeks post surgery than in serum drawn prior to surgery (Z = -3.366, P Conclusions The results presented here further validate the utility of t-TATI and s-TATI as prognostic biomarkers in patients with rectal cancer, independent of neoadjuvant RT.

  14. Lol p XI, a new major grass pollen allergen, is a member of a family of soybean trypsin inhibitor-related proteins.

    Science.gov (United States)

    van Ree, R; Hoffman, D R; van Dijk, W; Brodard, V; Mahieu, K; Koeleman, C A; Grande, M; van Leeuwen, W A; Aalberse, R C

    1995-05-01

    Monoclonal antibodies were obtained against an unknown allergen from Lolium perenne grass pollen. The allergen had an apparent molecular mass of 18 kd on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Earlier immunoblotting studies had shown that carbohydrate-specific IgG antibodies recognize an antigen of similar size. We sought to characterize the allergen biochemically and immunologically. The amino acid sequence of the allergen was determined by automated Edman degradation, and its monosaccharide composition was determined by gas chromatographic analysis. A panel of 270 grass pollen-positive sera was assessed in a RAST with the purified allergen. Protease digestion (proteinase K) and chemical deglycosylation (trifluoromethane sulfonic acid) were used to distinguish between carbohydrate and peptide epitopes for IgE antibodies. The allergen was shown to be a glycoprotein with a molecular mass of 16 kd, of which 8% is carbohydrate. Its amino acid sequence shares 32% homology with soybean trypsin inhibitor (Kunitz) but lacks its active site. No homology was found with known grass pollen allergens, hence it was designated Lol p XI. A high degree of homology (44%) was found with a tree pollen allergen, Ole e I, the major allergen of olive pollen. More than 65% of grass pollen-positive sera had IgE against Lol p XI. IgE reactivity was demonstrated both with the carbohydrate moiety and the peptide backbone. Lol p XI is a new major grass pollen allergen carrying an IgE-binding carbohydrate determinant. Lol p XI is structurally related to the major allergen from olive pollen.

  15. Defense response in non-genomic model species: methyl jasmonate exposure reveals the passion fruit leaves' ability to assemble a cocktail of functionally diversified Kunitz-type trypsin inhibitors and recruit two of them against papain.

    Science.gov (United States)

    Botelho-Júnior, Sylvio; Machado, Olga L T; Fernandes, Kátia V S; Lemos, Francisco J A; Perdizio, Viviane A; Oliveira, Antônia E A; Monteiro, Leandro R; Filho, Mauri L; Jacinto, Tânia

    2014-08-01

    Multiplicity of protease inhibitors induced by predators may increase the understanding of a plant's intelligent behavior toward environmental challenges. Information about defense mechanisms of non-genomic model plant passion fruit (Passiflora edulis Sims) in response to predator attack is still limited. Here, via biochemical approaches, we showed its flexibility to build-up a broad repertoire of potent Kunitz-type trypsin inhibitors (KTIs) in response to methyl jasmonate. Seven inhibitors (20-25 kDa) were purified from exposed leaves by chromatographic techniques. Interestingly, the KTIs possessed truncated Kunitz motif in their N-terminus and some of them also presented non-consensus residues. Gelatin-Native-PAGE established multiple isoforms for each inhibitor. Significant differences regarding inhibitors' activity toward trypsin and chymotrypsin were observed, indicating functional polymorphism. Despite its rarity, two of them also inhibited papain, and such bifunctionality suggests a recruiting process onto another mechanistic class of target protease (cysteine-type). All inhibitors acted strongly on midgut proteases from sugarcane borer, Diatraea saccharalis (a lepidopteran insect) while in vivo assays supported their insecticide properties. Moreover, the bifunctional inhibitors displayed activity toward midgut proteases from cowpea weevil, Callosobruchus maculatus (a coleopteran insect). Unexpectedly, all inhibitors were highly effective against midgut proteases from Aedes aegypti a dipteran insect (vector of neglected tropical diseases) opening new avenues for plant-derived PIs for vector control-oriented research. Our results reflect the KTIs' complexities in passion fruit which could be wisely exploited by influencing plant defense conditions. Therefore, the potential of passion fruit as source of bioactive compounds with diversified biotechnological application was strengthened.

  16. A novel spleen tyrosine kinase inhibitor blocks c-Jun N-terminal kinase-mediated gene expression in synoviocytes

    NARCIS (Netherlands)

    Cha, Hoon-Suk; Boyle, David L.; Inoue, Tomoyuki; Schoot, Reineke; Tak, Paul P.; Pine, Polly; Firestein, Gary S.

    2006-01-01

    Spleen tyrosine kinase (Syk) is a key regulator of cell signaling induced by cytokines or Fc receptor engagement. However, the role of Syk in rheumatoid arthritis (RA) is not known yet. We investigated the pathways activated by Syk in tumor necrosis factor-alpha (TNFalpha)-stimulated fibroblast-like

  17. Inter-alpha-trypsin inhibitor heavy chain 4: a novel biomarker for environmental exposure to particulate air pollution in patients with chronic obstructive pulmonary disease

    Directory of Open Access Journals (Sweden)

    Lee KY

    2015-04-01

    Full Text Available Kang-Yun Lee,1–3 Po-Hao Feng,1,2 Shu-Chuan Ho,4 Kai-Jen Chuang,5,6 Tzu-Tao Chen,2,3 Chien-Ling Su,2,4 Wen-Te Liu,2,4 Hsiao-Chi Chuang2,4 1Department of Internal Medicine, School of Medicine, College of Medicine, Taipei Medical University, 2Division of Pulmonary Medicine, Department of Internal Medicine, Shuang Ho Hospital, Taipei Medical University, 3Graduate Institute of Clinical Medicine, College of Medicine, Taipei Medical University, 4School of Respiratory Therapy, College of Medicine, Taipei Medical University, 5Department of Public Health, School of Medicine, College of Medicine, Taipei Medical University, 6School of Public Health, College of Public Health and Nutrition, Taipei Medical University, Taipei, Taiwan Abstract: Chronic obstructive pulmonary disease (COPD is a chronic inflammatory disease that is correlated with environmental stress. Particulate matter ≤10 µm (PM10 is considered to be a risk factor for COPD development; however, the effects of PM10 on the protein levels in COPD remain unclear. Fifty subjects with COPD and 15 healthy controls were recruited. Gene ontology analysis of differentially expressed proteins identified immune system process and binding as the most important biological process and molecular function, respectively, in the responses of PM10-exposed patients with COPD. Biomarkers for PM10 in COPD were identified and compared with the same in healthy controls and included proteoglycan 4 (PRG4, inter-alpha-trypsin inhibitor heavy chain 4 (ITIH4, and apolipoprotein F (APOF. PRG4 and ITIH4 were associated with a past 3-year PM10 exposure level. The receiver operating characteristic curve analysis showed that ITIH4 is a sensitive and specific biomarker for PM10 exposure (area under the curve [AUC] =0.690, P=0.015 compared with PRG4 (AUC =0.636, P=0.083, APOF (AUC =0.523, P=0.766, 8-isoprostane (AUC =0.563, P=0.405, and C-reactive protein (CRP; AUC =0.634, P=0.086. ITIH4 levels were correlated with CRP (r=0

  18. Effects of radiation therapy on tissue and serum concentrations of tumour associated trypsin inhibitor and their prognostic significance in rectal cancer patients

    International Nuclear Information System (INIS)

    Gaber, Alexander; Jirström, Karin; Stene, Christina; Hotakainen, Kristina; Nodin, Björn; Palmquist, Ingrid; Bjartell, Anders; Stenman, Ulf-Håkan; Jeppsson, Bengt; Johnson, Louis B

    2011-01-01

    We have previously demonstrated that elevated concentrations of tumour-associated trypsin inhibitor (TATI) in both tumour tissue (t-TATI) and in serum (s-TATI) are associated with a poor prognosis in colorectal cancer patients. It was also found that s-TATI concentrations were lower in patients with rectal cancer compared to patients with colon cancer. In this study, we investigated the effects of neoadjuvant radiotherapy (RT) on concentrations of t-TATI and s-TATI in patients with rectal cancer. TATI was analysed in serum, normal mucosa and tumour tissue collected at various time points in 53 rectal cancer patients enrolled in a case-control study where 12 patients received surgery alone, 20 patients 5 × 5 Gy (short-term) preoperative RT and 21 patients 25 × 2 Gy (long-term) preoperative RT. T-TATI was analysed by immunohistochemistry and s-TATI was determined by an immunofluorometric assay. Mann-Whitney U test and Wilcoxon Z (Z) test were used to assess t-TATI and s-TATI concentrations in relation to RT. Spearman's correlation (R) test was used to explore the associations between t-TATI, s-TATI and clinicopathological parameters. Overall survival (OS) according to high and low t-TATI and s-TATI concentrations was estimated by classification and regression tree analysis, Kaplan-Meier analysis and the log rank test. RT did not affect concentrations of t-TATI or s-TATI. In patients receiving short-term but not long-term RT, s-TATI concentrations were significantly higher 4 weeks post surgery than in serum drawn prior to surgery (Z = -3.366, P < 0.001). T-TATI expression correlated with male gender (R = 0.406, P = 0.008). High t-TATI expression in surgical specimens was associated with a significantly shorter OS (P = 0.045). S-TATI concentrations in serum drawn at all time points were associated with an impaired OS (P = 0.035 before RT, P = 0.001 prior to surgery, P = 0.043 post surgery). At all time points, s-TATI correlated with higher age (P < 0

  19. Ruptured Spleen

    Science.gov (United States)

    ... be caused by various underlying problems, such as mononucleosis and other infections, liver disease, and blood cancers. ... cause a ruptured spleen. For instance, people with mononucleosis — a viral infection that can cause an enlarged ...

  20. Wandering Spleen

    International Nuclear Information System (INIS)

    Al-Mashat, Faisal M.; Sibiany, Abdulrehman M.; Maimani, Abdulraouf A.; Alem, Fayad K.

    2004-01-01

    Cogenital malformations of the spleen are rare. We report 3 cases of wandering spleen presented as abominal or pelvi-abdominal mass. Two patients were suffering from chronic lower abdominal pain with thrombosed splenic pedicle and the third patient had an acute abdomen. All patients underwent splenectomies. Abdominal ultrasound, computerized tomography, Doppler's ultrasound, and radioisotpes studies were used to cpnfirm the diagnosis. The clinical, diagnostic and treatment modalities are discussed. (author)

  1. Trypsin from the pyloric caeca of bluefish (Pomatomus saltatrix).

    Science.gov (United States)

    Klomklao, Sappasith; Benjakul, Soottawat; Visessanguan, Wonnop; Kishimura, Hideki; Simpson, Benjamin K

    2007-12-01

    Trypsin was purified from the pyloric caeca of bluefish (Pomatomus saltatrix) by ammonium sulfate precipitation, acetone precipitation and soybean trypsin inhibitor-Sepharose 4B affinity chromatography. Bluefish trypsin migrated as a single band using both sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and native-PAGE and had a molecular mass of 28 kDa. The optima pH and temperature for the hydrolysis of benzoyl-dl-arginine-p-nitroanilide (BAPNA) were 9.5 and 55 degrees C, respectively. The enzyme was stable over a broad pH range (7 to 12), but was unstable at acidic pH, and at temperatures greater than 40 degrees C. The enzyme was inhibited by specific trypsin inhibitors: soybean trypsin inhibitor (SBTI), N-p-tosyl-l-lysine chloromethyl ketone (TLCK) and the serine protease inhibitor phenylmethyl sulfonylfluoride (PMSF). CaCl2 partially protected trypsin against activity loss at 40 degrees C, but NaCl (0 to 30%) decreased the activity in a concentration dependent manner. The N-terminal amino acid sequence of trypsin was determined as IVGGYECKPKSAPVQVSLNL and was highly homologous to other known vertebrate trypsins.

  2. Case report on the role of radiofrequency-assisted spleen-preserving surgery for splenic metastasis in the era of check-point inhibitors.

    Science.gov (United States)

    Mudan, Satvinder; Kumar, Jayant; Mafalda, Neves C; Kusano, Tomokazu; Reccia, Isabella; Zanallato, Artur; Dalgleish, Angus; Habib, Nagy

    2017-12-01

    An isolated splenic metastasis is a rare phenomenon noted in advanced stage melanoma. We report the role of radiofrequency (RF) -based splenic-preserving splenectomy in a patient with a solitary splenic metastasis from advanced stage melanoma that was managed with checkpoint inhibitors. We report a case of a 60-year-old man who presented with multiple lung metastases and a solitary splenic metastasis with advanced stage melanoma following excision of primary from his trunk 2.3 years back. Considering the diagnosis of advanced stage melanoma with multiple lung metastases and a solitary splenic metastasis, and its ongoing progressive nature. This case was discussed in the tumour board meeting. A decision was made to commence treatment with immunotherapy in the form of PD-1 inhibitor (programmed cell death 1 receptor) pembrolizumab. Follow-up restaging computer tomography (CT) scan of the abdomen and chest showed a significant reduction in the lung and chest wall lesions, but the splenic lesion remained unchanged. Given the lack of response to treatment in the splenic metastasis and the significant decrease in lung metastases, the multidisciplinary team decided that a partial splenectomy combined with continued immunotherapy treatment would be appropriate as the success of immunotherapy was imminent within the splenic preservation. The postoperative recovery was smooth and the patient was discharged from hospital on the sixth postoperative day with normal platelets and white blood cells. The histopathological analysis of the resected specimen showed a metastatic melanoma with negative margins.At 10-month follow-up after the splenic resection the patient had not experienced further tumour recurrences. Spleen-preserving resection for an isolated, solitary splenic metastasis of melanoma is a feasible approach as it not only preserves the ongoing efficacy of checkpoint inhibitors by preserving the physiological T cell milieu, but the immunomodulation properties of RF can

  3. Radioimmunoassay of trypsin

    International Nuclear Information System (INIS)

    Stagg, B.H.; Wood, T.P.

    1979-01-01

    This review describes the development and application of a novel test to determine levels of human immunoreactive trypsin, an enzyme produced solely by the pancreas, in biological fluids. Being organ-specific, the assay of immunoreactive trypsin should be an ideal marker of pancreatic function, and this is supported by the results of a number of clinical and research investigations. Use of this assay in studies of chronic pancreatitis, juvenile-onset diabetes, and cystic fibrosis, has yielded much valuable data, and it is expected that further research will lead to and improved understanding of these and other conditions associated with the pancreas in health and disease. (author)

  4. Smart PEGylation of trypsin.

    Science.gov (United States)

    Zarafshani, Zoya; Obata, Toshihiro; Lutz, Jean-François

    2010-08-09

    Thermoresponsive oligo(ethylene glycol)-based copolymers were investigated for trypsin conjugation. These copolymers have been synthesized by atom transfer radical polymerization of 2-(2-methoxyethoxy)ethyl methacrylate (MEO(2)MA) with oligo(ethylene glycol) methyl ether methacrylate (OEGMA(475), M(n) = 475 g.mol(-1)) at 60 degrees C in the presence of copper(I) chloride and 2,2'-bipyridyl. Two different ATRP initiators, containing succinimidyl ester moieties, were tested, namely, N-succinimidyl-2-bromopropionate and N-succinimidyl-2-bromoisobutyrate. In both cases, ATRP afforded well-defined polymers with a narrow molecular weight distribution and controlled chain-ends. However, the efficiency of initiation of the two initiators was lower than 1 and therefore the formed polymers exhibited a higher than expected mean degree of polymerization. Nevertheless, all types of polymers could be conjugated to trypsin. The conjugation reaction was performed in borax-HCl buffer. Sodium dodecyl sulfate poly(acrylamide) gel electrophoresis (SDS-PAGE) indicated that polymer/enzyme conjugates were obtained in all cases. However, (co)polymers initiated by N-succinimidyl-2-bromopropionate led to the best conjugation results. The formed P(MEO(2)MA-co-OEGMA(475))-trypsin conjugates were found to be thermoresponsive and moreover exhibited a higher enzymatic activity than unmodified trypsin.

  5. Free radical inactivation of trypsin

    International Nuclear Information System (INIS)

    Cudina, Ivana; Jovanovic, S.V.

    1988-01-01

    Reactivities of free radical oxidants, radical OH, Br2-anion radical and Cl 3 COO radical and a reductant, CO2-anion radical, with trypsin and reactive protein components were determined by pulse radiolysis of aqueous solutions at pH 7, 20 0 C. Highly reactive free radicals, radical OH, Br2-anion radical and CO2-anion radical, react with trypsin at diffusion controlled rates. Moderately reactive trichloroperoxy radical, k(Cl 3 COO radical + trypsin) preferentially oxidizes histidine residues. The efficiency of inactivation of trypsin by free radicals is inversely proportional to their reactivity. The yields of inactivation of trypsin by radical OH, Br2-anion radical and CO2-anion radical are low, G(inactivation) = 0.6-0.8, which corresponds to ∼ 10% of the initially produced radicals. In contrast, Cl 3 COO radical inactivates trypsin with ∼ 50% efficiency, i.e. G(inactivation) = 3.2. (author)

  6. Interaction of gallic acid with trypsin analyzed by spectroscopy

    Directory of Open Access Journals (Sweden)

    Hao Song

    2015-06-01

    Full Text Available The interactions between trypsin and gallic acid (GA were investigated by means of fluorescence spectroscopy, UV-vis absorption spectroscopy, resonance light scattering (RLS spectroscopy, synchronous fluorescence spectroscopy, and enzymatic inhibition assay. It was found that GA can cause the fluorescence quenching of trypsin during the process of formation of GA-trypsin complex, resulting in inhibition of trypsin activity (IC50 = 3.9 × 10−6 mol/L. The fluorescence spectroscopic data showed that the quenching efficiency can reach about 80%. The binding constants were 1.9371 × 104 L/mol, 1.8192 × 104 L/mol, and 1.7465 × 104 L/mol at three temperatures, respectively. The thermodynamic parameters revealed that hydrogen bonds, van der Waals, hydrophobic, and electrostatic interactions were involved in the binding process of GA to trypsin. Molecular modeling studies illustrated a specific display of binding information and explained most of the experiment phenomena. The microenvironments of tryptophan and tyrosine residue in trypsin were changed by the GA. Results indicated that GA was a strong quencher and inhibitor of trypsin.

  7. Efficiency of inactivation of trypsin inhibitory activity in some selected ...

    African Journals Online (AJOL)

    Trypsin inhibitor (TI) levels in the crop seeds varied between 0.0 in Adansonia digitata and 40.8 TIU/mg in Pterocarpus osun. Efficiency of inactivation of TI by autoclaving ranged from 58.1% in Millettia thonningii to 100% in Sesbania pachycarpa and Lonchocarpus. sericeus. It is concluded that the effect of heat treatment on ...

  8. Internal mobility of reactive-site-hydrolyzed recombinant Cucurbita maxima trypsin inhibitor-V characterized by NMR spectroscopy: evidence for differential stabilization of newly formed C- and N-termini.

    Science.gov (United States)

    Liu, J; Prakash, O; Huang, Y; Wen, L; Wen, J J; Huang, J K; Krishnamoorthi, R

    1996-09-24

    The solution structure and internal dynamics of the reactive-site (Lys44-Asp45 peptide bond) hydrolyzed form of recombinant Cucurbita maxima trypsin inhibitor-V (rCMTI-V*) were characterized by the application of two-dimensional 1H-15N NMR methods to the uniformly 15N-labeled protein. The 1H-15N chemical shift correlation spectra of rCMTI-V* were assigned, and the chemical shift data were compared with those available for rCMTI-V [Liu, J., Prakash, O., Cai, M., Gong, Y., Huang, Y., Wen, L., Wen, J. J., Huang, J.-K., & Krishnamoorthi, R. (1996) Biochemistry 35, 1516-1524] and CMTI-V* [Cai, M., Gong, Y., Prakash, O., & Krishnamoorthi, R. (1995) Biochemistry 34, 12087-12094] for which three-dimensional solution structures have been determined. It was deduced that the solution structure of rCMTI-V* was almost the same as that of CMTI-V*. 15N spin-lattice and spin-spin relaxation rate constants (R1 and R2, respectively) and ¿1H¿-15N steady-state heteronuclear Overhauser effects were measured for the peptide NH units and arginine and tryptophan N epsilon H groups in rCMTI-V*, and the model-free parameters [Lipari, G., & Szabo, A. (1982) J. Am. Chem. Soc. 104, 4546-4559, 4559-4570] were computed. Most of the backbone of rCMTI-V* is found to be highly constrained (S2 = 0.85), including the N-terminal residues 3-6 (S2 = 0.77). Residues 39-44, forming the C-terminal fragment of the binding loop, exhibit increased mobility (S2 = 0.51); however, the N-terminal segment (residues 46-48) retains rigidity as in the intact form (S2 = 0.83). The S2 values, 0.78 and 0.59, respectively, of Arg50 and Arg52 side chain NHs provide evidence not only for the conservation of the Arg hydrogen-bonds with the binding loop segments but also for the difference in strength between them. This is consistent with the earlier observation made from a study of rCMTI-V at two different pHs and its R50 and R52 mutants [Cai, M., Huang, Y., Prakash, O., Wen, L., Dunkelbarger, S. P., Huang, J.-K., Liu, J

  9. The retrorenal spleen

    International Nuclear Information System (INIS)

    Hopper, K.D.; Chantelois, A.E.

    1987-01-01

    An uncommon but potentially disastrous situation for invasive percutaneous renal procedures is the placement of the spleen behind the upper left renal pole. The authors termed this unique anatomic variant ''retrorenal spleen.'' Recently the authors reviewed the CT studies of 85 patients 16-85 years of age scanned in both the supine and prone positions. The relationship of the left kidney and spleen was evaluated. The overall frequency of retrorenal spleen was 12.7% for the supine and 17.9% for the prone studies. Careful correlation was made between the supine and prone studies with respect to the changing anatomic relationship of the kidney and spleen. In addition, the position of the suprarenal spleen was also evaluated in an effort to determine the safety of the subcostal approach

  10. Mechanisms involved in the chemical inhibition of the Eosin-sensitized photooxidation of trypsin

    Energy Technology Data Exchange (ETDEWEB)

    Rizzuto, F.; Spikes, J.D.

    1975-01-01

    A large series of compounds was screened for ability to protect trypsin from eosin-sensitized photodynamic inactivation. Eosin-sensitized photooxidation reactions of this type typically proceed via the triplet state of the dye and often involve singlet state oxygen as the oxidizing entity. In order to determine the mechanisms by which trypsin is protected from photoinactivation, a number of good protective agents (inhibitors) and some non-protective agents were selected for more detailed flash photolysis studies. Good inhibitors such as p-phenylenediamine, n-propyl gallate, serotonin creatinine sulfate and p-toluenediamine competed efficiently with oxygen and with trypsin for reaction with the triplet state of eosin. The inhibitors were shown to quench triplet eosin to the ground state and/or reduce triplet eosin to form the semireduced eosin radical and an oxidized form of the inhibitor. In the latter case, oxidized inhibitor could react by a reverse electron transfer reaction with the semireduced eosin radical to regenerate ground state eosin and the inhibitor. The good inhibitors also competed effectively with trypsin for oxidation by semioxidized eosin, thus giving another possible protective mechanism. Non-inhibitors such as halogen ions and the paramagnetic ions Co/sup + +/, Cu/sup + +/ and Mn/sup + +/ reacted only slowly with triplet and with semioxidized eosin. The primary pathway for the eosin-sensitized photooxidation of trypsin at pH 8.0 involved singlet oxygen, although semioxidized eosin may also participate.

  11. MORPHOMETRY OF SPLEEN

    Directory of Open Access Journals (Sweden)

    Radhika

    2016-03-01

    Full Text Available INTRODUCTION Spleen is organ of lymphatic system located on left side of abdominal cavity under diaphragm. It is a secondary lymphatic organ that plays an important role in cell mediated immunity. Foetal spleen is erythropoietic in nature. MATERIAL & METHODS Present study was done in 50 adult spleens and 50 foetal spleens. RESULTS Morphometric features like length, breadth, thickness & weight are measured. Length varied from 6.3 to 12.5 cm, breadth varied from 2.6 to 8.6 cm, thickness ranged from 2 cm to 4.6 cm, weight ranged from 65 g to 225 g. Average total length of spleen is 2.52 cm x 1.76 x 2 cm, weight 6.5 g. Shapes of spleens observed wedge shape spleen–48%, tetrahedral spleen–24%, triangular spleen-28%. Splenic notches on superior border & inferior border are observed. Incident of accessory spleen in 1% of cases. CONCLUSIONS Present knowledge of study may be helpful for surgeons in surgical procedures like splenectomy, resection of tumours and extirpation of cysts

  12. Computerized spleen volumetry

    International Nuclear Information System (INIS)

    Jahnke, T.; Mohring, R.; Schertel, L.

    1981-01-01

    We examined in experimental studies and clinical investigations on 34 patients in how far volumetry of the spleen can be carried out with a commonly available program, a whole-body computerized tomograph (SOMATOM) and an analytic equipment (EVALUSKOP). In this connection the authors tried to find also other ways of spleen volumetry by means of this unit combination. Our final result was that the given program for the usage of labelled areas presents itself as the best-suited technique for spleen volumetry which is also applicable in practice. (orig./MG) [de

  13. Overview of the Spleen

    Science.gov (United States)

    ... Neisseria meningitidis , and Haemophilus influenzae . Because of this risk, people receive vaccinations to help protect them from infection with these organisms. People should also be sure they receive the influenza vaccine every year, as is now ... the Spleen ...

  14. Laparoscopic Spleen Removal (Splenectomy)

    Science.gov (United States)

    ... Affairs and Humanitarian Efforts Login Laparoscopic Spleen Removal (Splenectomy) Patient Information from SAGES Download PDF Find a ... are suspected. What are the Advantages of Laparoscopic Splenectomy? Individual results may vary depending on your overall ...

  15. NMR studies of internal dynamics of serine proteinase protein inhibitors: Binding region mobilities of intact and reactive-site hydrolyzed Cucurbita maxima trypsin inhibitor (CMTI)-III of the squash family and comparison with those of counterparts of CMTI-V of the potato I family.

    Science.gov (United States)

    Liu, J; Gong, Y; Prakash, O; Wen, L; Lee, I; Huang, J K; Krishnamoorthi, R

    1998-01-01

    Serine proteinase protein inhibitors follow the standard mechanism of inhibition (Laskowski M Jr, Kato I, 1980, Annu Rev Biochem 49:593-626), whereby an enzyme-catalyzed equilibrium between intact (I) and reactive-site hydrolyzed inhibitor (I*) is reached. The hydrolysis constant, Khyd, is defined as [I*]/[I]. Here, we explore the role of internal dynamics in the resynthesis of the scissile bond by comparing the internal mobility data of intact and cleaved inhibitors belonging to two different families. The inhibitors studied are recombinant Cucurbita maxima trypsin inhibitor III (rCMTI-III; Mr 3 kDa) of the squash family and rCMTI-V (Mr approximately 7 kDa) of the potato I family. These two inhibitors have different binding loop-scaffold interactions and different Khyd values--2.4 (CMTI-III) and 9 (CMTI-V)--at 25 degrees C. The reactive-site peptide bond (P1-P1') is that between Arg5 and Ile6 in CMTI-III, and that between Lys44 and Asp45 in CMTI-V. The order parameters (S2) of backbone NHs of uniformly 15N-labeled rCMTI-III and rCMTI-III* were determined from measurements of 15N spin-lattice and spin-spin relaxation rates, and [1H]-15N steady-state heteronuclear Overhauser effects, using the model-free formalism, and compared with the data reported previously for rCMTI-V and rCMTI-V*. The backbones of rCMTI-III [(S2) = 0.71] and rCMTI-III* [(S2) = 0.63] are more flexible than those of rCMTI-V [(S2) = 0.83] and rCMTI-V* [(S2) = 0.85]. The binding loop residues, P4-P1, in the two proteins show the following average order parameters: 0.57 (rCMTI-III) and 0.44 (rCMTI-III*); 0.70 (rCMTI-V) and 0.40 (rCMTI-V*). The P1'-P4' residues, on the other hand, are associated with (S2) values of 0.56 (rCMTI-III) and 0.47 (rCMTI-III*); and 0.73 (rCMTI-V) and 0.83 (rCMTI-V*). The newly formed C-terminal (Pn residues) gains a smaller magnitude of flexibility in rCMTI-III* due to the Cys3-Cys20 crosslink. In contrast, the newly formed N-terminal (Pn' residues) becomes more flexible

  16. Correlation of binding-loop internal dynamics with stability and function in potato I inhibitor family: relative contributions of Arg(50) and Arg(52) in Cucurbita maxima trypsin inhibitor-V as studied by site-directed mutagenesis and NMR spectroscopy.

    Science.gov (United States)

    Cai, Mengli; Gong, Yu-Xi; Wen, Lisa; Krishnamoorthi, Ramaswamy

    2002-07-30

    The side chains of Arg(50) and Arg(52) at positions P(6)' and P(8)', respectively, anchor the binding loop to the protein scaffold by means of hydrogen bonds in Cucurbita maxima trypsin inhibitor-V (CMTI-V), a potato I family member. Here, we have investigated the relative contributions of Arg(50) and Arg(52) to the binding-loop flexibility and stability by determining changes in structure, dynamics, and proteolytic stability as a consequence of individually mutating them into an alanine. We have compared chemical shift assignments of main-chain hydrogens and nitrogens, and (1)H-(1)H interresidue nuclear Overhauser effects (NOEs) for the two mutants with those of the wild-type protein. We have also measured NMR longitudinal and transverse relaxation rates and (15)N-(1)H NOE enhancements for all backbone and side-chain NH groups and calculated the model-free parameters for R50A-rCMTI-V and R52A-rCMTI-V. The three-dimensional structures and backbone dynamics of the protein scaffold region remain very similar for both mutants, relative to the wild-type protein. The flexibility of the binding loop is increased in both R50A- and R52A-rCMTI-V. In R52A-rCMTI-V, the mean generalized order parameter () of the P(6)-P(1) residues of the binding loop (39-44) decreases to 0.68 +/- 0.02 from 0.76 +/- 0.04 observed for the wild-type protein. However, in R50A-rCMTI-V, the flexibility of the whole binding loop increases, especially that of the P(1)'-P(3)' residues (45-47), whose value drops dramatically to 0.35 +/- 0.03 from 0.68 +/- 0.03 determined for rCMTI-V. More strikingly, S(2) values of side-chain N epsilon Hs reveal that, in the R50A mutant, removal of the R50 hydrogen bond results in the loss of the R52 hydrogen bond too, whereas in R52A, the R50 hydrogen bond remains unaffected. Kinetic data on trypsin-catalyzed hydrolysis of the reactive-site peptide bond (P(1)-P(1)') suggest that the activation free energy barrier of the reaction at 25 degrees C is reduced by 2.1 kcal

  17. Liver and spleen scintigraphy

    International Nuclear Information System (INIS)

    Devries, D.F.

    1988-01-01

    Since the introduction of liver and spleen scintigraphy in the early 1950s, it has undergone considerable changes, the most notable being technetium 99m sulfur colloid, the gamma camera, and single photon emission computed tomography (SPECT). What is the role f liver-spleen scintigraphy in this high-technology society? This chapter attempts to address this question by looking at the radiopharmaceuticals, the technique, and most importantly, the application of scintigraphy to the diagnosis of focal and diffuse hepatic and splenic disease

  18. Radiology of the spleen

    International Nuclear Information System (INIS)

    Robertson, F.; Leander, P.; Ekberg, O.

    2001-01-01

    The spleen is generally not considered a challenge to the radiologist. Most often it poses a problem by anomalies or an irregular but normal contrast enhancement; however, a variety of inflammatory, infectious and neoplastic diseases may involve the spleen. CT and ultrasonography are screening modalities for the spleen. For problem solving, MR imaging can be helpful, especially due to its free choice of the imaging plane and because of the high resolution in contrast MR imaging. Splenic angiography as a diagnostic tool has generally been replaced by CT, ultrasound, or MR and is now used as an interventional method, e.g., in non-surgical management of patients with chronic idiopathic thrombocytopenia or in patients with splenic trauma. This article reviews the radiology of the spleen, including anatomy, embryology, splenomegaly, splenic injury, infarction, cysts, tumors, abscesses, sarcoidosis, and AIDS. Knowledge about the use of different imaging modalities and underlying gross and microscopic pathologic features leads to a better understanding of the radiologic findings. (orig.)

  19. Inhibitors

    Science.gov (United States)

    ... JM, and the Hemophilia Inhibitor Research Study Investigators. Validation of Nijmegen-Bethesda assay modifications to allow inhibitor ... webinars on blood disorders Language: English (US) Español (Spanish) File Formats Help: How do I view different ...

  20. Hemangiopericytoma of the spleen.

    Science.gov (United States)

    Illuminati, Giulio; Pizzardi, Giulia; Calio, Francesco; Pacilè, Maria A; Carboni, Fabio; Palumbo, Piergaspare; Vietri, Francesco

    2015-03-01

    Hemangiopericytoma of the spleen is a very rare tumor, with 14 isolated reports. It was our aim to review our experience and compare it with all the reported cases in an attempt to standardize surgical treatment, adjuvant treatment and follow-up protocol of this infrequent condition. A consecutive case series study, with a mean follow-up of 44 months. Five patients (mean age, 49 years) underwent simple splenectomy for hemangiopericytoma limited to the spleen followed by adriamycin-based chemotherapy in one patient. All the patients are alive and free from disease. For tumors confined to the spleen, simple splenectomy can be considered curative, without any need for further adjuvant treatment. On review of the medical literature, cure can still be achieved with complete resection of recurrences, when feasible, with adjuvant chemotherapy being also indicated. The slow-growing pattern of the tumor suggests a 10-year follow-up. Copyright © 2015 Surgical Associates Ltd. Published by Elsevier Ltd. All rights reserved.

  1. Paediatric Wandering Spleens in Malawi

    African Journals Online (AJOL)

    by a double layer of peritoneum. The wandering spleen is the rare description of an abnormally positioned spleen, which is thought to occur due to laxity, abnormality or absence of the aforementioned ligaments. The wandering spleen is noted to have a longer than normal pedicle, and because of its intraperitoneal location, ...

  2. The sequence and X-ray structure of the trypsin from Fusarium oxysporum.

    Science.gov (United States)

    Rypniewski, W R; Hastrup, S; Betzel, C; Dauter, M; Dauter, Z; Papendorf, G; Branner, S; Wilson, K S

    1993-06-01

    The trypsin from Fusarium oxysporum is equally homologous to trypsins from Streptomyces griseus, Streptomyces erythraeus and to bovine trypsin. A DFP (diisopropylfluorophosphate) inhibited form of the enzyme has been crystallized from 1.4 M Na2SO4, buffered with citrate at pH 5.0-5.5. The crystals belong to space group P2(1) with cell parameters a = 33.43 A, b = 67.65 A, c = 39.85 A and beta = 107.6 degrees. There is one protein molecule in the asymmetric unit. X-ray diffraction data to a resolution of 1.8 A were collected on film using synchrotron radiation. The structure was solved by molecular replacement using models of bovine and S. griseus trypsins and refined to an R-factor of 0.141. The overall fold is similar to other trypsins, with some insertions and deletions. There is no evidence of the divalent cation binding sites seen in other trypsins. The covalently bound inhibitor molecule is clearly visible.

  3. Crystallization, data collection and processing of the chymotrypsin–BTCI–trypsin ternary complex

    Energy Technology Data Exchange (ETDEWEB)

    Esteves, Gisele Ferreira; Teles, Rozeni Chagas Lima; Cavalcante, Nayara Silva; Neves, David; Ventura, Manuel Mateus [Laboratório de Biofísica, Instituto de Ciências Biológicas, Universidade de Brasília, 70910-900 Brasília-DF (Brazil); Barbosa, João Alexandre Ribeiro Gonçalves, E-mail: joao@lnls.br [Center for Structural Molecular Biology (CeBiME), Brazilian Synchrotron Light Laboratory (LNLS), CP 6192, 13083-970 Campinas-SP (Brazil); Freitas, Sonia Maria de, E-mail: joao@lnls.br [Laboratório de Biofísica, Instituto de Ciências Biológicas, Universidade de Brasília, 70910-900 Brasília-DF (Brazil)

    2007-12-01

    A ternary complex of the proteinase inhibitor (BTCI) with trypsin and chymotrypsin was crystallized and its crystal structure was solved by molecular replacement. A ternary complex of the black-eyed pea trypsin and chymotrypsin inhibitor (BTCI) with trypsin and chymotrypsin was crystallized by the sitting-drop vapour-diffusion method with 0.1 M HEPES pH 7.5, 10%(w/v) polyethylene glycol 6000 and 5%(v/v) 2-methyl-2,4-pentanediol as precipitant. BTCI is a small protein with 83 amino-acid residues isolated from Vigna unguiculata seeds and is able to inhibit trypsin and chymotrypsin simultaneously by forming a stable ternary complex. X-ray data were collected from a single crystal of the trypsin–BTCI–chymotrypsin ternary complex to 2.7 Å resolution under cryogenic conditions. The structure of the ternary complex was solved by molecular replacement using the crystal structures of the BTCI–trypsin binary complex (PDB code) and chymotrypsin (PDB code) as search models.

  4. Crystallization, data collection and processing of the chymotrypsin–BTCI–trypsin ternary complex

    International Nuclear Information System (INIS)

    Esteves, Gisele Ferreira; Teles, Rozeni Chagas Lima; Cavalcante, Nayara Silva; Neves, David; Ventura, Manuel Mateus; Barbosa, João Alexandre Ribeiro Gonçalves; Freitas, Sonia Maria de

    2007-01-01

    A ternary complex of the proteinase inhibitor (BTCI) with trypsin and chymotrypsin was crystallized and its crystal structure was solved by molecular replacement. A ternary complex of the black-eyed pea trypsin and chymotrypsin inhibitor (BTCI) with trypsin and chymotrypsin was crystallized by the sitting-drop vapour-diffusion method with 0.1 M HEPES pH 7.5, 10%(w/v) polyethylene glycol 6000 and 5%(v/v) 2-methyl-2,4-pentanediol as precipitant. BTCI is a small protein with 83 amino-acid residues isolated from Vigna unguiculata seeds and is able to inhibit trypsin and chymotrypsin simultaneously by forming a stable ternary complex. X-ray data were collected from a single crystal of the trypsin–BTCI–chymotrypsin ternary complex to 2.7 Å resolution under cryogenic conditions. The structure of the ternary complex was solved by molecular replacement using the crystal structures of the BTCI–trypsin binary complex (PDB code) and chymotrypsin (PDB code) as search models

  5. Structure basis 1/2SLPI and porcine pancreas trypsin interaction

    Energy Technology Data Exchange (ETDEWEB)

    Fukushima, Kei; Kamimura, Takashi; Takimoto-Kamimura, Midori, E-mail: m.kamimura@teijin.co.jp [Teijin Institute for Bio-Medical Research, 4-3-2 Asahigaoka, Hino-shi, Tokyo 191-8512 (Japan)

    2013-11-01

    1/2SLPI is a C-terminal domain of SLPI (secretory leukocyte protease inhibitor) which inhibits various serine proteases broadly. The present study is the first X-ray structural report on how 1/2SLPI with P1 Leu strongly inhibits trypsin and how it can inhibit multiple serine proteases. SLPI (secretory leukocyte protease inhibitor) is a 107-residue protease inhibitor which inhibits various serine proteases, including elastase, cathepsin G, chymotrypsin and trypsin. SLPI is obtained as a multiple inhibitor in lung defense and in chronic airway infection. X-ray crystal structures have so far reported that they are full-length SLPIs with bovine α-chymotrypsin and 1/2SLPI (recombinant C-terminal domain of SLPI; Arg58–Ala107) with HNE (human neutrophil elastase). To understand the role of this multiple inhibitory mechanism, the crystal structure of 1/2SLPI with porcine pancreas trypsin was solved and the binding modes of two other complexes compared. The Leu residue surprisingly interacts with the S1 site of trypsin, as with chymotrypsin and elastase. The inhibitory mechanism of 1/2SLPI using the wide primary binding site contacts (from P2′ to P5) with various serine proteases is discussed. These inhibitory mechanisms have been acquired in the evolution of the protection system for acute inflammatory diseases.

  6. HYDROLYSIS OF CHEESEWHEY PROTEINSWITH TRYPSIN, CHYMOTRYPSINAND CARBOXYPEPTIDASEA

    Directory of Open Access Journals (Sweden)

    M. F. CUSTÓDIO

    2009-01-01

    Full Text Available

    This work presents a method for adding value to cheese whey residues by whey proteins hydrolysis, using trypsin, chymotrypsin and carboxypeptidase A as catalysts. Sweet cheese whey was dialyzed and filtered in kaolin. Lactose and protein contents were analyzed after each step. The activities of bovine pancreas trypsin and chymotrypsin were measured at different pHs and temperatures. The optimal pH for the hydrolysis of whey proteins was 9.0 for both enzymes. Optima temperatures were 60ºC for trypsin, and 50ºC for chymotrypsin. Trypsin exhibited typical Michaelis-Menten behavior, but chymotrypsin did not. Electrophoretic analysis showed that neither trypsin nor chymotrypsin alone hydrolyzed whey proteins in less than three hours. Hydrolysis rates of -lactalbumin by trypsin, and of bovine serum albumin by chymotrypsin were low. When these enzymes were combined, however, all protein fractions were attacked and rates of hydrolysis were enhanced by one order of magnitude. The addition of carboxypeptidase A to the others enzymes did not improve the process yield.

  7. High feline trypsin-like immunoreactivity in a cat with pancreatitis and hepatic lipidosis.

    Science.gov (United States)

    Bruner, J M; Steiner, J M; Williams, D A; Van Alstine, W G; Blevins, W

    1997-06-15

    A 1.5-year-old domestic shorthair cat was examined because of vomiting and icterus. Clinicopathologic abnormalities included high alanine transaminase, alkaline phosphatase, and gamma-glutamyltransferase activities and high total bilirubin concentration. During abdominal ultrasonography, the left limb and body of the pancreas appeared hypoechoic, and a small quantity of peritoneal effusion was seen. The liver was diffusely hyperechoic, with echogenicity similar to that of the spleen, indicating hepatic lipidosis. Feline trypsin-like immunoreactivity was high, suggesting that the cat also had pancreatitis. The cat was treated with crystalloid fluids and was fed a protein-restricted diet via a percutaneous endoscopically placed gastrostomy tube. The cat's condition continued to deteriorate despite medical treatment, and it was euthanatized. Necropsy confirmed the clinical suspicion of acute pancreatitis and hepatic lipidosis. This case suggests that measurement of trypsin-like immunoreactivity may be useful in cats suspected of having pancreatitis.

  8. Isolated Amoebic Abscess of Spleen

    Directory of Open Access Journals (Sweden)

    Kaushik M

    2013-04-01

    Full Text Available Amoebic liver abscess is the most common extraintestinal manifestation of amoebiasis. Extrahepatic amoebic abscesses have occasionally been described in the lung, brain, and skin and presumably result from hematogenous spread. Isolated amoebic abscess of spleen has been reported scarcely in literature. We report here a case of isolated amoebic abscess of spleen.

  9. Chemical repair of trypsin-histidinyl radical

    International Nuclear Information System (INIS)

    Jovanovic, S.V.; Ruvarac, I.; Jankovic, I.; Josimovic, L.

    1991-01-01

    Oxyl radicals, such as hydroxyl, alkoxyl and peroxyl, react with biomolecules to produce bioradicals. Unless chemically repaired by suitable antioxidants, these bioradicals form stable products. This leads to loss of biological function of parent biomolecules with deleterious biological results, such as mutagenesis and cancer. Consequently, the understanding of the mechanisms of oxyl radical damage to biomolecules and chemical repair of such damage is crucial for the development of strategies for anticarcinogenesis and radioprotection. In this study the chemical repair of the histidinyl radical generated upon the trichloromethylperoxyl radical reaction with trypsin vas investigated by gamma radiolysis. The trypsin histidinyl radical is a resonance-stabilized heterocyclic free radical which was found to be unreactive with oxygen. The efficacy of the chemical repair of the trypsin-histidinyl radical by endogenous antioxidants which are electron donors (e.g. 5-hydroxytryptophan, uric acid) is compared to that of antioxidants which are H-atom donors (e. g. glutathione). 9 refs., 2 figs., 1 tab

  10. Enzymatic synthesis of gold nanoflowers with trypsin

    International Nuclear Information System (INIS)

    Li Linmei; Weng Jian

    2010-01-01

    A one-step and eco-friendly approach for the room-temperature synthesis of trypsin-mediated three-dimensional (3D) gold nanoflowers (AuNFs) with high colloidal stability is demonstrated. To prepare AuNFs, ascorbic acid (AA) was quickly added into the premixed solution of HAuCl 4 and trypsin at pH = 5.0. The results show that the molar ratio and feeding order of reactant agents, pH and reaction time play important roles in the formation of NFs. The growth mechanism of AuNFs is suggested as three steps: (1) immobilization of AuCl 4 - ions with a positively charged trypsin template, (2) spontaneous reduction of AuCl 4 - ions with AA in situ and capping Au 0 by 12 cysteines of trypsin, (3) reduction of more AuCl 4 - ions on the Au nuclei formed in the initial stages and anisotropic growth into AuNFs.

  11. The Spleen: A forgotten organ

    International Nuclear Information System (INIS)

    Castrillon German; Montoya Maria del Pilar; Echeverri Santiago

    2010-01-01

    The spleen has traditionally been regarded as an orphan organ. Its embryological development is described together with the digestive system, although it is not part of the gastrointestinal tract. Its main function is during the early fetal development when it produces both red and white blood cells, losing this function during the late fetal life. Nevertheless, the spleen continues to work as a filter for blood cells and also has important immune functions. As it is not necessary for the preservation of vital life functions, the spleen receives limited attention by both radiologists and clinicians. It is, however, an organ with multiple pathological conditions that have representation in the different diagnostic imaging modalities; radiologists, therefore, must be aware of these conditions and their radiological characteristics. This article provides a diagnostic approach to the most common diseases affecting the spleen using tomography and magnetic resonance.

  12. Trypsin from unicorn leatherjacket (Aluterus monoceros) pyloric caeca: purification and its use for preparation of fish protein hydrolysate with antioxidative activity.

    Science.gov (United States)

    Zamani, Abbas; Benjakul, Soottawat

    2016-02-01

    Fish proteases, especially trypsin, could be used to prepare fish protein hydrolysates with antioxidative activities. In this study, trypsin from the pyloric caeca of unicorn leatherjacket was purified by ammonium sulfate precipitation and soybean trypsin inhibitor (SBTI)-Sepharose 4B affinity chromatography. Hydrolysate from Indian mackerel protein isolate with different degrees of hydrolysis (20, 30 and 40% DH) was prepared using the purified trypsin, and antioxidative activities (1,1-diphenyl-2-picrylhydrazyl and 2,2'-azinobis(3-ethylbenzothiazoline-6-sulfonic acid) radical-scavenging activities, ferric-reducing antioxidant power and ferrous-chelating activity) of the hydrolysate were determined. Trypsin was purified 26.43-fold with a yield of 13.43%. The purified trypsin had a molecular weight (MW) of 23.5 kDa and optimal activity at pH 8.0 and 55 °C. It displayed high stability in the pH range of 6.0-11.0 and was thermally stable up to 50 °C. Both SBTI (0.05 mmol L(-1)) and N-p-tosyl-L-lysine-chloromethylketone (5 mmol L(-1)) completely inhibited trypsin activity. Antioxidative activities of the hydrolysate from Indian mackerel protein isolate increased with increasing DH up to 40% (P unicorn leatherjacket pyloric caeca was identified as trypsin based on its ability to hydrolyze a specific synthetic substrate and the response to specific trypsin inhibitors. The purified trypsin could hydrolyze Indian mackerel protein isolate, and the resulting hydrolysate exhibited antioxidative activity depending on its DH. © 2015 Society of Chemical Industry.

  13. Label-free electrical determination of trypsin activity by a silicon-on-insulator based thin film resistor.

    Science.gov (United States)

    Neff, Petra A; Serr, Andreas; Wunderlich, Bernhard K; Bausch, Andreas R

    2007-10-08

    A silicon-on-insulator (SOI) based thin film resistor is employed for the label-free determination of enzymatic activity. We demonstrate that enzymes, which cleave biological polyelectrolyte substrates, can be detected by the sensor. As an application, we consider the serine endopeptidase trypsin, which cleaves poly-L-lysine (PLL). We show that PLL adsorbs quasi-irreversibly to the sensor and is digested by trypsin directly at the sensor surface. The created PLL fragments are released into the bulk solution due to kinetic reasons. This results in a measurable change of the surface potential allowing for the determination of trypsin concentrations down to 50 ng mL(-1). Chymotrypsin is a similar endopeptidase with a different specificity, which cleaves PLL with a lower efficiency as compared to trypsin. The activity of trypsin is analyzed quantitatively employing a kinetic model for enzyme-catalyzed surface reactions. Moreover, we have demonstrated the specific inactivation of trypsin by a serine protease inhibitor, which covalently binds to the active site of the enzyme.

  14. Spleen: development and functional evaluation

    International Nuclear Information System (INIS)

    Sty, J.R.; Conway, J.J.

    1985-01-01

    Despite the fact that the spleen has multiple functions, only one has been widely used for evaluation of the organ by imaging techniques (phagocytosis of /sup 99m/Tc sulfur colloid). The usual splenic uptake of this radiocolloid can by used to determine the size, location, and integrity of the organ. A major use of splenic radiocolloid imaging has been in the study of congenital defects. Thus, eventration of the diaphragm, accessory spleens, splenogonadal fusion, the asplenia and polysplenia syndromes, and the wandering spleen are amenable to study by means of intravenously administered radiocolloid. Interference with the splenic uptake of radiocolloid can be either focal or generalized (as in functional asplenia). Imaging of the spleen has a major role in evaluating suspected trauma of the organ and in following its clinical course. The return of splenic function after splenectomy (splenosis or accessory spleens) can be documented by radionuclide imaging, and likely by hematologic techniques when the volume of tissue is sufficiently large. The detection of intrasplenic lesions is important in tumor staging and as an alerting sign to an ongoing process. 96 references

  15. Trypsin from viscera of vermiculated sailfin catfish, Pterygoplichthys disjunctivus, Weber, 1991: its purification and characterization.

    Science.gov (United States)

    Villalba-Villalba, Ana Gloria; Ramírez-Suárez, Juan Carlos; Valenzuela-Soto, Elisa Miriam; Sánchez, Guillermina García; Ruiz, Gisela Carvallo; Pacheco-Aguilar, Ramón

    2013-11-15

    Pterygoplichthys disjunctivus viscera trypsin was purified by fractionation with ammonium sulphate, gel filtration, affinity and ion exchange chromatography (DEAE-Sepharose). Trypsin molecular weight was approximately 27.5kDa according to SDS-PAGE, shown a single band in zymography. It exhibited maximal activity at pH 9.5 and 40°C, using N-benzoyl-dl-arginine-p-nitroanilide (BAPNA) as substrate. Enzyme was effectively inhibited by phenyl methyl sulphonyl fluoride (PMSF) (100%), N-α-p-tosyl-l-lysine chloromethyl ketone (TLCK) (85.4%), benzamidine (80.2%), and soybean trypsin inhibitor (75.6%) and partially inhibited by N-tosyl-l-phenylalanine chloromethyl ketone (TPCK) (10.3%), ethylendiaminetetraacetic acid (EDTA) (8.7%) and pepstatin A (1.2%). Enzyme activity was slightly affected by metal ions (Fe(2+)>Hg(2+)>Mn(2+)>K(+)>Mg(2+)>Li(+)>Cu(2+)). Trypsin activity decreased continuously as NaCl concentration increased (0-30%). Km and kcat values were 0.13mM and 1.46s(-1), respectively. Results suggest the enzyme have a potential application where room processing temperatures (25-35°C) or high salt (30%) concentration are needed, such as in fish sauce production. Copyright © 2013 Elsevier Ltd. All rights reserved.

  16. Toxic effects of Ricinus communis non proteic trypsin inhibitor for ...

    African Journals Online (AJOL)

    Marcelo Haro

    2015-10-21

    Oct 21, 2015 ... different growth stages, when the larvae burrow into the plant whorl causing ... or exposed to plant hormones (Rakwal et al., 2001). Plant proteinase ..... treatment, showing an evident deficit in larvae develop- ment. Pupae ...

  17. Optical studies on the purified eggplant trypsin inhibitor (Agricultural Chemistry)

    OpenAIRE

    金森, 正雄; 浅尾, 俊夫; 小垂, 真; 桂田, 昭彦; 伊吹, 文男; MASAO, KANAMORI; TOSHIO, ASAO; MAKOTO, KOTARU; AKIHIKO, KATSURADA; FUMIO, IBUKI; Laboratory of Nutritional and Food Chemistry, Department of Agricultural Chemistry, Faculty of Agriculture, Kyoto Prefectural University; Department of Food Science, Faculty of Home Economics, Mukogawa Women University; Present address, Laboratory of Food Chemistry, Faculty of Home Economics, Koka Women Junior College; Laboratory of Nutritional and Food Chemistry, Department of Agricultural Chemistry, Faculty of Agriculture, Kyoto Prefectural University; Laboratory of Nutritional and Food Chemistry, Department of Agricultural Chemistry, Faculty of Agriculture, Kyoto Prefectural University

    1980-01-01

    ナス果皮より均一に精製したトリプシン・インヒビターについてその光学的性質を検討した。各種pHにおいて, 紫外吸収スペクトルを測定したがpH 3.1から9.4の範囲では, チロシンおよびフェニールアラニン残基による同じスペルトルを示したが, pH 10以上ではチロシン残基のイオン化による著しい変化が認められた。またアミノ酸分析で確認されているようにトリプトフアンを含まないことは, 紫外吸収スペルトルからも明らかであった。275nmで励起した蛍光スペクトルが測定されたが, pH 3.1から9.4の範囲では蛍光強度は, ほゞ一定であり10以上で減少した。またCDスペクトルの測定結果はこのインヒビターが, ほとんどヘリックス構造やβ-構造を含まないことを示している。...

  18. Comparative analysis of novel autoantibody isotypes against citrullinated-inter-alpha-trypsin inhibitor heavy chain 3 (ITIH3)(542-556) peptide in serum from Taiwanese females with rheumatoid arthritis, primary Sjögren's syndrome and secondary Sjögren's syndrome in rheumatoid arthritis.

    Science.gov (United States)

    Liao, Chen-Chung; Chou, Pei-Lun; Cheng, Chao-Wen; Chang, Yu-Sheng; Chi, Wei-Ming; Tsai, Kai-Leun; Chen, Wei-Jung; Kung, Ting-Shuan; Tai, Chih-Chun; Lee, Kuan-Wei; Chen, You-Chia; Lin, Ching-Yu

    2016-06-01

    The purpose of this study was to discover and validate inter-alpha-trypsin inhibitor heavy chain H3 (ITIH3) as novel biomarkers, and evaluate autoantibody isotypes against an unmodified and citrullinated ITIH3(542-556) peptide among Taiwanese female patients with rheumatoid arthritis (RA), primary Sjögren's syndrome (pSS), secondary Sjögren's syndrome in rheumatoid arthritis (RA-sSS), and healthy controls (HCs). We used concanavalin A (Con A) affinity chromatography, 1-D SDS-PAGE, and label-free nano-LC-MS/MS to screen biomarker candidates (serum-derived Con A-captured proteins) and then identify PTMs of validated biomarkers (serum proteins) using pooled serum from 7 RA-sSS female patients and 7 age-matched HCs (the discovery set). Furthermore, the protein level and autoantibody isotype analyses were further validated against individual serum from 18 HCs, 18 RA, 18 pSS, and 18 RA-sSS patients (the validation set). Con A-bound ITIH3 was identified and validated as the only differentially expressed protein, which was elevated. Additionally, 2 novel PTMs in ITIH3 were identified and included citrullination at arginine-(546) and arginine-(556), and hexosamine at tryptophan-(558). Further, concentrations of anti-citrullinatd-ITIH3(542-556) peptide autoantibodies significantly increased in patients with RA, pSS, and RA-sSS compared to HCs. Especially, autoantibody IgM against the citrullinated-ITIH3(542-556) peptide showed better diagnostic performance in discriminating both RA versus pSS and pSS versus RA-sSS. By using comparative proteomic analysis of serum samples, the current study discovered and validates differentially expressed Con A-bound ITIH3 as a potential biomarker for secondary Sjögren's syndrome (SS) in rheumatoid arthritis (RA) patients and healthy controls (HCs). Besides, hexosamine and citrullination on ITIH3 were further identified. Through analyzing autoantibody isotypes against the citrullinated ITIH3 peptide, patients with RA, primary SS, and RA

  19. Cold adaptation, ca2+ dependency and autolytic stability are related features in a highly active cold-adapted trypsin resistant to autoproteolysis engineered for biotechnological applications.

    Directory of Open Access Journals (Sweden)

    Alvaro Olivera-Nappa

    Full Text Available Pig trypsin is routinely used as a biotechnological tool, due to its high specificity and ability to be stored as an inactive stable zymogen. However, it is not an optimum enzyme for conditions found in wound debriding for medical uses and trypsinization processes for protein analysis and animal cell culturing, where low Ca(2+ dependency, high activity in mild conditions and easy inactivation are crucial. We isolated and thermodynamically characterized a highly active cold-adapted trypsin for medical and laboratory use that is four times more active than pig trypsin at 10(° C and at least 50% more active than pig trypsin up to 50(° C. Contrary to pig trypsin, this enzyme has a broad optimum pH between 7 and 10 and is very insensitive to Ca(2+ concentration. The enzyme is only distantly related to previously described cryophilic trypsins. We built and studied molecular structure models of this trypsin and performed molecular dynamic calculations. Key residues and structures associated with calcium dependency and cryophilicity were identified. Experiments indicated that the protein is unstable and susceptible to autoproteolysis. Correlating experimental results and structural predictions, we designed mutations to improve the resistance to autoproteolysis and conserve activity for longer periods after activation. One single mutation provided around 25 times more proteolytic stability. Due to its cryophilic nature, this trypsin is easily inactivated by mild denaturation conditions, which is ideal for controlled proteolysis processes without requiring inhibitors or dilution. We clearly show that cold adaptation, Ca(2+ dependency and autolytic stability in trypsins are related phenomena that are linked to shared structural features and evolve in a concerted fashion. Hence, both structurally and evolutionarily they cannot be interpreted and studied separately as previously done.

  20. Fish trypsins: potential applications in biomedicine and prospects for production.

    Science.gov (United States)

    Jesús-de la Cruz, Kristal; Álvarez-González, Carlos Alfonso; Peña, Emyr; Morales-Contreras, José Antonio; Ávila-Fernández, Ángela

    2018-04-01

    In fishes, trypsins are adapted to different environmental conditions, and the biochemical and kinetic properties of a broad variety of native isoforms have been studied. Proteolytic enzymes remain in high demand in the detergent, food, and feed industries; however, our analysis of the literature showed that, in the last decade, some fish trypsins have been studied for the synthesis of industrial peptides and for specific biomedical uses as antipathogenic agents against viruses and bacteria, which have been recently patented. In addition, innovative strategies of trypsin administration have been studied to ensure that trypsins retain their properties until they exert their action. Biomedical uses require the production of high-quality enzymes. In this context, the production of recombinant trypsins is an alternative. For this purpose, E. coli -based systems have been tested for the production of fish trypsins; however, P. pastoris -based systems also seem to show great potential in the production of fish trypsins with higher production quality. On the other hand, there is a lack of information regarding the specific structures, biochemical and kinetic properties, and characteristics of trypsins produced using heterologous systems. This review describes the potential uses of fish trypsins in biomedicine and the enzymatic and structural properties of native and recombinant fish trypsins obtained to date, outlining some prospects for their study.

  1. Hydatid disease of the spleen

    International Nuclear Information System (INIS)

    Sinner, W.N. von; Stridbeck, H.

    1992-01-01

    Seven patients with hydatid disease of the spleen were examined by radiography, ultrasound, CT, and in one case MR imaging. The observations were confirmed by patho-anatomic findings except in 2 patients where high indirect hemagglutination tests confirmed the diagnosis. (orig./MG)

  2. Torsion of a wandering spleen

    African Journals Online (AJOL)

    No improvement was noted on detorsion of the vascular pedicle, and a splenectomy was performed. The spleen measured 120×90×55 mm and weighed 250 g. Histological examination of the organ identified significant haemorrhagic congestion associated with diffuse haemorrhagic necrosis, with no neoplasm or infiltrate.

  3. Performance comparison of three trypsin columns used in liquid chromatography?

    OpenAIRE

    ?lechtov?, Tereza; Gilar, Martin; Kal?kov?, Kv?ta; Moore, Stephanie M.; Jorgenson, James W.; Tesa?ov?, Eva

    2017-01-01

    Trypsin is the most widely used enzyme in proteomic research due to its high specificity. Although the in-solution digestion is predominantly used, it has several drawbacks, such as long digestion times, autolysis, and intolerance to high temperatures or organic solvents. To overcome these shortcomings trypsin was covalently immobilized on solid support and tested for its proteolytic activity. Trypsin was immobilized on bridge-ethyl hybrid silica sorbent with 300 ? pores, packed in 2.1 ? 30 m...

  4. Inactivation of proteinaceous protease inhibitors of soybeans by isolated fungi

    NARCIS (Netherlands)

    Meijer, M.M.T.; Spekking, W.T.J.; Sijtsma, L.; Bont, de J.A.M.

    1995-01-01

    Proteinaceous protease inhibitors, Kunitz Soybean Trypsin Inhibitor (KSTI) and Bowman Birk Inhibitor (BBI), in legume seeds reduce the digestibility of proteins in feed of monogastric animals. Enzymatic inactivation of these inhibitors will increase the nutritional value of the feed. The aim of this

  5. The trypsin-catalyzed hydrolysis of monomolecular films of lysylphosphatidylglycerol

    NARCIS (Netherlands)

    Gould, R.M.; Dawson, R.M.C.

    1972-01-01

    The hydrolysis by trypsin of the bacterial phospholipid, lysylphosphatidyl-glycerol has been studied at the air-water interface. High specific activity [14C]-lysylphosphatidylglycerol was prepared biosynthetically and the trypsin action followed by measuring the loss of surface radioactivity from a

  6. Torsion of wandering spleen and distal pancreas

    International Nuclear Information System (INIS)

    Sheflin, J.R.; Lee, C.M.; Kretchmar, K.A.

    1984-01-01

    Wandering spleen is the term applied to the condition in which a long pedicle allows the spleen to lie in an abnormal location. Torsion of a wandering spleen is an unusual cause of an acute abdomen and is rarely diagnosed preoperatively. Associated torsion of the distal pancreas is even more uncommon. The authors describe a patient with torsion of a wandering spleen and distal pancreas, who was correctly diagnosed, and define the merits of the imaging methods used. The initial examination should be 99 /sup m/Tc-sulfur colloid liner-spleen scanning

  7. Serum trypsin inhibitory capacity in hemodialysis patients

    International Nuclear Information System (INIS)

    Hashemi, Mohammad; Mehrabifar, Hamid; Homayooni, Fatemeh; Naderi, Mohammad; Montazerifar, Farzaneh; Ghavami, Saeid

    2009-01-01

    It has been established that overproduction of reactive oxygen species (ROS) occurs during hemodialysis causing oxidation of proteins. Alpha-1-antitrypsin is the major circulating anti-protease which contains methionine in the active site. The aim of the present study was to measure the level of serum trypsin inhibitory capacity (sTIC) in hemodialysis patients. This case-control study was performed in 52 hemodialysis patients and 49 healthy controls. sTIC was measured by enzymatic assay. The sTIC was significantly (P< 0.001) lower in hemodialysis patients (1.87 + - 0.67 micron mol/min/mL) than healthy controls (2.83 + - 0.44 micron mol/min/L). Reduction of sTIC may be due to the oxidation of methionine residue in the reactive site of alpha-1 antitrypsin. (author)

  8. [Ultrasound of spleen and retroperitoneum].

    Science.gov (United States)

    Salcedo Joven, I; Segura-Grau, A; Díaz Rodríguez, N; Segura-Cabral, J M

    2016-09-01

    Ultrasound provides data of extremely great value when studying spleen pathology, being diagnostic in splenomegaly and splenic trauma, as well as offering a good approach to the diagnosis of both benign and malignant focal pathology, particularly lymphoma. However, for the evaluation of adrenal and retroperitoneal diseases, other techniques such as CT or MRI are more suitable, even though ultrasound is still an excellent screening and monitoring method, as well as being useful in non-invasive therapeutic approaches. Copyright © 2014 Sociedad Española de Médicos de Atención Primaria (SEMERGEN). Publicado por Elsevier España, S.L.U. All rights reserved.

  9. Why Ser and not Thr brokers catalysis in the trypsin fold.

    Science.gov (United States)

    Pelc, Leslie A; Chen, Zhiwei; Gohara, David W; Vogt, Austin D; Pozzi, Nicola; Di Cera, Enrico

    2015-02-24

    Although Thr is equally represented as Ser in the human genome and as a nucleophile is as good as Ser, it is never found in the active site of the large family of trypsin-like proteases that utilize the Asp/His/Ser triad. The molecular basis of the preference of Ser over Thr in the trypsin fold was investigated with X-ray structures of the thrombin mutant S195T free and bound to an irreversible active site inhibitor. In the free form, the methyl group of T195 is oriented toward the incoming substrate in a conformation seemingly incompatible with productive binding. In the bound form, the side chain of T195 is reoriented for efficient substrate acylation without causing steric clash within the active site. Rapid kinetics prove that this change is due to selection of an active conformation from a preexisting ensemble of reactive and unreactive rotamers whose relative distribution determines the level of activity of the protease. Consistent with these observations, the S195T substitution is associated with a weak yet finite activity that allows identification of an unanticipated important role for S195 as the end point of allosteric transduction in the trypsin fold. The S195T mutation abrogates the Na(+)-dependent enhancement of catalytic activity in thrombin, activated protein C, and factor Xa and significantly weakens the physiologically important allosteric effects of thrombomodulin on thrombin and of cofactor Va on factor Xa. The evolutionary selection of Ser over Thr in trypsin-like proteases was therefore driven by the need for high catalytic activity and efficient allosteric regulation.

  10. Effect of synthetic adjuvants of biological activity of spleen proteins

    International Nuclear Information System (INIS)

    Kartasheva, A.L.; Yuferova, N.V.; Drozhennikov, V.A.; Orlova, E.B.; Perevezentseva, O.S.; Filatov, P.P.

    1981-01-01

    Intraperitoneal administration to mice of synthetic adjuvants of a polyanion type increases the spleen mass by 500% and rises the content of proteins with activity of inhibitor of DNAase 1. A protein fraction isolated from the spleen of treated animals administered to exposed (7.7 Gy) mice alone or in a combination with exogenous DNA increases survival up to 61.1 and 80.5%, respectively, as opposed to 36.6% in the case of administration of proteins from intact animals, or 8.3% in the control (no treatment). The protein fraction from treated animals administered to mice exposed to 5.1-5.5 Gy accelerates the recovery of hemopoesis and immune response better than proteins of intact animals

  11. MORPHOLOGICAL VARIATIONS OF SPLEEN: A CADAVERIC STUDY

    Directory of Open Access Journals (Sweden)

    Siva Chidambaram

    2015-07-01

    Full Text Available The Spleen is a large lymphoid organ situated in the left hypochondrial region having an important role in immunological and hematological functions of the human body. The aim of this study was to find the morphological variations of the spleen with respect to it’s a Shape, b Number of notches on its borders and c Presence of anomalous fissure on its surface. The Study was done on 60 formalin fixed cadaveric spleen from the Department of Anatomy, Narayana Medical College, Nellore, Andhra Pradesh. Out of 60 spleens we examined, the various shapes of the spleen were noted suc h as wedge shape (73.33%, triangular (13.33%, tetrahedral (6.67% and oval shape(6.67%.The number of spleen showing notches on its superior border was 38(63.33% and in inferior border it was 6(10%. Absence of splenic notch was observed in 10(16.67% s pleens and the remaining 6 spleens (10% shows notches on its both the borders. The anomalous splenic fissure was found in 4(6.67% spleens on its diaphragmatic surface. The knowledge of variations in the morphology of spleen are essential for physician, s urgeon, radiologist and forensic surgeon to differentiate it from the splenic pathology and splenic injury. In addition to this, it is also important for anatomist during routine classroom dissection and discussion.

  12. Immunoreactive trypsin and neonatalscreening for cystic fibrosis

    International Nuclear Information System (INIS)

    Travert, G.; Laroche, D.; Blandin, C.; Pasquet, C.

    1988-01-01

    Immunoreactive trypsin (IRT) was measured in dried blood spots from 160.822 five-day-old babies as a part of a regionwide neonatal screening program for cystic fibrosis. A second test was performed for 492 babies in whom blood IRT levels were found greater than 900 μg/l; retesting revealed persistent elevation in 55. Sweat testing confirmed cystic fibrosis in 43 babies, but results were normal in 12. During the course of this study, a total of 51 cystic fibrosis babies were identified: 43 by newborn screening, 6 because they had meconium ileus; so, early diagnosis was achieved in 49 cases out of 51. Two newborn babies did not have elevated IRT and they were missed by the screening test. Our results confirm that elevated blood IRT is characteristic of newborn babies with cystic fibrosis and show that this test has an excellent specificity (99.7%) and a good sensitivity (95%) when used as a neonatal screening test [fr

  13. Chemically modified, immobilized trypsin reactor with improved digestion efficiency

    NARCIS (Netherlands)

    Freije, J.R.; Mulder, P.P.; Werkman, W.; Rieux, L.; Niederlander, H.A G; Verpoorte, Sabeth; Bischoff, Rainer

    2005-01-01

    Tryptic digestion followed by identification using mass spectrometry is an important step in many proteomic studies. Here, we describe the preparation of immobilized, acetylated trypsin for enhanced digestion efficacy in integrated protein analysis platforms. Complete digestion of cytochrome c was

  14. Iron oxides in human spleen.

    Science.gov (United States)

    Kopáni, Martin; Miglierini, Marcel; Lančok, Adriana; Dekan, Július; Čaplovicová, Mária; Jakubovský, Ján; Boča, Roman; Mrazova, Hedviga

    2015-10-01

    Iron is an essential element for fundamental cell functions and a catalyst for chemical reactions. Three samples extracted from the human spleen were investigated by scanning (SEM) and transmission electron microscopy (TEM), Mössbauer spectrometry (MS), and SQUID magnetometry. The sample with diagnosis of hemosiderosis (H) differs from that referring to hereditary spherocytosis and the reference sample. SEM reveals iron-rich micrometer-sized aggregate of various structures-tiny fibrils in hereditary spherocytosis sample and no fibrils in hemochromatosis. Hematite and magnetite particles from 2 to 6 μm in TEM with diffraction in all samples were shown. The SQUID magnetometry shows different amount of diamagnetic, paramagnetic and ferrimagnetic structures in the tissues. The MS results indicate contribution of ferromagnetically split sextets for all investigated samples. Their occurrence indicates that at least part of the sample is magnetically ordered below the critical temperature. The iron accumulation process is different in hereditary spherocytosis and hemosiderosis. This fact may be the reason of different iron crystallization.

  15. Spleen-lung interface as diagnostic information

    International Nuclear Information System (INIS)

    DeLuca, S.A.; Kolodny, G.M.

    1975-01-01

    Left anterior, lateral, and posterior views on 50 consecutive /sup 99m/Tc-sulfur colloid lung scans were examined. Normal patients had continuity of activity between the left lung and the spleen on all three views. Patients with subphrenic abscess or large left pleural effusions showed no continuity between lung and spleen activity on any view, while other abnormalities, most commonly cardiomegaly, accounted for lack of lung-spleen continuity on the anterior view only. It is suggested that in all combined /sup 99m/Tc-sulfur colloid lung studies, the left side be examined as well as the right for abnormalities adjacent to the left diaphragm. (auth)

  16. [Syk inhibitors].

    Science.gov (United States)

    Kimura, Yukihiro; Chihara, Kazuyasu; Takeuchi, Kenji; Sada, Kiyonao

    2013-07-01

    Non-receptor type of protein-tyrosine kinase Syk (spleen tyrosine kinase) was isolated in the University of Fukui in 1991. Syk is known to be essential for the various physiological functions, especially in hematopoietic lineage cells. Moreover, ectopic expression of Syk by epigenetic changes is reported to cause retinoblastoma. Recently, novel Syk inhibitors were developed and its usefulness has been evaluated in the treatment of allergic rhinitis, rheumatoid arthritis, and idiopathic thrombocytopenic purpura. In this review, we will summarize the history, structure, and function of Syk, and then describe the novel Syk inhibitors and their current status. Furthermore, we will introduce our findings of the adaptor protein 3BP2 (c-Abl SH3 domain-binding protein-2), as a novel target of Syk.

  17. Changing spleen size after blunt abdominal trauma

    International Nuclear Information System (INIS)

    Goodman, L.R.; Aprahamian, C.

    1989-01-01

    The authors studied the incidence and significance of splenic enlargement on serial CT after abdominal trauma. Spleen size and density in 44 trauma patients were studied with serial, contrast-enhanced Ct. In 58% of the patients, ≥ 10% enlargement of the spleen was seen on follow-up scans. Ten patients had >50% enlargement. In several, the initial density of the spleen was less than that of the liver. Spleen density returned to normal on subsequent scans. Correlations between splenic changes and clinical parameters (such as blood replacement, hypotension, and various trauma indexes) were weak. The author's study indicated that serial splenic enlargement was a physiologic return to normal after major trauma, not a pathologic condition requiring splenectomy

  18. Inhibition of trypsin by condensed tannins and wine.

    Science.gov (United States)

    Gonçalves, Rui; Soares, Susana; Mateus, Nuno; de Freitas, Victor

    2007-09-05

    Phenolic compounds are abundant vegetable secondary metabolites in the human diet. The ability of procyanidin oligomers and wine polyphenols to inhibit trypsin activity was studied using a versatile and reliable in vitro method. The hydrolysis of the chromogenic substrate N-benzoyl-d,l-arginine-p-nitroanilide (BApNA) by trypsin was followed by spectrophotometry in the presence and absence of condensed tannins and wine. A clear relationship between the degree of polymerization of procyanidins and enzymatic inhibition was observed. Trypsin activity inhibition was also detected in several types of wine. In general, the inhibition increased with the concentration of phenolic compounds in wines. These results may be relevant when considering these compounds as antinutritional factors, thereby contributing to a reduced absorption of nutrients.

  19. Thermodynamic evaluation and modeling of proton and water exchange associated with benzamidine and berenil binding to ß-trypsin

    Directory of Open Access Journals (Sweden)

    M.T. Pereira

    2005-11-01

    Full Text Available Serine-proteases are involved in vital processes in virtually all species. They are important targets for researchers studying the relationships between protein structure and activity, for the rational design of new pharmaceuticals. Trypsin was used as a model to assess a possible differential contribution of hydration water to the binding of two synthetic inhibitors. Thermodynamic parameters for the association of bovine ß-trypsin (homogeneous material, observed 23,294.4 ± 0.2 Da, theoretical 23,292.5 Da with the inhibitors benzamidine and berenil at pH 8.0, 25ºC and with 25 mM CaCl2, were determined using isothermal titration calorimetry and the osmotic stress method. The association constant for berenil was about 12 times higher compared to the one for benzamidine (binding constants are K = 596,599 ± 25,057 and 49,513 ± 2,732 M-1, respectively; the number of binding sites is the same for both ligands, N = 0.99 ± 0.05. Apparently the driving force responsible for this large difference of affinity is not due to hydrophobic interactions because the variation in heat capacity (DCp, a characteristic signature of these interactions, was similar in both systems tested (-464.7 ± 23.9 and -477.1 ± 86.8 J K-1 mol-1 for berenil and benzamidine, respectively. The results also indicated that the enzyme has a net gain of about 21 water molecules regardless of the inhibitor tested. It was shown that the difference in affinity could be due to a larger number of interactions between berenil and the enzyme based on computational modeling. The data support the view that pharmaceuticals derived from benzamidine that enable hydrogen bond formation outside the catalytic binding pocket of ß-trypsin may result in more effective inhibitors.

  20. Purification and characterization of a novel trypsin-like protease from green-seeded chickpea (Cicer arientum).

    Science.gov (United States)

    Shamsi, Tooba Naz; Parveen, Romana; Sen, Priyankar; Fatima, Sadaf

    2017-05-28

    The present study describes the purification and physicochemical and biochemical characterization of trypsin-like protease from green-seeded chickpea (Cicer arientum). The crude extract of chickpea trypsin (CpT) was obtained by homogenization followed by differential ammonium sulfate precipitation. The CpT was purified by ion-exchange chromatography on diethylaminoethyl (DEAE) column, pre-equilibrated with 20 mM tris-CaCl 2 buffer (pH 8.2) with a flow rate of 0.5 mL min -1 . The molecular weight and purity of ∼23 kDa of CpT were determined by sodium dodecyl sulfate polyacrylamide gel electrophoresis. Activity of protease was determined using Nα-benzoyl-DL-arginine-p-nitroanilide as chromogenic substrate and CpT purified showed a specific inhibitor activity of 26978.7697 U mg -1 , fold purity of 9.8, and the yield of 70.2%. The characterization was performed for thermal stability, pH profile, and effect of various inhibitors on enzymatic activity. The protein isolated showed stability in the neutral to mild alkaline pH range and thermostability up to 50°C. CpT confirmed its serine nature as it was appreciably inhibited by serine protease inhibitors (maximum 6%), whereas metalloprotease inhibitors barely affected the activity of the enzyme (85%). To the best of our knowledge, it is first reported on purification of protease with trypsin-like properties, from this source.

  1. Orthosteric and Allosteric Regulation in Trypsin-Like Peptidases

    DEFF Research Database (Denmark)

    Kromann-Tofting, Tobias

    Trypsin-like serine peptidases play an important role in many physiological and pathophysiological processes, the latter including cardiovascular diseases and cancer. Binding of natural ligands to functional sites on the peptidase surface balances the level of activity and substrate specificity......-ray crystallography to determine crystal structures of active and inactive conformations of muPA, combined with biochemical analysis, elucidated an allosteric regulatory mechanism, which is now believed to be highly conserved in the trypsin-like serine peptidases. Targeting zymogen activation represents an attractive...

  2. Potential toxicity of phthalic acid esters plasticizer: interaction of dimethyl phthalate with trypsin in vitro.

    Science.gov (United States)

    Wang, Yaping; Zhang, Guowen; Wang, Langhong

    2015-01-14

    Dimethyl phthalate (DMP) is widely used as a plasticizer in industrial processes and has been reported to possess potential toxicity to the human body. In this study, the interaction between DMP and trypsin in vitro was investigated. The results of fluorescence, UV–vis, circular dichroism, and Fourier transform infrared spectra along with cyclic voltammetric measurements indicated that the remarkable fluorescence quenching and conformational changes of trypsin resulted from the formation of a DMP–trypsin complex, which was driven mainly by hydrophobic interactions. The molecular docking and trypsin activity assay showed that DMP primarily interacted with the catalytic triad of trypsin and led to the inhibition of trypsin activity. The dimensions of the individual trypsin molecules were found to become larger after binding with DMP by atomic force microscopy imaging. This study offers a comprehensive picture of DMP–trypsin interaction, which is expected to provide insights into the toxicological effect of DMP.

  3. Influence of surface-imprinted nanoparticles on trypsin activity.

    Science.gov (United States)

    Guerreiro, António; Poma, Alessandro; Karim, Kal; Moczko, Ewa; Takarada, Jessica; de Vargas-Sansalvador, Isabel Perez; Turner, Nicholas; Piletska, Elena; de Magalhães, Cristiana Schmidt; Glazova, Natalia; Serkova, Anastasia; Omelianova, Aleksandra; Piletsky, Sergey

    2014-09-01

    Here, the modulation of enzyme activity is presented by protein-imprinted nanoparticles produced using a solid-phase approach. Using trypsin as target, binding of the nanoparticles to the enzyme results in its inhibition or in stabilization, depending on the orientation of the immobilized enzyme used during imprinting. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  4. Immobilization of trypsin on sub-micron skeletal polymer monolith

    Energy Technology Data Exchange (ETDEWEB)

    Yao Chunhe [Beijing National Laboratory for Molecular Sciences, Key Laboratory of Analytical Chemistry for Living Biosystems, Institute of Chemistry, Chinese Academy of Sciences, Beijing 100190 (China); Graduate School, Chinese Academy of Sciences, Beijing 100049 (China); Qi Li, E-mail: qili@iccas.ac.cn [Beijing National Laboratory for Molecular Sciences, Key Laboratory of Analytical Chemistry for Living Biosystems, Institute of Chemistry, Chinese Academy of Sciences, Beijing 100190 (China); Hu Wenbin [Beijing National Laboratory for Molecular Sciences, Key Laboratory of Analytical Chemistry for Living Biosystems, Institute of Chemistry, Chinese Academy of Sciences, Beijing 100190 (China); Graduate School, Chinese Academy of Sciences, Beijing 100049 (China); Wang Fuyi [Beijing National Laboratory for Molecular Sciences, Key Laboratory of Analytical Chemistry for Living Biosystems, Institute of Chemistry, Chinese Academy of Sciences, Beijing 100190 (China); Yang Gengliang [College of Pharmacy, Hebei University, Baoding 071002 (China)

    2011-04-29

    A new kind of immobilized trypsin reactor based on sub-micron skeletal polymer monolith has been developed. Covalent immobilization of trypsin on this support was performed using the epoxide functional groups in either a one- or a multi-step reaction. The proteolytic activity of the immobilized trypsin was measured by monitoring the formation of N-{alpha}-benzoyl-L-arginine (BA) which is the digestion product of a substrate N-{alpha}-benzoyl-L-arginine ethyl ester (BAEE). Results showed that the digestion speed was about 300 times faster than that performed in free solution. The performance of such an enzyme reactor was further demonstrated by digesting protein myoglobin. It has been found that the protein digestion could be achieved in 88 s at 30 deg. C, which is comparable to 24 h digestion in solution at 37 {sup o}C. Furthermore, the immobilized trypsin exhibits increased stability even after continuous use compared to that in free solution. The present monolithic enzyme-reactor provides a promising platform for the proteomic research.

  5. A prospective randomized trial of Kotase ® (Bromelain + Trypsin) in ...

    African Journals Online (AJOL)

    International Journal of Medicine and Health Development. Journal Home · ABOUT THIS ... A prospective randomized trial of Kotase® (Bromelain + Trypsin) in the management of post-operative abdominal wounds at the University of Nigeria Teaching Hospital Enugu, Nigeria. Emmanuel R Ezeome, Aloy E Aghaji ...

  6. Solid solitary hamartoma of the spleen

    Directory of Open Access Journals (Sweden)

    Grubor Nikica

    2013-01-01

    Full Text Available Introduction. Hamartoma of the spleen is a rare, sometimes asymptomatic similar to hemangioma benign tumor of the spleen, which, owing to the new diagnostic imaging methods, is discovered with increasing frequency. It appears as solitary or multiple tumorous lesions. Case Outline. We present a 48-year-old woman in whom, during the investigation for Helicobacter pylori gastric infection and rectal bleeding, with ultrasonography, a mass 6.5×6.5 cm in diameter was discovered by chance within the spleen. Splenectomy was performed due to suspected lymphoma of the spleen. On histology, tumor showed to be of mixed cellular structure, with areas without white pulp, at places with marked dilatation of sinusoids and capillaries to the formation of „blood lakes“ between which broad hypercellular Billroth’s zones were present. Extramedullary hematopoiesis was found focally. The cells that covered vascular spaces were CD34+ and CD31+ and CD8- and CD21-. Conclusion. Hamartoma has to be taken into consideration always when well circumscribed hypervascular tumor within the spleen is found, particularly in children. Although the diagnosis of hamartoma may be suspected preoperatively, the exact diagnosis is established based on histological and immunohystochemistry examinations. Treatment is most often splenectomy and rarely a partial splenectomy is possible, which is recommended particularly in children.

  7. Spleen-preserving distal pancreatectomy in trauma.

    Science.gov (United States)

    Schellenberg, Morgan; Inaba, Kenji; Cheng, Vincent; Bardes, James M; Lam, Lydia; Benjamin, Elizabeth; Matsushima, Kazuhide; Demetriades, Demetrios

    2018-01-01

    Traumatic injuries to the distal pancreas are infrequent. Universally accepted recommendations about the need for routine splenectomy with distal pancreatectomy do not exist. The aims of this study were to compare outcomes after distal pancreatectomy and splenectomy versus spleen-preserving distal pancreatectomy, and to define the appropriate patient population for splenic preservation. All patients who underwent distal pancreatectomy (January 1, 2007, to December 31, 2014) were identified from the National Trauma Data Bank. Patients with concomitant splenic injury and those who underwent partial splenectomy were excluded. Demographics, clinical data, procedures, and outcomes were collected. Study groups were defined by surgical procedure: distal pancreatectomy and splenectomy versus spleen-preserving distal pancreatectomy. Baseline characteristics between groups were compared with univariate analysis. Multivariate analysis was performed with logistic and linear regression to examine differences in outcomes. Over the 8-year study period, 2,223 patients underwent distal pancreatectomy. After excluding 1,381 patients with concomitant splenic injury (62%) and 8 (pancreatectomy and splenectomy, those who underwent spleen-preserving distal pancreatectomy were younger (p pancreatectomy (p = 0.017). Complications, mortality, and intensive care unit LOS were not significantly different. In young patients after blunt trauma who are not severely injured, a spleen-preserving distal pancreatectomy should be considered to allow for conservation of splenic function and a shorter hospital LOS. In all other patients, the surgeon should not hesitate to remove the spleen with the distal pancreas. Therapy, level IV.

  8. Novel contrast agent for liver and spleen

    International Nuclear Information System (INIS)

    Seltzer, S.E.; Blau, M.; Adams, D.F.; Janoff, A.; Minchey, S.

    1991-01-01

    This paper determines whether the biodistribution and imaging characteristics of a liposome-encapsulated contrast agent, iotrolan-carrying interdigitation fusion (IF) vesicles, were acceptable for a liver-spleen CT contrast agent. IF vesicles with iotrolan in their aqueous phase were prepared by fusing small unilamellar liposomes into larger vesicles. The iodine-to-lipid ratio was 4.7. Biodistribution was measured with I-125 iotrolan-labeled IF vesicles in rats. CT imaging (Somatom Plus, Siemens Medical Systems) was performed in dogs. At the lowest dose (10 mg of iodine and 2.1 mg of lipid per kilogram) 72% of the ID was in the liver, 5% in spleen, and 1% in lungs at 1 hour. At the highest dose, (1,000 mg of iodine and 212 mg of lipid per kilogram), liver values were 68% ID, while spleen rose to 18%, lung 5%. Liver and spleen values stayed at peak for 24 hours then fell; the half-life was 6 days. In dogs, liver and spleen enhancement at 1 hour averaged 652 and 256 HU above baseline per gram of iodine per kilogram, respectively

  9. 21 CFR 524.2620 - Liquid crystalline trypsin, Peru balsam, castor oil.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 6 2010-04-01 2010-04-01 false Liquid crystalline trypsin, Peru balsam, castor... NEW ANIMAL DRUGS § 524.2620 Liquid crystalline trypsin, Peru balsam, castor oil. (a)(1) Specifications... delivered to the wound site contains 0.12 milligram of crystalline trypsin, 87.0 milligrams of Peru balsam...

  10. Function of the replanted spleen in dogs

    International Nuclear Information System (INIS)

    Velcek, F.T.; Kugaczewski, J.T.; Jongco, B.; Shaftan, G.W.; Rao, P.S.; Schiffman, G.; Kottmeier, P.K.

    1982-01-01

    The function of replanted splenic fragments was studied by comparing three groups of five dogs each, one group with intact spleens; one, post-splenectomy; and one with splenic replantation. Fifteen fragments were implanted into the omentum. Howell-Jolly bodies appeared after splenectomy but cleared in the replanted group after several months. 125 I-tagged attenuated pneumococcal clearance studies showed a significant difference between control and replanted group compared with the splenectomized group. The increase of pneumococcal antibody titers after vaccination differed significantly between the splenectomized and the replanted group. All replanted fragments were viable and showed growth over a 2-year period. These studies demonstrate that omental replantation of the canine spleen leads to the maintenance of certain functional splenic parameters comparable to the normal spleen which are significantly different from the splenectomized animal

  11. Function of the replanted spleen in dogs

    Energy Technology Data Exchange (ETDEWEB)

    Velcek, F.T.; Kugaczewski, J.T.; Jongco, B.; Shaftan, G.W.; Rao, P.S.; Schiffman, G.; Kottmeier, P.K.

    1982-06-01

    The function of replanted splenic fragments was studied by comparing three groups of five dogs each, one group with intact spleens; one, post-splenectomy; and one with splenic replantation. Fifteen fragments were implanted into the omentum. Howell-Jolly bodies appeared after splenectomy but cleared in the replanted group after several months. /sup 125/I-tagged attenuated pneumococcal clearance studies showed a significant difference between control and replanted group compared with the splenectomized group. The increase of pneumococcal antibody titers after vaccination differed significantly between the splenectomized and the replanted group. All replanted fragments were viable and showed growth over a 2-year period. These studies demonstrate that omental replantation of the canine spleen leads to the maintenance of certain functional splenic parameters comparable to the normal spleen which are significantly different from the splenectomized animal.

  12. Blunt trauma to the spleen: ultrasonographic findings

    Energy Technology Data Exchange (ETDEWEB)

    Doody, O. [Department of Radiology, Tallaght Hospital, Dublin (Ireland); Lyburn, D. [Department of Radiology, Cheltenham General Hospital (United Kingdom); Geoghegan, T. [Department of Radiology, Tallaght Hospital, Dublin (Ireland); Govender, P. [Department of Radiology, Tallaght Hospital, Dublin (Ireland); Monk, P.M. [Department of Radiology, Vancouver Hospital (Canada); Torreggiani, W.C. [Department of Radiology, Tallaght Hospital, Dublin (Ireland)]. E-mail: william.torreggiani@amnch.ie

    2005-09-01

    The spleen is the most frequently injured organ in adults who sustain blunt abdominal trauma. Splenic trauma accounts for approximately 25% to 30% of all intra-abdominal injuries. The management of splenic injury has undergone rapid change over the last decade, with increasing emphasis on splenic salvage and non-operative management. Identifying the presence and degree of splenic injury is critical in triaging the management of patients. Imaging is integral in the identification of splenic injuries, both at the time of injury and during follow-up. Although CT remains the gold standard in blunt abdominal trauma, US continues to play an important role in assessing the traumatized spleen. This pictorial review illustrates the various ultrasonographic appearances of the traumatized spleen. Correlation with other imaging is presented and complications that occur during follow-up are described.

  13. Laparoscopic Splenectomy in Patients With Spleen Injuries.

    Science.gov (United States)

    Ermolov, Aleksander S; Tlibekova, Margarita A; Yartsev, Peter A; Guliaev, Andrey A; Rogal, Mikhail M; Samsonov, Vladimir T; Levitsky, Vladislav D; Chernysh, Oleg A

    2015-12-01

    Spleen injury appears in 10% to 30% of abdominal trauma patients. Mortality among the patients in the last 20 years remains high (6% to 7%) and shows no tendency to decline. Nowadays nonoperative management is widely accepted management of patients with low-grade spleen injury, whereas management of patients with high-grade spleen injury (III and higher) is not so obvious. There are 3 methods exist in treatment of such patients: conservative (with or without angioembolization), spleen-preserving operations, and splenectomy. Today laparoscopic splenectomy is not a widely used operation and only few studies reported about successful use of laparoscopic splenectomy in patients with spleen injury.The aim of the study was to determine indications and contraindications for laparoscopic splenectomy in abdominal trauma patients and to analyze results of the operations. The study involved 42 patients with spleen injury grade III who were admitted in our institute in the years of 2010 to 2014. The patients were divided in 2 groups. Laparoscopic splenectomy was performed in 23 patients (group I) and "traditional" splenectomy was carried out in 19 patients (group II). There was no difference in the demographic data and trauma severity between the 2 groups. Noninvasive investigations, such as laboratory investigations, serial abdominal ultrasound examinations, x-ray in multiple views, and computed tomography had been performed before the decision about necessity of an operation was made. Patients after laparoscopic operations had better recovering conditions compared with patients with the same injury after "traditional" splenectomy. Neither surgery-related complications nor mortalities were registered in both groups. Laparoscopic splenectomy was more time-consuming operation than "traditional" splenectomy. We suggest that as experience of laparoscopic splenectomy is gained the operation time will be reduced. Laparoscopic splenectomy is a safe feasible operation in patients

  14. Interaction of methotrexate with trypsin analyzed by spectroscopic and molecular modeling methods

    Science.gov (United States)

    Wang, Yanqing; Zhang, Hongmei; Cao, Jian; Zhou, Qiuhua

    2013-11-01

    Trypsin is one of important digestive enzymes that have intimate correlation with human health and illness. In this work, the interaction of trypsin with methotrexate was investigated by spectroscopic and molecular modeling methods. The results revealed that methotrexate could interact with trypsin with about one binding site. Methotrexate molecule could enter into the primary substrate-binding pocket, resulting in inhibition of trypsin activity. Furthermore, the thermodynamic analysis implied that electrostatic force, hydrogen bonding, van der Waals and hydrophobic interactions were the main interactions for stabilizing the trypsin-methotrexate system, which agreed well with the results from the molecular modeling study.

  15. Sporamin-mediated resistance to beet cyst nematodes (Heterodera schachtii Schm.) is dependent on trypsin inhibitory activity in sugar beet (Beta vulgaris L.) hairy roots.

    Science.gov (United States)

    Cai, Daguang; Thurau, Tim; Tian, Yanyan; Lange, Tina; Yeh, Kai-Wun; Jung, Christian

    2003-04-01

    Sporamin, a sweet potato tuberous storage protein, is a Kunitz-type trypsin inhibitor. Its capability of conferring insect-resistance on transgenic tobacco and cauliflower has been confirmed. To test its potential as an anti-feedant for the beet cyst nematode (Heterodera schachtii Schm.), the sporamin gene SpTI-1 was introduced into sugar beet (Beta vulgaris L.) by Agrobacterium rhizogenes-mediated transformation. Twelve different hairy root clones expressing sporamin were selected for studying nematode development. Of these, 8 hairy root clones were found to show significant efficiency in inhibiting the growth and development of the female nematodes whereas 4 root clones did not show any inhibitory effects even though the SpTI-1 gene was regularly expressed in all of the tested hairy roots as revealed by northern and western analyses. Inhibition of nematode development correlated with trypsin inhibitor activity but not with the amount of sporamin expressed in hairy roots. These data demonstrate that the trypsin inhibitor activity is the critical factor for inhibiting growth and development of cyst nematodes in sugar beet hairy roots expressing the sporamin gene. Hence, the sweet potato sporamin can be used as a new and effective anti-feedant for controlling cyst nematodes offering an alternative strategy for establishing nematode resistance in crops.

  16. Role of trypsin in the replication of Avian metapneumovirus subtype C (strain MN-2a) and its entry into the Vero cells.

    Science.gov (United States)

    Paudel, Sarita; Shin, Hyun-Jin

    2015-12-01

    To understand the molecular mechanisms of Avian metapneumovirus (aMPV) and the requirements involved in the infection and fusion, trypsin treatment was done in the different stages of virus; before infection, during entry and after virus infection followed by aMPV infection. The growth kinetics of aMPV was compared in time dependent manner. The effect of trypsin was found in the later stage of aMPV infection increasing the numbers of infected cells with the significant higher titer of infectious virions to that of trypsin treated before infection, during entry and aMPV. A serine protease inhibitor reduced aMPV replication in a significant way, whereas cysteine peptidase (E-64), aspartic protease (pepstatin A), and metalloprotease (phosphoramidon) inhibitors had no effect on aMPV replication. Inoculation of aMPV on Vero cells expressing the membrane-associated protease TMPRSS2 resulted in higher virus titers than that inoculated on normal Vero cells and is statistically significant (p < 0.05). Also, an inhibitor of clathrin/caveolae-mediated endocytosis had no effect on virus progeny, indicating that aMPV does not use the endocytic pathway for entry but undergoes direct fusion. The effect of lysosomotropic agents was not significant, suggesting that aMPV does not require low-pH environment in endosomes to fuse its envelope with the plasma membrane. Copyright © 2015 Elsevier Ltd. All rights reserved.

  17. Syk inhibitors.

    Science.gov (United States)

    Chihara, Kazuyasu; Kimura, Yukihiro; Honjo, Chisato; Takeuchi, Kenji; Sada, Kiyonao

    2013-01-01

    Non-receptor type of protein-tyrosine kinase Syk (spleen tyrosine kinase) was isolated in University of Fukui in 1991. Syk is most highly expressed by haemopoietic cells and known to play crucial roles in the signal transduction through various immunoreceptors of the adaptive immune response. However, recent reports demonstrate that Syk also mediates other biological functions, such as innate immune response, osteoclast maturation, platelet activation and cellular adhesion. Moreover, ectopic expression of Syk by epigenetic changes is reported to cause retinoblastoma. Because of its critical roles on the cellular functions, the development of Syk inhibitors for clinical use has been desired. Although many candidate compounds were produced, none of them had progressed to clinical trials. However, novel Syk inhibitors were finally developed and its usefulness has been evaluated in the treatment of allergic rhinitis, rheumatoid arthritis and idiopathic thrombocytopenic purpura. In this review, we will summarize the history, structure and function of Syk, and then the novel Syk inhibitors and their current status. In addition, we will introduce our research focused on the functions of Syk on Dectin-1-mediated mast cell activation.

  18. Distal splenorenal shunt with partial spleen resection

    Directory of Open Access Journals (Sweden)

    Gajin Predrag

    2007-01-01

    Full Text Available Introduction: Hypersplenism is a common complication of portal hypertension. Cytopenia in hypersplenism is predominantly caused by splenomegaly. Distal splenorenal shunt (Warren with partial spleen resection is an original surgical technique that regulates cytopenia by reduction of the enlarged spleen. Objective. The aim of our study was to present the advantages of distal splenorenal shunt (Warren with partial spleen resection comparing morbidity and mortality in a group of patients treated by distal splenorenal shunt with partial spleen resection with a group of patients treated only by a distal splenorenal shunt. Method. From 1995 to 2003, 41 patients with portal hypertension were surgically treated due to hypersplenism and oesophageal varices. The first group consisted of 20 patients (11 male, mean age 42.3 years who were treated by distal splenorenal shunt with partial spleen resection. The second group consisted of 21 patients (13 male, mean age 49.4 years that were treated by distal splenorenal shunt only. All patients underwent endoscopy and assessment of oesophageal varices. The size of the spleen was evaluated by ultrasound, CT or by scintigraphy. Angiography was performed in all patients. The platelet and white blood cell count and haemoglobin level were registered. Postoperatively, we noted blood transfusion, complications and total hospital stay. Follow-up period was 12 months, with first checkup after one month. Results In the first group, only one patient had splenomegaly postoperatively (5%, while in the second group there were 13 patients with splenomegaly (68%. Before surgery, the mean platelet count in the first group was 51.6±18.3x109/l, to 118.6±25.4x109/l postoperatively. The mean platelet count in the second group was 67.6±22.8x109/l, to 87.8±32.1x109/l postoperatively. Concerning postoperative splenomegaly, statistically significant difference was noted between the first and the second group (p<0.05. Comparing the

  19. Comparative study of the binding of trypsin to caffeine and theophylline by spectrofluorimetry

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Ruiyong, E-mail: wangry@zzu.edu.cn [Department of Chemistry, Zhengzhou University, Zhengzhou 450001 (China); Kang, Xiaohui [Department of Chemistry, Zhengzhou University, Zhengzhou 450001 (China); Wang, Ruiqiang [The First Affiliated Hospital of Zhengzhou University, Zhengzhou 450052 (China); Wang, Rui; Dou, Huanjing; Wu, Jing; Song, Chuanjun [Department of Chemistry, Zhengzhou University, Zhengzhou 450001 (China); Chang, Junbiao, E-mail: changjunbiao@zzu.edu.cn [Department of Chemistry, Zhengzhou University, Zhengzhou 450001 (China)

    2013-06-15

    The interactions between trypsin and caffeine/theophylline were investigated by fluorescence spectroscopy, UV–visible absorption spectroscopy, resonance light scattering and synchronous fluorescence spectroscopy under mimic physiological conditions. The results revealed that the fluorescence quenching of trypsin by caffeine and theophylline was the result of the formed complex of caffeine–trypsin and theophylline–trypsin. The binding constants and thermodynamic parameters at three different temperatures were obtained. The hydrophobic interaction was the predominant intermolecular forces to stabilize the complex. Results showed that caffeine was the stronger quencher and bound to trypsin with higher affinity than theophylline. -- Highlights: ► The fluorescence of trypsin can be quenched by caffeine or theophylline via hydrophobic contacts. ► Caffeine binds to trypsin with higher affinity than theophylline. ► The influence of molecular structure on the binding aspects is reported.

  20. Trypsin Binding with Copper Ions Scavenges Superoxide: Molecular Dynamics-Based Mechanism Investigation

    Directory of Open Access Journals (Sweden)

    Xin Li

    2018-01-01

    Full Text Available Trypsin is a serine protease, which has been proved to be a novel superoxide scavenger. The burst of superoxide induced by polychlorinated biphenyls can be impeded by trypsin in both wild type and sod knockout mutants of Escherichia coli. The experimental results demonstrated that the activities of superoxide scavenging of trypsin were significantly accelerated by Cu ions. Also, with the addition of Cu ions, a new β-sheet (β7 transited from a random coil in the Cu(II-trypsin (TP system, which was favorable for the formation of more contacts with other sheets of trypsin. Residue–residue network analysis and the porcupine plots proved that the Cu ion in trypsin strengthened some native interactions among residues, which ultimately resulted in much greater stability of the Cu(II-TP system. Moreover, compact and stable trypsin structures with Cu ions might be responsible for significantly provoking the activity of superoxide scavenging.

  1. Comparative study of the binding of trypsin to caffeine and theophylline by spectrofluorimetry

    International Nuclear Information System (INIS)

    Wang, Ruiyong; Kang, Xiaohui; Wang, Ruiqiang; Wang, Rui; Dou, Huanjing; Wu, Jing; Song, Chuanjun; Chang, Junbiao

    2013-01-01

    The interactions between trypsin and caffeine/theophylline were investigated by fluorescence spectroscopy, UV–visible absorption spectroscopy, resonance light scattering and synchronous fluorescence spectroscopy under mimic physiological conditions. The results revealed that the fluorescence quenching of trypsin by caffeine and theophylline was the result of the formed complex of caffeine–trypsin and theophylline–trypsin. The binding constants and thermodynamic parameters at three different temperatures were obtained. The hydrophobic interaction was the predominant intermolecular forces to stabilize the complex. Results showed that caffeine was the stronger quencher and bound to trypsin with higher affinity than theophylline. -- Highlights: ► The fluorescence of trypsin can be quenched by caffeine or theophylline via hydrophobic contacts. ► Caffeine binds to trypsin with higher affinity than theophylline. ► The influence of molecular structure on the binding aspects is reported

  2. Trypsin diminishes the rat potency of polio serotype 3.

    Science.gov (United States)

    ten Have, R; Westdijk, J; Levels, L M A R; Koedam, P; de Haan, A; Hamzink, M R J; Metz, B; Kersten, G F A

    2015-11-01

    This study addresses observations made in view of testing in practice the guideline in the European Pharmacopoeia (EP) on omitting the rat potency test for release of polio containing vaccines. In general, use of the guideline is valid and the D-antigen ELISA can indeed be used as an in vitro alternative for the in vivo test. However, the set-up of the ELISA is crucial and should include detection of antigenic site 1 in polio serotype 3 as destruction of that site by trypsin results in a reduced rat potency. Antigenic site 1 in polio serotype 2 may also be modified by trypsin, but the cleavage of viral protein 1 (VP1) did not affect the rat potency. Therefore, any antigenic site, except site 1, can be used for detection of polio serotype 2. It is advised to include testing of the effect of trypsin treatment in the EP-guideline. This allows polio vaccine manufacturers to check whether their in-house ELISA needs improvement. Copyright © 2015 The International Alliance for Biological Standardization. Published by Elsevier Ltd. All rights reserved.

  3. Characterization of trypsin-derived peptides acrylamide-adducted hemoglobin

    International Nuclear Information System (INIS)

    Springer, D.L.; Goheen, S.C.; Edmonds, C.G.; McCulloch, M.; Sylvester, D.M.; Sander, C.; Bull, R.J.

    1991-01-01

    Even though there are a number of sources for human exposure to acrylamide, reliable biomarkers of exposure are not available. In an effort to develop such a biomarker, the authors are characterizing peptides derived from trypsin digests of acrylamide-adducted hemoglobin. For this, radiolabeled acrylamide was incubated with this, radiolabeled acrylamide was incubated with purified human hemoglobin (Ao) and decomposition products removed by dialysis. When the adducted hemoglobin was separated by reverse-phase HPLC, radioactivity eluted with the α and β subunits, suggesting covalent binding. Digestion of individual subunits with trypsin followed by reverse phase HPLC, indicated that most of the radioactivity associated with the α subunit co-eluted with a single peptide. Similar results were observed for the β subunit except that significant amounts of radioactivity eluted with the solvent front, suggesting that radioactivity was released by trypsin digestion. Currently, these preparation are under further characterization by electrospray ionization mass spectrometry. This approach will aid in the identification of the adducted will aid in the identification of the adducted peptide and subsequent preparation of an acrylamide-specific antibody

  4. Effects of Mucuna pruriens protease inhibitors on Echis carinatus venom.

    Science.gov (United States)

    Hope-Onyekwere, Nnadozie Stanley; Ogueli, Godwin Ifeanyi; Cortelazzo, Alessio; Cerutti, Helena; Cito, Annarita; Aguiyi, John C; Guerranti, Roberto

    2012-12-01

    The medicinal plant Mucuna pruriens, with reputed anti-snake venom properties has been reported to contain a kunitz-type trypsin inhibitor. This study was undertaken to further evaluate the protease inhibitory potential of gpMuc, a multiform glycoprotein, and other protein fractions from M. pruriens seeds against trypsin, chymotrypsin, Echis carinatus snake venom, ecarin and thrombin. The results showed that gpMuc inhibited both trypsin and chymotrypsin activities and was thermally stable, maintaining its trypsin inhibitory activity at temperatures of up to 50°C. Its structural conformation was also maintained at pH ranges of 4-7. Immunoreactivity study confirms that it contains protease-recognizing epitope on one of its isoforms. The whole protein extract of M. pruriens seeds inhibited prothrombin activation by ecarin and whole E. carinatus venom, and also thrombin-like activity using chromogenic assay. Copyright © 2012 John Wiley & Sons, Ltd.

  5. Imaging of the spleen: a proposed algorithm

    International Nuclear Information System (INIS)

    Shirkhoda, A.; McCartney, W.H.; Staab, E.V.; Mittelstaedt, C.A.

    1980-01-01

    The /sup 99m/Tc sulfur colloid scan is an effective initial method for evaluating splenic size, position, and focal or diffusely altered radionuclide uptake. Sonography is a useful next step in determining whether focal lesions are solid or cystic and the relation of the spleen to adjacent organs. In our opinion, computed tomography (CT) may be reserved for the few instances in which diagnostic questions remain unanswered after radionuclide scanning and sonography. Angiography is used primarily in splenic trauma. We evaluated 900 patients suspected of having liver-spleen abnormality. This experience, which led to a logically sequenced noninvasive imaging approach for evaluating suspected splenic pathology, is summarized and illustrated by several cases

  6. Imaging of the spleen: a proposed algorithm

    Energy Technology Data Exchange (ETDEWEB)

    Shirkhoda, A.; McCartney, W.H.; Staab, E.V.; Mittelstaedt, C.A.

    1980-07-01

    The /sup 99m/Tc sulfur colloid scan is an effective initial method for evaluating splenic size, position, and focal or diffusely altered radionuclide uptake. Sonography is a useful next step in determining whether focal lesions are solid or cystic and the relation of the spleen to adjacent organs. In our opinion, computed tomography (CT) may be reserved for the few instances in which diagnostic questions remain unanswered after radionuclide scanning and sonography. Angiography is used primarily in splenic trauma. We evaluated 900 patients suspected of having liver-spleen abnormality. This experience, which led to a logically sequenced noninvasive imaging approach for evaluating suspected splenic pathology, is summarized and illustrated by several cases.

  7. Localization to Chromosomes of Structural Genes for the Major Protease Inhibitors of Barley Grains

    DEFF Research Database (Denmark)

    Hejgaard, Jørn; Bjørn, S.E.; Nielsen, Gunnar Gissel

    1984-01-01

    Wheat-barley chromosome addition lines were compared by isoelectric focusing of protein extracts to identify chromosomes carrying loci for the major immunochemically distinct protease inhibitors of barley grains. Structural genes for the following inhibitors were localized: an inhibitor of both...... endogenous α-amylase 2 and subtilisin (ASI) on chromosome 2, two chymotrypsin/subtilisin inhibitors (CI-1 and CI-2) on chromosome 5 (long arm) and the major trypsin inhibitor (TI-1) on chromosome 3....

  8. Laparoscopic splenectomy for a simultaneous wandering spleen along with an ectopic accessory spleen. Case report and review of the literature

    Directory of Open Access Journals (Sweden)

    Antonia Rizzuto

    Full Text Available Background: Wandering spleen and accessory spleen are uncommon entity occurring during embryonic development. Wandering spleen results in an excessive mobility and migration of the spleen from its normal position in the left hypochondrium while accessory spleen is characterized by ectopic splenic masses or tissue disjointed from the main body of spleen.Due to the nonspecific and multiple symptoms the clinical diagnosis of both conditions is uncertain even with imaging techniques, such as CT and MRI. The coexistence of both diseases (wandering spleen ad accessory spleen is uncommon. Case report: A 17–year old European female with a history of minor beta thalassemia and recurrent attacks of abdominal pain. Pre- operative management consisted of routine laboratory tests, ultrasound, CT scan. An ectopic spleen along with an accessory spleen were diagnosed. After a multidisciplinary board a laparoscopic splenectomy was performed. Post-operative recovery was uneventful, and the patient was discharged on the 6th post-operative day with the indication to continue the therapy with low molecular weight heparin (LMWH for 30 days Conclusions: This case represents a simultaneous condition of wandering splenomegaly along with an ectopic wandering spleen. The coexistence of these two rare conditions is peculiar such as the age of the patient, as literature reports such diseases to affect children or more commonly people in the range of 20–40 years of age. Laparoscopic treatment for this particular condition is also unusual. Keywords: Ectopic spleen, Wandering spleen, Laparoscopic splenectomy

  9. Value of transoperative scintigraphy in the detection of accessory spleens

    International Nuclear Information System (INIS)

    Sezeur, A.; Goujard, F.; Labriolle-Vaylet, C.L. de; Wioland, M.; Douay, L.; Desmarquet, J.

    1990-01-01

    A case of accessory spleen, 1 cm in diameter, responsible for recurrence of an idiopathic thrombocytopenic purpura after splenectomy is reported. This case is original in that the accessory spleen could only be detected by transoperative scintigraphy. Transoperative scintigraphy is a simple method to be used when one or several unrecognized accessory spleens are responsible for recurrence of a blood disease after excision of the principal spleen [fr

  10. A wandering spleen presenting as a hypogastric mass: Case report ...

    African Journals Online (AJOL)

    ... the spleen. A 26-year-old woman was admitted to our hospital with vomiting and abdominal pain. Abdominal examination revealed a large ovoid hypogastric mass. A CT scan showed a wandering spleen in the hypogastric region. Exploratory laparotomy revealed an ischemic spleen. A total splenectomy was performed.

  11. Lectins as carriers: preparation and purification of a concanavalin A-trypsin conjugate

    International Nuclear Information System (INIS)

    Shier, W.T.

    1985-01-01

    The scheme for the preparation and purification of Con A-trypsin demonstrates the presence of Con A in the conjugate by affinity purification and by hemagglutination. The presence of Con A was demonstrated in two additional ways. The conjugate was prepared using trypsin labeled with iodine 125 and unlabeled Con A. The resulting conjugate containing 42,800 cpm/mg protein was used to demonstrate prolonged retention of the conjugated trypsin in mouse footpads. The presence of trypsin was also demonstrated in the conjugate by affinity chromatography and by the esterase activity characteristic of trypsin with tosyl-L-arginine methyl ester as substrate. Con A-trypsin was also assayed for proteolytic activity with a macromolecular substrate azocasein. A comparison of proteolytic activities with these two substrates indicated that 50-80% of the proteolytic activity observed with the small substrate is retained with macromolecular substrates

  12. Magnetic nanoparticles coated with polyaniline to stabilize immobilized trypsin

    Energy Technology Data Exchange (ETDEWEB)

    Maciel, J. C., E-mail: jackeline-maciel@hotmail.com [Universidade Federal de Roraima (Brazil); Mercês, A. A. D.; Cabrera, M. [Universidade Federal de Pernambuco, Laboratório de Imunopatologia Keizo Asami (Brazil); Shigeyosi, W. T. [Universidade Federal de São Carlos, Departamento de Física (Brazil); Souza, S. D. de; Olzon-Dionysio, M.; Fabris, J. D. [Universidade Federal dos Vales de Jequitinhonha e Mucuri (Brazil); Cardoso, C. A. [Universidade Federal de São Carlos, Departamento de Física (Brazil); Neri, D. F. M. [Universidade Federal do Vale do São Francisco (Brazil); Silva, M. P. C.; Carvalho, L. B. [Universidade Federal de Pernambuco, Laboratório de Imunopatologia Keizo Asami (Brazil)

    2016-12-15

    It is reported the synthesis of magnetic nanoparticles via the chemical co-precipitation of Fe {sup 3+} ions and their preparation by coating them with polyaniline. The electronic micrograph analysis showed that the mean diameter for the nanoparticles is ∼15 nm. FTIR, powder X-ray diffraction and Mössbauer spectroscopy were used to understand the chemical, crystallographic and {sup 57}Fe hyperfine structures for the two samples. The nanoparticles, which exhibited magnetic behavior with relatively high spontaneous magnetization at room temperature, were identified as being mainly formed by maghemite (γFe{sub 2}O{sub 3}). The coated magnetic nanoparticles (sample labeled “mPANI”) presented a real ability to bind biological molecules such as trypsin, forming the magnetic enzyme derivative (sample “mPANIG-Trypsin”). The amount of protein and specific activity of the immobilized trypsin were found to be 13±5 μg of protein/mg of mPANI (49.3 % of immobilized protein) and 24.1±0.7 U/mg of immobilized protein, respectively. After 48 days of storage at 4 {sup ∘}C, the activity of the immobilized trypsin was found to be 89 % of its initial activity. This simple, fast and low-cost procedure was revealed to be a promising way to prepare mPANI nanoparticles if technological applications addressed to covalently link biomolecules are envisaged. This route yields chemically stable derivatives, which can be easily recovered from the reaction mixture with a magnetic field and recyclable reused.

  13. Chitosan nanoencapsulated exogenous trypsin biomimics zymogen-like enzyme in fish gastrointestinal tract.

    Science.gov (United States)

    Kumari, Rakhi; Gupta, Subodh; Singh, Arvind R; Ferosekhan, S; Kothari, Dushyant C; Pal, Asim Kumar; Jadhao, Sanjay Balkrishna

    2013-01-01

    Exogenous proteolytic enzyme supplementation is required in certain disease conditions in humans and animals and due to compelling reasons on use of more plant protein ingredients and profitability in animal feed industry. However, limitations on their utility in diet are imposed by their pH specificity, thermolabile nature, inhibition due to a variety of factors and the possibility of intestinal damage. For enhancing the efficacy and safety of exogenous trypsin, an efficient chitosan (0.04%) nanoencapsulation-based controlled delivery system was developed. An experiment was conducted for 45 days to evaluate nanoencapsulated trypsin (0.01% and 0.02%) along with 0.02% bare trypsin and 0.4% chitosan nanoparticles against a control diet on productive efficiency (growth rate, feed conversion and protein efficiency ratio), organo-somatic indices, nutrient digestibility, tissue enzyme activities, hematic parameters and intestinal histology of the fish Labeo rohita. All the synthesized nanoparticles were of desired characteristics. Enhanced fish productive efficiency using nanoencapsulated trypsin over its bare form was noticed, which corresponded with enhanced (P<0.01) nutrient digestibility, activity of intestinal protease, liver and muscle tissue transaminases (alanine and aspartate) and dehydrogenases (lactate and malate), serum blood urea nitrogen and serum protein profile. Intestinal tissues of fish fed with 0.02% bare trypsin showed broadened, marked foamy cells with lipid vacuoles. However, villi were healthier in appearance with improved morphological features in fish fed with nanoencapsulated trypsin than with bare trypsin, and the villi were longer in fish fed with 0.01% nanoencapsulated trypsin than with 0.02% nanoencapsulated trypsin. The result of this premier experiment shows that nanoencapsulated trypsin mimics zymogen-like proteolytic activity via controlled release, and hence the use of 0.01% nanoencapsulated trypsin (in chitosan nanoparticles) over bare

  14. Chitosan nanoencapsulated exogenous trypsin biomimics zymogen-like enzyme in fish gastrointestinal tract.

    Directory of Open Access Journals (Sweden)

    Rakhi Kumari

    Full Text Available Exogenous proteolytic enzyme supplementation is required in certain disease conditions in humans and animals and due to compelling reasons on use of more plant protein ingredients and profitability in animal feed industry. However, limitations on their utility in diet are imposed by their pH specificity, thermolabile nature, inhibition due to a variety of factors and the possibility of intestinal damage. For enhancing the efficacy and safety of exogenous trypsin, an efficient chitosan (0.04% nanoencapsulation-based controlled delivery system was developed. An experiment was conducted for 45 days to evaluate nanoencapsulated trypsin (0.01% and 0.02% along with 0.02% bare trypsin and 0.4% chitosan nanoparticles against a control diet on productive efficiency (growth rate, feed conversion and protein efficiency ratio, organo-somatic indices, nutrient digestibility, tissue enzyme activities, hematic parameters and intestinal histology of the fish Labeo rohita. All the synthesized nanoparticles were of desired characteristics. Enhanced fish productive efficiency using nanoencapsulated trypsin over its bare form was noticed, which corresponded with enhanced (P<0.01 nutrient digestibility, activity of intestinal protease, liver and muscle tissue transaminases (alanine and aspartate and dehydrogenases (lactate and malate, serum blood urea nitrogen and serum protein profile. Intestinal tissues of fish fed with 0.02% bare trypsin showed broadened, marked foamy cells with lipid vacuoles. However, villi were healthier in appearance with improved morphological features in fish fed with nanoencapsulated trypsin than with bare trypsin, and the villi were longer in fish fed with 0.01% nanoencapsulated trypsin than with 0.02% nanoencapsulated trypsin. The result of this premier experiment shows that nanoencapsulated trypsin mimics zymogen-like proteolytic activity via controlled release, and hence the use of 0.01% nanoencapsulated trypsin (in chitosan

  15. Trypsin inhibitory activity of artemisinin and its biotransformed product

    International Nuclear Information System (INIS)

    Shahwar, D.; Raza, M.A.

    2013-01-01

    Summary: Artemisinin (1 ), a sesquiterpene lactone is an important constituent of anti-malarial drugs. In the present study, it was extracted from aerial parts of Artemisia roxburghiana Besser. Biotransformation of artemisinin ( 1 ) was carried out in the culture of Aspergillus niger GC-4 which yielded 5-hydroxy artemisinin (2 ) The structures of 1-2 were confirmed through spectral studies. Both compounds were screened against trypsin using colorimetric method. The biotransformed product 2 showed significant protease inhibitory activity with 53.5 +- 1.6% inhibition and IC/sub 50/ = 0.29 +- 0.02 mM as compared to artemisinin (20.4 +- 0.3% inhibition). (author)

  16. Human and bovine spinal disc mechanics subsequent to trypsin injection

    Directory of Open Access Journals (Sweden)

    Jeremy Alsup

    2017-10-01

    The Translational Potential of this Article: Preclinical testing of novel spinal devices is essential to the design validation and regulatory processes, but current testing techniques rely on cadaveric testing of primarily older spines with essentially random amounts of disc degeneration. The present work investigates the viability of using trypsin injections to create a more uniform preclinical model of disc degeneration from a mechanics perspective, for the purpose of testing spinal devices. Such a model would facilitate translation of new spinal technologies to clinical practice.

  17. Phosphoprotein phosphatase of bovine spleen cell nuclei: physicochemical properties

    International Nuclear Information System (INIS)

    Rezyapkin, V.I.; Leonova, L.E.; Komkova, A.I.

    1986-01-01

    The physicochemical properties of phosphoprotein phosphatase (EC 1.3.1.16) from bovine spleen cell nuclei were studied. The enzyme possesses broad substrate specificity and catalyzes the dephosphorylation of phosphocasein, ATP, ADP, and p-nitrophenyl phosphate (pNPP). K/sub m/ for ATP, ADP, and pNPP are equal to 0.44, 0.43, and 1.25 mM, respectively. M/sub r/ of the enzyme, according to the data of gel filtraction of Sephadex G-75 and electrophoresis in polyacrylamide gel of various concentrations is ∼ 33,000. In electrophoresis in the presence of SDS, two protein bands with M/sub r/ 12,000 and 18,000 are detected. In the enzyme molecule, acid amino acid residues predominate; two free SH groups and two disulfide bridges are detected. Phosphoprotein phosphatase is a glycoprotein, containing ∼ 22% carbonhydrates. The protein possesses a supplementary absorption maximum at 560 nm. Ammonium molybdate is a competitive inhibitor with K/sub i/ 0.37 μM, while sodium fluoride is a noncompetitive inhibitor with K/sub i/ 1.3 mM. Incubation in the presence of 2 mM phenylmethylsulfonyl fluoride for 25 h leads to a loss of ∼ 46% of the enzymatic activity. Ammonium molybdate, sodium fluoride, and PMSF are reversible inhibitors. Modifications of the SH groups, NH 2 groups, and histidine leads to a decrease in the enzymatic activity. Incubation of phosphoprotein phosphatase with [γ- 32 P]ATP leads to the incorporation of 0.33 mole 33 P per mole of the enzyme. The mechanism of hydrolysis of the phosphodiester bond, catalyzed by the enzyme, is discussed

  18. Highly Stable Trypsin-Aggregate Coatings on Polymer Nanofibers for Repeated Protein Digestion

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Byoung Chan; Lopez-Ferrer, Daniel; Lee, Sang-mok; Ahn, Hye-kyung; Nair, Sujith; Kim, Seong H.; Kim, Beom S.; Petritis, Konstantinos; Camp, David G.; Grate, Jay W.; Smith, Richard D.; Koo, Yoon-mo; Gu, Man Bock; Kim, Jungbae

    2009-04-01

    A stable and robust trypsin-based biocatalytic system was developed and demonstrated for proteomic applications. The system utilizes polymer nanofibers coated with trypsin aggregates for immobilized protease digestions. After covalently attaching an initial layer of trypsin to the polymer nanofibers, highly concentrated trypsin molecules are crosslinked to the layered trypsin by way of a glutaraldehyde treatment. This new process produced a 300-fold increase in trypsin activity compared with a conventional method for covalent trypsin immobilization and proved to be robust in that it still maintained a high level of activity after a year of repeated recycling. This highly stable form of immobilized trypsin was also resistant to autolysis, enabling repeated digestions of bovine serum albumin over 40 days and successful peptide identification by LC-MS/MS. Finally, the immobilized trypsin was resistant to proteolysis when exposed to other enzymes (i.e. chymotrypsin), which makes it suitable for use in “real-world” proteomic applications. Overall, the biocatalytic nanofibers with enzyme aggregate coatings proved to be an effective approach for repeated and automated protein digestion in proteomic analyses.

  19. Effects of bisphenol S on the structures and activities of trypsin and pepsin.

    Science.gov (United States)

    Wang, Yan-Qing; Zhang, Hong-Mei

    2014-11-19

    The effects of bisphenol S on the structures and activities of trypsin and pepsin were investigated by various methods like UV-visible absorbance, fluorescence, circular dichroism, and molecular docking. The secondary and tertiary structures of trypsin and pepsin were altered by bisphenol S binding, which resulted in the loosening of the skeletons of trypsin and pepsin. In addition, bisphenol S induced microenvironmental changes around tyrosine and tryptophan residues of trypsin and pepsin. The activity experimental results showed that the activity of pepsin decreases obviously with the increasing concentration of BPS, while the activity of trypsin does not change remarkably. The binding and thermodynamic parameters obtained by molecular docking and fluorescence spectroscopy showed that the bindings of bisphenol S to trypsin and pepsin were spontaneous processes and hydrogen bonding and hydrophobic interactions played a vital role in stabilizing the bisphenol S-trypsin and bisphenol S-pepsin complexes. The binding constants (K(A)) of bisphenol S with trypsin were 7.42 × 10(4) (298 K) and 5.91 × 10(4) L/mol (310 K), and those of pepsin were 5.78 × 10(4) (298 K) and 4.44 × 10(4) L/mol (310 K). Moreover, there was one main kind of binding site for bisphenol S on trypsin or pepsin.

  20. Discovery of natural mouse serum derived HIV-1 entry inhibitor(s).

    Science.gov (United States)

    Wei, M; Chen, Y; Xi, J; Ru, S; Ji, M; Zhang, D; Fang, Q; Tang, B

    Among rationally designed human immunodeficiency virus 1 (HIV-1) inhibitors, diverse natural factors have showed as potent anti-HIV activity in human blood. We have discovered that the boiled supernatant of healthy mouse serum could suppress HIV-1 entry, and exhibited reduced inhibitory activity after trypsin digestion. Further analysis demonstrated that only the fraction containing 10-25 K proteins could inhibit HIV-1 mediated cell-cell fusion. These results suggest that the 10-25 K protein(s) is novel natural HIV-1 entry inhibitor(s). Our findings provide important information about novel natural HIV entry inhibitors in mouse serum.

  1. The onset of hemoglobin synthesis in spleens of irradiated mice after bone marrow transplantation

    International Nuclear Information System (INIS)

    Ponka, P.; Fuchs, O.; Borova, J.; Necas, E.

    1977-01-01

    Messenger RNA (mRNA) for globin was isolated from spleens of irradiated mice in which erythroid differentiation was induced by a bone marrow graft. The globin mRNA was isolated either by means of sucrose gradients of reticulocyte polysomal RNA or by affinity chromatography of total spleen RNA on poly (U)-sepharose. The globin mRNA was tested in a wheat embryo cell-free system. The appearance of mRNA in the spleen erythroid colonies was correlated with other parameters of erythroid differentiation such as globin synthesis, activity of delta-aminolevulinic acid synthetase and iron uptake. Poly(A) containing mRNA did appear already on the 3rd day after grafting. However, significant translational activity of globin mRNA could be demonstrated only one day later together with increase in globin synthesis and delta-aminolevulinic acid synthetase and enhanced iron uptake. In the second part of this study mouse spleen cells rich in erythroid elements were incubated with a specific heme synthesis inhibitor (isonicotinic acid hydrazide, INH) and the synthesis of 9 S RNA was estimated. It was found that a 40-minute incubation with INH reduced uridine incorporation into 9 S RNA fraction by about 40%. (author)

  2. Desoxyribonucleic acid (DNA) synthesis in vitro by thymus and spleen cells of the rat after hyperthermia

    Energy Technology Data Exchange (ETDEWEB)

    Tempel, K.; Spath, A.

    1988-03-01

    The inhibition of the semiconservative and restorative DNA synthesis caused by hyperthermia (30 to 60 min, 43/sup 0/C) was significantly higher in spleen cells than in thymus cells. The DNA repair synthesis of thymus cells measured at 37/sup 0/C was increased by about two times the initial value after a pre-incubation of 30 to 90 min and 30 to 60 min, respectively, with 37 and 43/sup 0/C, respectively. Under the same conditions, the /sup 3/H-thymidine incorporation into the DNA of spleen cells diminished proportionally to the pre-incubation time after a pre-incubation of 30 and 45 min, respectively, with 43 and 37/sup 0/C, respectively. When hyperthermia and inhibitors of DNA synthesis or DNA repair (hydroxyurea, 1-..beta..-D-arabinofuranosylcytosine, 3', 5'-didesoxythymidine, and 3-aminobenzamide) were combined, overadditive effects - without cellspecific particularities - were seen only in the case of 3-aminobenzamide. Only in thymus cells, the inhibitor of DNA topoisomerase II novobiocin caused an overadditive reinforcement of the inhibition induced by hyperthermia of the semiconservative DNA synthesis. The stimulation of DNA repair synthesis in thymus cells caused by novobiocin with the aid of DNA polymerase ..beta.. could be compensated by hyperthermia. The sedimentation of thymus and spleen cell nucleoids was increased after hyperthermia. The results suggest a special importance of DNA topology and of the DNA polymerase ..beta.. activity for the cellular effect of hyperthermia.

  3. Relationship between number of spleen colonies and 125IdUrd incorporation into spleen and femur

    International Nuclear Information System (INIS)

    Inoue, T.; Bullis, J.E.; Cronkite, E.P.; Hubner, G.E.

    1983-01-01

    Graded numbers of bone marrow (BM) cells were injected into fatally irradiated mice. Eight days later the mice were given 3.0 μCi (1 Ci = 3.7 x 10 10 Bq) of 125 IdUrd to label proliferating cells in the spleen and BM. On day 9 the mice were killed and the spleens and femurs were removed for splenic colony assay and measurement of radioactivity in the spleen and femurs. The number of splenic colonies shows a linear relationship with dose of marrow cells injected from 10 4 to 10 5 cells. The slope of the curve of spleen colonies versus number of cells injected is 5 and below 10 4 there is a striking departure from the simple linearity. Below 2 x 10 3 cells injected, the logarithm of the observed colony yield is linear with logarithm of the number of cells injected. Poisson calculation of the average number of pluripotent stem cells that should be present with numbers of marrow cells injected below 2 x 10 3 followed closely the actual observations. The data show that there is no detectible proliferation in the BM until the dose of marrow cells exceeds 3.5 x 10 4 cells. Induction of cells into cycle increases the seeding into the BM, and thymidine cytocide drastically reduces seeding in the BM, leading us to conclude that the BM is repopulated almost exclusively by stem cells in DNA synthesis

  4. Chitosan nanoparticles-trypsin interactions: Bio-physicochemical and molecular dynamics simulation studies.

    Science.gov (United States)

    Salar, Safoura; Mehrnejad, Faramarz; Sajedi, Reza H; Arough, Javad Mohammadnejad

    2017-10-01

    Herein, we investigated the effect of the chitosan nanoparticles (CsNP) on the structure, dynamics, and activity of trypsin. The enzyme activity in complex with the nanoparticles slightly increased, which represents the interactions between the nanoparticles and the enzyme. The kinetic parameters of the enzyme, K m and k cat , increased after adding the nanoparticles, resulting in a slight increase in the catalytic efficiency (k cat /K m ). However, the effect of the nanoparticles on the kinetic stability of trypsin has not exhibited significant variations. Fluorescence spectroscopy did not show remarkable changes in the trypsin conformation in the presence of the nanoparticles. The circular dichroism (CD) spectroscopy results also revealed the secondary structure of trypsin attached to the nanoparticles slightly changed. Furthermore, we used molecular dynamics (MD) simulation to find more information about the interaction mechanisms between the nanoparticles and trypsin. The root mean square deviation (RMSD) of Cα atoms results have shown that in the presence of the nanoparticles, trypsin was stable. The simulation and the calculation of the binding free energy demonstrate that the nonpolar interactions are the most important forces for the formation of stable nanoparticle-trypsin complex. This study has explicitly elucidated that the nanoparticles have not considerable effect on the trypsin. Copyright © 2017. Published by Elsevier B.V.

  5. FRET-based modified graphene quantum dots for direct trypsin quantification in urine

    Energy Technology Data Exchange (ETDEWEB)

    Poon, Chung-Yan; Li, Qinghua [Department of Chemistry, Hong Kong Baptist University, Kowloon Tong, Hong Kong Special Administrative Region (Hong Kong); Zhang, Jiali; Li, Zhongping [Department of Chemistry, Hong Kong Baptist University, Kowloon Tong, Hong Kong Special Administrative Region (Hong Kong); Research Center of Environmental Science and Engineering, School of Chemistry and Chemical Engineering, Shanxi University, Taiyuan 030006 (China); Dong, Chuan [Research Center of Environmental Science and Engineering, School of Chemistry and Chemical Engineering, Shanxi University, Taiyuan 030006 (China); Lee, Albert Wai-Ming; Chan, Wing-Hong [Department of Chemistry, Hong Kong Baptist University, Kowloon Tong, Hong Kong Special Administrative Region (Hong Kong); Li, Hung-Wing, E-mail: hwli@hkbu.edu.hk [Department of Chemistry, Hong Kong Baptist University, Kowloon Tong, Hong Kong Special Administrative Region (Hong Kong)

    2016-04-21

    A versatile nanoprobe was developed for trypsin quantification with fluorescence resonance energy transfer (FRET). Here, fluorescence graphene quantum dot is utilized as a donor while a well-designed coumarin derivative, CMR2, as an acceptor. Moreover, bovine serum albumin (BSA), as a protein model, is not only served as a linker for the FRET pair, but also a fluorescence enhancer of the quantum dots and CMR2. In the presence of trypsin, the FRET system would be destroyed when the BSA is digested by trypsin. Thus, the emission peak of the donor is regenerated and the ratio of emission peak of donor/emission peak of acceptor increased. By the ratiometric measurement of these two emission peaks, trypsin content could be determined. The detection limit of trypsin was found to be 0.7 μg/mL, which is 0.008-fold of the average trypsin level in acute pancreatitis patient's urine suggesting a high potential for fast and low cost clinical screening. - Highlights: • A FRET-based biosensor was developed for direct quantification of trypsin. • Fast and sensitive screening of pancreatic disease was facilitated. • The direct quantification of trypsin in urine samples was demonstrated.

  6. In vitro assessment of phthalate acid esters-trypsin complex formation.

    Science.gov (United States)

    Chi, Zhenxing; Zhao, Jing; Li, Weiguo; Araghi, Arash; Tan, Songwen

    2017-10-01

    In this work, interactions of three phthalate acid esters (PAEs), including dimethyl phthalate (DMP), diethyl phthalate (DEP) and dibutyl phthalate (DBP), with trypsin have been studied in vitro, under simulated physiological conditions using multi-spectroscopic techniques and molecular modeling. The results show that these PAEs can bind to the trypsin, forming trypsin-PAEs complexes, mainly via hydrophobic interactions, with the affinity order of DMP > DEP > DBP. Binding to the PAEs is found to result in molecular deformation of trypsin. The modeling results suggest that only DBP can bind with the amino acid residues of the catalytic triad and S1 binding pocket of trypsin, leading to potential competitive enzyme inhibition. Copyright © 2017 Elsevier Ltd. All rights reserved.

  7. Identification and characterization of digestive serine proteases from inhibitor-resistant Helicoverpa zea larval midgut

    NARCIS (Netherlands)

    Volpicella, M.; Cordewener, J.H.G.; Jongsma, M.A.; Gallerani, R.; Ceci, L.R.; Beekwilder, M.J.

    2006-01-01

    Protease inhibitors mediate a natural form of plant defence against insects, by interfering with the digestive system of the insect. In this paper, affinity chromatography was used to isolate trypsins and chymotrypsins from Helicoverpa zea larvae, which had been raised on inhibitor-containing diet.

  8. [Delayed rupture of the spleen in a multiply injured patient].

    Science.gov (United States)

    Lică, I; Venter, M D; Mehic, R; Marian, R; Ionescu, G

    1997-01-01

    The authors present a case of delayed rupture of the spleen in a polytraumatised patient. This entity was defined as a late occurrence of signs and symptoms attributed to splenic injury not detected by diagnostic computed tomographic scanning during the initial examination. The mechanisms in which the delayed rupture of the spleen occurs are discussed and the conclusion is that the delayed rupture of the spleen represent a real clinical entity.

  9. Squash inhibitor family of serine proteinases

    International Nuclear Information System (INIS)

    Otlewski, J.; Krowarsch, D.

    1996-01-01

    Squash inhibitors of serine proteinases form an uniform family of small proteins. They are built of 27-33 amino-acid residues and cross-linked with three disulfide bridges. The reactive site peptide bond (P1-P1') is between residue 5 (Lys, Arg or Leu) and 6 (always Ile). High resolution X-ray structures are available for two squash inhibitors complexed with trypsin. NMR solution structures have also been determined for free inhibitors. The major structural motif is a distorted, triple-stranded antiparallel beta-sheet. A similar folding motif has been recently found in a number of proteins, including: conotoxins from fish-hunting snails, carboxypeptidase inhibitor from potato, kalata B1 polypeptide, and in some growth factors (e.g. nerve growth factor, transforming growth factor β2, platelet-derived growth factor). Squash inhibitors are highly stable and rigid proteins. They inhibit a number of serine proteinases: trypsin, plasmin, kallikrein, blood clotting factors: X a and XII a , cathepsin G. The inhibition spectrum can be much broadened if specific amino-acid substitutions are introduced, especially at residues which contact proteinase. Squash inhibitors inhibit proteinases via the standard mechanism. According to the mechanism, inhibitors are substrates which exhibit at neutral pH a high k cat /K m index for hydrolysis and resynthesis of the reactive site, and a low value of the hydrolysis constant. (author)

  10. Wandering spleen: a medical enigma, its natural history and rationalization.

    Science.gov (United States)

    Magowska, Anita

    2013-03-01

    Wandering spleen is a rare condition in which the spleen is not located in the left upper quadrant but is found lower in the abdomen or in the pelvic region because of the laxity of the peritoneal attachments. Many patients with wandering spleen are asymptomatic, hence the condition can be discovered only by abdominal examination or at a hospital emergency department if a patient is admitted to hospital because of severe abdominal pain, vomiting or obstipation. This article aims to provide a historical overview of wandering spleen diagnostics and surgical treatment supplemented with an analyses of articles on wandering spleen included in the PubMed database. One of the first clinical descriptions of a wandering spleen was written by Józef Dietl in 1854. The next years of vital importance are 1877 when A. Martin conducted the first splenectomy and in 1895 when Ludwik Rydygier carried out the first splenopexy to immobilize a wandering spleen. Since that time various techniques of splenectomy and splenopexy have been developed. Introducing medical technologies was a watershed in the development and treatment of wandering spleen, which is confirmed by the PubMed database. Despite the increased number of publications medical literature shows that a wandering spleen still remains a misdiagnosed condition, especially among children.

  11. An Unusual Reason for Gastric Variceal Hemorrhage: Wandering Spleen.

    Science.gov (United States)

    Köseoğlu, Hüseyin; Atalay, Roni; Büyükaşık, Naciye Şemnur; Canyiğit, Murat; Özer, Mehmet; Solakoğlu, Tevfik; Akın, Fatma Ebru; Bolat, Aylin Demirezer; Yürekli, Öykü Tayfur; Ersoy, Osman

    2015-12-01

    Wandering spleen is the displacement of the spleen due to the loss or weakening of the ligaments of the spleen and is seen very rarely with an incidence of less than 0.5 %. It can cause portal hypertension, but gastric variceal hemorrhage is a quite rare condition within the spectrum of this uncommon disease. We report a 22-year-old woman with wandering spleen presenting with life-threatening gastric variceal hemorrhage. Her diagnosis was made by computerized tomography. Endoscopic therapy was not adequate to stop the bleeding, and urgent splenectomy was performed. After surgery she has been well with no symptoms until now.

  12. Identification of a trypsin-like site associated with acetylcholinesterase by affinity labelling with (/sup 3/H)diisopropyl fluorophosphate

    Energy Technology Data Exchange (ETDEWEB)

    Small, D.H.; Chubb, I.W.

    1988-07-01

    In addition to its ability to hydrolyze acetylcholine, purified eel acetylcholinesterase possesses a trypsin-like endopeptidase activity. The tryptic activity is associated with a serine residue at a site that is distinct from the esteratic site. To label both the esteratic and tryptic sites, the enzyme was incubated with the serine hydrolase inhibitor (/sup 3/H)diisopropyl fluorophosphate. This compound labelled the protein in a biphasic manner, with both slow and rapid labelling kinetics. The time course of the rapid phase was similar to the time course of inactivation of the esteratic activity. The time course of the slow phase was similar to the time course of inactivation of the tryptic activity. Labelling of the nonesteratic site was inhibited by the trypsin inhibitor N alpha-p-tosyl-L-lysine chloromethyl ketone. The total number of sites labelled by (/sup 3/H)diisopropyl fluorophosphate on eel acetylcholinesterase was 2.6 mol/280,000 g protein, whereas the number of tryptic sites was less (0.52 mol/280,000 g). The results suggest that a subpopulation of acetylcholinesterase molecules may possess tryptic activity. Extensive chromatography of the purified enzyme by ion-exchange and gel filtration failed to separate the labelled tryptic component from acetylcholinesterase. On sodium dodecyl sulfate-polyacrylamide gels, the labelled tryptic component comigrated with a polypeptide of 50,000 molecular weight, which is a major proteolytic digestion product derived from the intact acetylcholinesterase monomer. Because of its localization in many noncholinergic peptide-containing cells, acetylcholinesterase could act as a neuropeptide processing enzyme in these cells.

  13. Effects of protease inhibitors on radiation transformation in vitro

    International Nuclear Information System (INIS)

    Kennedy, A.R.; Little, J.B.

    1981-01-01

    We have investigated the effects of three protease inhibitors, antipain, leupeptin, and soybean trypsin inhibitor, on the induction of oncogenic transformation in mouse C3H10T 1/2 cells by X-rays. The patterns of inhibition by the three protease inhibitors were different. Antipain was the most effective, having the ability to suppress completely radiation transformation as well as radiation transformation enhanced by the phorbol ester promoting agent 12-O-tetradecanoylphorbol-13-acetate. The fact that antipain could suppress transformation when present for only 1 day following irradiation suggests that an effect on a DNA repair process might be important in its action. Leupeptin was less effective than antipain in its inhibition of radiation transformation. Soybean trypsin inhibitor suppressed only the promotional effects of 12-O-tetradecanoylphorbol-13-acetate on transformation. Our results suggest that there may be more than one protease involved in carcinogenesis

  14. Salmon trypsin stimulates the expression of interleukin-8 via protease-activated receptor-2

    International Nuclear Information System (INIS)

    Larsen, Anett K.; Seternes, Ole-Morten; Larsen, Merethe; Aasmoe, Lisbeth; Bang, Berit

    2008-01-01

    In this study, we focus on salmon trypsin as an activator of inflammatory responses in airway cells in vitro. The rationale behind the investigation is that salmon industry workers are exposed to aerosols containing enzymes, which are generated during industrial processing of the fish. Knowing that serine proteases such as trypsin are highly active mediators with diverse biological activities, the stimulation of nuclear factor-kappa B (NF-κB) and interleukin (IL)-8 and the role of protease-activated receptors (PAR) in inflammatory signal mediation were investigated. Protease-activated receptors are considered important under pathological situations in the human airways, and a thorough understanding of PAR-induced cellular events and their consequences in airway inflammation is necessary. Human airway epithelial cells (A549) were exposed to trypsin isolated from fish (Salmo salar), and we observed that purified salmon trypsin could generate secretion of IL-8 in a concentration-dependent manner. Furthermore, we demonstrate that PAR-2 activation by salmon trypsin is coupled to an induction of NF-κB-mediated transcription using a PAR-2 transfected HeLa cell model. Finally, we show that the release of IL-8 from A549 following stimulation with purified salmon trypsin is mediated through activation of PAR-2 using specific small interfering RNAs (siRNAs). The results presented suggest that salmon trypsin, via activation of PAR-2, might influence inflammation processes in the airways if inhaled in sufficient amounts

  15. In vitro and in silico investigations of the binding interactions between chlorophenols and trypsin

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Yan-Qing, E-mail: wyqing76@126.com [Jiangsu Provincial Key Laboratory of Coastal Wetland Bioresources and Environmental Protection, Yancheng City 224002, Jiangsu Province (China); Institute of Applied Chemistry and Environmental Engineering, Yancheng Teachers University, Yancheng City 224002, Jiangsu Province (China); Tan, Chun-Yun [Institute of Applied Chemistry and Environmental Engineering, Yancheng Teachers University, Yancheng City 224002, Jiangsu Province (China); Zhuang, Shu-Lin [Institute of Environmental Science, College of Environmental and Resource Science, Zhejiang University, Hangzhou 310058 (China); Zhai, Peng-Zhan; Cui, Yun; Zhou, Qiu-Hua; Zhang, Hong-Mei [Institute of Applied Chemistry and Environmental Engineering, Yancheng Teachers University, Yancheng City 224002, Jiangsu Province (China); Fei, Zhenghao [Jiangsu Provincial Key Laboratory of Coastal Wetland Bioresources and Environmental Protection, Yancheng City 224002, Jiangsu Province (China); Institute of Applied Chemistry and Environmental Engineering, Yancheng Teachers University, Yancheng City 224002, Jiangsu Province (China)

    2014-08-15

    Graphical abstract: - Highlights: • Binding interactions of five chlorophenols with trypsin were investigated. • The number of chlorine atoms of chlorophenols partly affected the binding ability of them to trypsin. • Noncovalent interactions stabilized the trypsin–chlorophenols complexes. • There was the one main binding site of trypsin for chlorophenols. - Abstract: Being the first-degree toxic pollutants, chlorophenols (CP) have potential carcinogenic and mutagenic activity and toxicity. Since there still lacks studies on molecular interactions of chlorophenols with trypsin, one major binding target of many exogenous environmental pollutants, the binding interactions between five chlorophenols, 2-CP, 2,6-DCP, 2,4,6-TCP, 2,4,6-TCP, 2,3,4,6-TCP and PCP and trypsin were characterized by the combination of multispectroscopic techniques and molecular modeling. The chlorophenols bind at the one main site of trypsin and the binding induces the changes of microenvironment and global conformations of trypsin. Different number of chloride atoms significantly affects the binding and the binding constants K{sub A} ranks as K{sub A} (2-CP) < K{sub A} (2,6-DCP) ≈ K{sub A} (2,4,6-TCP) < K{sub A} (2,3,4,6-TCP) < K{sub A} (PCP). These chlorophenols interacts with trypsin mainly through hydrophobic interactions and via hydrogen bonding interactions and aromatic–aromatic π–π stacking interaction. Our results offer insights into the binding mechanism of chlorophenols with trypsin and provide important information for possible toxicity risk of chlorophenols to human health.

  16. Study of chicken liver and spleen by Moessbauer spectroscopy

    Energy Technology Data Exchange (ETDEWEB)

    Oshtrakh, M. I., E-mail: oshtrakh@mail.utnet.ru [Ural State Technical University-UPI, Division of Applied Biophysics, Faculty of Physical Techniques and Devices for Quality Control (Russian Federation); Milder, O. B.; Semionkin, V. A. [Ural State Technical University-UPI, Faculty of Experimental Physics (Russian Federation); Malakheeva, L. I. [Simbio Holding, Science Consultation Department (Russian Federation); Prokopenko, P. G. [Russian State Medical University, Faculty of Biochemistry (Russian Federation)

    2005-09-15

    A preliminary study of purified normal human liver ferritin, normal chicken liver and spleen tissues in lyophilized form showed differences in room temperature Moessbauer hyperfine parameters. An additional study of liver and spleen tissues with lower iron content from chicken with lymphoid leukemia indicated small differences between the quadrupole splittings in these samples compared with those in normal tissues.

  17. Study of chicken liver and spleen by Moessbauer spectroscopy

    International Nuclear Information System (INIS)

    Oshtrakh, M. I.; Milder, O. B.; Semionkin, V. A.; Malakheeva, L. I.; Prokopenko, P. G.

    2005-01-01

    A preliminary study of purified normal human liver ferritin, normal chicken liver and spleen tissues in lyophilized form showed differences in room temperature Moessbauer hyperfine parameters. An additional study of liver and spleen tissues with lower iron content from chicken with lymphoid leukemia indicated small differences between the quadrupole splittings in these samples compared with those in normal tissues.

  18. Evolution of the CT imaging findings of accessory spleen infarction

    International Nuclear Information System (INIS)

    Mendi, Resham; Abramson, Lisa P.; Pillai, Srikumar B.; Rigsby, Cynthia K.

    2006-01-01

    We report the case of a 12-year-old girl presenting with multiple episodes of left upper-quadrant pain caused by torsion of an accessory spleen. We present the CT findings of progression of accessory spleen infarction over the course of 7 days. (orig.)

  19. The wandering spleen: CT findings and possible pitfalls in diagnosis

    Energy Technology Data Exchange (ETDEWEB)

    Ben Ely, A.; Zissin, R.; Copel, L.; Vasserman, M.; Hertz, M.; Gottlieb, P.; Gayer, G

    2006-11-15

    Aim: To report the CT features of wandering spleen, a rare condition which can be incidentally detected as an abdominal or pelvic mass or can present with torsion, causing an acute abdomen. Materials and methods: The CT studies of seven patients, two children and five adults, with wandering spleen were reviewed. CT was performed urgently in three patients for acute abdomen, and electively in four. Results: CT findings of wandering spleen included absence of the spleen in its normal position and a mass located elsewhere in the abdomen or pelvis, i.e. an ectopic spleen, enhancing homogeneously in four cases and failing partially or completely to enhance in the other three, indicating infarction. A 'whirl' appearance representing the twisted splenic pedicle was seen in the three cases with torsion. Urgent splenectomy confirmed infarction secondary to torsion. Conclusion: The possible diagnosis of wandering spleen should be kept in mind when CT shows the spleen to be absent from its usual position and a mass is found elsewhere in the abdomen or pelvis. When, in addition, a 'whirl' or partial or no enhancement of this mass are seen in a case presenting with acute abdomen, torsion of a wandering spleen is a likely diagnosis.

  20. Analogues of Cucurbita maxima trypsin inhibitor III (CMTI-III) with elastase inhibitory activity.

    Science.gov (United States)

    Rózycki, J; Kupryszewski, G; Rolka, K; Ragnarsson, U; Zbyryt, T; Krokoszyńska, I; Wilusz, T

    1994-04-01

    Three new CMTI-III analogues containing the Val residue in the reactive site (position 5) were synthesized by the solid-phase method. The analogues displayed an elastase inhibitory activity. It is shown that the removal of the N-terminal Arg residue and the introduction of the Gly-Pro-Gln tripeptide in the region 23-25 decreases the antielastase activity by two orders of magnitude. The removal of the disulfide bridge in positions 16-28 and the substitution of Ala for Cys16 and Gly for Cys28 decreases the activity (measured as Ka with HLE) by five orders of magnitude as compared with [Val5]CMTI-III.

  1. Genus .i.Nostoc./i. - a source of novel trypsin inhibitors

    Czech Academy of Sciences Publication Activity Database

    Drápalová, P.; Štys, D.; Lukešová, Alena; Kopecký, Jiří

    2008-01-01

    Roč. 127, č. 1 (2008), s. 61-82 ISSN 1864-1318 R&D Projects: GA MŠk ME 874 Institutional research plan: CEZ:AV0Z60660521; CEZ:AV0Z50200510 Keywords : cyanobacteria * LC/MS * Nostoc Subject RIV: EH - Ecology, Behaviour

  2. Toxic effects of Ricinus communis non-protein trypsin inhibitor on ...

    African Journals Online (AJOL)

    AFRICAN JOURNALS ONLINE (AJOL) · Journals · Advanced Search · USING AJOL ... Digestibility (AD), Efficiency of Ingested Food Conversion (EIC), Efficiency of Digested Food Conversion (EDC) and the Metabolic Cost (100 - EDC).

  3. Study of ultrasonic imagine of spleen in patients with leukemia

    International Nuclear Information System (INIS)

    Zheng Hui; Zhou Chunyan; Jiang Ju; Luo Liying; Huang Yanhong

    2011-01-01

    To investigate spleen ultrasonic imagine in patients with leukemia and to provide basis information for preventing and treat disease,the spleens imaging of 158 patients with leukemia were detected by B mode ultrasonicgraphy and the data of clinical medical examination were analyzed.The results showed that the spleens' ultrasonic imagine of patients with leukemia were not related to the degree of anemia.The ultrasonic imagines of spleen in patients with chronic leukemia were different to the other kinds of leukemia.The ultrasonic imagine of spleens in leukemia patients are related to types and development of leukemia.The B-ultrasound screening should be used to help clinical diagnosis and treatment of patients with leukemia. (authors)

  4. Human and bovine spinal disc mechanics subsequent to trypsin injection.

    Science.gov (United States)

    Alsup, Jeremy; Bishop, Timothy; Eggett, Dennis; Bowden, Anton E

    2017-10-01

    To investigate the biomechanical effects of injections of a protease on the characteristics of bovine coccygeal and human lumbar disc motion segments. Mechanics of treated tissues were measured immediately after injection and 3 h after injection. Motion segments underwent axial rotation and flexion-extension loading. Stiffness and neutral zone parameters experienced significant changes over time, with bovine tissues more strongly affected than human cadaver tissues. This was true in both axial rotation and flexion-extension. The treatment type significantly affected the neutral zone measurements in axial rotation. Hysteresis parameters were impacted by control injections. The extrapolation of bovine coccygeal motion testing results to human lumbar disc mechanics is not yet practical. The injected treatment may have a smaller impact on disc mechanics than time in testing. Viscoelasticity of human lumbar discs may be impacted by any damage to the annulus fibrosis induced by needlestick. Preclinical testing of novel spinal devices is essential to the design validation and regulatory processes, but current testing techniques rely on cadaveric testing of primarily older spines with essentially random amounts of disc degeneration. The present work investigates the viability of using trypsin injections to create a more uniform preclinical model of disc degeneration from a mechanics perspective, for the purpose of testing spinal devices. Such a model would facilitate translation of new spinal technologies to clinical practice.

  5. The use of poly(ethylene terephthalate)-poly(aniline) composite for trypsin immobilisation

    Energy Technology Data Exchange (ETDEWEB)

    Caramori, S.S. [Laboratorio de Quimica de Proteinas, Departamento de Bioquimica e Biologia Molecular, Instituto de Ciencias Biologicas, Universidade Federal de Goias, Cx. Postal 131, 74001-970 Goiania-GO (Brazil)], E-mail: samanthabio@hotmail.com; Fernandes, K.F. [Laboratorio de Quimica de Proteinas, Departamento de Bioquimica e Biologia Molecular, Instituto de Ciencias Biologicas, Universidade Federal de Goias, Cx. Postal 131, 74001-970 Goiania-GO (Brazil)], E-mail: katia@icb.ufg.br

    2008-08-01

    This paper presents trypsin immobilisation on strips of poly(ethylene terephthalate)-poly(aniline), activated with glutaraldehyde (PET-PANIG) composite. The photomicrography of the material showed changes corresponding to the chemical modifications produced in the steps of synthesis. The immobilisation process was very efficient under optimal conditions (18.6%). The immobilised and free enzyme presented the same pH and temperature optimum. PET-PANIG-trypsin was able to hydrolyse casein, albumin, gelatine, and skimmed milk. Km{sub app} value for PET-PANIG-trypsin was very close to Km of the free enzyme for casein. Immobilised trypsin showed higher stability than the free enzyme, with 100% activity after 14 days of storage at 4 deg. C and 100% operational stability after 4 cycles of use.

  6. Bioconjugation of trypsin onto gold nanoparticles: Effect of surface chemistry on bioactivity

    International Nuclear Information System (INIS)

    Hinterwirth, Helmut; Lindner, Wolfgang; Lämmerhofer, Michael

    2012-01-01

    Highlights: ► Size and spacer affect bioactivity of nanoparticulate trypsin reactor. ► Increase of GNP's size increases activity of bound trypsin. ► Increase of spacer length increases amount and activity of immobilized enzyme by factor 6. ► Decrease of digestion time up to less than 1 h when trypsin immobilized onto GNPs. ► Reduced auto-digestion compared to trypsin in-solution. - Abstract: The systematic study of activity, long-time stability and auto-digestion of trypsin immobilized onto gold nanoparticles (GNPs) is described in this paper and compared to trypsin in-solution. Thereby, the influence of GNP's size and immobilization chemistry by various linkers differing in lipophilicity/hydrophilicity and spacer lengths was investigated with regard to the bioactivity of the conjugated enzyme. GNPs with different sizes were prepared by reduction and simultaneous stabilization with trisodium citrate and characterized by UV/vis spectra, dynamic light scattering (DLS), ζ-potential measurements and transmission electron microscopy (TEM). GNPs were derivatized by self-assembling of bifunctional thiol reagents on the nanoparticle (NP) surface via dative thiol-gold bond yielding a carboxylic acid functionalized surface. Trypsin was either attached directly via hydrophobic and ionic interactions onto the citrate stabilized GNPs or immobilized via EDC/NHS bioconjugation onto the carboxylic functionalized GNPs, respectively. The amount of bound trypsin was quantified by measuring the absorbance at 280 nm. The activity of bound enzyme and its Michaelis Menten kinetic parameter K m and v max were measured by the standard chromogenic substrate N α -Benzoyl-DL-arginine 4-nitroanilide hydrochloride (BApNA). Finally, digestion of a standard protein mixture with the trypsin-conjugated NPs followed by analysis with LC–ESI-MS and successful MASCOT search demonstrated the applicability of the new heterogenous nano-structured biocatalyst. It could be shown that the

  7. Bioconjugation of trypsin onto gold nanoparticles: Effect of surface chemistry on bioactivity

    Energy Technology Data Exchange (ETDEWEB)

    Hinterwirth, Helmut; Lindner, Wolfgang [Department of Analytical Chemistry, University of Vienna, Waehringerstrasse 38, 1090 Vienna (Austria); Laemmerhofer, Michael, E-mail: michael.laemmerhofer@uni-tuebingen.de [Department of Analytical Chemistry, University of Vienna, Waehringerstrasse 38, 1090 Vienna (Austria)

    2012-07-06

    Highlights: Black-Right-Pointing-Pointer Size and spacer affect bioactivity of nanoparticulate trypsin reactor. Black-Right-Pointing-Pointer Increase of GNP's size increases activity of bound trypsin. Black-Right-Pointing-Pointer Increase of spacer length increases amount and activity of immobilized enzyme by factor 6. Black-Right-Pointing-Pointer Decrease of digestion time up to less than 1 h when trypsin immobilized onto GNPs. Black-Right-Pointing-Pointer Reduced auto-digestion compared to trypsin in-solution. - Abstract: The systematic study of activity, long-time stability and auto-digestion of trypsin immobilized onto gold nanoparticles (GNPs) is described in this paper and compared to trypsin in-solution. Thereby, the influence of GNP's size and immobilization chemistry by various linkers differing in lipophilicity/hydrophilicity and spacer lengths was investigated with regard to the bioactivity of the conjugated enzyme. GNPs with different sizes were prepared by reduction and simultaneous stabilization with trisodium citrate and characterized by UV/vis spectra, dynamic light scattering (DLS), {zeta}-potential measurements and transmission electron microscopy (TEM). GNPs were derivatized by self-assembling of bifunctional thiol reagents on the nanoparticle (NP) surface via dative thiol-gold bond yielding a carboxylic acid functionalized surface. Trypsin was either attached directly via hydrophobic and ionic interactions onto the citrate stabilized GNPs or immobilized via EDC/NHS bioconjugation onto the carboxylic functionalized GNPs, respectively. The amount of bound trypsin was quantified by measuring the absorbance at 280 nm. The activity of bound enzyme and its Michaelis Menten kinetic parameter K{sub m} and v{sub max} were measured by the standard chromogenic substrate N{sub {alpha}}-Benzoyl-DL-arginine 4-nitroanilide hydrochloride (BApNA). Finally, digestion of a standard protein mixture with the trypsin-conjugated NPs followed by analysis with

  8. Disorder of G2-M Checkpoint Control in Aniline-Induced Cell Proliferation in Rat Spleen.

    Directory of Open Access Journals (Sweden)

    Jianling Wang

    Full Text Available Aniline, a toxic aromatic amine, is known to cause hemopoietic toxicity both in humans and animals. Aniline exposure also leads to toxic response in spleen which is characterized by splenomegaly, hyperplasia, fibrosis and the eventual formation of tumors on chronic in vivo exposure. Previously, we have shown that aniline exposure leads to iron overload, oxidative DNA damage, and increased cell proliferation, which could eventually contribute to a tumorigenic response in the spleen. Despite our demonstration that cell proliferation was associated with deregulation of G1 phase cyclins and increased expression of G1 phase cyclin-dependent kinases (CDKs, molecular mechanisms, especially the regulation of G2 phase and contribution of epigenetic mechanisms in aniline-induced splenic cellular proliferation remain largely unclear. This study therefore, mainly focused on the regulation of G2 phase in an animal model preceding a tumorigenic response. Male Sprague-Dawley rats were given aniline (0.5 mmol/kg/day in drinking water or drinking water only (controls for 30 days, and expression of G2 phase cyclins, CDK1, CDK inhibitors and miRNAs were measured in the spleen. Aniline treatment resulted in significant increases in cell cycle regulatory proteins, including cyclins A, B and CDK1, particularly phosphor-CDK1, and decreases in CDK inhibitors p21 and p27, which could promote the splenocytes to go through G2/M transition. Our data also showed upregulation of tumor markers Trx-1 and Ref-1 in rats treated with aniline. More importantly, we observed lower expression of miRNAs including Let-7a, miR-15b, miR24, miR-100 and miR-125, and greater expression of CDK inhibitor regulatory miRNAs such as miR-181a, miR-221 and miR-222 in the spleens of aniline-treated animals. Our findings suggest that significant increases in the expression of cyclins, CDK1 and aberrant regulation of miRNAs could lead to an accelerated G2/M transition of the splenocytes, and

  9. Accessory spleen compromising response to splenectomy for idiopathic thrombocytopenic purpura

    International Nuclear Information System (INIS)

    Ambriz, P.; Munoz, R.; Quintanar, E.; Sigler, L.; Aviles, A.; Pizzuto, J.

    1985-01-01

    Accessory spleens were sought in 28 patients who had undergone splenectomy for chronic idiopathic thrombocytopenic purpura (ITP), using a variety of techniques. Abdominal scintigraphy with autologous erythrocytes labeled with Tc-99m and opsonized with anit-D IgG (radioimmune method) proved to be most useful, clearly demonstrating one or more accessory spleens in 12 cases (43%). Computed tomography (CT) was also helpful. Four out of five patients demonstrated an increased platelet count following surgery, the effectiveness of which was illustrated by the radioimmune scan. Patients who have had splenectomy for chronic ITP should be scanned using radioimmune techniques and CT to determine whether an accessory spleen is present

  10. [Objective assessment of trauma severity in patients with spleen injuries].

    Science.gov (United States)

    Alekseev, V S; Ivanov, V A; Alekseev, S V; Vaniukov, V P

    2013-01-01

    The work presents an analysis of condition severity of 139 casualties with isolated and combined spleen injuries on admission to a surgical hospital. The assessment of condition severity was made using the traditional gradation and score scale VPH-SP. The degree of the severity of combined trauma of the spleen was determined by the scales ISS. The investigation showed that the scale ISS and VPH-SP allowed objective measurement of the condition severity of patients with spleen trauma. The score assessment facilitated early detection of the severe category of the patients, determined the diagnostic algorithm and the well-timed medical aid.

  11. [Lymphangiomatosis of the spleen. Report of a clinical case].

    Science.gov (United States)

    Talarico, C; Cerasoli, V; Mancini, B; Mulieri, G; Cancellario D'Alena, F; Montemurro, L; Verna, F

    2000-01-01

    Lymphangiomatosis confined to the spleen is a very are condition. The authors in this article describes one new case and briefly reviews the literature. In this case, after the exclusion of an hydatidosis of the spleen, a total splenectomy was performed. The histologic findings confirmed the lymphangiomatosis of the spleen. The authors emphasize the surgical strategy in splenic lymphangiomyomatosis, infact the total splenectomy is mandatory, because the splenic parenchyma is nearly completely substitute by the cysts. For this reason is preferably, before surgery, to perform the antibateric profilaxis against the OPSI.

  12. Protease purification and characterization of a serine protease inhibitor from Egyptian varieties of soybean seeds and its efficacy against Spodoptera littoralis

    Directory of Open Access Journals (Sweden)

    El-latif Ashraf Oukasha Abd

    2015-01-01

    Full Text Available Serine inhibitors have been described in many plant species and are universal throughout the plant kingdom. Trypsin inhibitors are the most common type. In the present study, trypsin and chymotrypsin inhibitory activity was detected in the seed flour extracts of four Egyptian varieties of soybean (Glycine max. The soybean variety, Giza 22, was found to have higher trypsin and chymotrypsin inhibitory potential compared to other tested soybean varieties. For this reason, Giza 22 was selected for further purification studies which used ammonium sulphate fractionation and DEAE-Sephadex A-25 column. Soybean purified proteins showed a single band on SDS-PAGE corresponding to a molecular mass of 17.9 kDa. The purified inhibitor was stable at temperatures below 60°C and was active at a wide range of pH, from 2 to 12 pH. The kinetic analysis revealed a non-competitive type of inhibition against trypsin and chymotrypsin enzymes. The inhibitor constant (Ki values suggested that the inhibitor has higher affinity toward a trypsin enzyme than to a chymotrypsin enzyme. Purified inhibitor was found to have deep and negative effects on the mean larval weight, larval mortality, pupation, and mean pupal weight of Spodoptera littoralis. It may be concluded, that soybean protease inhibitor gene(s could be potential targets for those future studies which are concerned with developing insect resistant transgenic plants

  13. Isolation, expression and characterization of a novel dual serine protease inhibitor, OH-TCI, from king cobra venom.

    Science.gov (United States)

    He, Ying-Ying; Liu, Shu-Bai; Lee, Wen-Hui; Qian, Jin-Qiao; Zhang, Yun

    2008-10-01

    Snake venom Kunitz/BPTI members are good tools for understanding of structure-functional relationship between serine proteases and their inhibitors. A novel dual Kunitz/BPTI serine proteinase inhibitor named OH-TCI (trypsin- and chymotrypsin-dual inhibitor from Ophiophagus hannah) was isolated from king cobra venom by three chromatographic steps of gel filtration, trypsin affinity and reverse phase HPLC. OH-TCI is composed of 58 amino acid residues with a molecular mass of 6339Da. Successful expression of OH-TCI was performed as the maltose-binding fusion protein in E. coli DH5alpha. Much different from Oh11-1, the purified native and recombinant OH-TCI both had strong inhibitory activities against trypsin and chymotrypsin although the sequence identity (74.1%) between them is very high. The inhibitor constants (K(i)) of recombinant OH-TCI were 3.91 x 10(-7) and 8.46 x10(-8)M for trypsin and chymotrypsin, respectively. To our knowledge, it was the first report of Kunitz/BPTI serine proteinase inhibitor from snake venom that had equivalent trypsin and chymotrypsin inhibitory activities.

  14. The Spleen Revisited: An Overview on Magnetic Resonance Imaging

    Directory of Open Access Journals (Sweden)

    João Palas

    2013-01-01

    Full Text Available Despite being well visualized by different cross-sectional imaging techniques, the spleen is many times overlooked during the abdominal examination. The major reason is the low frequency of splenic abnormalities, the majority consisting of incidental findings. There has been a steady increase in the number of performed abdominal magnetic resonance imaging (MRI studies; therefore, it is important to be familiar to the major MRI characteristics of disease processes involving the spleen, in order to interpret the findings correctly, reaching whenever possible the appropriate diagnosis. The spleen may be involved in several pathologic conditions like congenital diseases, trauma, inflammation, vascular disorders and hematologic disorders, benign and malignant tumors, and other disease processes that focally or diffusely affect the spleen. This paper presents a description and representative MRI images for many of these disorders.

  15. Sonographic determination of spleen to left kidney ratio among Igbo ...

    African Journals Online (AJOL)

    The weight and height of the subjects were obtained with the participants wearing light ... Results: Measurement of spleen and left kidney lengths were reliable within and between .... obtained in the coronal plane passing through the renal.

  16. RNA Sequence of Spleen of Newcastle Disease Infected Chickens

    Data.gov (United States)

    US Agency for International Development — At 21 days of age, chickens were infected with Newcastle Disease virus (or a mock injection as controls), and spleens were harvested at 2 and 6 days post infection....

  17. Effect of low dose radiation on apoptosis in mouse spleen

    International Nuclear Information System (INIS)

    Chen Dong; Liu Jiamei; Chen Aijun; Liu Shuzheng

    1999-01-01

    Objective: To study the effect of whole body irradiation (WBI) with different doses of X-ray on apoptosis in mouse spleen. Methods: Time course changes and dose-effect relationship of apoptosis in mouse spleen induced by WBI were observed with transmission electron microscopy (TEM) qualitatively and TUNEL method semi-quantitatively. Results: Many typical apoptotic lymphocytes were found by TEM in mouse spleen after WBI with 2 Gy. No marked alterations of ultrastructure were found following WBI with 0.075 Gy. It was observed by TUNEL that the apoptosis of splenocytes increased after high dose radiation and decreased following low dose radiation (LDR). The dose-effect relationship of radiation-induced apoptosis showed a J-shaped curve. Conclusion: The effect of different doses of ionizing radiation on apoptosis in mouse spleen was distinct. And the decrease of apoptosis after LDR is considered a manifestation of radiation hormesis

  18. Gastric volvulus associated with wandering spleen in adult: Case report

    Directory of Open Access Journals (Sweden)

    Oktay Büyükaşık

    2012-07-01

    Full Text Available Gastric volvulus associated with wandering spleen is a rare diagnosis in adult ages. So far, only two cases have been reported in the literature. 82 year old male patients admitted to emergency department with complaint of nausea, vomiting and constipation. Physical examination and computerize tomography detected a big solid mass with regular contour which is full filling abdominal left lower quadrant. Exploratory laparotomy revealed a wandering spleen in sizes of 13x13x15 cm in the mentioned region. The spleen which had two masses on was partially ischemic. The stomach had rotated through cardiopyloric axis due to long pedicle of the spleen and adhesions neighborhood to corpoantral junction. Thus gastric passage was partialy obstructed. Splenectomy and anterior gastropexy were applied. The patient was discharged in health at 6th day postoperatively.

  19. Torsion of a Wandering Spleen Presenting as Acute Abdomen

    International Nuclear Information System (INIS)

    Chauhan, Narvir Singh; Kumar, Satish

    2016-01-01

    Wandering spleen is a rare condition which if uncorrected, can result in torsion and infarction. Clinical presentation of a wandering spleen can vary from asymptomatic abdominal mass to acute abdominal pain. Radiological investigations play a pivotal role in diagnosis as the clinical diagnosis is usually impossible. We present a case of wandering spleen with torsion and complete infarction that occurred in a 32-year-old multiparous female. The diagnosis was established preoperatively on colour Doppler and CT of the abdomen with subsequent confirmation on surgery. Wandering spleen is a rare clinical condition which can present as acute abdomen. An increased awareness of this entity together with the timely use of ultrasound and CT of the abdomen can play an important role in preoperative diagnosis and surgical management

  20. Modification of radioresponse of chick spleen with vitamin E treatment

    International Nuclear Information System (INIS)

    Rana, K.; Malhotra, N.

    1995-01-01

    Seven days old white leghorn male chicks were exposed to 2.25 Gy whole body gamma radiations with and without vitamin E and studied for histopathological changes in the spleen for a period of twenty eight days postirradiation. The results reveal that the radiation-induced depletion of lymphocytic population in the lymphoid region and the damage to the tissue architecture is comparatively less and reparation of the spleen faster in the vitamin E treated irradiated chicks. (author). 12 refs., 9 figs

  1. Investigation of the human spleen by X-ray microanalysis

    International Nuclear Information System (INIS)

    Kopani, M.; Jakubovsky, J.; Polak, S.

    2001-01-01

    Qualitative and quantitative topographic analysis using X-ray fluorescence (XRF), X-ray powder diffraction (XRD) and scanning electron microscopy was performed in tissue samples of rat and human spleens. The presence of silico-aluminium and silico-calcareous particles of various sizes could be seen. The presence of the inorganic substances mentioned in the human red pulp cords is assumed to be a consequence of the purifying function of the spleen. (Authors)

  2. Torsion of wandering spleen in patient with horseshoe kidney

    International Nuclear Information System (INIS)

    Molski, St.; Zurada, A.; Meder, G.; Lasek, W.

    2005-01-01

    Wandering spleen is rare pathology, mostly occurring in young women. Disease may be congenital or acquired. Absence or laxity of ligaments leads to spleen pathologic mobility and may cause torsion of its pedicle, resulting in ischemia or infarct even haemorrhagic shock and patients death. We report a case of young woman previously diagnosed (and treated nonoperative) with wandering spleen who presented acute abdomen after minor blunt trauma. She was evaluated with abdominal ultrasound (US) and spiral computed tomography (CT). Torsion of splenic pedicle and splenic rupture was diagnosed and a horseshoe kidney as well. Laparotomy followed by splenectomy confirmed the existence of an intrapelvic torsioned wandering spleen. The only definitive treatment of wandering spleen is operative since nonoperative treatment is associated with high complication rate. Earlier diagnosis of wandering spleen in asymptomatic patients lets to direct diagnosis when patient starts to present with acute abdomen. CT and abdominal US play most important role in diagnosing splenic pedicle torsion. To our knowledge this is a first case of torsion of splenic pedicle in patient with horseshoe kidney. We consider this coincidence to be a congenital defect as both conditions may develop in second month gestation. (author)

  3. Percutaneous radiofrequency ablation of spleen for the treatment of hypersplenism

    International Nuclear Information System (INIS)

    Wu Yuxuan; Zhang Yanfang; Zheng Xuefen; Zhang Yuanhua; Kong Jian; Shen Xinying; Dou Yongchong

    2009-01-01

    Objective: To summarize the clinical effect and experience of CT-guided radiofrequency ablation (RFA) of spleen by using cool-tip electrodes in the treatment of hypersplenism in patients with liver cirrhosis and portal hypertension. Methods: CT-guided RFA of spleen by using cool-tip electrodes was performed in 15 patients with hypersplenism associated with liver cirrhosis and portal hypertension. The routine blood count was studied both before and after the procedure. Enhanced CT or MR scanning was reexamined after the treatment to determine the ablated volume of the spleen. The results were statistically analyzed. Results: The ablated volume of the spleen accounted for (31.0 ± 4.6)% of the whole spleen. Before the treatment the platelet count was (62 ± 9.8) x 10 9 /L. One month after the treatment, the platelet count was increased to (96 ± 11) x 10 9 /L, which was significantly higher than that before the treatment (P<0.05). One patient developed portal thrombosis four months after RFA, and no other serious complications occurred. Conclusion: CT-guided radiofrequency ablation of spleen by using cool-tip electrodes is an effective and safe treatment for hypersplenism in patients with liver cirrhosis and portal hypertension. (authors)

  4. Functional assessment of hepatocytes after transplantation into rat spleen

    International Nuclear Information System (INIS)

    Woods, R.J.; Fuller, B.J.; Attenburrow, V.D.; Nutt, L.H.; Hobbs, K.E.

    1982-01-01

    The retention of structural integrity and metabolic function by isolated hepatocytes after ectopic transplantation has been investigated in autografted rats. Rats were partially hepatectomized and isolated hepatocytes prepared from the excised liver lobes were implanted into their spleens. Histological examination of the spleens 7 or more weeks after implantation revealed aggregates of hepatocytes in the red pulp. Two tests of biochemical function were applied to the hepatocytes after transplantation. In the first the hepatobiliary imaging agent technetium-99m N-[N'-(2, 6-dimethylphenyl)carbamoylmethyl]iminodiacetic acid (99mTc HIDA), which was shown to be avidly taken up by isolated hepatocytes in vitro, was infused into the tail veins of autograft and control rats. Radioactivity accumulating in the spleens of autografted rats was markedly greater than that in controls implanted with lethally damaged cells or in nontransplanted rats. In the second the presence of bilirubin metabolites was sought in autograft spleens after intravenous infusion of bilirubin. Both mono- and diglucuronides of bilirubin were recovered from the spleens of autograft rats but no conjugates were recovered from the spleens of unoperated controls. We conclude that after autotransplantation isolated hepatocytes retain their morphology and at least some of their functional activities

  5. CT numbers of liver and spleen in normal children

    International Nuclear Information System (INIS)

    Kim, Young Kim

    2002-01-01

    To determine the mean liver CT numbers, and differences between liver and spleen, and liver and back muscle CT numbers in normal children, and to correlate the findings with sex and age. One hundred and five normal children aged 2-14 years underwent pre-contrast CT scanning. Mean CT numbers of the liver, spleen, and back muscles were calculated, as well as the differences in CT numbers between the liver and spleen (liver-spleen CT numbers), and between the liver-back muscle CT numbers were 70.22±6.51 HU, 53.28±3.57 HU, 17.13±6.57 HU, and 11.88±5.94 HU, respectively. Mean liver CT numbers and the difference between liver and back muscle CT numbers were not different by age. By sex, all the CT numbers did not vary according to age. The sex of a subject did not affect the CT number. The children's mean liver CT number was 70.22±6.51 HU and the difference between liver and spleen CT numbers was 17.13±6.57 HU. Younger children had higher liver CT and liver-spleen CT numbers than older children. No CT numbers varied according to sex

  6. [Torsion of wandering spleen in a teenager: about a case].

    Science.gov (United States)

    Dème, Hamidou; Akpo, Léra Géraud; Fall, Seynabou; Badji, Nfally; Ka, Ibrahima; Guèye, Mohamadou Lamine; Touré, Mouhamed Hamine; Niang, El Hadj

    2016-01-01

    Wandering or migrating spleen is a rare anomaly which is usually described in children. Complications, which include pedicle torsion, are common and can be life-threatening. We report the case of a 17 year-old patient with a long past medical history of epigastric pain suffering from wandering spleen with chronic torsion of the pedicle. The clinical picture was marked by spontaneously painful epigastric mass, evolved over the past 48 hours. Abdominal ultrasound objectified heterogeneous hypertrophied ectopic spleen in epigastric position and a subcapsular hematoma. Doppler showed a torsion of splenic pedicle which was untwisted 2 turns and a small blood stream on the splenic artery. Abdominal CT scan with contrast injection showed a lack of parenchymal enhancement of large epigastric ectopic spleen and a subcapsular hematoma. The diagnosis of wandering spleen with chronic torsion of the pedicle complicated by necrosis and subcapsular hematoma was confirmed. The patient underwent splenectomy. The postoperative course was uneventful. We here discuss the contribution of ultrasound and CT scan in the diagnosis of wandering spleen with chronic torsion of the pedicle.

  7. Assessment and partial purification of serine protease inhibitors from Rhipicephalus (Boophilus annulatuslarvae

    Directory of Open Access Journals (Sweden)

    Sedigheh Nabian

    Full Text Available Ticks are rich sources of serine protease inhibitors, particularly those that prevent blood clotting and inflammatory responses during blood feeding. The tick Rhipicephalus (Boophlus annulatusis an important ectoparasite of cattle. The aims of this study were to characterize and purify the serine protease inhibitors present in R. (B. annulatus larval extract. The inhibitors were characterized by means of one and two-dimensional reverse zymography, and purified using affinity chromatography on a trypsin-Sepharose column. The analysis on one and two-dimensional reverse zymography of the larval extract showed trypsin inhibitory activity at between 13 and 40 kDa. Through non-reducing SDS-PAGE and reverse zymography for proteins purified by trypsin-Sepharose affinity chromatography, some protein bands with molecular weights between 13 and 34 kDa were detected. Western blotting showed that five protein bands at 48, 70, 110, 130 and 250 kDa reacted positively with immune serum, whereas there was no positive reaction in the range of 13-40 kDa. Serine protease inhibitors from R. (B. annulatus have anti-trypsin activity similar to inhibitors belonging to several other hard tick species, thus suggesting that these proteins may be useful as targets in anti-tick vaccines.

  8. Sonographic determination of normal spleen size in an adult African population

    Energy Technology Data Exchange (ETDEWEB)

    Mustapha, Zainab; Tahir, Abdulrahman [Department of Radiology, University of Maiduguri Teaching Hospital, Maiduguri, Borno State (Nigeria); Tukur, Maisaratu [Department of Human Physiology, University of Maiduguri, Maiduguri, Borno State (Nigeria); Bukar, Mohammed [Department of Obstetrics and Gynaecology, University of Maiduguri Teaching Hospital, Maiduguri, Borno State (Nigeria); Lee, Wai-Kit, E-mail: leewk33@hotmail.co [Department of Medical Imaging, St. Vincent' s Hospital, University of Melbourne, 41 Victoria Parade, Fitzroy, Victoria 3065 (Australia)

    2010-07-15

    Objective: The purpose of this study was to determine the normal range of spleen size in an adult African population, and compare the findings to published data to determine any correlation with ethnicity. Materials and methods: Three hundred and seventy-four African adults without conditions that can affect the spleen or splenic abnormalities were evaluated with ultrasonography. Spleen length, width and thickness were measured and spleen volume calculated. Spleen size was correlated with age, gender, height, weight, and body mass index. Results: The mean spleen volume was 120 cm{sup 3}. Spleen volume correlated with spleen width (r = 0.85), thickness (r = 0.83) and length (r = 0.80). Men had a larger mean spleen volume than women. No correlation was found between spleen volume and age, weight, height, or body mass index. Conclusion: Mean spleen volume in African adults is smaller than data from Western sources, and cannot be explained by difference in body habitus.

  9. Molecular glues for manipulating enzymes: trypsin inhibition by benzamidine-conjugated molecular glues† †Electronic supplementary information (ESI) available: Synthesis of TEG–BA, Gluen–BA, mGluen–BA and Gluen–Ph; 1H NMR, 13C NMR, MALDI-TOF MS, electronic absorption, and CD spectra; zeta potential distributions; SLS plots; DLS histograms; and related experimental procedures. See DOI: 10.1039/c5sc00524h Click here for additional data file.

    Science.gov (United States)

    Mogaki, Rina

    2015-01-01

    Water-soluble bioadhesive polymers bearing multiple guanidinium ion (Gu+) pendants at their side-chain termini (Gluen–BA, n = 10 and 29) that were conjugated with benzamidine (BA) as a trypsin inhibitor were developed. The Gluen–BA molecules are supposed to adhere to oxyanionic regions of the trypsin surface, even in buffer, via a multivalent Gu+/oxyanion salt-bridge interaction, such that their BA group properly blocks the substrate-binding site. In fact, Glue10–BA and Glue29–BA exhibited 35- and 200-fold higher affinities for trypsin, respectively, than a BA derivative without the glue moiety (TEG–BA). Most importantly, Glue10–BA inhibited the protease activity of trypsin 13-fold more than TEG–BA. In sharp contrast, mGlue27–BA, which bears 27 Gu+ units along the main chain and has a 5-fold higher affinity than TEG–BA for trypsin, was inferior even to TEG–BA for trypsin inhibition. PMID:28706668

  10. Rapid and Efficient Protein Digestion using Trypsin Coated Magnetic Nanoparticles under Pressure Cycles

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Byoungsoo; Lopez-Ferrer, Daniel; Kim, Byoung Chan; Na, Hyon Bin; Park, Yong Il; Weitz, Karl K.; Warner, Marvin G.; Hyeon, Taeghwan; Lee, Sang-Won; Smith, Richard D.; Kim, Jungbae

    2011-01-01

    Trypsin-coated magnetic nanoparticles (EC-TR/NPs), prepared via a simple crosslinking of the enzyme to magnetic nanoparticles, were highly stable and could be easily captured using a magnet after the digestion was complete. EC-TR/NPs showed a negligible loss of trypsin activity after multiple uses and continuous shaking, while a control sample of covalently-attached trypsin on NPs resulted in a rapid inactivation under the same conditions due to the denaturation and autolysis of trypsin. Digestions were carried out on a single model protein, a five protein mixture, and a whole mouse brain proteome, and also compared for digestion at atmospheric pressure and 37 ºC for 12 h, and in combination with pressure cycling technology (PCT) at room temperature for 1 min. In all cases, the EC-TR/NPs performed equally as well or better than free trypsin in terms of the number of peptide/protein identifications and reproducibility across technical replicates. However, the concomitant use of EC-TR/NPs and PCT resulted in very fast (~1 min) and more reproducible digestions.

  11. Action of trypsin on structural changes of collagen fibres from sea cucumber (Stichopus japonicus).

    Science.gov (United States)

    Liu, Zi-Qiang; Tuo, Feng-Yan; Song, Liang; Liu, Yu-Xin; Dong, Xiu-Ping; Li, Dong-Mei; Zhou, Da-Yong; Shahidi, Fereidoon

    2018-08-01

    Trypsin, a representative serine proteinase, was used to hydrolyse the collagen fibres from sea cucumber (Stichopus japonicus) to highlight the role of serine proteinase in the autolysis of sea cucumber. Partial disaggregation of collagen fibres into collagen fibrils upon trypsin treatment occurred. The trypsin treatment also caused a time-dependent release of water-soluble glycosaminoglycans and proteins. Therefore, the degradation of the proteoglycan bridges between collagen fibrils might account for the disaggregation of collagen fibrils. For trypsin-treated collagen fibres (72 h), the collagen fibrils still kept their structural integrity and showed characteristic D-banding pattern, and the dissolution rate of hydroxyproline was just 0.21%. Meanwhile, Fourier transform infrared analysis showed the collagen within trypsin-treated collagen fibres (72 h) still retaining their triple-helical conformation. These results suggested that serine proteinase participated in the autolysis of S. japonicus body wall by damaging the proteoglycan bridges between collagen fibrils and disintegrating the latter. Copyright © 2018 Elsevier Ltd. All rights reserved.

  12. Purification, partial characterization, and immunological relationships of multiple low molecular weight protease inhibitors of soybean

    Energy Technology Data Exchange (ETDEWEB)

    Hwang, D L.R.; Lin, K T.D.; Yang, W K; Foard, D E

    1977-01-01

    Five protease inhibitors, I-V, in the molecular weight range 7000-8000 were purified from Tracy soybeans by ammonium sulfate precipitation, gel filtration on Sephadex G-100 and G-75, and column chromatography on DEAE-cellulose. In common with previously described trypsin inhibitors from legumes, I-V have a high content of half-cystine and lack tryptophan. By contrast with other legume inhibitors, inhibitor II contains 3 methionine residues. Isoelectric points range from 6.2 to 4.2 in order from inhibitor I to V. Molar ratios (inhibitor/enzyme) for 50% trypsin inhibition are I = 4.76, II = 1.32, III = 3.22, IV = 2.17, V = 0.97. Only V inhibits chymotrypsin significantly (molar ratio = 1.33 for 50% inhibition). The sequence of the first 16 N-terminal amino acid residues of inhibitor V is identical to that of the Bowman-Birk inhibitor; all other observations also indicate that inhibitor V and Bowman-Birk are identical. The first 20 N-terminal amino acid residues of inhibitor II show high homology to those of Bowman-Birk inhibitor, differing by 1 deletion and 5 substitutions. Immunological tests show that inhibitors I through IV are fully cross-reactive with each other but are distinct from inhibitor V.

  13. Detection of five tumor markers in lung cancer by trypsin digestion of sputum method

    International Nuclear Information System (INIS)

    Lin Min; Nong Tianlei; Liu Daying

    2011-01-01

    To explore the detection of five tumor markers by trypsin digestion of sputum in the diagnosis of lung cancer, the samples of sputum in patients with lung cancer and benign lung disease were digested by trypsin and used to measure five tumor markers. The results showed that the sputum were well digested by 6% trypsin at pH8 and no affect on the determination of tumor markers. The CEA, CA125, CA153, CA211 and NSE levels in lung cancer group were significantly higher than that of in benign group (P<0.05). The sputum CEA and CA125 levels were significantly higher than that of the serum levels (P<0.05). The detection of sputum CEA, CA125, CA153, CA211 and NSE levels have clinical value in the diagnosis of lung cancer. When combined with other diagnostic methods,it might be helpful for further diagnosis in non confirmed lung cancer patients. (authors)

  14. Robust Trypsin Coating on Electrospun Polymer Nanofibers in Rigorous Conditions and Its Uses for Protein Digestion

    Energy Technology Data Exchange (ETDEWEB)

    Ahn, Hye-Kyung; Kim, Byoung Chan; Jun, Seung-Hyun; Chang, Mun Seock; Lopez-Ferrer, Daniel; Smith, Richard D.; Gu, Man Bock; Lee, Sang-Won; Kim, Beom S.; Kim, Jungbae

    2010-12-15

    An efficient protein digestion in proteomic analysis requires the stabilization of proteases such as trypsin. In the present work, trypsin was stabilized in the form of enzyme coating on electrospun polymer nanofibers (EC-TR), which crosslinks additional trypsin molecules onto covalently-attached trypsin (CA-TR). EC-TR showed better stability than CA-TR in rigorous conditions, such as at high temperatures of 40 °C and 50 °C, in the presence of organic co-solvents, and at various pH's. For example, the half-lives of CA-TR and EC-TR were 0.24 and 163.20 hours at 40 ºC, respectively. The improved stability of EC-TR can be explained by covalent-linkages on the surface of trypsin molecules, which effectively inhibits the denaturation, autolysis, and leaching of trypsin. The protein digestion was performed at 40 °C by using both CA-TR and EC-TR in digesting a model protein, enolase. EC-TR showed better performance and stability than CA-TR by maintaining good performance of enolase digestion under recycled uses for a period of one week. In the same condition, CA-TR showed poor performance from the beginning, and could not be used for digestion at all after a few usages. The enzyme coating approach is anticipated to be successfully employed not only for protein digestion in proteomic analysis, but also for various other fields where the poor enzyme stability presently hampers the practical applications of enzymes.

  15. CT findings in children with blunt trauma in the spleen

    International Nuclear Information System (INIS)

    Nishiguchi, Hiroyasu; Shimizu, Toshihisa; Ohmura, Makoto; Kawai, Naoki; Tauchi, Hayato; Hayakawa, Masao; Nishio, Yoshinori; Watanabe, Shinsuke.

    1991-01-01

    We evaluated CT findings in 19 children with blunt injuries in the spleen. CT demonstrated laceration of the spleen in 7 children, rupture of the spleen in 7, and splenic hematoma in 5. Leakage of the contrast medium was observed in 3 children, of whom 1 was treated by arterial embolization. Laparotomy was performed in 3 children (15.8%) other than the 3 showing contrast medium leakage; hemostasis by compression was performed in 1 with laceration, and splenectomy in 2 with rupture. Late splenic rupture or abscess did not occur in any child. One child (5.3%) died of complicating injuries. Many of children with blunt splenic injuries can be successfully treated with conservative treatment, and CT scanning is useful for evaluating the degree of splenic injuries and complicating injuries. (author)

  16. CT findings in children with blunt trauma in the spleen

    Energy Technology Data Exchange (ETDEWEB)

    Nishiguchi, Hiroyasu; Shimizu, Toshihisa; Ohmura, Makoto; Kawai, Naoki; Tauchi, Hayato; Hayakawa, Masao; Nishio, Yoshinori (Kyoto Second Red Cross Hospital (Japan)); Watanabe, Shinsuke

    1991-09-01

    We evaluated CT findings in 19 children with blunt injuries in the spleen. CT demonstrated laceration of the spleen in 7 children, rupture of the spleen in 7, and splenic hematoma in 5. Leakage of the contrast medium was observed in 3 children, of whom 1 was treated by arterial embolization. Laparotomy was performed in 3 children (15.8%) other than the 3 showing contrast medium leakage; hemostasis by compression was performed in 1 with laceration, and splenectomy in 2 with rupture. Late splenic rupture or abscess did not occur in any child. One child (5.3%) died of complicating injuries. Many of children with blunt splenic injuries can be successfully treated with conservative treatment, and CT scanning is useful for evaluating the degree of splenic injuries and complicating injuries. (author).

  17. Giant Accessory Right-Sided Suprarenal Spleen in Thalassaemia

    Directory of Open Access Journals (Sweden)

    A. Arra

    2013-01-01

    Full Text Available An accessory spleen is defined as ectopic splenic tissue that develops due to failure of fusion of cells during embryonic development as they migrate from the midline to the left upper quadrant. While benign, complications may arise which include trauma, torsion, or infarction of the ectopic tissue. Additionally, patients who have had a splenectomy secondary to treatment for previous pathology such as a haematological malignancy or idiopathic thrombocytopenia purpura may experience persistent symptoms due to the accessory splenic tissue. The presence of an accessory spleen is therefore of significant diagnostic and therapeutic importance. To the best of the authors' knowledge, this case is the second and largest reported case of a giant right suprarenal accessory spleen and highlights the difficulty in differentiation of these masses from malignant adrenal tumours.

  18. Candidiasis of the liver and spleen in childhood

    International Nuclear Information System (INIS)

    Miller, J.H.; Greenfield, L.D.; Wald, B.R.

    1982-01-01

    Four children with acute leukemia and surgically documented candidiasis of the liver and/or spleen were examined with a combination of diagnostic imaging modalities including /sup 99m/Tc-sulfur colloid and /sup 67/Ga- citrate scintigraphy, gray-scale ultrasound, and body computed tomography (CT). Abnormalities were detected in every individual examined. /sup 99m/Tc-sulfur colloid scintigraphy revealed ''cold'' areas in the liver or spleen. With /sup 67/Ga scintigraphy, these areas were ''cold'' in some individuals and ''hot'' in others. Gray-scale ultrasound demonstrated hypoechoic lesions with central areas of increased echogenicity in hepatic involvement, and hypoechoic replacement of the spleen in splenic involvement. CT in one patient revealed low-density areas without contrast enhancement within the hepatic parenchyma and unsuspected renal involvement

  19. Candidiasis of the liver and spleen in childhood

    International Nuclear Information System (INIS)

    Miller, J.H.; Greenfield, L.D.; Wald, B.R.

    1982-01-01

    Four children with acute leukemia and surgically documented candidiasis of the liver and/or spleen were examined with a combination of diagnostic imaging modalities including 99 mTc-sulfur colloid and 67 Ga-citrate scintigraphy, gray-scale ultrasound, and body computed tomography (CT). Abnormalities were detected in every individual examined. 99 mTc-sulfur colloid scintigraphy revealed cold areas in the liver or spleen. With 67 Ga scintigraphy, these areas were cold in some individuals and hot in others. Gray-scale ultrasound demonstrated hypoechoic lesions with central areas of increased echogenicity in hepatic involvement, and hypoechoic replacement of the spleen in splenic involvement. CT in one patient revealed low-density areas without contrast enhancement within the hepatic parenchyma and unsuspected renal involvement

  20. Candidiasis of the liver and spleen in childhood

    Energy Technology Data Exchange (ETDEWEB)

    Miller, J.H. (Childrens Hospital of Los Angeles, CA); Greenfield, L.D.; Wald, B.R.

    1982-02-01

    Four children with acute leukemia and surgically documented candidiasis of the liver and/or spleen were examined with a combination of diagnostic imaging modalities including /sup 99m/Tc-sulfur colloid and /sup 67/Ga- citrate scintigraphy, gray-scale ultrasound, and body computed tomography (CT). Abnormalities were detected in every individual examined. /sup 99m/Tc-sulfur colloid scintigraphy revealed ''cold'' areas in the liver or spleen. With /sup 67/Ga scintigraphy, these areas were ''cold'' in some individuals and ''hot'' in others. Gray-scale ultrasound demonstrated hypoechoic lesions with central areas of increased echogenicity in hepatic involvement, and hypoechoic replacement of the spleen in splenic involvement. CT in one patient revealed low-density areas without contrast enhancement within the hepatic parenchyma and unsuspected renal involvement.

  1. Candidiasis of the liver and spleen in childhood

    Energy Technology Data Exchange (ETDEWEB)

    Miller, J.H.; Greenfield, L.D.; Wald, B.R.

    1982-02-01

    Four children with acute leukemia and surgically documented candidiasis of the liver and/or spleen were examined with a combination of diagnostic imaging modalities including /sup 99/mTc-sulfur colloid and /sup 67/Ga-citrate scintigraphy, gray-scale ultrasound, and body computed tomography (CT). Abnormalities were detected in every individual examined. /sup 99/mTc-sulfur colloid scintigraphy revealed cold areas in the liver or spleen. With /sup 67/Ga scintigraphy, these areas were cold in some individuals and hot in others. Gray-scale ultrasound demonstrated hypoechoic lesions with central areas of increased echogenicity in hepatic involvement, and hypoechoic replacement of the spleen in splenic involvement. CT in one patient revealed low-density areas without contrast enhancement within the hepatic parenchyma and unsuspected renal involvement.

  2. Semiautomated spleen volumetry with diffusion-weighted MR imaging.

    Science.gov (United States)

    Lee, Jeongjin; Kim, Kyoung Won; Lee, Ho; Lee, So Jung; Choi, Sanghyun; Jeong, Woo Kyoung; Kye, Heewon; Song, Gi-Won; Hwang, Shin; Lee, Sung-Gyu

    2012-07-01

    In this article, we determined the relative accuracy of semiautomated spleen volumetry with diffusion-weighted (DW) MR images compared to standard manual volumetry with DW-MR or CT images. Semiautomated spleen volumetry using simple thresholding followed by 3D and 2D connected component analysis was performed with DW-MR images. Manual spleen volumetry was performed on DW-MR and CT images. In this study, 35 potential live liver donor candidates were included. Semiautomated volumetry results were highly correlated with manual volumetry results using DW-MR (r = 0.99; P volumetry was significantly shorter compared to that of manual volumetry with DW-MR (P volumetry with DW-MR images can be performed rapidly and accurately when compared with standard manual volumetry. Copyright © 2011 Wiley Periodicals, Inc.

  3. Cancer of the colon spleen angle. Presentation of a case

    International Nuclear Information System (INIS)

    Martinez Sanchez, Yariana; De la Rosa Perez, Nereida; Barcelo Casanova, Renato E

    2010-01-01

    The colon cancer is currently an important public health problem in developed countries. It is the fourth most common cancer in the world. We report the case of a 65-years-old, black, female patient, assisting our consultation with dyspeptic disturbances as the unique symptom, without known risk factors. We indicated a colon by enema and a distal narrowing was observed at the colon spleen angle, at the same zone of the physiologic narrowing at that level. A colonoscopy was carried out diagnosing a left colon tumor near the spleen angle. It was operated with segmental resection of the spleen angle and a biopsy was made. Pathologic anatomy informed a well-differentiated colon adenocarcinoma

  4. High-gradient magnetic affinity separation of trypsin from porcine pancreatin

    DEFF Research Database (Denmark)

    Hubbuch, Jürgen; Thomas, Owen R. T.

    2002-01-01

    We introduce a robust and scale-flexible approach to macromolecule purification employing tailor-made magnetic adsorbents and high-gradient magnetic separation technology adapted from the mineral processing industries. Detailed procedures for the synthesis of large quantities of low-cost defined......-scale studies approximate to95% of the endogenous trypsin present in a crude porcine pancreatin feedstock was recovered with a purification factor of approximate to4.1 at the expense of only a 4% loss in a-amylase activity. Efficient recovery of trypsin from the same feedstock was demonstrated at a vastly...

  5. Bowman-Birk inhibitor affects pathways associated with energy metabolism in Drosophila melanogaster

    Science.gov (United States)

    Bowman-Birk inhibitor (BBI) is toxic when fed to certain insects, including the fruit fly, Drosophila melanogaster. Dietary BBI has been demonstrated to slow growth and increase insect mortality by inhibiting the digestive enzymes trypsin and chymotrypsin, resulting in a reduced supply of amino acid...

  6. The forgotten organ: Contrast enhanced sonography of the spleen

    International Nuclear Information System (INIS)

    Goerg, Christian

    2007-01-01

    Objective: Ultrasound contrast agents in conjunction with contrast specific imaging techniques, are increasingly accepted in clinical use for diagnostic imaging in several organs. Contrast enhanced sonography (CES) of second-generation contrast media have shown a spleen-specific uptake of the microbubble contrast agent. The aim of this review is to illustrate indications for the use of CES in patients with suspected (peri-)splenic pathology. Methods: This review based on the experience of transcutaneous CES in 200 patients with (peri-)splenic pathology diagnosed by B-mode sonography at an internal medicine center. CES studies were performed with a contrast-devoted unit (Acuson, Sequoia, Siemens medical solution) that had contrast-specific, continuous-mode software. A low mechanical index was used. A sulfur hexafluoride-based microbubble contrast medium (Sonovue, Bracco SpA, Milan, Italy) was injected. Results: On our experience, there are several clinical conditions which may show an diagnostic advantage of CES in comparison to B-mode US. CES should be performed to investigate: (1) the perisplenic tumor to diagnose or exclude accessory spleen, (2) the small-sized spleen to diagnose functional asplenia/hyposplenia, (3) the inhomogenous spleen of unknown cause to diagnose focal lesions within the spleen, (4) the incidentally found hypoechoic splenic tumor to diagnose high vascular splenic hemangioma, (5) focal lesions suspect for splenic abscess, hematoma, infarction to confirme diagnosis, and (6) patients with abdominal trauma to diagnose or exclude splenic injuriy. Conclusion: CES is of diagnostic value in several clinical circumstances to diagnose accessory spleen, functional asplenia, small-sized splenic involvement, high vascular splenic hemangioma, and vascular splenic pathology like splenic infarction, splenic abscess, and splenic laceration

  7. The forgotten organ: Contrast enhanced sonography of the spleen

    Energy Technology Data Exchange (ETDEWEB)

    Goerg, Christian [Medizinische Universitaetsklinik, Baldingerstrasse, 35043 Marburg/Lahn (Germany)], E-mail: goergc@med.uni-marburg.de

    2007-11-15

    Objective: Ultrasound contrast agents in conjunction with contrast specific imaging techniques, are increasingly accepted in clinical use for diagnostic imaging in several organs. Contrast enhanced sonography (CES) of second-generation contrast media have shown a spleen-specific uptake of the microbubble contrast agent. The aim of this review is to illustrate indications for the use of CES in patients with suspected (peri-)splenic pathology. Methods: This review based on the experience of transcutaneous CES in 200 patients with (peri-)splenic pathology diagnosed by B-mode sonography at an internal medicine center. CES studies were performed with a contrast-devoted unit (Acuson, Sequoia, Siemens medical solution) that had contrast-specific, continuous-mode software. A low mechanical index was used. A sulfur hexafluoride-based microbubble contrast medium (Sonovue, Bracco SpA, Milan, Italy) was injected. Results: On our experience, there are several clinical conditions which may show an diagnostic advantage of CES in comparison to B-mode US. CES should be performed to investigate: (1) the perisplenic tumor to diagnose or exclude accessory spleen, (2) the small-sized spleen to diagnose functional asplenia/hyposplenia, (3) the inhomogenous spleen of unknown cause to diagnose focal lesions within the spleen, (4) the incidentally found hypoechoic splenic tumor to diagnose high vascular splenic hemangioma, (5) focal lesions suspect for splenic abscess, hematoma, infarction to confirme diagnosis, and (6) patients with abdominal trauma to diagnose or exclude splenic injuriy. Conclusion: CES is of diagnostic value in several clinical circumstances to diagnose accessory spleen, functional asplenia, small-sized splenic involvement, high vascular splenic hemangioma, and vascular splenic pathology like splenic infarction, splenic abscess, and splenic laceration.

  8. Biomechanical response of human spleen in tensile loading.

    Science.gov (United States)

    Kemper, Andrew R; Santago, Anthony C; Stitzel, Joel D; Sparks, Jessica L; Duma, Stefan M

    2012-01-10

    Blunt splenic injuries are most frequently caused as a result of motor vehicle collisions and are associated with high mortality rates. In order to accurately assess the risk of automotive related spleen injuries using tools such as finite element models, tissue level tolerance values and suitable material models must be developed and validated based on appropriate biomechanical data. This study presents a total of 41 tension tests performed on spleen parenchyma coupons and 29 tension tests performed on spleen capsule/parenchyma coupons. Standard dog-bone coupons were obtained from fresh human spleen and tested within 48 h of death. Each coupon was tested once to failure at one of the four loading rates to investigate the effects of rate dependence. Load and acceleration data were obtained at each of the specimen grips. High-speed video and optical markers placed on the specimens were used to measure local displacement. Failure stress and strain were calculated at the location of failure in the gage length of the coupon. The results of the study showed that both the spleen parenchyma and the capsule are rate dependent, with higher loading rates yielding higher failure stresses and lower failure strains. The results also show that the failure stress of the splenic capsule is significantly greater than that of the underlying parenchyma. Overall, this study provides novel biomechanical data that demonstrate the rate dependent tissue level tolerance values of human spleen tissue in tensile loading, which can aid in the improvement of finite element models used to assess injury risk in blunt trauma. Published by Elsevier Ltd.

  9. Robotic spleen-preserving distal pancreatectomy. A case report.

    Science.gov (United States)

    Vasilescu, C; Sgarbura, O; Tudor, S; Herlea, V; Popescu, I

    2009-01-01

    Distal pancreatectomy (DP) is the removal of the pancreatic tissue at the left side of the superior mesenteric vein and it is traditionally approached by an open or laparoscopic exposure. Preservation of the spleen is optional but appears to have a better immunological outcome. We present the case of a 53-year old patient with a 2.4/2.2 tumor located in the tail of the pancreas, with high tumour marker values for whom we decided to perform a robotic spleen-preserving distal pancreatectomy (RSPDP). The postoperative outcome was satisfactory. In conclusion, we recommend this type of approach for small pancreatic tail lesions.

  10. Incidental Detection of Temporary Focal FDG Retention in the Spleen

    Energy Technology Data Exchange (ETDEWEB)

    Park, Youn Joon; Lee, Jai Hyuen; Jee, Keum Nahn; Namgung, Hwan [Dankook Univ. College of Medicine, Seoul (Korea, Republic of)

    2011-06-15

    F 18 fluorodeoxyglucose (FDG) positron emission tomography (PET)/computed tomography (CT) is a valuable tool in discriminating malignancy from benign lesion. But because various false positive results reduce the diagnostic specificity, nuclear medicine physicians should be familiar with possible false positive cases. Although many cases of high FDG uptake mimicking malignancy have been reported, temporary intense focal FDG uptake of normal spleen has not been reported previously. We report herein a phenomenon of temporary intense focal FDG uptake of normal spleen without evidence of metastasis in a 46 year old woman with a history of anal cancer.

  11. Spleen and liver enlargement in a patient with rheumatoid arthritis.

    Science.gov (United States)

    Bedoya, María Eugenia; Ceccato, Federico; Paira, Sergio

    2015-01-01

    We describe the case of a 51-year-old woman with a seropositive, erosive, and non-nodular rheumatoid arthritis of 15 year of evolution. The patient had poor compliance with medical visits and treatment. She came to the clinic with persistent pancytopenia and spleen and liver enlargement. Liver and bone marrow biopsies were carried out and amyloidosis, neoplasias and infections were ruled out. We discuss the differential diagnosis of pancytopenia and spleen and liver enlargement in a long-standing rheumatoid arthritis patient. Copyright © 2014 Elsevier España, S.L.U. All rights reserved.

  12. Gastric Volvulus and Wandering Spleen: A Rare Surgical Emergency

    Directory of Open Access Journals (Sweden)

    Georgios Lianos

    2013-01-01

    Full Text Available Gastric volvulus is a rare but potentially life-threatening clinical entity due to possible gastric necrosis. A wandering spleen may also be associated with gastric volvulus. Patients presenting with the triad epigastralgia, vomiting followed by retching, and difficulty or inability to pass a nasogastric tube into the stomach are likely to have gastric volvulus. The operating surgeon should include this rare entity in the differential diagnosis when dealing with a patient with such a clinical profile. Herein, we present a case of gastric volvulus associated with a wandering spleen in a 28-year-old Caucasian woman and we provide a brief review of the literature on this issue.

  13. Cystic lymphangioma of the spleen: US-CT-MRI correlation

    Energy Technology Data Exchange (ETDEWEB)

    Bezzi, M.; Spinelli, A.; Pierleoni, M.; Andreoli, G.M. [Dept. of Radiology, University of Rome ' La Sapienza' (Italy)

    2001-07-01

    A case of a surgically confirmed cystic lymphangioma of the spleen is presented. Preoperative imaging consisted of US, contrast-enhanced CT and MRI, all showing a multiloculated lesion with small cystic cavities divided by thin septa, corresponding to dilated lymphatic spaces. Preoperative studies correlated well with the pathologic findings. Cystic lymphangioma of the spleen is a very rare condition and is usually solitary and asymptomatic. Large lymphangiomas may be an indication for splenectomy, since the risk of rupture is high even from minor abdominal trauma. Preoperative diagnosis may be achieved with correlated noninvasive imaging. (orig.)

  14. Characterization and pharmacological properties of a novel multifunctional Kunitz inhibitor from Erythrina velutina seeds.

    Directory of Open Access Journals (Sweden)

    Richele J A Machado

    Full Text Available Inhibitors of peptidases isolated from leguminous seeds have been studied for their pharmacological properties. The present study focused on purification, biochemical characterization and anti-inflammatory and anticoagulant evaluation of a novel Kunitz trypsin inhibitor from Erythrina velutina seeds (EvTI. Trypsin inhibitors were purified by ammonium sulfate (30-60%, fractionation followed by Trypsin-Sepharose affinity chromatography and reversed-phase high performance liquid chromatography. The purified inhibitor showed molecular mass of 19,210.48 Da. Furthermore, a second isoform with 19,228.16 Da was also observed. The inhibitor that showed highest trypsin specificity and enhanced recovery yield was named EvTI (P2 and was selected for further analysis. The EvTI peptide fragments, generated by trypsin and pepsin digestion, were further analyzed by MALDI-ToF-ToF mass spectrometry, allowing a partial primary structure elucidation. EvTI exhibited inhibitory activity against trypsin with IC50 of 2.2×10(-8 mol.L(-1 and constant inhibition (Ki of 1.0×10(-8 mol.L(-1, by a non-competitive mechanism. In addition to inhibit the activity of trypsin, EvTI also inhibited factor Xa and neutrophil elastase, but do not inhibit thrombin, chymotrypsin or peptidase 3. EvTI was investigated for its anti-inflammatory and anti-coagulant properties. Firstly, EvTI showed no cytotoxic effect on human peripheral blood cells. Nevertheless, the inhibitor was able to prolong the clotting time in a dose-dependent manner by using in vitro and in vivo models. Due to anti-inflammatory and anticoagulant EvTI properties, two sepsis models were here challenged. EvTI inhibited leukocyte migration and specifically acted by inhibiting TNF-α release and stimulating IFN-α and IL-12 synthesis. The data presented clearly contribute to a better understanding of the use of Kunitz inhibitors in sepsis as a bioactive agent capable of interfering in blood coagulation and inflammation.

  15. Purification and partial characterisation of a trypsin from the processing waste of the silver mojarra (Diapterus rhombeus).

    Science.gov (United States)

    Silva, Janilson F; Espósito, Talita S; Marcuschi, Marina; Ribeiro, Karina; Cavalli, Ronaldo O; Oliveira, Vitor; Bezerra, Ranilson S

    2011-12-01

    An alkaline peptidase was purified from the viscera of the silver mojarra (Diapterus rhombeus) in a three-step process: heat treatment, ammonium sulphate fractionation and molecular exclusion chromatography (Sephadex® G-75), with final specific activity 86-fold higher than the enzyme extract and yield of 22.1%. The purified enzyme had an estimated molecular mass of 26.5kDa and NH2-terminal amino acid sequence IVGGYECTMHSEAHE. Higher enzyme activity was observed at pH 8.5 and between 50 and 55°C. The enzyme was completely inactivated after 30min at 55°C and it was significantly more stable at alkaline pH. Km, Kcat and Kcat·Km(-1) values, using BApNA as substrate, were 0.266mM, 0.93s(-1) and 3.48mM(-1)s(-1), respectively. Enzyme activity increased in the presence of the ions (1mM) K(+), Li(+) and Ca(2+), but was inhibited by Fe(2+), Cd(2+), Cu(2+), Al(3+), Hg(2+), Zn(2+) and Pb(2+) as well as by the trypsin inhibitors TLCK and benzamidine. Copyright © 2011 Elsevier Ltd. All rights reserved.

  16. [The primary structure of the alpha-amylase inhibitor Hoe 467A from Streptomyces tendae 4158. A new class of inhibitors].

    Science.gov (United States)

    Aschauer, H; Vértesy, L; Nesemann, G; Braunitzer, G

    1983-10-01

    The native or modified alpha-amylase inhibitor Hoe 467A - isolated from the culture medium of Streptomyces tendae 4158 - and overlapping peptides were degraded by the automatic Edman technique. The oxidized or aminoethylated or oxidized and maleoylated inhibitor was digested with trypsin and the native inhibitor with pepsin. Further digestion with Staphylococcus aureus proteinase was also carried out. After peptic digestion two cystin peptides were isolated, which allowed the establishment of the disulfide bonds. The alpha-amylase inhibitor is a polypeptid consisting of 74 amino-acid residues with a molecular mass of 7958 Da. The inhibitor is composed of all naturally occurring amino acids except methionine and phenylalanine and shows no sequence homology to known inhibitors. The clinical and pharmacological importance in respect to the inhibitors ability for inactivation of human salivary and pancreatic alpha-amylase is discussed. Especially the proteinase resistance of the inhibitor enables a clinical application in human (e.g. Diabetes mellitus) per os.

  17. Conformational Network and Residence Time Estimation of Trypsin-Benzamidine Unbinding Pathways

    OpenAIRE

    Dickson, Alex; Lotz, Samuel D.

    2016-01-01

    In this poster we present results from molecular dynamics sampling of benzamidine unbinding from trypsin. We give background on the weighted ensemble technique used (WExplore) and the Markovian state model construction. Our network shows three unique unbinding pathways including a never before observed unbinding pathway. We also estimate residence time to within one order of magnitude to the experimental value.

  18. Quantitative proteomics reveals the kinetics of trypsin-catalyzed protein digestion.

    Science.gov (United States)

    Pan, Yanbo; Cheng, Kai; Mao, Jiawei; Liu, Fangjie; Liu, Jing; Ye, Mingliang; Zou, Hanfa

    2014-10-01

    Trypsin is the popular protease to digest proteins into peptides in shotgun proteomics, but few studies have attempted to systematically investigate the kinetics of trypsin-catalyzed protein digestion in proteome samples. In this study, we applied quantitative proteomics via triplex stable isotope dimethyl labeling to investigate the kinetics of trypsin-catalyzed cleavage. It was found that trypsin cleaves the C-terminal to lysine (K) and arginine (R) residues with higher rates for R. And the cleavage sites surrounded by neutral residues could be quickly cut, while those with neighboring charged residues (D/E/K/R) or proline residue (P) could be slowly cut. In a proteome sample, a huge number of proteins with different physical chemical properties coexists. If any type of protein could be preferably digested, then limited digestion could be applied to reduce the sample complexity. However, we found that protein abundance and other physicochemical properties, such as molecular weight (Mw), grand average of hydropathicity (GRAVY), aliphatic index, and isoelectric point (pI) have no notable correlation with digestion priority of proteins.

  19. Effects of tannic acid on trypsin and leucine aminopeptidase activities in gypsy moth larval midgut

    Directory of Open Access Journals (Sweden)

    Mrdaković Marija

    2013-01-01

    Full Text Available The effects of allelochemical stress on genetic variations in the specific activities of gypsy moth digestive enzymes (trypsin and leucine aminopeptidase and relative midgut mass (indirect measure of food consumption, as well as variability in their plasticity, were investigated in fifth instar gypsy moths originating from two populations with different trophic adaptations (oak and locust-tree forests. Thirty-two full-sib families from the Quercus population and twenty-six full-sib families from the Robinia population were reared on an artificial diet with or without supplementation with tannic acid. Between population differences were observed as higher average specific activity of trypsin and relative midgut mass in larvae from the Robinia population. Significant broad-sense heritabilities were observed for the specific activity of trypsin in the control state, and for specific activity of leucine aminopeptidase in a stressful environment. Significantly lower heritability for relative midgut mass was recorded in larvae from the Robinia population reared under stressful conditions. Significant variability of trypsin plasticity in larvae from both populations and significant variability of leucine aminopeptidase plasticity in larvae from the Robinia population point to the potential for the evolution of enzyme adaptive plastic responses to the presence of stressor. Non-significant across-environment genetic correlations do not represent a constraint for the evolution of enzyme plasticity. [Projekat Ministarstva nauke Republike Srbije, br. 173027

  20. Biochemical Characterization of An Arginine-Specific Alkaline Trypsin from Bacillus licheniformis.

    Science.gov (United States)

    Gong, Jin-Song; Li, Wei; Zhang, Dan-Dan; Xie, Min-Feng; Yang, Biao; Zhang, Rong-Xian; Li, Heng; Lu, Zhen-Ming; Xu, Zheng-Hong; Shi, Jin-Song

    2015-12-17

    In the present study, we isolated a trypsin-producing strain DMN6 from the leather waste and identified it as Bacillus licheniformis through a two-step screening strategy. The trypsin activity was increased up to 140 from 20 U/mL through culture optimization. The enzyme was purified to electrophoretic homogeneity with a molecular mass of 44 kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and the specific activity of purified enzyme is 350 U/mg with Nα-Benzoyl-L-arginine ethylester as the substrate. The optimum temperature and pH for the trypsin are 65 °C and pH 9.0, respectively. Also, the enzyme can be significantly activated by Ba(2+). This enzyme is relatively stable in alkaline environment and displays excellent activity at low temperatures. It could retain over 95% of enzyme activity after 180 min of incubation at 45 °C. The distinguished activity under low temperature and prominent stability enhance its catalytic potential. In the current work, the open reading frame was obtained with a length of 1371 nucleotides that encoded a protein of 456 amino acids. These data would warrant the B. licheniformis trypsin as a promising candidate for catalytic application in collagen preparation and leather bating through further protein engineering.

  1. Evaluation of the possible proteomic application of trypsin from Streptomyces griseus

    Czech Academy of Sciences Publication Activity Database

    Štosová, T.; Šebela, M.; Řehulka, Pavel; Šedo, O.; Havliš, J.; Zdráhal, Z.

    2008-01-01

    Roč. 376, č. 1 (2008), s. 94-102 ISSN 0003-2697 Institutional research plan: CEZ:AV0Z40310501 Keywords : MALDI-TOF MS * Streptomyces griseus * trypsin Subject RIV: CB - Analytical Chemistry, Separation Impact factor: 3.088, year: 2008

  2. Biochemical Characterization of An Arginine-Specific Alkaline Trypsin from Bacillus licheniformis

    Directory of Open Access Journals (Sweden)

    Jin-Song Gong

    2015-12-01

    Full Text Available In the present study, we isolated a trypsin-producing strain DMN6 from the leather waste and identified it as Bacillus licheniformis through a two-step screening strategy. The trypsin activity was increased up to 140 from 20 U/mL through culture optimization. The enzyme was purified to electrophoretic homogeneity with a molecular mass of 44 kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and the specific activity of purified enzyme is 350 U/mg with Nα-Benzoyl-l-arginine ethylester as the substrate. The optimum temperature and pH for the trypsin are 65 °C and pH 9.0, respectively. Also, the enzyme can be significantly activated by Ba2+. This enzyme is relatively stable in alkaline environment and displays excellent activity at low temperatures. It could retain over 95% of enzyme activity after 180 min of incubation at 45 °C. The distinguished activity under low temperature and prominent stability enhance its catalytic potential. In the current work, the open reading frame was obtained with a length of 1371 nucleotides that encoded a protein of 456 amino acids. These data would warrant the B. licheniformis trypsin as a promising candidate for catalytic application in collagen preparation and leather bating through further protein engineering.

  3. The treatment of spleen injuries: a retrospective study.

    Science.gov (United States)

    Dehli, Trond; Bågenholm, Anna; Trasti, Nora Christine; Monsen, Svein Arne; Bartnes, Kristian

    2015-10-29

    Hemorrhage after blunt trauma is a major contributor to death after trauma. In the abdomen, an injured spleen is the most frequent cause of major bleeding. Splenectomy is historically the treatment of choice. In 2007, non-operative management (NOM) with splenic artery embolization (SAE) was introduced in our institution. The indication for SAE is hemodynamically stable patients with extravasation of contrast, or grade 3-5 spleen injury according to the Abbreviated Organ Injury Scale 2005, Update 2008. We wanted to examine if the introduction of SAE increased the rate of salvaged spleens in our trauma center. All patients discharged with the diagnosis of splenic injury in the period 01.01.2000 - 31.12.2013 from the University Hospital of North Norway Tromsø were included in the study. Patients admitted for rehabilitation purposes or with an iatrogenic injury were excluded. A total of 109 patients were included in the study. In the period 2000-7, 20 of 52 patients were splenectomized. During 2007-13, there were 6 splenectomies and 24 SAE among 57 patients. The reduction in splenectomies is significant (p < 0.001). There is an increase in the rate of treated patients (splenectomy and SAE) from 38 to 53 % in the two time periods, but not significantly (p = 0.65). The rate of salvaged spleens has increased after the introduction of SAE in our center. The study is registered at www.clinicaltrials.gov with the identification number NCT01965548.

  4. [Spleen injuries in Spain: at what point are we?].

    Science.gov (United States)

    Jiménez Fuertes, Montiel; Costa Navarro, David; Jover Navalón, José María; Turégano Fuentes, Fernando; Ceballos Esparragón, José; Yuste, Pedro; Sánchez Tocino, Juan María; Navarro Soto, Salvador; Montmany, Sandra

    2013-11-01

    Management of spleen trauma has changed over last decades, although there is no data on its treatment in Spain. The aim of this study is to determine the characteristics of spleen injuries in adults with severe abdominal injuries and how we manage them. A prospective study using the databases of six Spanish hospitals: Gregorio Marañón Hospital, Virgen de la Vega Hospital, Torrevieja Hospital, Getafe Hospital, Doce de Octubre Hospital and Corporació Sanitària Parc Taulí. A total of 566 patients who had sustained spleen injuries were analyzed (448 males and 118 females), most of them were due to blunt trauma (94%), and the most frequent mechanism of injury was motor vehicle accident. The mean Injury Severity Score (ISS) was 25.2. The initial treatment was surgical in 56.6% of the patients (85.3% total splenectomy and 14.7% other conservative surgical procedures, of which 4.6% finally failed and required total splenectomy). The remaining 43.4% were initially managed conservatively, but 6.5% of them finally required surgical splenectomy, and in 8.8% angio-embolization was performed. In Spain, management of spleen trauma is mainly surgical (particularly splenectomy). Angio-embolization and conservative surgical procedures are now hardly used. Copyright © 2011 AEC. Published by Elsevier Espana. All rights reserved.

  5. Torsion of the spleen with incomplete infarction: case report

    International Nuclear Information System (INIS)

    Lernau, O.Z.; Baron, J.; Nissan, S.

    1977-01-01

    Torsion and infarction of a ''wandering spleen'' is a rare disease which is often confused with other acute abdominal crises. A correct preoperative diagnosis, when made, has usually been determined by arteriographic studies. A child is described in whom changes in the TcSC scan made a correct diagnosis possible by non-invasive methods

  6. Spontaneous rupture of the spleen after infectious mononucleosis

    DEFF Research Database (Denmark)

    Gulstad, Mikkel Bak; Thomsen, Henrik

    2013-01-01

    Non-traumatic rupture of the spleen (NRS) is a rare but serious complication to infectious mononucleosis (IM) and it is important to have in mind, when patients have IM. Although splenectomy has been advocated as the appropriate treatment for this problem, the trend goes towards conservative...

  7. Hydatid disease of the spleen; Ultrasonography, CT and MR imaging

    Energy Technology Data Exchange (ETDEWEB)

    Sinner, W.N. von; Stridbeck, H. (Dept. of Diagnostic Radiology, King Faisal Specialist Hospital, and Research Center, Riyadh (Saudi Arabia) Lund Univ. Hospital (Sweden))

    1992-09-01

    Seven patients with hydatid disease of the spleen were examined by radiography, ultrasound, CT, and in one case MR imaging. The observations were confirmed by patho-anatomic findings except in 2 patients where high indirect hemagglutination tests confirmed the diagnosis. (orig./MG).

  8. Scintigraphic evidence of transplanted hepatocytes in spleen and liver

    International Nuclear Information System (INIS)

    Henne-Bruns, D.; Kremer, B.; Gramminger, K.; Broelsch, C.

    1986-01-01

    In rats suffering from hepatic enzymatic deficiency transplanted hepatocytes could be evidenced scintigraphically in liver, spleen and granulomas. In pigs, however, it is very difficult to demonstrate transplanted hepatocytes by scintiscanning because of the thickness of the tissues and the high background radiation in large animals

  9. Morphology and some biomechanical properties of human liver and spleen

    Czech Academy of Sciences Publication Activity Database

    Stingl, J.; Bača, V.; Čech, V.; Kovanda, J.; Kovandová, H.; Mandys, Václav; Rejmontová, J.; Sosna, B.

    2002-01-01

    Roč. 24, - (2002), s. 285-289 ISSN 0930-1038 Institutional research plan: CEZ:AV0Z5039906 Keywords : Human liver and spleen Subject RIV: FE - Other Internal Medicine Disciplines Impact factor: 0.252, year: 2002

  10. Primary hemangiosarcoma of the spleen with angioscintigraphic demonstration of metastases

    DEFF Research Database (Denmark)

    Hermann, G G; Fogh, J; Graem, N

    1984-01-01

    A case of primary hemangiosarcoma of the spleen in a 48-year-old woman is presented. Twenty-eight months after splenectomy the patient developed a severe anemia of the microangiopathic type, thrombocytopenia, and a leukoerythroblastic peripheral blood picture. In contrast to x-ray and conventional...

  11. Sonographic Determination of Spleen to Left Kidney Ratio among ...

    African Journals Online (AJOL)

    Background: Clinical determination of mild splenomegaly is notoriously inaccurate. Objectives: To determine sonographically the spleen to left kidney ratio according to age and somatometric parameters among school age children in a tropical environment. Methods: A cross sectional study and convenience sampling were ...

  12. Oligopeptidase B from Serratia proteamaculans. III. Inhibition analysis. Specific interactions with metalloproteinase inhibitors.

    Science.gov (United States)

    Mikhailova, A G; Khairullin, R F; Kolomijtseva, G Ya; Rumsh, L D

    2012-03-01

    Inhibition of the novel oligopeptidase B from Serratia proteamaculans (PSP) by basic pancreatic trypsin inhibitor, Zn2+ ions, and o- and m-phenanthroline was investigated. A pronounced effect of calcium ions on the interaction of PSP with inhibitors was demonstrated. Inversion voltamperometry and atomic absorption spectrometry revealed no zinc ions in the PSP molecule. Hydrophobic nature of the enzyme inhibition by o- and m-phenanthroline was established.

  13. Mutations in serine protease inhibitor Kazal type 1 are strongly associated with chronic pancreatitis

    OpenAIRE

    Drenth, J P H; te Morsche, R; Jansen, J B M J

    2002-01-01

    Background: Although chronic pancreatitis is associated with risk factors such as alcoholism, hyperparathyroidism, and hypertriglyceridaemia, little is known of the actual aetiology of the disease. It is thought that inappropriate activation of trypsinogen causes pancreatitis, and indeed in cases of hereditary pancreatitis mutations of cationic trypsinogen (PRSS1) have been described. As serine protease inhibitor Kazal type 1 (SPINK1) is a potent natural inhibitor of pancreatic trypsin activi...

  14. Improved purification process of β- and α-trypsin isoforms by ion-exchange chromatography

    Directory of Open Access Journals (Sweden)

    Alexandre Martins Costa Santos

    2008-08-01

    Full Text Available The purpose of this work was to improve the separation and yield of pure β- and α-trypsin isoforms by ion-exchange chromatography and to characterize some physical-chemical properties of these isoforms. Purification of trypsin isoforms was performed by ion-exchange chromatography in 0.1 mol/L tris-HC buffer, pH 7.10 at 4ºC. The sample loading, salt concentration, flow rate and pH of mobile phase were varied to determine their effects on the resolution of the separation. The resolution was optimized mainly between β- and α-trypsin. Pure isoforms were obtained by chromatographying 100 mg of commercial trypsin during seven days, yielding 51 mg of high purity β-trypsin and 13 mg of α-trypsin partially pure, with small amounts of contaminating of ψ-trypsin. Thus, time and resolution of purification were optimized yielding large amounts of pure active enzymes that are useful for several research areas and biotechnology.O propósito deste trabalho foi melhorar a separação e o rendimento das isoformas puras β- e α-tripsina por meio de cromatografia de troca iônica e caracterizar algumas propriedades físico-químicas dessas isoformas. A purificação de isoformas de tripsina foi realizada em SE Sephadex, com tampão tris-HCl, pH 7,10 a 4ºC. A quantidade de amostra, a concentração salina, o fluxo e o pH da fase móvel foram variados para determinar o efeito sobre a resolução da separação. A resolução foi otimizada principalmente entre β- e α-tripsina, utilizando o pH 7,10 a 4ºC. Isoformas puras foram obtidas a partir de 100 mg de tripsina comercial bovina depois de sete dias de cromatografia, fornecendo 51,0 mg de β-tripsina totalmente pura e 13,0 mg de α-tripsina parcialmente pura, com quantidades pequenas de contaminação por ψ-Tripsina. Assim, tempo e resolução da purificação foram otimizados redendo grandes quantidades de enzimas puras e ativas que são úteis em várias áreas de pesquisa e ciências biotecnológicas.

  15. The role of proteases, endoplasmic reticulum stress and SERPINA1 heterozygosity in lung disease and alpha-1 anti-trypsin deficiency.

    LENUS (Irish Health Repository)

    Greene, Catherine M

    2012-02-01

    The serine proteinase inhibitor alpha-1 anti-trypsin (AAT) provides an antiprotease protective screen throughout the body. Mutations in the AAT gene (SERPINA1) that lead to deficiency in AAT are associated with chronic obstructive pulmonary diseases. The Z mutation encodes a misfolded variant of AAT that is not secreted effectively and accumulates intracellularly in the endoplasmic reticulum of hepatocytes and other AAT-producing cells. Until recently, it was thought that loss of antiprotease function was the major cause of ZAAT-related lung disease. However, the contribution of gain-of-function effects is now being recognized. Here we describe how both loss- and gain-of-function effects can contribute to ZAAT-related lung disease. In addition, we explore how SERPINA1 heterozygosity could contribute to smoking-induced chronic obstructive pulmonary diseases and consider the consequences.

  16. Rare Case of an Epithelial Cyst in an Intrapancreatic Accessory Spleen Treated by Robot-Assisted Spleen Preserving Distal Pancreatectomy.

    Science.gov (United States)

    van Dijck, Willemijn P M; Groot, Vincent P; Brosens, Lodewijk A A; Hagendoorn, Jeroen; Rinkes, Inne H M Borel; van Leeuwen, Maarten S; Molenaar, I Quintus

    2016-01-01

    Epithelial cyst in an intrapancreatic accessory spleen (ECIPAS) is exceedingly rare with only 57 cases reported since the first publication in 1980. Comprehensive clinical and diagnostic features remain to be clarified. We present a case of ECIPAS in a 21-year-old Philippine woman who was admitted with right upper quadrant abdominal pain. A cystic lesion in the pancreatic tail was discovered and evaluated by computed tomography and magnetic resonance images. Based on clinical and radiological features a solid pseudopapillary neoplasm was suspected. The patient underwent robot-assisted spleen preserving distal pancreatectomy. Pathological evaluation revealed a 26 mm intrapancreatic accessory spleen with a 16 mm cyst, lined by multilayered epithelium in the tail of the pancreas. The postoperative course was uneventful. Differentiating ECIPAS from (pre)malignant cystic pancreatic neoplasms based on clinical and radiological features remains difficult. When typical radiological signs can be combined with scintigraphy using Technetium-99m labelled colloid or Technetium-99m labelled erythrocytes, which can identify the solid component of the lesion as splenic tissue, it should be possible to make the right diagnosis noninvasively. When pancreatectomy is inevitable due to symptoms or patient preference, minimally invasive laparoscopic or robot-assisted spleen preserving distal pancreatectomy should be considered.

  17. Rare Case of an Epithelial Cyst in an Intrapancreatic Accessory Spleen Treated by Robot-Assisted Spleen Preserving Distal Pancreatectomy

    Directory of Open Access Journals (Sweden)

    Willemijn P. M. van Dijck

    2016-01-01

    Full Text Available Epithelial cyst in an intrapancreatic accessory spleen (ECIPAS is exceedingly rare with only 57 cases reported since the first publication in 1980. Comprehensive clinical and diagnostic features remain to be clarified. We present a case of ECIPAS in a 21-year-old Philippine woman who was admitted with right upper quadrant abdominal pain. A cystic lesion in the pancreatic tail was discovered and evaluated by computed tomography and magnetic resonance images. Based on clinical and radiological features a solid pseudopapillary neoplasm was suspected. The patient underwent robot-assisted spleen preserving distal pancreatectomy. Pathological evaluation revealed a 26 mm intrapancreatic accessory spleen with a 16 mm cyst, lined by multilayered epithelium in the tail of the pancreas. The postoperative course was uneventful. Differentiating ECIPAS from (premalignant cystic pancreatic neoplasms based on clinical and radiological features remains difficult. When typical radiological signs can be combined with scintigraphy using Technetium-99m labelled colloid or Technetium-99m labelled erythrocytes, which can identify the solid component of the lesion as splenic tissue, it should be possible to make the right diagnosis noninvasively. When pancreatectomy is inevitable due to symptoms or patient preference, minimally invasive laparoscopic or robot-assisted spleen preserving distal pancreatectomy should be considered.

  18. Rare Case of an Epithelial Cyst in an Intrapancreatic Accessory Spleen Treated by Robot-Assisted Spleen Preserving Distal Pancreatectomy

    NARCIS (Netherlands)

    van Dijck, Willemijn P M; Groot, Vincent P; Brosens, Lodewijk A A; Hagendoorn, Jeroen; Rinkes, Inne H M Borel; van Leeuwen, Maarten S; Molenaar, I Quintus

    2016-01-01

    Epithelial cyst in an intrapancreatic accessory spleen (ECIPAS) is exceedingly rare with only 57 cases reported since the first publication in 1980. Comprehensive clinical and diagnostic features remain to be clarified. We present a case of ECIPAS in a 21-year-old Philippine woman who was admitted

  19. B-mode and contrast-enhanced sonographic assessment of accessory spleen in the dog.

    Science.gov (United States)

    Rossi, Federica; Rabba, Silvia; Vignoli, Massimo; Haers, Hendrik; Terragni, Rossella; Saunders, Jimmy H

    2010-01-01

    Four dogs with an accessory spleen are described. The accessory spleens appeared as a round-to-triangular structure located in the perisplenic area. They were homogeneous and isoechoic with the adjacent spleen. Contrast-enhanced ultrasound was performed using a second generation microbubble contrast medium (sulfur hexafluoride). The type and timing of enhancement of the accessory spleen was similar to that of the parent spleen. Contrast-enhanced ultrasound is a noninvasive modality useful in distinguishing an accessory spleen from a mass of another origin.

  20. Physical-chemical characterization and stability study of alpha-trypsin at ph 3.0 by differential scanning calorimetry

    Energy Technology Data Exchange (ETDEWEB)

    Santos, A.M.C.; Santana, M.A.; Gomide, F.T.F.; Oliveira, J.S.; Vilas Boas, F.A.S.; Santoro, M.M.; Teixera, K.N. [Universidade Federal de Minas Gerais (UFMG), Belo Horizonte, MG (Brazil). Inst. de Ciencias Biologicas (ICB). Dept. de Bioquimica e Imunologia; Miranda, A.A.C.; Biondi, I. [Universidade Estadual de Feira de Santana (UEFS), BA (Brazil). Dept. de Ciencias Biologicas; Vasconcelos, A.B.; Bemquerer, M.P. [EMBRAPA Recursos Geneticos e Biotecnologia, Brasilia, DF (Brazil). Parque Estacao Biologica (PqEB)

    2008-07-01

    Full text: {alpha}-Trypsin is a serine-protease with a polypeptide chain of 223 amino acid residues and six disulfide bridges. It is a globular protein with predominance of antiparallel {beta}-sheet secondary structure and it has two domains with similar structures. In the present work, a stability study of {alpha}-trypsin in the acid pH range was performed and physical-chemical denaturation parameters were measured by using differential scanning calorimetry (DSC). The {alpha}-trypsin has a shelf-life (t{sub 95%}) of about ten months at pH 3.0 and 4 deg C and its hydrolysis into the {psi}-trypsin isoform is negligible during six months as monitored by mass spectrometry (Micromass Q-ToF). The observed {delta}H{sub cal}/{delta}H{sub vH} ratio is close to unity for {alpha}-trypsin denaturation, which suggests the occurrence of a two-state transition, devoid of molten-globule intermediates. At pH 3.0, {alpha}-trypsin unfolded with T{sub m} 325.9 K and {delta}H= 99.10 kcal mol{sup -1}, and the change in heat capacity between the native and unfolded forms of the protein was estimated to be 1.96 {+-} 0.18 kcal mol{sup -1} K{sup -1}. The stability of {alpha}-trypsin calculated at 298 K and at pH 3.0 was {delta}G{sub U} = 6.10 kcal mol{sup -1}. These values are in the range expected for a small globular protein. These results show that the thermodynamic parameters for unfolding of {beta}-trypsin do not change substantially after its conversion to {alpha}-trypsin.

  1. Dual core quantum dots for highly quantitative ratiometric detection of trypsin activity in cystic fibrosis patients

    Science.gov (United States)

    Castelló Serrano, Iván; Stoica, Georgiana; Matas Adams, Alba; Palomares, Emilio

    2014-10-01

    We present herein two colour encoded silica nanospheres (2nanoSi) for the fluorescence quantitative ratiometric determination of trypsin in humans. Current detection methods for cystic fibrosis diagnosis are slow, costly and suffer from false positives. The 2nanoSi proved to be a highly sensitive, fast (minutes), and single-step approach nanosensor for the screening and diagnosis of cystic fibrosis, allowing the quantification of trypsin concentrations in a wide range relevant for clinical applications (25-350 μg L-1). Furthermore, as trypsin is directly related to the development of cystic fibrosis (CF), different human genotypes, i.e. CF homozygotic, CF heterozygotic, and unaffected, respectively, can be determined using our 2nanoSi nanospheres. We anticipate the 2nanoSi system to be a starting point for non-invasive, easy-to-use and cost effective ratiometric fluorescent biomarkers for recessive genetic diseases like human cystic fibrosis. In a screening program in which the goal is to detect disease and also the carrier status, early diagnosis could be of great help.We present herein two colour encoded silica nanospheres (2nanoSi) for the fluorescence quantitative ratiometric determination of trypsin in humans. Current detection methods for cystic fibrosis diagnosis are slow, costly and suffer from false positives. The 2nanoSi proved to be a highly sensitive, fast (minutes), and single-step approach nanosensor for the screening and diagnosis of cystic fibrosis, allowing the quantification of trypsin concentrations in a wide range relevant for clinical applications (25-350 μg L-1). Furthermore, as trypsin is directly related to the development of cystic fibrosis (CF), different human genotypes, i.e. CF homozygotic, CF heterozygotic, and unaffected, respectively, can be determined using our 2nanoSi nanospheres. We anticipate the 2nanoSi system to be a starting point for non-invasive, easy-to-use and cost effective ratiometric fluorescent biomarkers for

  2. Scintigraphy of liver and spleen in vinyl chloride workers

    Energy Technology Data Exchange (ETDEWEB)

    Biersack, H J; San Luis, T Jr; Lange, C E; Thelen, M; Veltman, G; Winkler, C [Bonn Univ. (Germany, F.R.). Inst. fuer Klinische und Experimentelle Nuklearmedizin; Bonn Univ. (Germany, F.R.). Klinik und Poliklinik der Haut- und Geschlechtskrankheiten; Bonn Univ. (Germany, F.R.). Radiologische Klinik)

    1977-10-01

    In 152 VC-exposed workers of whom 124 were employed in the PVC-production and 28 in VC-processing plants, liver and spleen imaging was performed using sup(99m)Tc-sulphur colloid and /sup 197/Hg-BMHP. In 101 (= 81%) of the 124 workers of the PVC-production plant and in 18 (= 64%) workers of PVC-processing factories pathological liver and spleen scintigrams were found. The most frequent pathological change in the scintigraphic image was an increase in splenic colloid accumulation, when compared with the liver uptake. Three angiosarcomas of the liver were detected through circumscribed defects of colloid accumulation. Sequential liver scintigraphy was done in 15 cases. In 7 patients with esophageal varices, considerable decrease in portal venous blood flow was demonstrated. - As a result of our investigations it can be stated that scintigraphically detectable changes are sensitive indicators of VC-induced lesions of the liver including liver fibrosis, portal hypertension and angiosarcoma.

  3. Inflammatory pseudo tumor of the spleen. Radiologic findings

    International Nuclear Information System (INIS)

    Folgueral, M.; Naranjo, G.; Grasa, J.

    1998-01-01

    The inflammatory splenic pseudo tumor is an uncommon benign lesion of unknown etiology that usually is detected incidentally in patients being examined for other disorders. We present the ultrasound, computed tomography (CT) and magnetic resonance (MR) findings in two patients (a 56-year-old woman and 67-year-old man) with inflammatory pseudo tumor of the spleen. Ultrasound images of the inflammatory splenic pseudo tumor show a hypoechoic mass with or without calcifications. In CT, the mass presents a low attenuation that is slightly enhanced by intravenous injection of an iodinated contrast medium. MR images show an isointense spleen in T1-weighted spin-echo sequences that is hypointense in T2-weighted sequences. The differential diagnosis should consider other more common splenic masses. but the definitive diagnosis can not be based on imaging methods alone. In the both cases, the final diagnosis was reached after splenectomy and pathological study. (Author) 8 refs

  4. Primary Angiosarcoma of the Spleen: An Oncological Enigma

    Directory of Open Access Journals (Sweden)

    Myoteri Despoina

    2014-01-01

    Full Text Available Introduction. Primary splenic angiosarcoma is an extremely unusual neoplasm originating from sinusoidal vascular endothelium. Surgical extirpation is the mainstay of treatment of this highly malignant disease. Case Presentation. An 82-year-old woman was admitted with left pleural effusion and a palpable left upper quadrant abdominal mass, secondary to splenomegaly by two large splenic tumors. Classic open splenectomy was performed and angiosarcoma of the spleen was the final histopathological diagnosis, which was primary since no other disease site was revealed. Discussion. The incidence of the disease is 0.14–0.23 cases per million, with slight male predominance. Etiology is not established and clinical presentation may confuse even experienced physicians. Imaging modalities cannot differentiate the lesion from other vascular splenic neoplasms and the correct diagnosis is mainly set after histopathological examination of the resected spleen. As with other sarcomas, surgery is the only curative approach, while chemo- and radiotherapy have poor results. Prognosis remains dismal.

  5. ALTERED HISTOLOGY OF THE THYMUS AND SPLEEN IN CONTAMINANT-EXPOSED JUVENILE AMERICAN ALLIGATORS

    Science.gov (United States)

    Morphological difference in spleen and thymus are closely related to functional immune differences. Hormonal regulation of the immune system has been demonstrated in reptilian splenic and thymic tissue. Spleens and thymus were obtained from juvenile alligators at two reference si...

  6. Wound-induced proteinase inhibitor in Salix viminalis and its association with defence against insects

    Energy Technology Data Exchange (ETDEWEB)

    Saarikoski, P. [Swedish Univ. of Agricultural Sciences, Uppsala (Sweden). Dept. of Forest Genetics

    1997-09-01

    For successful traditional breeding, the plant material has to be screened for genetic variation for the desired traits. By screening Salix clones for wound-induced proteinase inhibitor (PI) activity and ethylene evolution, it was possible to identify variation for both characters among the Salix clones tested. However, no correlation was observed with insect and pathogen resistance. Since there was no simple relationship between wound-induced ethylene production, accumulation of PI and pest resistance, a more systematic investigation of Salix PIs was begun. A gene (swin1.1) encoding a 21 kDa trypsin inhibitor with characteristics of Kunitz-type of PI was sequenced. The trypsin inhibitor encoded by the isolated swin1.1 gene was shown to be functional in vitro and exhibit specificity for trypsin. It is therefore likely that this PI is involved in the plant defence in Salix, since many insects have trypsin as their major digestive protease. In further support of this view, in bio-tests with poplar the mortality of the first instar larvae (Lymantria dispar) was significantly increased, both after application of the trypsin inhibitor encoded by swin1.1 directly on poplar leaves and after feeding the larvae with transgenic poplar over-expressing the swin1.1 gene. In Salix, the swin1.1 gene was shown to be induced by mechanical wounding, insect feeding and by treatment with the signalling substances salicylic and jasmonic acid. The locally wound-induced response (mechanical and insect) was greater than the systemic response. Other swin1 gene family members were also differentially expressed after the inductive treatment. 187 refs., 3 figs., 2 tabs.

  7. Pancreatitis-associated fluid collections involving the spleen

    International Nuclear Information System (INIS)

    Vick, C.W.; Simeone, J.F.; Ferrucci, J.T. Jr.; Wittenberg, J.; Mueller, P.R.; Harvard Medical School, Boston, MA

    1981-01-01

    The clinical and radiographic features of 2 patients with dissecting pancreatitis-associated fluid collections involving the spleen are described. A typical appearance of left upper quadrant fluid collection lateral to the splenic pulp was observed by ultrasonography (US) or computed body tomography (CBT). Although these findings are nonspecific, a left upper quadrant fluid collection may be characterized definitively by US/CBT-guided needle aspiration. (orig.)

  8. Spleens and holoendemic malaria in West New Guinea.

    Science.gov (United States)

    METSELAAR, D

    1956-01-01

    The author describes the results obtained in recent malaria surveys in West New Guinea, where what is essentially holoendemic malaria prevails. However, the spleen-rate in adults differs markedly from what is regarded as normal under holoendemic conditions according to the definition put forward at the Malaria Conference in Equatorial Africa in 1950. The author therefore concludes that that definition is not properly applicable to New Guinea.

  9. Postnatal development of the spleen in Didelphis virginiana.

    Science.gov (United States)

    Cutts, J H; Krause, W J

    1982-01-01

    The postnatal development of the spleen has been examined in 85 opossums ranging in age from newborn to adult. At birth the spleen consists of a well vascularized mass of mesenchymal tissue and lacks lymphatic tissue or any evidence of haemopoietic activity. Haemopoiesis is evident at seven days, increases to a maximum at about two to three weeks and thereafter gradually declines. Although production of granulocytes has disappeared by 60 days postnatum, a small degree of erythropoiesis and megakaryocyte formation continues throughout life. Lymphatic tissue appears by the third week, but germinal centres do not appear until after weaning. A feature of the spleen during the first three to four days is the presence of a population of primitive 'blast' cells. Images Fig. 1 Fig. 2 Fig. 3 Fig. 4 Fig. 5 Fig. 6 Fig. 7 Fig. 8 Fig. 9 Fig. 10 Fig. 11 Fig. 12 Fig. 13 Fig. 14 Fig. 15 Fig. 16 Fig. 17 Fig. 18 Fig. 19 Fig. 20 Fig. 21 Fig. 22 Fig. 23 Fig. 24 PMID:7153176

  10. Data from quantitative label free proteomics analysis of rat spleen

    Directory of Open Access Journals (Sweden)

    Khadar Dudekula

    2016-09-01

    Full Text Available The dataset presented in this work has been obtained using a label-free quantitative proteomic analysis of rat spleen. A robust method for extraction of proteins from rat spleen tissue and LC-MS-MS analysis was developed using a urea and SDS-based buffer. Different fractionation methods were compared. A total of 3484 different proteins were identified from the pool of all experiments run in this study (a total of 2460 proteins with at least two peptides. A total of 1822 proteins were identified from nine non-fractionated pulse gels, 2288 proteins and 2864 proteins were identified by SDS-PAGE fractionation into three and five fractions respectively. The proteomics data are deposited in ProteomeXchange Consortium via PRIDE PXD003520, Progenesis and Maxquant output are presented in the supported information. The generated list of proteins under different regimes of fractionation allow assessing the nature of the identified proteins; variability in the quantitative analysis associated with the different sampling strategy and allow defining a proper number of replicates for future quantitative analysis. Keywords: Spleen, Rat, Protein extraction, Label-free quantitative proteomics

  11. Return to play after liver and spleen trauma.

    Science.gov (United States)

    Juyia, Rushad F; Kerr, Hamish A

    2014-05-01

    Sport-related spleen and liver injuries pose a challenge for the physician. Although rare, these injuries can have serious and even life-threatening outcomes if not accurately diagnosed and managed in a timely fashion. Currently, there are no evidence-based guidelines on duration and intensity of restricted activity and return to play after spleen and liver injury. In addition, there is controversy on follow-up imaging after injury. PubMed was searched using the terms splenic or spleen and trauma and hepatic or liver and trauma from 1980 to 2013. The citations from sentinel papers were also reviewed. Clinical review. Level 3. Ultrasound is ideal in the unstable athlete. Nonoperative management of blunt splenic and hepatic injuries is recommended for hemodynamically stable patients regardless of injury grade, patient age, or presence of associated injuries. Follow-up imaging is not routinely recommended unless clinically indicated. Athletes may engage in light activity for the first 3 months after injury and then gradual return to unrestricted activity as tolerated. High-level athletes may choose splenectomy or serial imaging for faster return to play. Intravenous contrast-enhanced computed tomography is the diagnostic imaging modality of choice in stable athletes with blunt abdominal trauma. C.

  12. High dispersity of carbon nanotubes diminishes immunotoxicity in spleen.

    Science.gov (United States)

    Lee, Soyoung; Khang, Dongwoo; Kim, Sang-Hyun

    2015-01-01

    From the various physiochemical material properties, the chemical functionalization order of single-walled carbon nanotubes (swCNTs) has not been considered as a critical factor for modulating immunological responses and toxicological aspects in drug delivery applications. Although most nanomaterials, including carbon nanotubes, are specifically accumulated in spleen, few studies have focused on spleen immunotoxicity. For this reason, this study demonstrated that the dispersity of swCNTs significantly influenced immunotoxicity in vitro and in vivo. For cytotoxicity of swCNTs, MTT assay, reactive oxygen species production, superoxide dismutase activity, cellular uptake, and confocal microscopy were used in macrophages. In the in vivo study, female BALB/c mice were intravenously administered with 1 mg/kg/day of swCNTs for 2 weeks. The body weight, organ weight, hematological change, reverse-transcription polymerase chain reaction, and lymphocyte population were evaluated. Different orders of chemical functionalization of swCNTs controlled immunotoxicity. In short, less-dispersed swCNTs caused cytotoxicity in macrophages and abnormalities in immune organs such as spleen, whereas highly dispersed swCNTs did not result in immunotoxicity. This study clarified that increasing carboxyl groups on swCNTs significantly mitigated immunotoxicity in vitro and in vivo. Our findings clarified the effective immunotoxicological factors of swCNTs by increasing dispersity of swCNTs and provided useful guidelines for the effective use of nanomaterials.

  13. Radioimmunoassay of trypsin in dried blood importance for the neonatal detection of cystic fibrosis

    International Nuclear Information System (INIS)

    Travert, G.; Mustin, C.; Fernandez, Y.

    1981-01-01

    The demonstration of very high levels of immunoreactive trypsin in the blood of newborn infants with cystic fibrosis has provided a new way of detecting the disease soon after birth. A radioimmunoassay of trypsin in the eluate of blood dried on filter paper has now been developed. The sensitivity and accuracy of the method, as well as the good correlation observed between the values obtained and those of the conventional plasma assay, indicate that it is reliable and well adapted to the newborn. The new assay can easily be inserted into the present system of neonatal disease detection. A preliminary assessment of more than 5000 tests enables the authors to report an early diagnosis of proven cystic fibrosis and to discuss an essential aspect of mass-detection methods: the indicence of false-positive results [fr

  14. Activity of trypsin-like enzymes and gelatinases in rats with doxorubicin cardiomyopathy

    OpenAIRE

    Iu. А. Gordiienko; Ya. V. Babets; А. О. Kulinich; А. І. Shevtsova; G. О. Ushakova

    2014-01-01

    Activity of trypsin-like enzymes (ATLE) and gelatinases A and B were studied in the blood plasma and extracts from cardiac muscle, cerebral cortex and cerebellum of rats with cardiomyopathy caused by anthracycline antibiotic doxorubicin against the background of preventive application of corvitin and α-ketoglutarate. ATLE significantly increased in blood plasma and extracts from cerebral cortex but decreased in extracts from cardiac muscle and cerebellum in doxorubicin cardiomyopathy (DCMP). ...

  15. Structural and Chemical Characterization of Silica Spheres before and after Modification by Silanization for Trypsin Immobilization

    Directory of Open Access Journals (Sweden)

    Eduardo F. Barbosa

    2017-01-01

    Full Text Available In the last decades, silica particles of a variety of sizes and shapes have been characterized and chemically modified for several applications, from chromatographic separation to dental supplies. The present study proposes the use of aminopropyl triethoxysilane (APTS silanized silica particles to immobilize the proteolytic enzyme trypsin for the development of a bioreactor. The major advantage of the process is that it enables the polypeptides hydrolysis interruption simply by removing the silica particles from the reaction bottle. Silanized silica surfaces showed significant morphological changes at micro- and nanoscale level. Chemical characterization showed changes in elemental composition, chemical environment, and thermal degradation. Their application as supports for trypsin immobilization showed high immobilization efficiency at reduced immobilization times, combined with more acidic conditions. Indirect immobilization quantification by reversed-phase ultrafast high performance liquid chromatography proved to be a suitable approach due to its high linearity and sensitivity. Immobilized trypsin activities on nonmodified and silanized silica showed promising features (e.g., selective hydrolysis for applications in proteins/peptides primary structure elucidation for proteomics. Silanized silica system produced some preferential targeting peptides, probably due to the hydrophobicity of the nanoenvironment conditioned by silanization.

  16. Influence of carbohydrates on the interaction of procyanidin B3 with trypsin.

    Science.gov (United States)

    Gonçalves, Rui; Mateus, Nuno; De Freitas, Victor

    2011-11-09

    The biological properties of procyanidins, in particular their inhibition of digestive enzymes, have received much attention in the past few years. Dietary carbohydrates are an environmental factor that is known to affect the interaction of procyanidins with proteins. This work aimed at understanding the effect of ionic food carbohydrates (polygalacturonic acid, arabic gum, pectin, and xanthan gum) on the interaction between procyanidins and trypsin. Physical-chemical techniques such as saturation transfer difference-NMR (STD-NMR) spectroscopy, fluorescence quenching, and nephelometry were used to evaluate the interaction process. Using STD-NMR, it was possible to identify the binding of procyanidin B3 to trypsin. The tested carbohydrates prevented the association of procyanidin B3 and trypsin by a competition mechanism in which the ionic character of carbohydrates and their ability to encapsulate procyanidins seem crucial leading to a reduction in STD signal and light scattering and to a recovery of the proteins intrinsic fluorescence. On the basis of these results, it was possible to grade the carbohydrates in their aggregation inhibition ability: XG > PA > AG ≫ PC. These effects may be relevant since the coingestion of procyanidins and ionic carbohydrates are frequent and furthermore since these might negatively affect the antinutritional properties ascribed to procyanidins in the past.

  17. Radio-immuno-assay for trypsin in newborn-screening for cystic fibrosis

    International Nuclear Information System (INIS)

    Sander, J.; Niehaus, C.

    1982-01-01

    In 4,956 infants the concentration of immunoreactive trypsin was measured in dried blood on filter paper using a double antibody radioimmuno assay. About 90% of all results were below 40 ng/ml. In 13 infants the concentration of immunoreactive trypsin exceeded 80 ng/ml. These infants were examined clinically, including sweationtophoresis. We found three children suffering from cystic fibrosis. One further child showing an elevated concentration of chloride in the sweat (60 mval/ml) could not be reexamined. The concentration of immunoreactive trypsin in the cystic fibrosis children was 230, 297, and in one case 108 ng/ml at the 76th day of life, whereas the values for the 9 other children were between 80 and 154 ng/ml. We believe these results justify to use this test for a much higher number of infants, especially because it is inexpensive and can easily be added to existing newborn screening programs for inborn errors of metabolism. (orig.) [de

  18. Trypsin level in gallbladder bile and ductitis and width of the cystic duct.

    Science.gov (United States)

    Vracko, J; Wiechel, K L

    2000-01-01

    The change from laparotomy to laparoscopy for cholecystectomy has raised the question of how to manage concomitant bile duct stones. The present-day interest--and controversy--has focused on a transcystic approach reported to be feasible in 66-96% of cases, but without explaining the necessary prerequisite: the widening of the cystic duct. The cystic duct, wide mainly in patients with bile duct stones, has been reported to be highly variable: from strictured to very wide. The present study aims at comparing the trypsin level in the gallbladder bile and the cystic duct morphology and width in patients with and without bile duct stones. A prospective series of 63 gallstone patients, 30 with and 33 without bile duct stones (controls), underwent cholecystectomy and bile duct clearance. The study includes the trypsin level in the gallbladder bile, the width and morphology of the cystic duct, and the size of the gallstones. The patients with bile duct stones had, in contrast to the controls, higher trypsin levels in the gallbladder bile (P extraction feasible.

  19. Immobilization of trypsin on miniature incandescent bulbs for infrared-assisted proteolysis

    Energy Technology Data Exchange (ETDEWEB)

    Ge, Huimin; Bao, Huimin; Zhang, Luyan; Chen, Gang, E-mail: gangchen@fudan.edu.cn

    2014-10-03

    Highlights: • Trypsin was immobilized on miniature incandescent bulbs via chitosan coating. • The bulbs acted as enzymatic reactors and the generators of infrared radiation. • The bulb bioreactors were successfully employed in infrared-assisted proteolysis. • The proteolysis could accomplish within 5 min with high sequence coverages. - Abstract: A novel efficient proteolysis approach was developed based on trypsin-immobilized miniature incandescent bulbs and infrared (IR) radiation. Trypsin was covalently immobilized in the chitosan coating on the outer surface of miniature incandescent bulbs with the aid of glutaraldehyde. When an illuminated enzyme-immobilized bulb was immersed in protein solution, the emitted IR radiation could trigger and accelerate heterogeneous protein digestion. The feasibility and performance of the novel proteolysis approach were demonstrated by the digestion of hemoglobin (HEM), cytochrome c (Cyt-c), lysozyme (LYS), and ovalbumin (OVA) and the digestion time was significantly reduced to 5 min. The obtained digests were identified by MALDI-TOF-MS with the sequence coverages of 91%, 77%, 80%, and 52% for HEM, Cyt-c, LYS, and OVA (200 ng μL{sup −1} each), respectively. The suitability of the prepared bulb bioreactors to complex proteins was demonstrated by digesting human serum.

  20. Trypsin digest protocol to analyze the retinal vasculature of a mouse model.

    Science.gov (United States)

    Chou, Jonathan C; Rollins, Stuart D; Fawzi, Amani A

    2013-06-13

    Trypsin digest is the gold standard method to analyze the retinal vasculature (1-5). It allows visualization of the entire network of complex three-dimensional retinal blood vessels and capillaries by creating a two-dimensional flat-mount of the interconnected vascular channels after digestion of the non-vascular components of the retina. This allows one to study various pathologic vascular changes, such as microaneurysms, capillary degeneration, and abnormal endothelial to pericyte ratios. However, the method is technically challenging, especially in mice, which have become the most widely available animal model to study the retina because of the ease of genetic manipulations (6,7). In the mouse eye, it is particularly difficult to completely remove the non-vascular components while maintaining the overall architecture of the retinal blood vessels. To date, there is a dearth of literature that describes the trypsin digest technique in detail in the mouse. This manuscript provides a detailed step-by-step methodology of the trypsin digest in mouse retina, while also providing tips on troubleshooting difficult steps.

  1. Preparation and characterization of magnetic levan particles as matrix for trypsin immobilization

    Energy Technology Data Exchange (ETDEWEB)

    Maciel, J.C. [Programa de Pos-Graduacao em Ciencias Biologicas, Universidade Federal de Pernambuco, Cidade Universitaria, 50670-901 Recife, PE (Brazil); Andrad, P.L. [Programa de Pos-Graduacao em Ciencia de Materiais, Universidade Federal de Pernambuco, Cidade Universitaria, 50679-901 Recife, PE (Brazil); Neri, D.F.M., E-mail: davidfmneri@yahoo.com.br [Universidade Federal do Vale do Sao Francisco, 56304-205 Petrolina, PE (Brazil); Carvalho, L.B. [Departamento de Bioquimica, Universidade Federal de Pernambuco, Cidade Universitaria, 50679-901 Recife, PE (Brazil); Cardoso, C.A. [Departamento de Fisica, Universidade Federal de Sao Carlos, 13565-905 Sao Carlos, PE (Brazil); Calazans, G.M.T. [Departamento de Antibioticos, Universidade Federal de Pernambuco, Cidade Universitaria, 50670-901 Recife, PE (Brazil); Albino Aguiar, J. [Departamento de Fisica, Universidade Federal de Pernambuco, Cidade Universitaria, 50679-901 Recife, PE (Brazil); Silva, M.P.C. [Departamento de Bioquimica, Universidade Federal de Pernambuco, Cidade Universitaria, 50679-901 Recife, PE (Brazil)

    2012-04-15

    Magnetic levan was synthesized by co-precipitating D-fructofuranosyl homopolysaccharide with a solution containing Fe{sup 2+} and Fe{sup 3+} in alkaline conditions at 100 Degree-Sign C. The magnetic levan particles were characterized by scanning electron microscopy (SEM), magnetization measurements, X-ray diffractometry (XRD) and infrared spectroscopy (IR). Afterwards, magnetic levan particles were functionalized by NaIO{sub 4} oxidation and used as matrices for trypsin covalent immobilization. Magnetite and magnetic levan particles were both heterogeneous in shape and levan-magnetite presented bigger sizes compared to magnetite according to SEM images. Magnetic levan particles exhibited a magnetization 10 times lower as compared to magnetite ones, probably, due to the coating layer. XRD diffractogram showed that magnetite is the dominant phase in the magnetic levan. Infrared spectroscopy showed characteristics absorption bands of levan and magnetite (O-H, C-O-C and Fe-O bonds). The immobilized trypsin derivative was reused 10 times and lost 16% of its initial specific activity only. Therefore, these magnetic levan particles can be proposed as an alternative matrices for enzyme immobilization. - Highlights: Black-Right-Pointing-Pointer The magnetic levan particles presented larger size variation than magnetite particles due to the changes produced by coating. Black-Right-Pointing-Pointer The utilization of magnetic levan particles showed to be efficacious for immobilization of enzymes as trypsin. Black-Right-Pointing-Pointer Magnetic particles can be planned as other matrix for immobilization of biomolecule in various division processes in biotechnology.

  2. Wandering spleen associated with omphalocele in a neonate: An unusual case with non-operative management

    Directory of Open Access Journals (Sweden)

    Daisuke Masui

    2017-09-01

    Full Text Available Wandering spleen with a history of omphalocele is extremely rare. We encountered a male baby with wandering spleen associated with omphalocele. This case of wandering spleen in a neonate was diagnosed by ultrasound and computed tomography scans after surgery for omphalocele. Our case was able to be managed non-operatively due to the lack of any findings suggesting torsion of the spleen and its asymptomatic status. We herein report the clinical presentation as well as the treatment options regarding wandering spleen associated with omphalocele.

  3. Transplantation of homologous bone marrow cells to lethally irradiated mice: changes in the spleen

    Energy Technology Data Exchange (ETDEWEB)

    Viktora, L; Hach, P; Zoubkova, M

    1975-01-01

    Bone marrow cell suspensions were administered intravenously to lethally irradiated mice. The number of colonies in the spleen and the regeneration of hematopoietic tissue in the spleen were studied on the 9th day after irradiation and transplantation. From a comparison of the histological picture and weight of the spleens, the authors conclude that the degree of regeneration of hematopoiesis in the spleen after irradiation and transplantation is reflected in the weight of the spleen as well as in the number of hematopoietic colonies.

  4. Thermostable trypsin conjugates immobilized to biogenic magnetite show a high operational stability and remarkable reusability for protein digestion

    Science.gov (United States)

    Pečová, M.; Šebela, M.; Marková, Z.; Poláková, K.; Čuda, J.; Šafářová, K.; Zbořil, R.

    2013-03-01

    In this work, magnetosomes produced by microorganisms were chosen as a suitable magnetic carrier for covalent immobilization of thermostable trypsin conjugates with an expected applicability for efficient and rapid digestion of proteins at elevated temperatures. First, a biogenic magnetite was isolated from Magnetospirillum gryphiswaldense and its free surface was coated with the natural polysaccharide chitosan containing free amino and hydroxy groups. Prior to covalent immobilization, bovine trypsin was modified by conjugating with α-, β- and γ-cyclodextrin. Modified trypsin was bound to the magnetic carriers via amino groups using 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide and N-hydroxysulfosuccinimide as coupling reagents. The magnetic biomaterial was characterized by magnetometric analysis and electron microscopy. With regard to their biochemical properties, the immobilized trypsin conjugates showed an increased resistance to elevated temperatures, eliminated autolysis, had an unchanged pH optimum and a significant storage stability and reusability. Considering these parameters, the presented enzymatic system exhibits properties that are superior to those of trypsin forms obtained by other frequently used approaches. The proteolytic performance was demonstrated during in-solution digestion of model proteins (horseradish peroxidase, bovine serum albumin and hen egg white lysozyme) followed by mass spectrometry. It is shown that both magnetic immobilization and chemical modification enhance the characteristics of trypsin making it a promising tool for protein digestion.

  5. Label-Free Fluorescent Detection of Trypsin Activity Based on DNA-Stabilized Silver Nanocluster-Peptide Conjugates

    Directory of Open Access Journals (Sweden)

    Cai-Xia Zhuo

    2016-11-01

    Full Text Available Trypsin is important during the regulation of pancreatic exocrine function. The detection of trypsin activity is currently limited because of the need for the substrate to be labeled with a fluorescent tag. A label-free fluorescent method has been developed to monitor trypsin activity. The designed peptide probe consists of six arginine molecules and a cysteine terminus and can be conjugated to DNA-stabilized silver nanoclusters (DNA-AgNCs by Ag-S bonding to enhance fluorescence. The peptide probe can also be adsorbed to the surface of graphene oxide (GO, thus resulting in the fluorescence quenching of DNA-AgNCs-peptide conjugate because of Förster resonance energy transfer. Once trypsin had degraded the peptide probe into amino acid residues, the DNA-AgNCs were released from the surface of GO, and the enhanced fluorescence of DNA-AgNCs was restored. Trypsin can be determined with a linear range of 0.0–50.0 ng/mL with a concentration as low as 1 ng/mL. This label-free method is simple and sensitive and has been successfully used for the determination of trypsin in serum. The method can also be modified to detect other proteases.

  6. Long-term following-up of viability of spleen autotransplants in the Beagle canine model.

    Science.gov (United States)

    Sajtos, Erika; Balint, Anita; Brath, Endre; Nemeth, Norbert; Peto, Katalin; Kovacs, Judit; Galuska, Laszlo; Varga, Jozsef; Fodor, Zoltan; Furka, Istvan; Miko, Iren

    2012-02-01

    To examine the possible late complications of splenectomy or spleen autotransplantation in large laboratory animal model, in which we need non-invasive or minimal-invasive methods for long-term monitoring of the experimental animals. Experimental groups of beagle dogs were: non-operated control, sham-operated control, splenectomy, spleen autotransplantation with 5 or 10 spleen-chips taken into the greater omentum (Furka's technique). Prior to operations, on the 1(st) postoperative week, monthly till the 6(th) as well as in the 9(th) and 12(th) month, hemorheological examinations were performed. In postoperative 12(th) month colloid scintigraphy and diagnostic laparoscopy were carried out. At the end of the investigation comparative morphological examinations were performed, too. From the 4(th)-5(th) postoperative month filtration function of spleen-autotransplants showed particular restoration compared to splenectomy group. However, the functional results did not reach the values of the control or sham-operated groups. Sham-operated control's scintigraphy nicely showed activity in the spleen. In spleen autotransplantation-groups scintigraphy indicated well the activity of spleen-chips. During diagnostic laparoscopy spleen-chips with their blood supply were found. Histologically, the structure of spleen-autotransplants was similar to normal splenic tissue. The autotransplants are regenerated, their functions have been partly restored, and thus spleen autotransplantation may prevent the possible complications of splenectomy. These parameters and the presented investigative protocol are suitable for long-term following-up of viability of the spleen-autotransplants.

  7. Management of Blunt Trauma to the Spleen (Part 1

    Directory of Open Access Journals (Sweden)

    Seyed Abbas Banani

    2010-06-01

    Full Text Available AbstractSpleen is the most frequent solid organ to be injured in bluntabdominal trauma. Considering its important role in providingimmunity and preventing infection by a variety of mechanisms,every attempt should be made, if possible, to salvagethe traumatized spleen at any age particularly in children. Afterprimary resuscitation, mandatory requirements for nonoperativemanagement include absence of homodynamic instability,lack of associated major organ injury, admission inthe intensive care unit for high-grade splenic injury and in theward for milder types with close monitoring. About two thirdof the patients would respond to non-operative management.In most patients, failure of non-operative measures usuallyoccur within 12 hours of management. Determinant role ofabdominal sonography or computed tomography, and in selectedcases, diagnostic peritoneal lavage, for appropriate decisioncannot be overemphasized. However, the high status ofclinical judgment would not be replaced by any paraclinicalinvestigations. When operation is unavoidable, if possible,spleen saving procedures (splenorrhaphy or partial splenectomyshould be tried. In case of total splenectomy, autotransplantation,preferably in the omental pouch, may lead toreturn of immunity, at least partially, to prevent or reduce thechance of subsequent infection. Although total splenectomywith autograft is immunologically superior to total splenectomy-only procedure, these patients should also be protectedby vaccination and daily antibiotic for certain period of time.The essential steps for prevention of overwhelming infectionafter total splenectomy are not only immunization and administrationof daily antibiotic (up to 5 years of age or one year inolder children, but include education and information aboutthis dangerous complication. When non-operative managementis successful, the duration of activity restriction (inweeks is almost equal to the grade of splenic injury plus 2.Iran J Med Sci 2010

  8. Laparoscopic splenectomy for spontaneous rupture of the spleen

    Directory of Open Access Journals (Sweden)

    Pinky M Thapar

    2016-01-01

    Full Text Available Laparoscopic splenectomy is a gold standard for management of planned benign splenic pathologies. Spontaneous rupture of the spleen (SRS leading to acute abdomen occurs in only 1% of all splenic ruptures. Laparoscopic splenectomy in traumatic and atraumatic rupture due to intra-splenic pathology is reported. We present the first reported case of laparoscopic splenectomy in a 23-year-old male who presented with hemoperitoneum due to idiopathic or SRS. The procedure was safely accomplished with slight modified technique and minimum usage of advanced gadgets.

  9. Mucormycosis resulting in a pseudoaneurysm in the spleen

    Energy Technology Data Exchange (ETDEWEB)

    Nevitt, P.C.; Das Narla, L. [Medical College of Virginia, Virginia Commonwealth Univ., Richmond (United States); Hingsbergen, E.A. [Children' s Radiologic Institute, Children' s Hospital, Columbus, OH (United States)

    2001-02-01

    Mucormycosis is an uncommon and frequently fatal fungal infection. It characteristically affects patients with diabetes mellitus or patients with severe immunosuppression. The hallmark of mucormycosis infection is tissue infarction and vascular invasion. We present clinical data and imaging studies of a 16 year-old child with acute lymphoblastic leukemia complicated by disseminated mucormycosis resulting in a pseudoaneurysm of the spleen. This was successfully managed by a combination of systemic antifungal therapy (Amphotericin B) and surgery (splenectomy). This entity has not been described in the literature. (orig.)

  10. Mucormycosis resulting in a pseudoaneurysm in the spleen

    International Nuclear Information System (INIS)

    Nevitt, P.C.; Das Narla, L.; Hingsbergen, E.A.

    2001-01-01

    Mucormycosis is an uncommon and frequently fatal fungal infection. It characteristically affects patients with diabetes mellitus or patients with severe immunosuppression. The hallmark of mucormycosis infection is tissue infarction and vascular invasion. We present clinical data and imaging studies of a 16 year-old child with acute lymphoblastic leukemia complicated by disseminated mucormycosis resulting in a pseudoaneurysm of the spleen. This was successfully managed by a combination of systemic antifungal therapy (Amphotericin B) and surgery (splenectomy). This entity has not been described in the literature. (orig.)

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

    International Nuclear Information System (INIS)

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

    1974-11-01

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

  12. [Spleen autotransplant. Natural history and description of a case].

    Science.gov (United States)

    Ceccherini, E; Sereni, P; Ferrari, F; Fagioli Zucchi, A; Croce, F; Di Maggio, G; Vattimo, A; Mancini, S

    1989-09-30

    After considering the natural history of spleen auto-transplant, a clinical case followed up for seven months with instrumental (echography, scintigraphy) and humoral (Jolly bodies, Heinz bodies, reticulocytes, platelets, complement, immune globulin) examinations has been considered so as to verify "take" and function. One months after reimplantation the patient was again operated on for the onset of an intestinal occlusion due to adherences. On that occasion it was possible to control that the implant had taken. It is concluded that personally used parameters proved to be well correlated and that scintigraphy and echography are two complementary, effective techniques for monitoring auto-transplants.

  13. The spleen in the sickling disorders: an update

    International Nuclear Information System (INIS)

    Khatib, Rana; Sarnaik, Sharada A.; Rabah, Raja

    2009-01-01

    In early life, patients with sickle cell disease (SCD) can have acute, life-threatening emergencies related to splenic hypofunction (overwhelming bacterial sepsis), as well as anemic crises from acute splenic sequestration because of sudden pooling of blood in the spleen. The landmark penicillin prophylaxis study in 1985 showed a remarkable decrease in mortality from sepsis in young children with SCD who were treated with oral penicillin prophylaxis compared to placebo. Since that study, newborns are screened for SCD and placed on oral penicillin prophylaxis in nearly all of the United States, as well as in other countries where the disease is highly prevalent. The previously described permanent, complete and nearly universal ''autosplenectomy'' emerging by late childhood or early adulthood is now challenged by recent findings of reversibility of splenic dysfunction by the antisickling drug hydroxyurea or by successful allogeneic stem cell transplantation, even in older patients. Imaging techniques for hypofunction of the spleen are the most commonly used modalities to guide the clinician in decisions regarding medical or surgical management. (orig.)

  14. Data from quantitative label free proteomics analysis of rat spleen.

    Science.gov (United States)

    Dudekula, Khadar; Le Bihan, Thierry

    2016-09-01

    The dataset presented in this work has been obtained using a label-free quantitative proteomic analysis of rat spleen. A robust method for extraction of proteins from rat spleen tissue and LC-MS-MS analysis was developed using a urea and SDS-based buffer. Different fractionation methods were compared. A total of 3484 different proteins were identified from the pool of all experiments run in this study (a total of 2460 proteins with at least two peptides). A total of 1822 proteins were identified from nine non-fractionated pulse gels, 2288 proteins and 2864 proteins were identified by SDS-PAGE fractionation into three and five fractions respectively. The proteomics data are deposited in ProteomeXchange Consortium via PRIDE PXD003520, Progenesis and Maxquant output are presented in the supported information. The generated list of proteins under different regimes of fractionation allow assessing the nature of the identified proteins; variability in the quantitative analysis associated with the different sampling strategy and allow defining a proper number of replicates for future quantitative analysis.

  15. Elastography of the normal canine liver, spleen and kidneys.

    Science.gov (United States)

    Holdsworth, Andrew; Bradley, Kate; Birch, Sally; Browne, William J; Barberet, Virginie

    2014-01-01

    Elastography is a simple, expedient and noninvasive technique that may be used to assess the elasticity or stiffness of a tissue, in conjunction with traditional B-mode ultrasonography. Quantitative assessment of tissue stiffness can be made which involves measurement of the shear wave velocity within the tissue of interest. The goal of this study was to assess the feasibility of elastography for clinical use in the abdomen of conscious small animals and to investigate factors that affect shear wave velocity measurement. Elastography was performed on the liver, spleen, and kidneys of 15 dogs at predefined depths within the parenchyma. Breed, age, gender, neuter status, and weight were documented for each animal. Depth at which measurements were taken had a significant negative relationship with the shear wave velocity value obtained. Individual dog effects, such as weight and gender, also appeared to have a significant effect on the shear wave velocity measurement for specific organs; weight had a significant positive effect on the shear wave velocity for each of the organs examined, whereas the effect of gender was inconsistent between organs (having a positive effect for the liver and a negative effect for the spleen). It is hoped that these results may act as a baseline to guide further work into the field of elastography in companion animals. © 2014 American College of Veterinary Radiology.

  16. The spleen in the sickling disorders: an update

    Energy Technology Data Exchange (ETDEWEB)

    Khatib, Rana; Sarnaik, Sharada A. [Wayne State University School of Medicine, Children' s Hospital of Michigan, Carmen and Ann Adams Department of Pediatrics, Detroit, MI (United States); Rabah, Raja [Wayne State University School of Medicine, Department of Pathology, Detroit, MI (United States)

    2009-01-15

    In early life, patients with sickle cell disease (SCD) can have acute, life-threatening emergencies related to splenic hypofunction (overwhelming bacterial sepsis), as well as anemic crises from acute splenic sequestration because of sudden pooling of blood in the spleen. The landmark penicillin prophylaxis study in 1985 showed a remarkable decrease in mortality from sepsis in young children with SCD who were treated with oral penicillin prophylaxis compared to placebo. Since that study, newborns are screened for SCD and placed on oral penicillin prophylaxis in nearly all of the United States, as well as in other countries where the disease is highly prevalent. The previously described permanent, complete and nearly universal ''autosplenectomy'' emerging by late childhood or early adulthood is now challenged by recent findings of reversibility of splenic dysfunction by the antisickling drug hydroxyurea or by successful allogeneic stem cell transplantation, even in older patients. Imaging techniques for hypofunction of the spleen are the most commonly used modalities to guide the clinician in decisions regarding medical or surgical management. (orig.)

  17. Spleen lymphocyte function modulated by a cocoa-enriched diet.

    Science.gov (United States)

    Ramiro-Puig, E; Pérez-Cano, F J; Ramírez-Santana, C; Castellote, C; Izquierdo-Pulido, M; Permanyer, J; Franch, A; Castell, M

    2007-09-01

    Previous studies have shown the down-regulating in vitro effect of cocoa flavonoids on lymphocyte and macrophage activation. In the present paper, we report the capacity of a long-term rich cocoa diet to modulate macrophage cytokine secretion and lymphocyte function in young rats. Weaned rats received natural cocoa (4% or 10% food intake), containing 32 mg flavonoids/g, for 3 weeks. Spleen immune function was then evaluated through the analysis of lymphocyte composition, their proliferative response and their ability to secrete cytokines and Ig. In addition, the status of activated peritoneal macrophages was established through tumour necrosis factor (TNF)-alpha secretion. The richest cocoa diet (10%) caused a reduction of TNF-alpha secretion by peritoneal macrophages showing anti-inflammatory activity. Similarly, although a 10% cocoa diet increased lymphocyte proliferation rate, it down-regulated T helper 2 (Th2)-related cytokines and decreased Ig secretion. These changes were accompanied by an increase in spleen B cell proportion and a decrease in Th cell percentage. In summary, these results demonstrate the functional activity of a cocoa-high dosage in down-regulating the immune response that might be beneficial in hypersensitivity and autoimmunity.

  18. Decreasing the amount of trypsin in in-gel digestion leads to diminished chemical noise and improved protein identifications.

    Science.gov (United States)

    Hu, Mo; Liu, Yanhua; Yu, Kaiwen; Liu, Xiaoyun

    2014-09-23

    Pre-fractionation by gel electrophoresis is often combined with liquid chromatography-mass spectrometry (LC-MS) for large-scale profiling of complex protein samples. An essential component of this widely applied proteomic platform is in-gel protein digestion. In nearly two decades of practicing this approach, an extremely high level of trypsin has been utilized due to the consideration of slow enzyme diffusion into the gel matrix. Here we report that trypsin autolysis products contribute to the bulk of chemical noise in in-gel digestion and remarkably we found evidence that the amount of trypsin can be slashed by an order of magnitude with comparable digestion performance. By revising perhaps the most critical element of this decade-old digestion protocol, the proteomics community relying on gel separation prior to LC-MS analysis will benefit instantly from much lowered cost due to enzyme expenditure. More importantly, substantially reduced chemical noise (i.e., trypsin self-cleavage products) as a result of less enzyme usage translates into more protein identifications when limited amounts of samples are the interest of interrogation. In-gel digestion is one of the most widely used methods in proteomics. An exceedingly high level of trypsin has been utilized due to the consideration of slow enzyme diffusion into the gel matrix. This requirement has been faithfully kept in nearly two decades of practicing this approach. Here we report that trypsin concentration can be slashed by at least an order of magnitude while still providing comparable digestion performance. Thus the proteomics community relying on gel separation prior to LC-MS analysis will benefit instantly from much lowered enzyme cost. More importantly, substantially reduced chemical noise (i.e., trypsin autolysis products) due to less enzyme usage translates into ~30% more protein identifications when limited amounts of protein samples are analyzed. Copyright © 2014 Elsevier B.V. All rights reserved.

  19. Organization of the expanded cumulus-extracellular matrix in preovulatory follicles: arole for inter-alpha-trypsin inhibitor.

    Czech Academy of Sciences Publication Activity Database

    Nagyová, Eva

    2015-01-01

    Roč. 49, č. 1 (2015), s. 37-45 ISSN 1210-0668 R&D Projects: GA ČR GA305/05/0960 Institutional support: RVO:67985904 Keywords : cumulus expansion * cumulus-extracellular matrix * hyaluronan Subject RIV: ED - Physiology

  20. Covalent transfer of interalpha-trypsin inhibitor heavy chains to hyaluronan in porcine oocyte-cumulus complexes cultured in vitro

    Czech Academy of Sciences Publication Activity Database

    Nagyová, Eva; Camaioni, A.; Procházka, Radek; Salustri, A.

    2003-01-01

    Roč. 68, - (2003), s. 362 ISSN 0006-3363. [ SSR Annual meeting/36./. 19.07.2003-22.07.2003, Cincinnati] Institutional research plan: CEZ:AV0Z5045916 Keywords : porcine oocyte Subject RIV: ED - Physiology

  1. Trypsin as enhancement in cyclical tracheal decellularization: Morphological and biophysical characterization

    Energy Technology Data Exchange (ETDEWEB)

    Giraldo-Gomez, D.M., E-mail: davidmauro2008@gmail.com [Posgrado en Ciencia e Ingeniería de Materiales, Universidad Nacional Autónoma de México (UNAM), Unidad de Posgrado Edificio “C” 1er Piso, Circuito de Posgrados, Avenida Universidad 3000, Ciudad Universitaria, Coyoacán, C.P. 04510, México D. F., México (Mexico); Instituto de Investigaciones en Materiales, Universidad Nacional Autónoma de México (UNAM), Circuito Exterior, Avenida Universidad 3000, Ciudad Universitaria, Coyoacán, C.P. 04510, México D.F., México (Mexico); Leon-Mancilla, B. [Departamento de Cirugía, Facultad de Medicina, Universidad Nacional Autónoma de México (UNAM), Edificio “D” Planta Baja, Circuito Interior, Avenida Universidad 3000, Ciudad Universitaria, Coyoacán, C.P. 04510, México D.F., México (Mexico); Del Prado-Audelo, M.L. [Instituto de Investigaciones en Materiales, Universidad Nacional Autónoma de México (UNAM), Circuito Exterior, Avenida Universidad 3000, Ciudad Universitaria, Coyoacán, C.P. 04510, México D.F., México (Mexico); and others

    2016-02-01

    There are different types of tracheal disorders (e.g. cancer, stenosis and fractures). These can cause respiratory failure and lead to death of patients. Several attempts have been made for trachea replacement in order to restore the airway, including anastomosis and implants made from synthetic or natural materials. Tracheal allotransplantation has shown high rejection rates, and decellularization has emerged as a possible solution. Decellularization involves the removal of antigens from cells in the organ or tissue, leaving a matrix that can be used as 3D cell-scaffold. Although this process has been used for tracheal replacement, it usually takes at least two months and time is critical for patients with tracheal disorders. Therefore, there is necessary to develop a tracheal replacement process, which is not only effective, but also quick to prepare. The aim of this research was to develop a faster trachea decellularization protocol using Trypsin enzyme and Ethylenediaminetetraacetic acid (EDTA) as decellularization agents. Three protocols of cyclic trachea decellularization (Protocols A, B, and C) were compared. Following Protocol A (previously described in the literature), 15 consecutive cycles were performed over 32 days. Protocol B (a variation of Protocol A) — EDTA being added — with 15 consecutive cycles performed over 60 days. Finally, Protocol C, with the addition of Trypsin as a decellularization agent, 5 consecutive cycles being performed over 10 days. For the three protocols, hematoxylin–eosin (H&E) staining and DNA residual content quantification were performed to establish the effectiveness of the decellularization process. Scanning Electron Microscopy (SEM) was used to observe the changes in porosity and microarrays. To evaluate the structural matrices integrity, Thermogravimetric Analysis (TGA) and biomechanical test were used. None of the protocols showed significant alteration or degradation in the components of the extracellular matrix

  2. Differential antibody production by adherent and nonadherent spleen cells transferred to irradiated and cyclophosphamide-treated recipient mice

    International Nuclear Information System (INIS)

    Albright, J.F.; Deitchman, J.W.; Hassell, S.A.; Ozato, K.

    1975-01-01

    Mouse spleen cells were separated into adherent (Ad) and nonadherent (Nad) populations by incubation in plastic petri dishes. Adherent, Nad and unfractionated cell preparations (UCP) were transferred into syngeneic recipient mice that had been either irradiated or cyclophosphamide (CY) treated and the adoptive humoral Ab responses were studied by assessment of hemolytic Ab-forming cells (PFC) or humoral serum Ab production. Adherent cells failed to produce PFC in irradiated recipients, but functioned vigorously in CY-treated recipients. Nonadherent cells generated PFC in either type of host, as did UCP. Studies of comparative responses in CY-treated recipients revealed that: (a) Ad-cells generated 2 / 3 the number of PFC given by equivalent numbers of transferred Nad cells and UCP; (b) per equivalent numbers of transferred cells the Ad fraction generated 5 times more and 16 times more Ab than did the Nad cells and UCP, respectively. Spleen cells taken from mice 6 hr after CY treatment failed to respond to the mitogens phytohemagglutinin and bacterial lipopolysaccharide, showing that all cells were temporarily incapable of proliferation. Transfer of spleen cells from donor mice 16 hr after CY treatment, into thymectomized, irradiated, bone marrow-reconstituted recipients revealed substantial T-helper cell activity. We conclude that: (a) Ad preparations lacked T cells that were supplied by CY-treated recipients although T cell proliferation was temporarily inhibited in the latter; (b) B cells present in the Ad fraction were removed from some type of inhibitor of Ab synthesis and/or secretion, the production of which may be associated with T cells present in Nad preparations and UCP; (c) T-helper cells were only transiently affected by CY

  3. A novel poly(deep eutectic solvent)-based magnetic silica composite for solid-phase extraction of trypsin

    International Nuclear Information System (INIS)

    Xu, Kaijia; Wang, Yuzhi; Li, Yixue; Lin, Yunxuan; Zhang, Haibao; Zhou, Yigang

    2016-01-01

    Novel poly(deep eutectic solvent) grafted silica-coated magnetic microspheres (Fe 3 O 4 @SiO 2 -MPS@PDES) were prepared by polymerization of choline chloride-itaconic acid (ChCl-IA) and γ-MPS-modified magnetic silica composites, and were characterized by vibrating sample magnetometer (VSM), Fourier transform infrared spectrometry (FT-IR), X-ray photoelectron spectra (XPS), thermal gravimetric analysis (TGA) and transmission electron microscope (TEM). Then the synthetic Fe 3 O 4 @SiO 2 -MPS@PDES microspheres were applied for the magnetic solid-phase extraction (MSPE) of trypsin for the first time. After extraction, the concentration of trypsin in the supernatant was determined by a UV–vis spectrophotometer. Single factor experiments were carried out to investigate the effects of the extraction process, including the concentration of trypsin, the ionic strength, the pH value, the extraction time and the temperature. Experimental results showed the extraction capacity could reach up to 287.5 mg/g under optimized conditions. In comparison with Fe 3 O 4 @SiO 2 -MPS, Fe 3 O 4 @SiO 2 -MPS@PDES displayed higher extraction capacity and selectivity for trypsin. According to the regeneration studies, Fe 3 O 4 @SiO 2 -MPS@PDES microspheres can be recycled six times without significant loss of its extraction capacity, and retained a high extraction capacity of 233 mg/g after eight cycles. Besides, the activity studies also demonstrated that the activity of the extracted trypsin was well retained. Furthermore, the analysis of real sample revealed that the prepared magnetic microspheres can be used to purify trypsin in crude bovine pancreas extract. These results highlight the potential of the proposed Fe 3 O 4 @SiO 2 -MPS@PDES-MSPE method in separation of biomolecules. - Highlights: • A strategy for solid-phase extraction of trypsin based on poly(deep eutectic solvent) modified magnetic silica microspheres. • Fe 3 O 4 @SiO 2 -MPS@PDES showed higher extraction capacity

  4. A novel poly(deep eutectic solvent)-based magnetic silica composite for solid-phase extraction of trypsin

    Energy Technology Data Exchange (ETDEWEB)

    Xu, Kaijia [State Key Laboratory of Chemo/Biosensing and Chemometrics, College of Chemistry and Chemical Engineering, Hunan University, Changsha, 410082 (China); Wang, Yuzhi, E-mail: wyzss@hnu.edu.cn [State Key Laboratory of Chemo/Biosensing and Chemometrics, College of Chemistry and Chemical Engineering, Hunan University, Changsha, 410082 (China); Li, Yixue; Lin, Yunxuan; Zhang, Haibao [State Key Laboratory of Chemo/Biosensing and Chemometrics, College of Chemistry and Chemical Engineering, Hunan University, Changsha, 410082 (China); Zhou, Yigang [Department of Microbiology, College of Basic Medicine, Central South University, Changsha, 410083 (China)

    2016-11-23

    Novel poly(deep eutectic solvent) grafted silica-coated magnetic microspheres (Fe{sub 3}O{sub 4}@SiO{sub 2}-MPS@PDES) were prepared by polymerization of choline chloride-itaconic acid (ChCl-IA) and γ-MPS-modified magnetic silica composites, and were characterized by vibrating sample magnetometer (VSM), Fourier transform infrared spectrometry (FT-IR), X-ray photoelectron spectra (XPS), thermal gravimetric analysis (TGA) and transmission electron microscope (TEM). Then the synthetic Fe{sub 3}O{sub 4}@SiO{sub 2}-MPS@PDES microspheres were applied for the magnetic solid-phase extraction (MSPE) of trypsin for the first time. After extraction, the concentration of trypsin in the supernatant was determined by a UV–vis spectrophotometer. Single factor experiments were carried out to investigate the effects of the extraction process, including the concentration of trypsin, the ionic strength, the pH value, the extraction time and the temperature. Experimental results showed the extraction capacity could reach up to 287.5 mg/g under optimized conditions. In comparison with Fe{sub 3}O{sub 4}@SiO{sub 2}-MPS, Fe{sub 3}O{sub 4}@SiO{sub 2}-MPS@PDES displayed higher extraction capacity and selectivity for trypsin. According to the regeneration studies, Fe{sub 3}O{sub 4}@SiO{sub 2}-MPS@PDES microspheres can be recycled six times without significant loss of its extraction capacity, and retained a high extraction capacity of 233 mg/g after eight cycles. Besides, the activity studies also demonstrated that the activity of the extracted trypsin was well retained. Furthermore, the analysis of real sample revealed that the prepared magnetic microspheres can be used to purify trypsin in crude bovine pancreas extract. These results highlight the potential of the proposed Fe{sub 3}O{sub 4}@SiO{sub 2}-MPS@PDES-MSPE method in separation of biomolecules. - Highlights: • A strategy for solid-phase extraction of trypsin based on poly(deep eutectic solvent) modified magnetic silica

  5. Inibição da tripsina de bicho-mineiro do cafeeiro por um fator não-protéico presente em extratos de folhas de mamona Coffee leaf miner trypsin inhibition with castor bean leaf extracts mediated by a non-protein agent

    Directory of Open Access Journals (Sweden)

    Guilherme Duarte Rossi

    2010-04-01

    Full Text Available Inibidores de tripsina representam uma estratégia de controle de insetos e, por isso, a identificação e caracterização desses inibidores são etapas muito importantes para que novas formas de controle de pragas sejam desenvolvidas. Os inibidores de tripsina atuam na digestão primária de proteínas e comprometem o processo digestivo por completo, reduzindo a disponibilidade de aminoácidos ao inseto. A incorporação de inibidores de tripsina na dieta de insetos-praga é uma forma de controle cuja eficácia foi verificada por diferentes autores. Este projeto foi conduzido a fim de se observar a eficiência de extratos de folhas de mamona na inibição "in vitro" de proteinases do tipo tripsina do bicho-mineiro do cafeeiro. Após testes realizados com os extratos de folhas de mamona não-fervidos e fervidos com e sem a adição de β-mercaptoetanol 0,2% (v/v e mediante precipitações com acetona, verificou-se que o inibidor é uma molécula termoresistente e não-protéica. Desta forma, iniciou-se um processo de purificação da molécula inibidora por meio de cromatografia de adsorção com posterior análise em espectrômetro de massas. Os resultados dos testes de inibição indicaram a presença de um inibidor de tripsina eficaz contra o bicho-mineiro do cafeeiro nos extratos de folhas de mamona capaz de inibir 2,48 + 0,15 UTI, o que representa aproximadamente 40% de inibição. Em testes realizados com tripsina bovina observou-se que o extrato de folhas de mamona não apresenta poder de inibição sobre essa enzima.Trypsin inhibitors stand for a strategy of insect control and, therefore, the identification and characterization of these inhibitors are very important steps for new forms of pest control to be developed. Trypsin inhibitors act in the primary digestion of proteins and endanger the digestive process wholly, reducing the availability of aminoacids to the insect. The incorporation of trypsin inhibitors in the diet of pest

  6. The immobilisation of trypsin and glucose oxidase onto natural rubber-g.co-HEMA - high energy radiation derived copolymeric support systems

    International Nuclear Information System (INIS)

    Devi, S.; Guthrie, J.T.; Beddows, C.G.

    1990-01-01

    Natural rubber has been grafted with 2-HEMA by three different methods each involving Co(60)γ-radiation as the initiation source. The grafted samples were used in the immobilisation of glucose oxidase and trypsin. Optimisation of immobilisation involving trypsin was studied with regard to the pH and the type of crosslinking agent. It was observed that the immobilised enzyme had superior stability over a wider pH range when compared to the free trypsin. The retention of activity demonstrated by the immobilised trypsin was significant. That of immobilised glucose oxidase was far from being satisfactory. (author)

  7. Pathomorphology of spleen lymphocyte apoptosis in large dose 60Co γ-irradiated mice

    International Nuclear Information System (INIS)

    Gao Linlu; Cui Yufang; Yang Hong; Xia Guowei; Peng Ruiyun; Gao Yabing; Wang Dewen

    2000-01-01

    Objective: The aim of the authors was to investigate the pathomorphology changes of spleen lymphocyte apoptosis after 60 Co γ-irradiation. Methods: The mice were irradiated with 6, 9, 12, 15 and 20 Gy of 60 Co γ-rays. At different times after irradiation, the mice were sacrificed and the pathological changes of spleen lymphocyte were observed by light and transmission electron microscopies. Results: Spleen lymphocyte decreased evidently and the peak of apoptosis in spleen lymphocyte was dependent on radiation dose and the time after irradiation. Conclusion: After γ-irradiation with large doses, pathological changes of spleen lymphocyte apoptosis in mice can be divided into obviously different stages. The main causes of death of spleen lymphocytes are different in different dose groups

  8. Role of nuclear medicine imaging in differential diagnosis of accessory spleens in patients after splenectomy

    International Nuclear Information System (INIS)

    D’Amico, Andrea; Cofalik, Anna; Przeorek, Cesary; Gawlik, Tomasz; Olczyk, Tomasz; Kalemba, Michał; Modorowska, Alicja; Turska-d’Amico, Maria; Bobek-Billewicz, Barbara; Jarzab, Barbara

    2012-01-01

    More than 10% of healthy population has one or more accessory spleens. The most common location is the hilum of the spleen or area near the tail of the pancreas. The radiological appearance of accessory spleens in oncologic patients who underwent splenectomy can be misinterpreted as a recurrence, especially in the case of compensatory growth of an accessory spleen in successive radiological examinations. We present the cases of three patients who underwent splenectomy for gastric carcinoid, gastric adenocarcinoma and cancer of the left adrenal gland, respectively. CT examination and/or PET-CT scan revealed suspicious findings in the left upper abdomen. In one patient, the dimensional increase of this finding in successive examinations was initially considered suggestive for cancer recurrence. Scintigraphy with 99m Tc-nanocolloid was able to confirm the presence of an accessory spleen in all these patients. Splenic scintigraphy is an economical, accessible and accurate tool in differential diagnosis of accessory spleens in patients after splenectomy

  9. Wandering spleen, gastric and pancreatic volvulus and right-sided descending and sigmoid colon.

    Science.gov (United States)

    Flores-Ríos, Enrique; Méndez-Díaz, Cristina; Rodríguez-García, Esther; Pérez-Ramos, Tania

    2015-10-01

    Wandering spleen is a rare condition, characterized by a mobile spleen that is attached only by an elongated vascular pedicle, allowing it to migrate to any part of the abdomen or pelvis. Mesenteroaxial gastric volvulus usually occurs in children and may be associated with wandering spleen. Both entities result from abnormal laxity or absence of the peritoneal attachments due to abnormal fusion of the peritoneal mesenteries. Pancreatic volvulus is a very rare anomaly, with only a few isolated case reports described in association with wandering spleen. Anomalous right sided descending and sigmoid colon is a very rare entity and its association with wandering spleen has not been previously reported. We report a case of wandering spleen associated with mesenteroaxial gastric volvulus, pancreatic volvulus and rightward shift of the splenic flexure of the colon and right sided descending and sigmoid colon in a young female.

  10. Genetically pyramiding protease-inhibitor genes for dual broad-spectrum resistance against insect and phytopathogens in transgenic tobacco.

    Science.gov (United States)

    Senthilkumar, Rajendran; Cheng, Chiu-Ping; Yeh, Kai-Wun

    2010-01-01

    Protease inhibitors provide a promising means of engineering plant resistance against attack by insects and pathogens. Sporamin (trypsin inhibitor) from sweet potato and CeCPI (phytocystatin) from taro were stacked in a binary vector, using pMSPOA (a modified sporamin promoter) to drive both genes. Transgenic tobacco lines of T0 and T1 generation with varied inhibitory activity against trypsin and papain showed resistance to both insects and phytopathogens. Larvae of Helicoverpa armigera that ingested tobacco leaves either died or showed delayed growth and development relative to control larvae. Transgenic tobacco-overexpressing the stacked genes also exhibited strong resistance against bacterial soft rot disease caused by Erwinia carotovora and damping-off disease caused by Pythium aphanidermatum. Thus, stacking protease-inhibitor genes, driven by the wound and pathogen responsive pMSPOA promoter, is an effective strategy for engineering crops to resistance against insects and phytopathogens.

  11. [Organ-preserving method in the surgical treatment of the spleen injuries].

    Science.gov (United States)

    Khripun, A I; Alimov, A N; Salikov, A V; Priamikov, A D; Alimov, V A; Sukiasian, A A; Popov, T V; Urvantseva, O M

    2014-01-01

    The authors have experience in organ-preserving operations for spleen rupture with the splenic artery ligation in 156 casualties. They consider that such operations let to preserve the spleen, to avoid the postoperative rebleeding and ischemia of pancreas tail and body. Also it is accompanied by the low indications of lethality and postoperative complications. The authors consider that this operation is alternative to splenectomy and other techniques of organ-preserving operations in case of spleen trauma.

  12. GM-CSF augments the immunosuppressive capacity of neonatal spleen cells in vitro

    International Nuclear Information System (INIS)

    Morrissey, P.J.; Ireland, R.

    1991-01-01

    Addition of exogenous granulocyte-macrophage colony stimulating factor (GM-CSF) to cultures of adult murine spleen cells with sheep red blood cells (SRBC) results in an augmented plaque forming cell (PFC) response. The influence of GM-CSF on the ability of neonatal spleen cells to suppress the anti-SRBC plaque forming response of adult spleen cells was tested by adding GM-CSF to cultures of neonatal and adult spleen cells. The suppressive capacity of the neonatal spleen cells was augmented by exogenous GM-CSF. The augmented suppression of the neonatal spleen cells was dependent on a G-10 adherent population since the addition of GM-CSF to cultures containing G-10 passed neonatal spleen cells resulted in an augmented PFC response and not suppression. Neonatal splenic glass adherent cells were also capable of suppressing the response. Neonatal spleen cells or purified neonatal glass adherent spleen cells cultured in the presence of GM-CSF had markedly increased levels of PGE2 in the culture supernatant. Neonatal spleen cells cultured with GM-CSF had increased numbers of morphologically identifiable macrophages after 48 hr of culture. Both irradiation and G-10 passage of the neonatal spleen diminished the numbers of macrophages formed in response to GM-CSF, and both of these manipulations resulted in reversal of suppression in response to GM-CSF. Thus, the augmented suppressive capacity of neonatal spleen cells in response to GM-CSF is probably mediated by its ability to drive monocyte to macrophage differentiation as well as increase the suppressive capacity of the existing neonatal splenic macrophages by increasing their production of PGE2

  13. Torsion of the accessory spleen with infarction : CT features in a case report

    Energy Technology Data Exchange (ETDEWEB)

    Yun, Jung Kyung; Lee, Jun Sik; Kim, Mee Eun; Pyun, Hae Wook; Lee, Il Gi; Lee, Jong Gil; Kim, Hee Jin; Kim, Ik Su [Fatima Hospital, Taegu (Korea, Republic of)

    2000-05-01

    Torsion of the accessory spleen is a rare entity that can have variable clinical presentations. We report case involving an 11-year-old boy with severe abdominal pain and a mass that was found to be due to infarction of the accessory spleen, which was twisted on its pedicle. CT revealed a low-attenuating mass with peripheral inflammatory changes in the left upper abdomen. The mass was pathologically confirmed as torsion of the accessory spleen with infarction. (author)

  14. Radioisotope spleen scan in patients with splenic injury

    International Nuclear Information System (INIS)

    Mishalany, H.G.; Miller, J.H.; Woolley, M.M.

    1982-01-01

    The technetium /sup 99m/Tc sulfur colloid liver-spleen scan is a valuable aid in diagnosis and treatment of patients with splenic injury. After reviewing the charts of 47 patients who were ill as a result of splenic trauma, we came to the following conclusions: (1) the scan identified the injury, accurately mapped its extent, and indicated the presence or absence of associated liver injuries; (2) the scans were useful in following the extent and rate of healing of the splenic injury; (3) the scan is an indirect measurement of of return of splenic fuction; (4) the procedure can be performed in a reasonable time frame with no serious morbidity; and (5) the indications, contraindications, and timing of scans are now reasonably well established

  15. Radioisotope spleen scan in patients with splenic injury

    Energy Technology Data Exchange (ETDEWEB)

    Mishalany, H.G.; Miller, J.H.; Woolley, M.M.

    1982-09-01

    The technetium /sup 99m/Tc sulfur colloid liver-spleen scan is a valuable aid in diagnosis and treatment of patients with splenic injury. After reviewing the charts of 47 patients who were ill as a result of splenic trauma, we came to the following conclusions: (1) the scan identified the injury, accurately mapped its extent, and indicated the presence or absence of associated liver injuries; (2) the scans were useful in following the extent and rate of healing of the splenic injury; (3) the scan is an indirect measurement of of return of splenic fuction; (4) the procedure can be performed in a reasonable time frame with no serious morbidity; and (5) the indications, contraindications, and timing of scans are now reasonably well established.

  16. Post-Traumatic Pseudocyst of the Spleen: Sclerotherapy with Ethanol

    International Nuclear Information System (INIS)

    Voelk, Markus; Rogler, Gerhard; Strotzer, Michael; Lock, Guntram; Manke, Christoph; Feuerbach, Stefan

    1999-01-01

    We report a case of successful percutaneous treatment of a chronic post-traumatic splenic pseudocyst using alcohol as the sclerosing agent. A 26-year-old man presented with a symptomatic cystic mass located in the spleen. Aspiration of 300 ml of fluid was only temporarily effective, and therefore a drainage catheter was placed 3 days later. After histopathologic and microbiologic exclusion of a malignant or infectious origin, local sclerotherapy with alcohol was performed because of recurrence after percutaneous drainage. This therapy was repeated six times within 2 weeks. Two weeks later, the remaining volume was determined to be 16 ml. Six months after treatment the cyst was no longer visible. To our knowledge this is the first case of a chronic post-traumatic splenic cyst treated with alcohol. Percutaneous sclerotherapy of a symptomatic post-traumatic splenic pseudocyst may be an alternative to surgical treatment

  17. Anti-fibrinolytic and anti-microbial activities of a serine protease inhibitor from honeybee (Apis cerana) venom.

    Science.gov (United States)

    Yang, Jie; Lee, Kwang Sik; Kim, Bo Yeon; Choi, Yong Soo; Yoon, Hyung Joo; Jia, Jingming; Jin, Byung Rae

    2017-10-01

    Bee venom contains a variety of peptide constituents, including low-molecular-weight protease inhibitors. While the putative low-molecular-weight serine protease inhibitor Api m 6 containing a trypsin inhibitor-like cysteine-rich domain was identified from honeybee (Apis mellifera) venom, no anti-fibrinolytic or anti-microbial roles for this inhibitor have been elucidated. In this study, we identified an Asiatic honeybee (A. cerana) venom serine protease inhibitor (AcVSPI) that was shown to act as a microbial serine protease inhibitor and plasmin inhibitor. AcVSPI was found to consist of a trypsin inhibitor-like domain that displays ten cysteine residues. Interestingly, the AcVSPI peptide sequence exhibited high similarity to the putative low-molecular-weight serine protease inhibitor Api m 6, which suggests that AcVSPI is an allergen Api m 6-like peptide. Recombinant AcVSPI was expressed in baculovirus-infected insect cells, and it demonstrated inhibitory activity against trypsin, but not chymotrypsin. Additionally, AcVSPI has inhibitory effects against plasmin and microbial serine proteases; however, it does not have any detectable inhibitory effects on thrombin or elastase. Consistent with these inhibitory effects, AcVSPI inhibited the plasmin-mediated degradation of fibrin to fibrin degradation products. AcVSPI also bound to bacterial and fungal surfaces and exhibited anti-microbial activity against fungi as well as gram-positive and gram-negative bacteria. These findings demonstrate the anti-fibrinolytic and anti-microbial roles of AcVSPI as a serine protease inhibitor. Copyright © 2017 Elsevier Inc. All rights reserved.

  18. Role of the spleen in cyclophosphamide-induced hematosuppression and extramedullary hematopoiesis in mice.

    Science.gov (United States)

    Wang, Yuli; Meng, Qinggang; Qiao, Haiquan; Jiang, Hongchi; Sun, Xueying

    2009-05-01

    Extramedullary hematopoiesis (EMH) is induced in spleens due to various diseases. The aim of this study is to investigate the role of spleen in cyclophosphamide (CTX)-induced hematosuppression and EMH in mice. Balb/c mice were IP injected with 300 mg/kg CTX 2 weeks after splenectomy or sham operation and randomly sacrificed 1, 3, 7, 14, and 21 days after injection. Blood samples were collected, and spleens were weighed, histologically analyzed, and then used for flow cytometry. There were significant differences in white blood count, red blood count, platelet numbers and hemoglobin concentration between the splenectomized and sham-operated mice after CTX injection. The cellularity of the spleen was reduced 3 days following CTX treatment but then rose 7 days after CTX treatment. The numbers of colony-forming units in the spleen reached a peak 7 days after CTX injection, then declined. Flow cytometry demonstrated the percentage of CD34(+) and CD117(+) cells in the spleen increased 7 days after CTX injection, indicating the hematopoietic stem and progenitor cells in the spleen. The study indicates that EMH occurs as a compensatory reaction to CTX-induced hematosuppression in the murine spleen, implying that conservation of the spleen may promote the recovery of cancer patients from chemotherapy-induced hematosuppression.

  19. Ehrlichia canis morulae and DNA detection in whole blood and spleen aspiration samples.

    Science.gov (United States)

    Faria, Joice Lara Maia; Dagnone, Ana Sílvia; Munhoz, Thiago Demarchi; João, Carolina Franchi; Pereira, Wanderson Adriano Biscola; Machado, Rosângela Zacarias; Tinucci-Costa, Mirela

    2010-01-01

    The aim of this study was to compare the detection of Ehrlichia canis morulae and DNA by nPCR in whole blood and spleen aspiration. The sample included 40 dogs showing thrombocytopenia associated to clinical signs suggestive of canine ehrlichiosis. Morulae detection showed that in 35 of the dogs studied, 17 had morulae in spleen tissue, and two in buffy coat smears. E. canis DNA was detected in 29/40 blood samples. We verified that morulae detection is more efficient in cytological preparations from spleen aspiration. On the other hand, nPCR on spleen and blood samples were equally efficient for disease diagnosis.

  20. Interactions between Biliverdin, Oxidative Damage, and Spleen Morphology after Simulated Aggressive Encounters in Veiled Chameleons.

    Directory of Open Access Journals (Sweden)

    Michael W Butler

    Full Text Available Stressors frequently increase oxidative damage--unless organisms simultaneously mount effective antioxidant responses. One putative mitigative mechanism is the use of biliverdin, an antioxidant produced in the spleen during erythrocyte degradation. We hypothesized that both wild and captive-bred male veiled chameleons (Chamaeleo calyptratus, which are highly aggressive to conspecifics, would respond to agonistic displays with increased levels of oxidative damage, but that increased levels of biliverdin would limit this increase. We found that even just visual exposure to a potential combatant resulted in decreased body mass during the subsequent 48-hour period, but that hematocrit, biliverdin concentration in the bile, relative spleen size, and oxidative damage in plasma, liver, and spleen were unaffected. Contrary to our predictions, we found that individuals with smaller spleens exhibited greater decreases in hematocrit and higher bile biliverdin concentrations, suggesting a revision to the idea of spleen-dependent erythrocyte processing. Interestingly, individuals with larger spleens had reduced oxidative damage in both the liver and spleen, demonstrating the spleen's importance in modulating oxidative damage. We also uncovered differences in spleen size and oxidative damage between wild and captive-bred chameleons, highlighting environmentally dependent differences in oxidative physiology. Lastly, we found no relationship between oxidative damage and biliverdin concentration, calling into question biliverdin's antioxidant role in this species.

  1. Radiobiological studies of cells in multicellular spheroids using a sequential trypsinization technique

    International Nuclear Information System (INIS)

    Giesbrecht, J.L.; Wilson, W.R.; Hill, R.P.

    1981-01-01

    The radiation response of V79 Chinese hamster cells grown as multicellular spheroids has been investigated by determining survival curves for treatment under a variety of different oxygen concentrations. Spheroids were irradiated under fully oxygenated conditions in air-equilibrated medium at 37 0 C, in medium exposed to lower oxygen tension (5% O 2 ) for times varying from 1 hr to 3 days, or under anoxic conditions. For comparison with the spheroids, using identical treatment conditions, V79 cells were grown in suspension as a subconfluent monolayer attached to Sephadex (microcarrier) beads and irradiated under fully oxygenated or anoxic conditions. The radiation response of cells at different depths within the spheroid was investigated by using a sequential trypsinization technique developed to remove eight or nine shells of cells successively from the spheroid surface. When irradiation was given under fully oxygenated conditions the outer few cell layers were more sensitive than the inner cells, a finding which is not understood. As expected the inner cells in spheroids irradiated in air (at 37 0 C) or in 5% O 2 are more resistant than the outer cells. For an acute exposure to 5% O 2 (1 hr) in the inner cells displayed full radiobiological hypoxia; however, for chronic exposures to low oxygen this was not the case. These results with the sequential trypsinization procedure suggest that the radiation response of cells in spheroids is more complex than anticipted

  2. Kinetic Dissection of the Pre-existing Conformational Equilibrium in the Trypsin Fold*

    Science.gov (United States)

    Vogt, Austin D.; Chakraborty, Pradipta; Di Cera, Enrico

    2015-01-01

    Structural biology has recently documented the conformational plasticity of the trypsin fold for both the protease and zymogen in terms of a pre-existing equilibrium between closed (E*) and open (E) forms of the active site region. How such plasticity is manifested in solution and affects ligand recognition by the protease and zymogen is poorly understood in quantitative terms. Here we dissect the E*-E equilibrium with stopped-flow kinetics in the presence of excess ligand or macromolecule. Using the clotting protease thrombin and its zymogen precursor prethrombin-2 as relevant models we resolve the relative distribution of the E* and E forms and the underlying kinetic rates for their interconversion. In the case of thrombin, the E* and E forms are distributed in a 1:4 ratio and interconvert on a time scale of 45 ms. In the case of prethrombin-2, the equilibrium is shifted strongly (10:1 ratio) in favor of the closed E* form and unfolds over a faster time scale of 4.5 ms. The distribution of E* and E forms observed for thrombin and prethrombin-2 indicates that zymogen activation is linked to a significant shift in the pre-existing equilibrium between closed and open conformations that facilitates ligand binding to the active site. These findings broaden our mechanistic understanding of how conformational transitions control ligand recognition by thrombin and its zymogen precursor prethrombin-2 and have direct relevance to other members of the trypsin fold. PMID:26216877

  3. Comparison of mesenchymal stem cells released from poly(N-isopropylacrylamide) copolymer film and by trypsinization

    International Nuclear Information System (INIS)

    Yang Lei; Liu Tianqing; Song Kedong; Jiang Lili; Wu Shuang; Guo Wenhua; Cheng Fang; Lu, Jian R

    2012-01-01

    Temperature-responsive platforms containing poly(N-isopropylacrylamide) (PNIPAAm) have been developed as an effective substitute for enzymatic treatment to recover adherent cells, but it remains unclear whether this alternative harvesting method tends to support stem cells preserving them being primitive. This study mainly investigated the biological properties of mesenchymal stem cells derived from rat bone marrow and human adipose tissue (BM-MSCs and AT-MSCs) after being cultured on PNIPAAm copolymer films and recovered by temperature drop, and compared the cells harvested from glass coverslips with trypsinization as controls. The experimental results demonstrated that after three serial passages, the released MSCs from thermal liftoff showed no significant differences in cell morphology, immunophenotype and osteogenesis for BM-MSCs or adipogenesis for AT-MSCs, but had higher viability, stronger proliferation and higher adipogenic differentiation for BM-MSCs or higher osteogenic differentiation for AT-MSCs compared with the trypsinization group. Besides, more proteins remained around or within the cell membranes upon temperature drop. It is concluded that cell detachment with more extracellular matrix proteins facilitates the maintenance of membrane proteins, and accordingly preserves MSC properties related to viability, proliferation and differentiation to some extent. This indicates that the PNIPAAm copolymer films and their matching cooling treatment can be used as effective alternatives to the existing culture substrates and traditional enzymatic digestion for MSCs. (paper)

  4. Effect of detergents, trypsin, and bivalent metal ions on interfacial activation and functioning of phospholipase D.

    Science.gov (United States)

    Madyarov, Sh R

    2014-07-01

    The effects of detergents, trypsin, and bivalent metal ions on production of phosphatidic and lysophosphatidic acids by the action of phospholipase D (PLD) on lecithin and lysolecithin were studied. It was found that these reaction products and dodecyl sulfate ions activate PLD, whereas other anionic detergents are less effective. A protective effect of the functioning enzyme against its hydrolytic inactivation by trypsin was found. Bivalent metal ions can be arranged in the following sequence by their ability to activate PLD in the hydrolysis of lecithin and lysolecithin: Ca2+>Sr2+>Ba2+>Mg2+. These results are considered in relation to a proposed mechanism of activation and functioning of PLD with the participation of clusters of phosphatidates and lysophosphatidates. Such Me2+-induced formation of rafts or microdomains from the products of hydrolysis of phospholipids can rationalize not only PLD activation and self-regulation, but also the action of this mechanism on other components and properties of biomembranes. PLD and other lipolytic enzymes can be classified as lateral vector enzymes.

  5. A novel poly(deep eutectic solvent)-based magnetic silica composite for solid-phase extraction of trypsin.

    Science.gov (United States)

    Xu, Kaijia; Wang, Yuzhi; Li, Yixue; Lin, Yunxuan; Zhang, Haibao; Zhou, Yigang

    2016-11-23

    Novel poly(deep eutectic solvent) grafted silica-coated magnetic microspheres (Fe 3 O 4 @SiO 2 -MPS@PDES) were prepared by polymerization of choline chloride-itaconic acid (ChCl-IA) and γ-MPS-modified magnetic silica composites, and were characterized by vibrating sample magnetometer (VSM), Fourier transform infrared spectrometry (FT-IR), X-ray photoelectron spectra (XPS), thermal gravimetric analysis (TGA) and transmission electron microscope (TEM). Then the synthetic Fe 3 O 4 @SiO 2 -MPS@PDES microspheres were applied for the magnetic solid-phase extraction (MSPE) of trypsin for the first time. After extraction, the concentration of trypsin in the supernatant was determined by a UV-vis spectrophotometer. Single factor experiments were carried out to investigate the effects of the extraction process, including the concentration of trypsin, the ionic strength, the pH value, the extraction time and the temperature. Experimental results showed the extraction capacity could reach up to 287.5 mg/g under optimized conditions. In comparison with Fe 3 O 4 @SiO 2 -MPS, Fe 3 O 4 @SiO 2 -MPS@PDES displayed higher extraction capacity and selectivity for trypsin. According to the regeneration studies, Fe 3 O 4 @SiO 2 -MPS@PDES microspheres can be recycled six times without significant loss of its extraction capacity, and retained a high extraction capacity of 233 mg/g after eight cycles. Besides, the activity studies also demonstrated that the activity of the extracted trypsin was well retained. Furthermore, the analysis of real sample revealed that the prepared magnetic microspheres can be used to purify trypsin in crude bovine pancreas extract. These results highlight the potential of the proposed Fe 3 O 4 @SiO 2 -MPS@PDES-MSPE method in separation of biomolecules. Copyright © 2016 Elsevier B.V. All rights reserved.

  6. A potential human hepatocellular carcinoma inhibitor from Bauhinia purpurea L. seeds: from purification to mechanism exploration.

    Science.gov (United States)

    Fang, Evandro Fei; Bah, Clara Shui Fern; Wong, Jack Ho; Pan, Wen Liang; Chan, Yau Sang; Ye, Xiu Juan; Ng, Tzi Bun

    2012-02-01

    A 20-kDa Kunitz-type trypsin-chymotrypsin inhibitor, Bauhinia purpurea trypsin inhibitor (BPLTI), has been isolated from the seeds of B. purpurea L. by using liquid chromatography procedures that involved ion exchange chromatography on Sp-Sepharose and Mono S and gel filtration on Superdex 75. BPLTI demonstrated protease inhibitory activities of 7226 BAEE units/mg and 65 BTEE units/mg toward trypsin and α-chymotrypsin, respectively. BPLTI was relatively thermal (0-60°C) and pH (3-10) stable and its activity could be decreased by dithiothreitol treatment. BPLTI exhibited a wide spectrum of anti-proliferative and pro-apoptotic activities especially on human hepatocellular carcinoma Hep G2 cells. However, it was devoid of a significant antiproliferative effect on immortal human hepatic WRL 68 cells. We show here that BPLTI stimulates apoptosis in Hep G2 cells, including (1) evoking DNA damage including the production of chromatin condensation and apoptotic bodies; (2) induction of cell apoptosis/necrosis; (3) mitochondrial membrane depolarization; and (4) increasing the production of cytokines. Taken together, our findings show for the first time that purified protease inhibitor from B. purpurea L. seeds is a promising candidate for the treatment of human hepatocellular carcinoma.

  7. Use of the neutron diffraction - H/D exchange technique to determine the conformational dynamics of trypsin

    International Nuclear Information System (INIS)

    Kossiakoff, A.A.

    1982-01-01

    Reported here are studies analyzing the extent and nature of the inherent conformational fluctuations in trypsin by neutron diffraction - hydrogen exchange techniques. The pattern of exchange investigates systematic relationships between exchangeable sites and the structural and chemical properties of the molecule. Our findings that pH 7, 20 0 and 1 year of soaking all sites of trypsin are fully exchanged except those which are especially well protected by the structure. Essentially all the sites in which the peptide hydrogens are bonded directly to water molecules - either in the bulk solvent regions or in interior clusters - are fully exchanged. 41 references, 10 figures

  8. Docking, thermodynamics and molecular dynamics (MD) studies of a non-canonical protease inhibitor, MP-4, from Mucuna pruriens.

    Science.gov (United States)

    Kumar, Ashish; Kaur, Harmeet; Jain, Abha; Nair, Deepak T; Salunke, Dinakar M

    2018-01-12

    Sequence and structural homology suggests that MP-4 protein from Mucuna pruriens belongs to Kunitz-type protease inhibitor family. However, biochemical assays showed that this protein is a poor inhibitor of trypsin. To understand the basis of observed poor inhibition, thermodynamics and molecular dynamics (MD) simulation studies on binding of MP-4 to trypsin were carried out. Molecular dynamics simulations revealed that temperature influences the spectrum of conformations adopted by the loop regions in the MP-4 structure. At an optimal temperature, MP-4 achieves maximal binding while above and below the optimum temperature, its functional activity is hampered due to unfavourable flexibility and relative rigidity, respectively. The low activity at normal temperature is due to the widening of the conformational spectrum of the Reactive Site Loop (RSL) that reduces the probability of formation of stabilizing contacts with trypsin. The unique sequence of the RSL enhances flexibility at ambient temperature and thus reduces its ability to inhibit trypsin. This study shows that temperature influences the function of a protein through modulation in the structure of functional domain of the protein. Modulation of function through appearance of new sequences that are more sensitive to temperature may be a general strategy for evolution of new proteins.

  9. Solution Structure of a Novel C2-Symmetrical Bifunctional Bicyclic Inhibitor Based on SFTI-1

    International Nuclear Information System (INIS)

    Jaulent, Agnes M.; Brauer, Arnd B. E.; Matthews, Stephen J.; Leatherbarrow, Robin J.

    2005-01-01

    A novel bifunctional bicyclic inhibitor has been created that combines features both from the Bowman-Birk inhibitor (BBI) proteins, which have two distinct inhibitory sites, and from sunflower trypsin inhibitor-1 (SFTI-1), which has a compact bicyclic structure. The inhibitor was designed by fusing together a pair of reactive loops based on a sequence derived from SFTI-1 to create a backbone-cyclized disulfide-bridged 16-mer peptide. This peptide has two symmetrically spaced trypsin binding sites. Its synthesis and biological activity have been reported in a previous communication [Jaulent and Leatherbarrow, 2004, PEDS 17, 681]. In the present study we have examined the three-dimensional structure of the molecule. We find that the new inhibitor, which has a symmetrical 8-mer half-cystine CTKSIPP'I' motif repeated through a C 2 symmetry axis also shows a complete symmetry in its three-dimensional structure. Each of the two loops adopts the expected canonical conformation common to all BBIs as well as SFTI-1. We also find that the inhibitor displays a strong and unique structural identity, with a notable lack of minor conformational isomers that characterise most reactive site loop mimics examined to date as well as SFTI-1. This suggests that the presence of the additional cyclic loop acts to restrict conformational mobility and that the deliberate introduction of cyclic symmetry may offer a general route to locking the conformation of β-hairpin structures

  10. Ex vivo perfusion of human spleens maintains clearing and processing functions.

    Science.gov (United States)

    Buffet, Pierre A; Milon, Geneviève; Brousse, Valentine; Correas, Jean-Michel; Dousset, Bertrand; Couvelard, Anne; Kianmanesh, Reza; Farges, Olivier; Sauvanet, Alain; Paye, François; Ungeheuer, Marie-Noëlle; Ottone, Catherine; Khun, Huot; Fiette, Laurence; Guigon, Ghislaine; Huerre, Michel; Mercereau-Puijalon, Odile; David, Peter H

    2006-05-01

    The spleen plays a central role in the pathophysiology of several potentially severe diseases such as inherited red cell membrane disorders, hemolytic anemias, and malaria. Research on these diseases is hampered by ethical constraints that limit human spleen tissue explorations. We identified a surgical situation--left splenopancreatectomy for benign pancreas tumors--allowing spleen retrieval at no risk for patients. Ex vivo perfusion of retrieved intact spleens for 4 to 6 hours maintained a preserved parenchymal structure, vascular flow, and metabolic activity. Function preservation was assessed by testing the ability of isolated-perfused spleens to retain Plasmodium falciparum-infected erythrocytes preexposed to the antimalarial drug artesunate (Art-iRBCs). More than 95% of Art-iRBCs were cleared from the perfusate in 2 hours. At each transit through isolated-perfused spleens, parasite remnants were removed from 0.2% to 0.23% of Art-iRBCs, a proportion consistent with the 0.02% to 1% pitting rate previously established in artesunate-treated patients. Histologic analysis showed that more than 90% of Art-iRBCs were retained and processed in the red pulp, providing the first direct evidence of a zone-dependent parasite clearance by the human spleen. Human-specific physiologic or pathophysiologic mechanisms involving clearing or processing functions of the spleen can now be experimentally explored in a human tissue context.

  11. Thermal ablation for partial splenectomy hemostasis, spleen trauma, splenic metastasis and hypersplenism.

    Science.gov (United States)

    Duan, Ya-Qi; Liang, Ping

    2013-05-01

    Many studies have been conducted on splenic thermal ablation for partial splenectomy hemostasis, spleen trauma, splenic metastasis and hypersplenism. In this article, we review the evolution and current status of radiofrequency and microwave ablation in the treatment of spleen diseases. All publications from 1990 to 2011 on radiofrequency and microwave ablation for partial splenectomy hemostasis, spleen trauma, splenic metastasis and hypersplenism were retrieved by searching PubMed. Thermal ablation in the spleen for partial splenectomy hemostasis, spleen trauma, splenic metastasis and hypersplenism can preserve part of the spleen and maintain splenic immunologic function. Thermal ablation for assisting hemostasis in partial splenectomy minimizes blood loss during operation. Thermal ablation for spleen trauma reduces the number of splenectomy and the amount of blood transfusion. Thermal ablation for splenic metastasis is minimally invasive and can be done under the guidance of an ultrasound, which helps shorten the recovery time. Thermal ablation for hypersplenism increases platelet (PLT) and white blood cell (WBC) counts and improves liver function. It also helps to maintain splenic immunologic function and even improves splenic immunologic function in the short-term. In conclusion, thermal ablative approaches are promising for partial splenectomy hemostasis, spleen trauma, splenic metastasis and hypersplenism. In order to improve therapeutic effects, directions for future studies may include standardized therapeutic indications, prolonged observation periods and enlarged sample sizes.

  12. Detection of early gamma-postirradiation effects in murine spleen by proton NMR relaxation times.

    Science.gov (United States)

    Zebrowska, G; Lewa, C J; Ramee, M P; Husson, F; De Certaines, J D

    2001-01-01

    It was our aim to evaluate the potential of proton relaxation times for the early detection of radiation-induced spleen changes. Female Swiss mice were irradiated with doses ranging from 0.05 Gy to 4 Gy. The body weight, the spleen weight and the spleen water content of single animals were determined. Measurements of longitudinal (T1) and transversal (T2) proton relaxation times of the spleen samples were performed in a 0.47 T spectrometer. Histological examinations of the control and irradiated organs were performed. NMR measurements during the first five days after irradiation showed that total body gamma-irradiation with doses from 1.5 Gy to 4 Gy results in decreasing T1 of the murine spleen. Significant shortening in T2 was observed for the spleen of animals irradiated with a dose of 4 Gy. Histological examinations demonstrated subnormal architecture in slices derived from animals irradiated with 2 Gy and 4 Gy. The fluctuations of the spleen T1 and T2 of irradiated mice are correlated with relative spleen weight and can be used to estimate radiation induced changes in this organ.

  13. Epithelial cyst of the spleen with squamous metaplasia: a rare entity.

    Science.gov (United States)

    Shanthi, Vissa; Reddy, Vengala Chidananda; Rao, Nandam Mohan; Grandhi, Bhavana; Kona, Suneetha

    2014-04-01

    Epithelial splenic cysts are uncommon lesions which occur the spleen. The aetiopathogenesis of these cysts is not clear. We are reporting a case of an epithelial cyst which occurred in the spleen in a 40-year-old female, which was multi loculated and which had flattened lining epithelium. Some foci showed squamous metaplasia.

  14. Effects of boron on structure and antioxidative activities of spleen in rats.

    Science.gov (United States)

    Hu, Qianqian; Li, Shenghe; Qiao, Enmei; Tang, Zhongtao; Jin, Erhui; Jin, Guangming; Gu, Youfang

    2014-04-01

    In order to determine the relationship between boron and development of the spleen, especially in the promoting biological effects, we examined the effects of different levels of boron on weight, organ index, microstructure, and antioxidative activities of the spleen in rats. Sprague-Dawley (SD) rats were selected and treated with different concentrations of boron, and then, the organs were resected and weighed. One half of the tissue was fixed and embedded in paraffin to observe tissue structure changes. The other half of the tissue was homogenated for determining the antioxidant activities. The results showed that 40 mg/L of boron could increase weight, organ indexes, and antioxidant capacity of spleens and improve the spleen tissue structure, while the boron concentration above 80 mg/L could decrease weight, organ indexes, and antioxidant capacity of spleens and damage the spleen tissue structure. The higher the concentration, the more serious the damage was. Especially at the concentration of 640 mg/L, it could significantly inhibit the development of the spleen and even exhibit toxic effect. Hence, low boron concentration played a protective role in the development of the spleen, while high boron concentration could damage the organs and even produce toxic effect.

  15. An epidermoid cyst of accessory spleen simulating tumors of the tail of pancreas

    Directory of Open Access Journals (Sweden)

    Amit Kumar Sinha

    2015-07-01

    Full Text Available An epidermoid cyst of accessory spleen, a rare condition may present as pseudocyst of pancreas and other cystic tumors of the pancreas. This case report along with the review of literature attributes some clinical features and investigative pattern to differentiate between epidermoid cyst of accessory spleen and other cystic tumor of pancreas.

  16. Two different mechanisms of immune-complex trapping in the mouse spleen during immune responses

    NARCIS (Netherlands)

    Yoshida, K.; van den Berg, T. K.; Dijkstra, C. D.

    1993-01-01

    The capacity of immune-complex (IC) trapping was examined using purified horse radish peroxidase (HRP)-anti-HRP (PAP) on frozen sections of mouse spleen in vitro. We investigated the trapping mechanisms by applying the IC with or without fresh mouse serum added on the spleen sections of naive as

  17. Accessory spleen presenting as acute abdomen: A case report and operative management

    Directory of Open Access Journals (Sweden)

    A. Landmann

    2016-09-01

    Full Text Available Accessory spleens are found in 10–30% of patients and are asymptomatic. Rarely, torsion of an accessory spleen can cause abdominal pain and acute abdomen. We present the case of an 8-year-old girl who arrives to the emergency room with left upper quadrant abdominal pain. CT scan revealed a non-enhancing soft tissue mass and multiple small splenules. Laparoscopy revealed a torsed accessory spleen and malrotation. Accessory spleen is a common congenital anomaly that is frequently asymptomatic. Rarely, an accessory spleen may become torsed around its vascular pedicle resulting in severe abdominal pain. Treatment is surgical resection. Torsion of accessory splenic tissue is a rare cause of acute abdomen in pediatric patients.

  18. Observation on Therapeutic Effect of the Depression of Heart-spleen Deficiency with Wuling Capsule

    International Nuclear Information System (INIS)

    Li Jinbin; Liu Ping

    2010-01-01

    To evaluate the effect on the treatment of depression belong to the type of heart-spleen deficiency with Wuling capsule, 37 patients were assigned into two groups: the deficiency of both the heart and spleen group (I) and the non deficiency of both the heart and spleen group (II). The efficacy of two groups was surveyed and compared after taken Wuling capsule 2 and 4 weeks,respectively. After treatment, there was a difference (P 0.05). The satisfactory effects were showed on various kinds of depressions using wuling capsules,while deficiency of both the heart and spleen group effects were better than that of the non deficiency of both the heart and spleen group. (authors)

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

    International Nuclear Information System (INIS)

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

    1986-01-01

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

  20. Trypsin inhibitory activity in blood and cervical mucus of sheep following chronic gamma irradiation

    International Nuclear Information System (INIS)

    Molnarova, M.; Arendarcik, J.; Pastorova, B.

    1982-01-01

    The effect was investigated of chronic gamma irradiation for a period of 7 days to a total dose of 6.7 Gy. A decrease was found in TIA (trypsin inhibition activity) of blood plasma to 73.6% of the value prior to irradiation. The low-molecular fraction of TIA increased to 194.6%. Later in the experiment the values decreased. The TIA dynamics of the cervical mucus had the reverse character. The values increased; at the 16th day after irradiation they increased up to 392% of the initial values. A comparison with previous experiments shows that acute local irradiation with an almost three-fold dose of the hypothalamo-pituitary area and of the ovaries does not induce significant changes in the blood plasma TIA. (M.D.)

  1. Influence of Tableting on the Conformation and Thermal Stability of Trypsin as a Model Protein

    DEFF Research Database (Denmark)

    Klukkert, Marten; Van De Weert, Marco; Fanø, Mathias

    2015-01-01

    was performed to determine the Tm as well as the folding reversibility after thermal denaturation of the reconstituted samples. It was found that compacted samples showed reduced activity accompanied by an altered secondary structure. Conformational changes that occur in the solid state were partially...... reversible upon tablet reconstitution. Aqueous-state IR spectroscopy combined with partial least squares was shown to be a powerful tool to follow irreversible structural changes and evaluate sample bioactivity. Besides its conformation, the thermal stability of trypsin was altered as a result of the applied...... compaction pressure, indicated by a reduced folding reversibility. In conclusion, this study reveals that tableting can have a negative impact on the biological quality of protein APIs....

  2. RAMA casein zymography: Time-saving and highly sensitive casein zymography for MMP7 and trypsin.

    Science.gov (United States)

    Yasumitsu, Hidetaro; Ozeki, Yasuhiro; Kanaly, Robert A

    2016-11-01

    To detect metalloproteinase-7 (MMP7), zymography is conducted using a casein substrate and conventional CBB stain. It has disadvantages because it is time consuming and has low sensitivity. Previously, a sensitive method to detect MMP7 up to 30 pg was reported, however it required special substrates and complicated handlings. RAMA casein zymography described herein is rapid, sensitive, and reproducible. By applying high-sensitivity staining with low substrate conditions, the staining process is completed within 1 h and sensitivity was increased 100-fold. The method can detect 10 pg MMP7 by using commercially available casein without complicated handlings. Moreover, it increases detection sensitivity for trypsin. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  3. Activity of trypsin-like enzymes and gelatinases in rats with doxorubicin cardiomyopathy

    Directory of Open Access Journals (Sweden)

    Iu. А. Gordiienko

    2014-12-01

    Full Text Available Activity of trypsin-like enzymes (ATLE and gelatinases A and B were studied in the blood plasma and extracts from cardiac muscle, cerebral cortex and cerebellum of rats with cardiomyopathy caused by anthracycline antibiotic doxorubicin against the background of preventive application of corvitin and α-ketoglutarate. ATLE significantly increased in blood plasma and extracts from cerebral cortex but decreased in extracts from cardiac muscle and cerebellum in doxorubicin cardiomyopathy (DCMP. In addition, a significant increase of activity of both gelatinases in plasma and tissue extracts was observed. Preventive administration of corvitin and α-ketoglutarate resulted in differently directed changes of activity of the above mentioned enzymes in heart and brain tissues. Obtained data confirm the hypothesis about activation of proteolysis under the influence of anthracycline antibiotics and testify to selective effect of corvitin and α-ketoglutarate on ATLE and gelatinases.

  4. Activity of trypsin-like enzymes and gelatinases in rats with doxorubicin cardiomyopathy.

    Science.gov (United States)

    Gordiienko, Iu A; Babets, Ya V; Kulinich, A O; Shevtsova, A I; Ushakova, G O

    2014-01-01

    Activity of trypsin-like enzymes (ATLE) and gelatinases A and B were studied in the blood plasma and extracts from cardiac muscle, cerebral cortex and cerebellum of rats with cardiomyopathy caused by anthracycline antibiotic doxorubicin against the background of preventive application of corvitin and α-ketoglutarate. ATLE significantly increased in blood plasma and extracts from cerebral cortex but decreased in extracts from cardiac muscle and cerebellum in doxorubicin cardiomyopathy (DCMP). In addition, a significant increase of activity of both gelatinases in plasma and tissue extracts was observed. Preventive administration of corvitin and α-ketoglutarate resulted in differently directed changes of activity of the above mentioned enzymes in heart and brain tissues. Obtained data confirm the hypothesis about activation of proteolysis under the influence of anthracycline antibiotics and testify to selective effect of corvitin and α-ketoglutarate on ATLE and gelatinases.

  5. The spleen-to-liver ratios in hepatic diseases

    International Nuclear Information System (INIS)

    Vorne, M.; Jurvelin, J.; Vaehaetalo, S.; Himanka, E.

    1984-01-01

    We compared light pen (LPEN) and Region of Interest (ROI) computer methods in determining spleen-to-liver (S/L) ratios both in anterior and posterior images in various liver diseases. The S/L ratio was independent of age or type of colloid used (equal particle size provided). Results with corresponding LPEN and ROI programs did not differ significantly from each other. The sensitivity and specificity were tested and the anterior view yielded somewhat better results than the posterior view but the best results were obtained when both projections were used. The sensitivity for all liver diseases was 60% and the corresponding specificity 93%. In hepatocellular diseases the sensitivity was 80-100%, but the S/L ratio had only 37% sensitivity for hepatic metastases. Hepatomegaly in the anterior view was found in 67% of fatty liver cases, in 25% of cirrhosis cases, in 20% of hepatitis and in 25% of metastatic livers. Splenomegaly was noted in 39-54% of patients with hepatocellular diseases but only in 4-10% of metastatic diseases. (orig.) [de

  6. Prediction of trypsin/molecular fragment binding affinities by free energy decomposition and empirical scores

    Science.gov (United States)

    Benson, Mark L.; Faver, John C.; Ucisik, Melek N.; Dashti, Danial S.; Zheng, Zheng; Merz, Kenneth M.

    2012-05-01

    Two families of binding affinity estimation methodologies are described which were utilized in the SAMPL3 trypsin/fragment binding affinity challenge. The first is a free energy decomposition scheme based on a thermodynamic cycle, which included separate contributions from enthalpy and entropy of binding as well as a solvent contribution. Enthalpic contributions were estimated with PM6-DH2 semiempirical quantum mechanical interaction energies, which were modified with a statistical error correction procedure. Entropic contributions were estimated with the rigid-rotor harmonic approximation, and solvent contributions to the free energy were estimated with several different methods. The second general methodology is the empirical score LISA, which contains several physics-based terms trained with the large PDBBind database of protein/ligand complexes. Here we also introduce LISA+, an updated version of LISA which, prior to scoring, classifies systems into one of four classes based on a ligand's hydrophobicity and molecular weight. Each version of the two methodologies (a total of 11 methods) was trained against a compiled set of known trypsin binders available in the Protein Data Bank to yield scaling parameters for linear regression models. Both raw and scaled scores were submitted to SAMPL3. Variants of LISA showed relatively low absolute errors but also low correlation with experiment, while the free energy decomposition methods had modest success when scaling factors were included. Nonetheless, re-scaled LISA yielded the best predictions in the challenge in terms of RMS error, and six of these models placed in the top ten best predictions by RMS error. This work highlights some of the difficulties of predicting binding affinities of small molecular fragments to protein receptors as well as the benefit of using training data.

  7. Effect of sorbitol and glycerol on the stability of trypsin and difference between their stabilization effects in the various solvents.

    Science.gov (United States)

    Pazhang, Mohammad; Mehrnejad, Faramarz; Pazhang, Yaghub; Falahati, Hanieh; Chaparzadeh, Nader

    2016-01-01

    The effect of glycerol and sorbitol on the stability of porcine pancreas trypsin was investigated in this work. Molecular dynamics simulation and thermostability results showed that trypsin has two flexible regions, and polyols (sorbitol and glycerol) stabilize the enzyme by decreasing the flexibility of these regions. Radial distribution function results exhibited that sorbitol and glycerol were excluded from the first water layer of the enzyme, therefore decrease the flexibility of the regions by preferential exclusion. Also, results showed that the stabilization effect of sorbitol is more than glycerol. This observation could be because of the larger decrease in the fluctuations of trypsin in the presence of sorbitol. We also examined the role of solvent's hydrophobicity in enzyme stabilization by sorbitol and glycerol. To do so, the thermostability of trypsin was evaluated in the presence of solvents with different hydrophobicity (methanol, ethanol, isopropanol and n-propanol) in addition to the polyols. Our results depicted that glycerol is a better stabilizer than sorbitol in the presence of hydrophobic solvents (n-propanol), whereas sorbitol is a better stabilizer than glycerol in the presence of hydrophilic solvents (methanol). © 2015 International Union of Biochemistry and Molecular Biology, Inc.

  8. A capillary monolithic trypsin reactor for efficient protein digestion in online and offline coupling to ESI and MALDI mass spectrometry.

    Science.gov (United States)

    Spross, Jens; Sinz, Andrea

    2010-02-15

    We describe the preparation of a capillary trypsin immobilized monolithic enzyme reactor (IMER) for a rapid and efficient digestion of proteins down to the femtomole level. Trypsin was immobilized on a poly(glycidyl methacrylate-co-acrylamide-co-ethylene glycol dimethycrylate) monolith using the glutaraldehyde technique. Digestion efficiencies of the IMER were evaluated using model proteins and protein mixtures as well as chemically cross-linked lysozyme regarding the addition of denaturants and increasing digestion temperature. The trypsin IMER described herein is applicable for the digestion of protein mixtures. Even at a 1000-fold molar excess of one protein, low-abundance proteins are readily identified, in combination with MS/MS analysis. An online setup of the IMER with reversed phase nano-HPLC separation and nano-ESI-MS/MS analysis was established. The great potential of the trypsin IMER for proteomics applications comprise short digestion times in the range of seconds to minutes, in addition to improved digestion efficiencies, compared to in-solution digestion.

  9. Changes in antigenicity of porcine serum albumin in gamma-irradiated sausage extract by treatment with pepsin and trypsin

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Koth-Bong-Woo-Ri; Song, Eu-Jin [Department of Food Science and Technology/Institute of Food Science, Pukyong National University, Busan 608-737 (Korea, Republic of); Lee, So-Young [Traditional Food Research Group, Korea Food Research Institute, Seongnam 463-746 (Korea, Republic of); Park, Jin-Gyu [Radiation Food Science and Biotechnology, Advanced Radiation Technology Institute, Korea Atomic Energy Research Institute, Jeongup 580-185 (Korea, Republic of); Lee, Ju-Woon [National Federation of Fisheries Cooperatives, Fisheries Economic Institute, Seoul 138-827 (Korea, Republic of); Byun, Myung-Woo [Department of Culinary Nutrition, Woosong University, Daejon 300-718 (Korea, Republic of); Ahn, Dong-Hyun, E-mail: dhahn@pknu.ac.kr [Department of Food Science and Technology/Institute of Food Science, Pukyong National University, Busan 608-737 (Korea, Republic of)

    2011-11-15

    Pork is known as an allergenic food with porcine serum albumin (PSA, 66 kDa) representing the major allergen. This study was conducted to investigate the change in antigenicity of PSA in gamma-irradiated sausage extract treated with pepsin and trypsin. Sausage products (A and B) were irradiated at 1, 3, 10, and 20 kGy. After irradiation, sausage proteins were extracted and digested with pepsin (1:200, 30 min) and trypsin (1:300, 5, 30, 60, 90, and 120 min). The binding ability of PSA in extracts of the irradiated sausages (A and B) decreased by over 3 kGy relative to the binding ability of PSA in extracts of intact sausages and showed no notable differences when the dose of radiation ranged from 3 to 20 kGy. After treatment with pepsin and trypsin, the binding ability of PSA in extracts of the irradiated sausages was decreased more relative to that of intact sausages and showed no significant differences when the period of trypsin treatment is increased or when the dose of irradiation is increased. The sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) results indicated that there was no visible change in the intensity of the PSA band in extracts of the irradiated sausages. After pepsin and trypsin treatment, the intensity of PSA band faded with increasing doses of irradiation. In conclusion, antigenicity of PSA in pork sausages could be reduced by gamma irradiation. - Highlights: > Change in antigenicity of PSA in irradiated sausage extract (ISE) was examined. > Binding ability of PSA in ISE was decreased compared to intact extract. > Binding ability of PSA in ISE after enzyme treatments was also further decreased. > Intensity of PSA band in ISE after enzyme treatments became weak.

  10. Changes in antigenicity of porcine serum albumin in gamma-irradiated sausage extract by treatment with pepsin and trypsin

    International Nuclear Information System (INIS)

    Kim, Koth-Bong-Woo-Ri; Song, Eu-Jin; Lee, So-Young; Park, Jin-Gyu; Lee, Ju-Woon; Byun, Myung-Woo; Ahn, Dong-Hyun

    2011-01-01

    Pork is known as an allergenic food with porcine serum albumin (PSA, 66 kDa) representing the major allergen. This study was conducted to investigate the change in antigenicity of PSA in gamma-irradiated sausage extract treated with pepsin and trypsin. Sausage products (A and B) were irradiated at 1, 3, 10, and 20 kGy. After irradiation, sausage proteins were extracted and digested with pepsin (1:200, 30 min) and trypsin (1:300, 5, 30, 60, 90, and 120 min). The binding ability of PSA in extracts of the irradiated sausages (A and B) decreased by over 3 kGy relative to the binding ability of PSA in extracts of intact sausages and showed no notable differences when the dose of radiation ranged from 3 to 20 kGy. After treatment with pepsin and trypsin, the binding ability of PSA in extracts of the irradiated sausages was decreased more relative to that of intact sausages and showed no significant differences when the period of trypsin treatment is increased or when the dose of irradiation is increased. The sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) results indicated that there was no visible change in the intensity of the PSA band in extracts of the irradiated sausages. After pepsin and trypsin treatment, the intensity of PSA band faded with increasing doses of irradiation. In conclusion, antigenicity of PSA in pork sausages could be reduced by gamma irradiation. - Highlights: → Change in antigenicity of PSA in irradiated sausage extract (ISE) was examined. → Binding ability of PSA in ISE was decreased compared to intact extract. → Binding ability of PSA in ISE after enzyme treatments was also further decreased. → Intensity of PSA band in ISE after enzyme treatments became weak.

  11. Spleen size changes in children with homozygous β-thalassaemia in relation to blood transfusion

    International Nuclear Information System (INIS)

    Karpathios, Th.; Antypas, A.; Dimitriou, P.; Nicolaidou, P.; Fretzayas, A.; Thomaidis, Th.; Matsaniotis, N.

    1982-01-01

    18 thalassaemic children, aged 3.5 to 13 years comprise our clinical material. In 14 of them, clinically elicited spleen markings, haematocrit, blood platelet count and red cell morphology were studied daily for a whole period between 2 transfusions. In 10 patients considerable changes in spleen size were noticed. According to our clinical observations the spleen size starts decreasing 1 to 3 d after blood transfusion up to the 10th posttransfusion day fluctuating thereafter to reach its maximum size again prior to the next blood transfusion. The decrease of spleen size was followed by an increase of haematocrit and blood platelet count and vice versa. 4 additional children were studied clinically only twice: prior to and 7 to 10 d after blood transfusion. A definite decrease of the spleen size following blood transfusion was observed. Spleen and liver sup(99m)Tc-sulfur colloid uptake was studied in 10 of the above children prior to and 7 to 10 d after blood transfusion. Statistically significant post-transfusion increase of the spleen uptake was demonstrated. Our findings suggest that (a) splenic size is relevant to blood volume sequestrated int this organ, (b) splenic radioactive uptake increases with its post-transfusion reductin in size. (author)

  12. Influence of apoptosis on liver and spleen resistance in dogs with visceral leishmaniosis.

    Science.gov (United States)

    Moreira, Pamela Rodrigues Reina; Franciscato, Douglas Augusto; Rossit, Sabrina Micelli; Munari, Danísio Prado; Vasconcelos, Rosemeri de Oliveira

    2016-01-01

    The aim of this study was to evaluate apoptosis and parasite load in the liver and spleen of dogs with visceral leishmaniosis (VL), using immunohistochemistry. Liver and spleen samples from 71 dogs with VL were used. The parasite load in the spleen and liver showed significant difference between organs in infected group (P=0.0219). The density of the parasite load in the spleen (median=2.4) was higher than liver (median=0.8). Immunodetection of apoptotic cells was predominant in lymphocytes and differ between the infected and control group in spleen (P=0.0307) and liver (P=0.0346). There was a significant correlation between apoptosis and parasite load (P = 0.0084; r=0.3104) only in the spleen of the infected group, where it was observed that, when increasing the number of apoptotic cells increases the parasitic load. It was concluded that the liver and spleen of infected dogs presented greater numbers of cells undergoing apoptosis (lymphocytes) than the control group, thus suggesting that this process may be contributing towards the survival of Leishmania in these organs, because lymphocyte in apoptosis did not have the ability to present and recognize the antigen, allowing the survival of the parasite.

  13. Factors influencing liver and spleen volume changes after donor hepatectomy for living donor liver transplantation

    Energy Technology Data Exchange (ETDEWEB)

    Bae, Ji Hee; Ryeom, Hunku; Song, Jung Hup [Kyungpook National University Hospital, Daegu (Korea, Republic of)

    2013-11-15

    To define the changes in liver and spleen volumes in the early postoperative period after partial liver donation for living-donor liver transplantation (LDLT) and to determine factors that influence liver and spleen volume changes. 27 donors who underwent partial hepatectomy for LDLT were included in this study. The rates of liver and spleen volume change, measured with CT volumetry, were correlated with several factors. The analyzed factors included the indocyanine green (ICG) retention rate at 15 minutes after ICG administration, preoperative platelet count, preoperative liver and splenic volumes, resected liver volume, resected-to-whole liver volume ratio (LV{sub R}/LV{sub W}), resected liver volume to the sum of whole liver and spleen volume ratio [LV{sub R}/(LV{sub W} + SV{sub 0})], and pre and post hepatectomy portal venous pressures. In all hepatectomy donors, the volumes of the remnant liver and spleen were increased (increased rates, 59.5 ± 50.5%, 47.9 ± 22.6%). The increment rate of the remnant liver volume revealed a positive correlation with LV{sub R}/LV{sub W} (r = 0.759, p < 0.01). The other analyzed factors showed no correlation with changes in liver and spleen volumes. The spleen and remnant liver volumes were increased at CT volumetry performed 2 weeks after partial liver donation. Among the various analyzed factors, LV{sub R}/LV{sub W} influences the increment rate of the remnant liver volume.

  14. Alzheimer's disease pathological lesions activate the spleen tyrosine kinase.

    Science.gov (United States)

    Schweig, Jonas Elias; Yao, Hailan; Beaulieu-Abdelahad, David; Ait-Ghezala, Ghania; Mouzon, Benoit; Crawford, Fiona; Mullan, Michael; Paris, Daniel

    2017-09-06

    The pathology of Alzheimer's disease (AD) is characterized by dystrophic neurites (DNs) surrounding extracellular Aβ-plaques, microgliosis, astrogliosis, intraneuronal tau hyperphosphorylation and aggregation. We have previously shown that inhibition of the spleen tyrosine kinase (Syk) lowers Aβ production and tau hyperphosphorylation in vitro and in vivo. Here, we demonstrate that Aβ-overexpressing Tg PS1/APPsw, Tg APPsw mice, and tau overexpressing Tg Tau P301S mice exhibit a pathological activation of Syk compared to wild-type littermates. Syk activation is occurring in a subset of microglia and is age-dependently increased in Aβ-plaque-associated dystrophic neurites of Tg PS1/APPsw and Tg APPsw mice. In Tg Tau P301S mice, a pure model of tauopathy, activated Syk occurs in neurons that show an accumulation of misfolded and hyperphosphorylated tau in the cortex and hippocampus. Interestingly, the tau pathology is exacerbated in neurons that display high levels of Syk activation supporting a role of Syk in the formation of tau pathological species in vivo. Importantly, human AD brain sections show both pathological Syk activation in DNs around Aβ deposits and in neurons immunopositive for pathological tau species recapitulating the data obtained in transgenic mouse models of AD. Additionally, we show that Syk overexpression leads to increased tau accumulation and promotes tau hyperphosphorylation at multiple epitopes in human neuron-like SH-SY5Y cells, further supporting a role of Syk in the formation of tau pathogenic species. Collectively, our data show that Syk activation occurs following Aβ deposition and the formation of tau pathological species. Given that we have previously shown that Syk activation also promotes Aβ formation and tau hyperphosphorylation, our data suggest that AD pathological lesions may be self-propagating via a Syk dependent mechanism highlighting Syk as an attractive therapeutic target for the treatment of AD.

  15. Palliative Spleen Irradiation: Can we Standardize its Technique?

    International Nuclear Information System (INIS)

    NAZMY, M.S.; RADWAN, A.; MOKHTAR, M.

    2008-01-01

    To explore the pattern of practice of palliative splenic irradiation (PSI) at the National Cancer Institute (NCI), Cairo University. Patients and Methods: The medical records of patients referred for PSI during the time period from 1990 to 2005 were retrospectively reviewed. We compared the three most common planning techniques (two parallel opposing, single direct field, anterior and lateral fields). Results: Eighteen patients who received PSI were identified. Thirteen patients were diagnosed as CML and 5 as CLL. The mean age of the patients was 44 (±16) years and the majority were men (60%). Spleen enlargement was documented in all cases. The single direct anterior field was the most commonly used technique. The dose per fraction ranged from 25 c Gy to 100 c Gy. The total dose ranged from 125 c Gy to 1200 c Gy and the median was 200 c Gy (mean 327 c Gy). There was no significant difference between CML and CLL patients regarding the dose level. Three out of 5 CLL patients and only one out of 13 CML patients received re-irradiation. All patients showed subjective improvement regarding pain and swelling. There was a significant increase in the hemoglobin level and a significant decrease in the WBC count. The single direct field shows variations in the dose from 56 to 102%; however, it is the simplest and the best regarding the dose to the surrounding normal tissues especially the kidney and the liver. Conclusion: PSI has a significant palliative benefit. Although the most widely accepted technique is the 2 parallel opposing anterior-posterior fields, single anterior field is also considered as a suitable option. Higher doses are needed for CLL patients compared to CML patients

  16. Tie2 Expressing Monocytes in the Spleen of Patients with Primary Myelofibrosis.

    Directory of Open Access Journals (Sweden)

    Rita Campanelli

    Full Text Available Primary myelofibrosis (PMF is a Philadelphia-negative (Ph- myeloproliferative disorder, showing abnormal CD34+ progenitor cell trafficking, splenomegaly, marrow fibrosis leading to extensive extramedullary haematopoiesis, and abnormal neoangiogenesis in either the bone marrow or the spleen. Monocytes expressing the angiopoietin-2 receptor (Tie2 have been shown to support abnormal angiogenic processes in solid tumors through a paracrine action that takes place in proximity to the vessels. In this study we investigated the frequency of Tie2 expressing monocytes in the spleen tissue samples of patients with PMF, and healthy subjects (CTRLs, and evaluated their possible role in favouring spleen angiogenesis. We show by confocal microscopy that in the spleen tissue of patients with PMF, but not of CTRLs, the most of the CD14+ cells are Tie2+ and are close to vessels; by flow cytometry, we found that Tie2 expressing monocytes were Tie2+CD14lowCD16brightCDL62-CCR2- (TEMs and their frequency was higher (p = 0.008 in spleen tissue-derived mononuclear cells (MNCs of patients with PMF than in spleen tissue-derived MNCs from CTRLs undergoing splenectomy for abdominal trauma. By in vitro angiogenesis assay we evidenced that conditioned medium of immunomagnetically selected spleen tissue derived CD14+ cells of patients with PMF induced a denser tube like net than that of CTRLs; in addition, CD14+Tie2+ cells sorted from spleen tissue derived single cell suspension of patients with PMF show a higher expression of genes involved in angiogenesis than that found in CTRLs. Our results document the enrichment of Tie2+ monocytes expressing angiogenic genes in the spleen of patients with PMF, suggesting a role for these cells in starting/maintaining the pathological angiogenesis in this organ.

  17. Computed tomography of the spleen and liver in sickle cell disease

    International Nuclear Information System (INIS)

    Magid, D.; Fishman, E.K.; Siegelman, S.S.

    1984-01-01

    The spleen was assessed in 10 patients with sickle cell disease studied with computed tomography (CT) for abdominal pain and/or unexplained fever. Patients with homozygous sickle cell anemia were found to have small, densely calcified spleens with occasional low-density infarcts. Five of six had hepatomegaly, and there was one case each of hepatic abscess, infarcts, and hemochromatosis. All patients with heterozygous sickle cell disease were found to have splenomegaly, with a variety of findings including acute hemorrhage, acute and chronic infarcts, rupture, and possible sequestration. It was concluded that CT is useful for evaluating the status of the spleen and liver in symptomatic patients with sickle cell disease

  18. Inflammatory pseudotumor of the spleen: report of a case in a child

    International Nuclear Information System (INIS)

    Dardanelli, Esteban; Cermeno, Claudia; Rizzi, Ana Maria; Felipe, Laura; Goldberg, Alberto; Moguillansky, Silvia

    2003-01-01

    We report a case of inflammatory pseudotumor of the spleen in a 4 years old child. Clinical findings were limited to diffuse abdominal pain and a palpable mass at the left hypochondrium. Ultrasonography and CT revealed a solid homogeneous mass in the lower pole of the spleen, with irregular enhancement with IV contrast. Laparoscopic splenectomy was performed. The diagnosis was histological. The pseudotumor of the spleen is extremely rare, especially in children, with only 4 cases (including our own) reported in the literature. Our patient constitutes the youngest case ever reported. (author)

  19. Active suppression of in vitro reactivity of spleen cells after BCG treatment

    International Nuclear Information System (INIS)

    Orbach-Arbouys, S.; Poupon, M.F.

    1978-01-01

    It was found that spleen cells from mice injected i.v. with large doses of BCG responded to PHA stimulation less intensely than did normal spleen cells. It was shown that nylon wool column purified BCG treated T cells also had a low PHA reactivity. Unfractionated spleen cells, adherent cells or T-enriched populations from BCG treated mice, when added to normal T cells lowered their PHA reactivity. When the same BCG treated cell populations were added to tumor cells in vitro, they inhibited their growth. (author)

  20. Recurrent spleen enlargement during cyclic granulocyte-macrophage colony-stimulating factor therapy for myelodysplastic syndrome

    International Nuclear Information System (INIS)

    Delmer, A.; Karmochkine, M.; Cadiou, M.; Gerhartz, H.; Zittoun, R.

    1990-01-01

    A 65-year-old woman with refractory anemia with excess of blasts received sequential courses of granulocyte-macrophage colony-stimulating factor therapy (GM-CSF) and low-dose cytosine arabinoside. Each course of GM-CSF induced a rapid and tremendous increase in leukocyte count as well as in spleen size, 111-indium chloride scanning suggested a myeloid metaplasia of the spleen. This observation suggests that in some patients the granulopoietic response to the myeloid growth factor stimulation may be predominant in the spleen

  1. Knowledge of the anatomy and physiology of the spleen throughout Antiquity and the Early Middle Ages.

    Science.gov (United States)

    Paraskevas, George K; Koutsouflianiotis, Konstantinos N; Nitsa, Zoi; Demesticha, Theano; Skandalakis, Panagiotis

    2016-01-01

    The evolution of knowledge regarding the anatomy and physiology of the spleen throughout Antiquity and the Early Middle Ages is described, and general perceptions about this organ during different eras along this time line are presented. The original words of great physicians from the period of time stretching from Ancient Egypt to the Avicennan era are quoted and discussed to demonstrate how knowledge of the spleen has evolved and to present the theories that dominated each era. Furthermore, theories about illnesses relating to the spleen are reported, which show how this organ was perceived-in terms of its function and anatomy-during each era.

  2. Regeneration of hemopoietic precursor cells in spleen organ cultures from irradiated mice: influence of genotype of cells injected and of the spleen microenvironment

    International Nuclear Information System (INIS)

    von Melchner, H.; Lieschke, G.J.

    1981-01-01

    The regeneration of hemopoietic precursor cells was monitored in spleen organ cultures from lethally irradiated mice injected with 10(7) normal syngeneic or allogeneic bone marrow cells. The important role of the microenvironment in supporting hemopoiesis was confirmed by the failure of mutant Sl/Sld spleens to support CFC regeneration in organ cultures. However, the extent and quality of the CFC regeneration was clearly dependent on the genetic properties of the injected cells. Evidence for this was obtained from the regeneration patterns of various CFC types in organ cultured spleens derived from different mouse donor-recipient strain combinations that maintained the differences in the bone marrow frequency of various CFC types characteristic of the donor strain

  3. Regeneration of hemopoietic precursor cells in spleen organ cultures from irradiated mice: influence of genotype of cells injected and of the spleen microenvironment

    International Nuclear Information System (INIS)

    von Melchner, H.; Lieschke, G.J.

    1981-01-01

    The regeneration of hemopoietic precursor cells (colony-forming cells, CFC) was monitored in spleen organ cultures from lethally irradiated mice injected with 10(7) normal syngeneic or allogeneic bone marrow cells. The important role of the microenvironment in supporting hemopoiesis was confirmed by the failure of mutant S1/S1d spleens to support CFC regeneration in organ cultures. However, the extent and quality of the CFC regeneration was clearly dependent on the genetic properties of the injected cells. Evidence for this was obtained from the regeneration patterns of various CFC types in organ cultured spleens derived from different mouse donor-recipient strain combinations (CBA/CBA, CBA/C57BL, CBA/BALB/c, C57BL/C57BL, C57BL/CBA, C57BL/BALB/c) that maintained the differences in the bone marrow frequency of various CFC types characteristic of the donor strain

  4. Development of a rapid high-efficiency scalable process for acetylated Sus scrofa cationic trypsin production from Escherichia coli inclusion bodies.

    Science.gov (United States)

    Zhao, Mingzhi; Wu, Feilin; Xu, Ping

    2015-12-01

    Trypsin is one of the most important enzymatic tools in proteomics and biopharmaceutical studies. Here, we describe the complete recombinant expression and purification from a trypsinogen expression vector construct. The Sus scrofa cationic trypsin gene with a propeptide sequence was optimized according to Escherichia coli codon-usage bias and chemically synthesized. The gene was inserted into pET-11c plasmid to yield an expression vector. Using high-density E. coli fed-batch fermentation, trypsinogen was expressed in inclusion bodies at 1.47 g/L. The inclusion body was refolded with a high yield of 36%. The purified trypsinogen was then activated to produce trypsin. To address stability problems, the trypsin thus produced was acetylated. The final product was generated upon gel filtration. The final yield of acetylated trypsin was 182 mg/L from a 5-L fermenter. Our acetylated trypsin product demonstrated higher BAEE activity (30,100 BAEE unit/mg) than a commercial product (9500 BAEE unit/mg, Promega). It also demonstrated resistance to autolysis. This is the first report of production of acetylated recombinant trypsin that is stable and suitable for scale-up. Copyright © 2015 Elsevier Inc. All rights reserved.

  5. Atomic-scale investigation of the interactions between tetrabromobisphenol A, tetrabromobisphenol S and bovine trypsin by spectroscopies and molecular dynamics simulations

    International Nuclear Information System (INIS)

    Ding, Keke; Zhang, Huanxin; Wang, Haifei; Lv, Xuan; Pan, Liumeng; Zhang, Wenjing; Zhuang, Shulin

    2015-01-01

    Highlights: • The interaction of TBBPA/TBBPS with bovine trypsin was deciphered for the first time. • The fluorescence of bovine trypsin was quenched in a concentration-dependent mode. • TBBPA and TBBPS bind at the ANS binding site with distinct binding modes. • TBBPS has a higher binding affinity toward bovine trypsin than TBBPA. • Our in vitro and in silico approach is helpful to assess risk of TBBPA-related BFRs. - Abstract: Tetrabromobisphenol A (TBBPA) and its replacement alternative tetrabromobisphenol S (TBBPS) are used widely as brominated flame retardants (BFRs). However, the potential risk of their effects on bovine trypsin remains largely unknown. We investigated the effects of TBBPA and TBBPS to bovine trypsin by the fluorescence spectroscopy, circular dichroism and molecular dynamics (MD) simulations. They statically quenched the intrinsic fluorescence of bovine trypsin in a concentration-dependent mode and caused slight red-shifted fluorescence. The short and long fluorescence lifetime decay components of bovine trypsin were both affected, partly due to the disturbed microenvironmental changes of Trp215. The β-sheet content of bovine trypsin was significantly reduced from 82.4% to 75.7% and 76.6% by TBBPA and TBBPS, respectively, possibly impairing the physiological function of bovine trypsin. TBBPA and TBBPS bind at the 8-anilinonaphthalene-1-sulfonate (ANS) binding site with an association constant of 1.09 × 10 4 M −1 and 2.41 × 10 4 M −1 at 298 K, respectively. MD simulations revealed that van der Waals interactions and hydrogen bond interactions are dominant for TBBPA, whereas electrostatic interactions are critical for TBBPS. Our in vitro and in silico studies are beneficial to the understanding of risk assessment and future design of environmental benign BFRs.

  6. Quantification of trypsin with a radioimmunoassay in herring larvae (Clupea harengus L.) compared with a highly sensitive fluorescence technique to determine tryptic enzyme activity

    International Nuclear Information System (INIS)

    Ueberschaer, B.; Pedersen, B.H.; Hjelmeland, K.

    1993-01-01

    Enzymatic activity and quantity of the protease trypsin were measured in individual herring larvae (Clupea harengus L.). The enzymatic activity assay was done by a fluorescence technique, and a radioimmunoassay was used for quantification of trypsin. The results are compared and the differences between the techniques discussed. Both methods have similar results, as high or low values in trypsin quantity were reflected in high or low values of tryptic activity. Quantity and activity were linearly and positively correlated, but small differences between methods were found at the lowest detection limits. Both techniques reflect the high variability between individual larvae. (orig.)

  7. The effect of bound to dialdehudecellulose protein concentration on the activity of immobilized trypsin after γ-irradiation and in process of storage

    International Nuclear Information System (INIS)

    Belov, A.A.; Ryl'tsev, V.V.; Ignatyuk, T.E.; Filatov, V.N.

    1994-01-01

    It is found the complex effect of the bound enzyme concentration on the proteolytic activity of trypsin immobilized to dialdehydecellulose (preriodate oxidation) after γ-irradiation and in process of storage. It is shown the occurance of three stages of immobilized enzyme inactivation in process of immobilization and storage. The velocity of inactivation did not depend on bound trypsin concentration. The ratio of proteolytic activity of samples before and after γ-irradiation was increased with the increase of immobilized to carrier enzyme concentration and was not change (in range of experiment error) in process of storage. The results were compared with that of cryctlline trypsin

  8. Biopotency of serine protease inhibitors from cowpea (Vigna unguiculata) seeds on digestive proteases and the development of Spodoptera littoralis (Boisduval).

    Science.gov (United States)

    Abd El-latif, Ashraf Oukasha

    2015-05-01

    Serine protease inhibitors (PIs) have been described in many plant species and are universal throughout the plant kingdom, where trypsin inhibitors is the most common type. In the present study, trypsin and chymotrypsin inhibitory activity was detected in the seed flour extracts of 13 selected cultivars/accessions of cowpea. Two cowpea cultivars, Cream7 and Buff, were found to have higher trypsin and chymotrypsin inhibitory potential compared to other tested cultivars for which they have been selected for further purification studies using ammonium sulfate fractionation and DEAE-Sephadex A-25 column. Cream7-purified proteins showed two bands on sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) corresponding to molecular mass of 17.10 and 14.90 kDa, while the purified protein from Buff cultivar showed a single band corresponding mass of 16.50 kDa. The purified inhibitors were stable at temperature below 60°C and were active at wide range of pH from 2 to 12. The kinetic analysis revealed noncompetitive type of inhibition for both inhibitors against both enzymes. The inhibitor constant (Ki ) values suggested high affinity between inhibitors and enzymes. Purified inhibitors were found to have deep and negative effects on the mean larval weight, larval mortality, pupation, and mean pupal weight of Spodoptera littoralis, where Buff PI was more effective than Cream7 PI. It may be concluded that cowpea PI gene(s) could be potential insect control protein for future studies in developing insect-resistant transgenic plants. © 2014 Wiley Periodicals, Inc.

  9. The use of trypsin-like blood activity as a marker of pulmonary fibrosis severity.

    Directory of Open Access Journals (Sweden)

    V. V. Rodionova

    2018-05-01

    Full Text Available Purpose – to determine changes in the trypsin-like blood activity and its relationship with acute phase indices of inflammation in patients with pulmonary fibrosis (PF as a marker of course severity and prognosis of the disease. Materials and Methods: The study included 18 patients: 15 (83% women and 3 (17% men, mean age 53±2.5 years. The control group included 15 practically healthy persons. All the examined patients (n=18 were divided into two groups: with mild or moderately severe PF – 8 (44.4% patients (group I, severe PF – 10 (55.6% of patients (group II. Duration of the disease - from 1 month. up to 4 years. Patients underwent clinic-laboratory and anthropometric studies, a mMRC questionnaire was used, blood saturation was measured, lung radiography in 2 projections, echocardiography, and if necessary a high-resolution computer tomography etc were performed. Results and discussion: More than half of the patients were overweight (44,4% or had obesity of І-ІІІ st. (27.8%. The severity of dyspnea according to mMRC scale was 3.0 (3.0-4.0 points in patients of group II and 2.5 (2.0-3.0 points in patients of group I. There was a decrease in C-reactive protein (CRP in group I and a tendency to increase in patients of group II. When analyzing the indicator of trypsin-like blood activity (TLA, it was found that the median TLA at the beginning of the observation was twice as high as in healthy individuals, direct correlation was established between the level of TLA and the severity of the disease course. After treatment the level of TLA decreased in group I patients. In the severe course of PF, the average TLA level remained high. The progredient course of LF is characterized by severe clinical symptoms, a significant increase in TLA, CRP, reduced O2 saturation, a lack of response to treatment and an unfavorable prognosis. The TLA index can be used as a biochemical marker of the severity of PF along with the CRPindex, O2 saturation and

  10. Detection of viral infection by immunofluorescence in formalin-fixed tissues, pretreated with trypsin

    Directory of Open Access Journals (Sweden)

    O. M. Barth

    1988-06-01

    Full Text Available The presence of viral antigen in sections from formalin-fixed and paraffin-embedded human tissues was demonstrated by trypsin digestion followed by direct or indirect immunofluorescence. The specimens may be used for retrospective diagnosis. The immunofluorescence technique has to be adapted to the suspected virus infection on the basis of previous histopathology study. Variations of trypsin concentration time and temperature of incubation, expose different viral antigens and have to be previously tested for each unknown system. For measles virus detection in lung a stronger digestion has to be applied as compared to adenovirus or respiratory disease viruses in the same tisue. Flavivirus in liver tissue needs a weaker digestion. The reproducibility of the method makes it useful as a routine technique in diagnosis of virus infection.A presença de antígeno viral em cortes de tecidos humanos fixados em formol e emblocados em parafina foi demonstrada pela digestão com tripsina foi demonstrada pela ingestão com tripsina seguida de imunofluorescência direta ou indireta. Os espécimens podem ser utilizados para diagnoses retrospectivas. A técnica da imunofluorescência deve ser adaptada à infecção viral suspeita segundo diagnosie histopatológica prévia. Os parâmetros para a digestão do tecido pela tripsina, relacionados à concentração, duração de atuação e temperatura, expõem diferentes antígenos virais e devem ser previamente testados para cada sistema a ser estabelecido. Uma digestão mais intensa deve ser aplicada para a detecção do vírus do sarampo em tecido pulmonar do que para adenovírus ou vírus respiratório sincicial no mesmo tecido. Por outro lado, o vírus da febre amarela em tecido de fígado necessita de uma digestão mais fraca.

  11. Spleen artery embolization increases the success of nonoperative management following blunt splenic injury

    Directory of Open Access Journals (Sweden)

    Isaac Chun-Jen Chen

    2011-08-01

    Conclusion: Performance of SAE for the patients with blunt splenic injury could increase the successful rate of NOM significantly and safely. An algorithm including the angioembolization might be beneficial in the management of patients with blunt spleen trauma.

  12. Imaging of liver and spleen candidiasis in patients with acute leukemia

    International Nuclear Information System (INIS)

    Seino, Yasuo; Tamakawa, Y.; Kato, T.; Kimura, Y.; Miyazaki, S.; Miura, R.; Ishida, H.

    1988-01-01

    Four patients with acute leukemia were found to have candidal abscess of liver and spleen. CT and US showed hepatosplenomegaly and microabscess. These findings might be useful in diagnosis of visceral candidiasis. (author)

  13. Imaging of liver and spleen candidiasis in patients with acute leukemia

    Energy Technology Data Exchange (ETDEWEB)

    Seino, Yasuo; Tamakawa, Y.; Kato, T.; Kimura, Y.; Miyazaki, S.; Miura, R.; Ishida, H.

    1988-01-01

    Four patients with acute leukemia were found to have candidal abscess of liver and spleen. CT and US showed hepatosplenomegaly and microabscess. These findings might be useful in diagnosis of visceral candidiasis.

  14. Dose to the liver and spleen in pediatric patients undergoing technetium-99m sulfur colloid scans

    International Nuclear Information System (INIS)

    Thomas, S.R.; Purdom, R.C.; Kereiakes, J.G.; Gelfand, M.J.; Maxon, H.R.

    1979-01-01

    Quantitative conjugate view external counting techniques were applied to determine radiation dose to the liver and spleen in pediatric patients undergoing /sup 99m/Tc-sulfur colloid (Tc-SC) liver scans. The effective half-life of /sup99m/Tc-SC was 5.8 +- 0.23 hours and 5.2 +- 0.68 hours in the liver and spleen, respectively. Dose per administered activity ranged from 0.34 to 0.63 rad/mCi (92 to 170 μGy/MBq) for the liver and 0.35 to 1.96 rad/mCi (95.0 to 530.0 μGy/MBq) for the spleen. The spleen to liver dose ratio ranged from 1.0 to 4.9. These values are compared with results extrapolated from published adult data to the pediatric population

  15. Scintigraphic diagnosis and computed tomographic localization of an accessory spleen following relapse of chronic immune thrombocytopaenia

    International Nuclear Information System (INIS)

    Cardaci, G.T.; Blake, M.P.

    1992-01-01

    Chronic immune thrombocytopaenia is an immunologically mediated disorder resulting in disordered platelet kinetics and potentially life-threatening disease. Failure of medical therapy is an indication for splenectomy, and responses are seen in 80% of patients following this procedure. An important cause of relapse following splenectomy is the presence of an accessory spleen. A patient with Hodgkin's Disease developed chronic immune thrombocytopaenia despite previous splenectomy. A remission was induced with immunosuppressive therapy, but he later relapsed. An accessory spleen was detected using 99 m Tc denatured red blood cells and localized using computed tomography. Resection of the accessory spleen resulted in clinical remission. As accessory spleens are often small in size, combined modality imaging is recommended in the evaluation of this disorder. 15 refs., 2 figs

  16. Compositions, Protease Inhibitor and Gelling Property of Duck Egg Albumen as Affected by Salting

    Science.gov (United States)

    2018-01-01

    Chemical compositions, trypsin inhibitory activity, and gelling properties of albumen from duck egg during salting of 30 days were studied. As the salting time increased, moisture content decreased, the salt content and surface hydrophobicity increased (psalting time of 30 days (psalting of 30 days. Based on texture profile analysis, hardness, springiness, gumminess, chewiness, and resilience of albumen gel decreased with increasing salting time. Conversely, salted albumen gels exhibited higher cohesiveness and adhesiveness, compared to those of fresh albumen. Scanning electron microscopic study revealed that gel of salted albumen showed the larger voids and less compactness. In general, salting lowered trypsin inhibitory activity and gelling property of albumen from duck egg to some extent. Nevertheless, the salted albumen with the remaining inhibitor could be an alternative additive for surimi or other meat products to prevent proteolysis. PMID:29725221

  17. Loss of Sympathetic Nerves in Spleens from Patients with End Stage Sepsis

    Directory of Open Access Journals (Sweden)

    Donald B. Hoover

    2017-12-01

    Full Text Available The spleen is an important site for central regulation of immune function by noradrenergic sympathetic nerves, but little is known about this major region of neuroimmune communication in humans. Experimental studies using animal models have established that sympathetic innervation of the spleen is essential for cholinergic anti-inflammatory responses evoked by vagal nerve stimulation, and clinical studies are evaluating this approach for treating inflammatory diseases. Most data on sympathetic nerves in spleen derive from rodent studies, and this work has established that remodeling of sympathetic innervation can occur during inflammation. However, little is known about the effects of sepsis on spleen innervation. Our primary goals were to (i localize noradrenergic nerves in human spleen by immunohistochemistry for tyrosine hydroxylase (TH, a specific noradrenergic marker, (ii determine if nerves occur in close apposition to leukocytes, and (iii determine if splenic sympathetic innervation is altered in patients who died from end stage sepsis. Staining for vesicular acetylcholine transporter (VAChT was done to screen for cholinergic nerves. Archived paraffin tissue blocks were used. Control samples were obtained from trauma patients or patients who died after hemorrhagic stroke. TH + nerves were associated with arteries and arterioles in all control spleens, occurring in bundles or as nerve fibers. Individual TH + nerve fibers entered the perivascular region where some appeared in close apposition to leukocytes. In marked contrast, spleens from half of the septic patients lacked TH + nerves fibers and the average abundance of TH + nerves for the septic group was only 16% of that for the control group (control: 0.272 ± 0.060% area, n = 6; sepsis: 0.043 ± 0.026% area, n = 8; P < 0.005. All spleens lacked cholinergic innervation. Our results provide definitive evidence for the distribution of noradrenergic

  18. Aniline-induced nitrosative stress in rat spleen: Proteomic identification of nitrated proteins

    International Nuclear Information System (INIS)

    Fan Xiuzhen; Wang Jianling; Soman, Kizhake V.; Ansari, G.A.S.; Khan, M. Firoze

    2011-01-01

    Aniline exposure is associated with toxicity to the spleen which is characterized by splenomegaly, hyperplasia, fibrosis, and a variety of sarcomas on chronic exposure in rats. However, mechanisms by which aniline elicits splenotoxic responses are not well understood. Earlier we have shown that aniline exposure leads to increased nitration of proteins in the spleen. However, nitrated proteins remain to be characterized. Therefore, in the current study using proteomic approaches, we focused on characterizing the nitrated proteins in the spleen of aniline-exposed rats. Aniline exposure led to increased tyrosine nitration of proteins, as determined by 2D Western blotting with anti-3-nitrotyrosine specific antibody, compared to the controls. The analyzed nitrated proteins were found in the molecular weight range of 27.7 to 123.6 kDa. A total of 37 nitrated proteins were identified in aniline-treated and control spleens. Among them, 25 were found only in aniline-treated rats, 11 were present in both aniline-treated and control rats, while one was found in controls only. The nitrated proteins identified mainly represent skeletal proteins, chaperones, ferric iron transporter, enzymes, nucleic acids binding protein, and signaling and protein synthesis pathways. Furthermore, aniline exposure led to significantly increased iNOS mRNA and protein expression in the spleen, suggesting its role in increased reactive nitrogen species formation and contribution to increased nitrated proteins. The identified nitrated proteins provide a global map to further investigate alterations in their structural and functional properties, which will lead to a better understanding of the role of protein nitration in aniline-mediated splenic toxicity. - Highlights: → Proteomic approaches are used to identify nitrated proteins in the spleen. → Twenty five nitrated proteins were found only in the spleen of aniline-treated rats. → Aniline exposure led to increased iNOS mRNA and protein

  19. Role of the spleen in the maturation of B-lymphocytes

    International Nuclear Information System (INIS)

    Strober, S.

    1976-01-01

    Spleen cells from unimmunized Lewis rats were fractionated by 1 x g velocity sedimentation and assayed for their ability to restore the adoptive primary antibody response to horse spleen ferritin in irradiated syngeneic recipients given T cell supplementation. Large, medium and small cell fractions all showed virgin B cell activity. Similar studies with thoractic duct cells show that virgin B cell activity is restricted to the small and medium cells. Large spleen cells produced a 2-ME sensitive adoptive antibody response which persisted for 21 days. All antibody produced by small cells at 21 days was 2-ME resistant. Examination of spleen cells during recovery from sublethal irradiation showed that virgin B cell activity was first detected at 14 days, and is confined to the large cell fraction. Experiments with congenic rats which differ at the immunoglobulin (Ig) light chain allotype showed that small cells from the bone marrow injected intravenously can transform into large Ig-bearing cells in the spleen. The relationship of the subclasses of virgin B cells in the spleen to B cell maturation is discussed

  20. EFFECT OF SEDATION ON CONTRAST-ENHANCED ULTRASONOGRAPHY OF THE SPLEEN IN HEALTHY DOGS.

    Science.gov (United States)

    Rossi, Federica; Fina, Caroline; Stock, Emmelie; Vanderperren, Katrien; Duchateau, Luc; Saunders, Jimmy H

    2016-05-01

    Contrast-enhanced ultrasound of the spleen enables the dynamic assessment of the perfusion of this organ, however, both subjective and quantitative evaluation can be strongly influenced by sedative agent administration. The purpose of this prospective, experimental study was to test effects of two sedative agents on splenic perfusion during contrast-enhanced ultrasound of the spleen in a sample of healthy dogs. Contrast-enhanced ultrasound of the spleen was repeated in six healthy Beagles following a cross-over study design comparing three protocols: awake, butorphanol 0.2 mg/Kg intramuscular (IM), and dexmedetomidine 500 μg/m(2) IM. After intravenous injection of a phospholipid stabilized sulfur hexafluoride microbubble solution (SonoVue®, Bracco Imaging, Milano, Italy), the enhancement intensity and perfusion pattern of the splenic parenchyma were assessed and perfusion parameters were calculated. Normal spleen was slightly heterogeneous in the early phase, but the parenchyma was homogeneous at a later phase. Sedation with butorphanol did not modify perfusion of the spleen. Dexmedetomidine significantly reduced splenic enhancement, providing diffuse parenchymal hypoechogenicity during the entire examination. Measured parameters were significantly modified, with increased arrival time (AT; (dogs. Short-term and diffuse heterogeneity of the spleen in the early venous phase was determined to be a normal finding. © 2016 American College of Veterinary Radiology.

  1. Age dependence of spleen- and muscle-corrected hepatic signal enhancement on hepatobiliary phase gadoxetate MRI

    Energy Technology Data Exchange (ETDEWEB)

    Matoori, Simon [Paracelsus Medical University Salzburg, Department of Radiology, Salzburg (Austria); Hirslanden Clinic St. Anna, Clinical Research Group, Lucerne (Switzerland); Froehlich, Johannes M. [Hirslanden Clinic St. Anna, Clinical Research Group, Lucerne (Switzerland); ETH Zurich, Department of Chemistry and Applied Biosciences, Institute of Pharmaceutical Sciences, Zurich (Switzerland); Cantonal Hospital Winterthur, Department of Radiology, Winterthur (Switzerland); Breitenstein, Stefan [Cantonal Hospital Winterthur, Department of Surgery, Clinic for Visceral and Thoracic Surgery, Winterthur (Switzerland); Doert, Aleksis [Cantonal Hospital Winterthur, Department of Radiology, Winterthur (Switzerland); Pozdniakova, Viktoria [Stavanger University Hospital, Department of Radiology, Stavanger (Norway); Koh, Dow-Mu [Royal Marsden Hospital, Department of Radiology, Surrey, England (United Kingdom); Gutzeit, Andreas [Paracelsus Medical University Salzburg, Department of Radiology, Salzburg (Austria); Hirslanden Clinic St. Anna, Clinical Research Group, Lucerne (Switzerland); Cantonal Hospital Winterthur, Department of Radiology, Winterthur (Switzerland)

    2016-06-15

    To identify correlations of signal enhancements (SE) and SE normalized to reference tissues of the spleen, kidney, liver, musculus erector spinae (MES) and ductus hepatocholedochus (DHC) on hepatobiliary phase gadoxetate-enhanced MRI with patient age in non-cirrhotic patients. A heterogeneous cohort of 131 patients with different clinical backgrounds underwent a standardized 3.0-T gadoxetate-enhanced liver MRI between November 2008 and June 2013. After exclusion of cirrhotic patients, a cohort of 75 patients with no diagnosed diffuse liver disease was selected. The ratio of signal intensity 20 min post- to pre-contrast administration (SE) in the spleen, kidney, liver, MES and DHC, and the SE of the kidney, liver and DHC normalized to the reference tissues spleen or MES were compared to patient age. Patient age was inversely correlated with the liver SE normalized to the spleen and MES SE (both p < 0.001) and proportionally with the SE of the spleen (p = 0.043), the MES (p = 0.030) and the kidney (p = 0.022). No significant correlations were observed for the DHC (p = 0.347) and liver SE (p = 0.606). The age dependence of hepatic SE normalized to the enhancement in the spleen and MES calls for a cautious interpretation of these quantification methods. (orig.)

  2. Preliminary experimental study on splenic hemodynamics of radiofrequency ablation for the spleen.

    Science.gov (United States)

    Baba, Yasutaka; Hayashi, Sadao; Nagasato, Kohei; Higashi, Michiyo; Yoshiura, Takashi

    2017-08-01

    To test the splenic blood flow change after radiofrequency ablation (RFA) of the spleen in a porcine experimental model. Six pigs underwent RFA of the spleen via laparotomy. During the procedure of RFA, clamping of splenic artery (one) and both splenic artery/vein (one) was also performed. Measurement of blood flow of both splenic artery (SA) and splenic vein (SV) with flow-wire at pre- and post-RFA of the spleen was also performed. Ablated splenic lesions were created as estimating ∼50% area of the spleen in all pigs. Resected specimens reveal not only the coagulated necrosis but also the congestion of the spleen. On the SA hemodynamics, maximum peak velocity (MPV) changed from 37 ± 7 to 24 ± 8 cm/s (normal), 11 to 10 cm/s (clamp of the SA), and 12 to 7.5 cm/s (clamp of both SA/SV), respectively. On the SV hemodynamic, MPV changed from 15 ± 5 to 13 ± 4 cm/s (normal), 17 to 15 cm/s (clamp of the SA), and 17 to 26 cm/s (clamp of both SA/SV), respectively. RFA of the spleen could induce coagulation necrosis and reduce the splenic arterial blood flow.

  3. Age dependence of spleen- and muscle-corrected hepatic signal enhancement on hepatobiliary phase gadoxetate MRI

    International Nuclear Information System (INIS)

    Matoori, Simon; Froehlich, Johannes M.; Breitenstein, Stefan; Doert, Aleksis; Pozdniakova, Viktoria; Koh, Dow-Mu; Gutzeit, Andreas

    2016-01-01

    To identify correlations of signal enhancements (SE) and SE normalized to reference tissues of the spleen, kidney, liver, musculus erector spinae (MES) and ductus hepatocholedochus (DHC) on hepatobiliary phase gadoxetate-enhanced MRI with patient age in non-cirrhotic patients. A heterogeneous cohort of 131 patients with different clinical backgrounds underwent a standardized 3.0-T gadoxetate-enhanced liver MRI between November 2008 and June 2013. After exclusion of cirrhotic patients, a cohort of 75 patients with no diagnosed diffuse liver disease was selected. The ratio of signal intensity 20 min post- to pre-contrast administration (SE) in the spleen, kidney, liver, MES and DHC, and the SE of the kidney, liver and DHC normalized to the reference tissues spleen or MES were compared to patient age. Patient age was inversely correlated with the liver SE normalized to the spleen and MES SE (both p < 0.001) and proportionally with the SE of the spleen (p = 0.043), the MES (p = 0.030) and the kidney (p = 0.022). No significant correlations were observed for the DHC (p = 0.347) and liver SE (p = 0.606). The age dependence of hepatic SE normalized to the enhancement in the spleen and MES calls for a cautious interpretation of these quantification methods. (orig.)

  4. Laue diffraction as a tool in dynamic studies: Hydrolysis of a transiently stable intermediate in catalysis by trypsin

    Energy Technology Data Exchange (ETDEWEB)

    Singer, P.T.; Berman, L.E.; Cai, Z.; Mangel, W.F.; Jones, K.W.; Sweet, R.M. (Brookhaven National Lab., Upton, NY (United States)); Carty, R.P. (State Univ. of New York, Brooklyn, NY (United States). Dept. of Biochemistry); Schlichting, I. (Brandeis Univ., Waltham, MA (United States). Rosenstiel Basic Medical Science Center); Stock, A. (Center for Advanced Biotechnology and Medicine, Piscataway, NJ (Un

    1992-01-01

    A transiently stable intermediate in trypsin catalysis, guanidinobenzyol-Ser-195 trypsin, can be trapped and then released by control of the pH in crystals of the enzyme. This effect has been investigated by static and dynamic white-beam Laue crystallography. Comparison of structures determined before and immediately after a pH jump reveals the nature of concerted changes that accompany activation of the enzyme. Careful analysis of the results of several structure determinations gives information about the reliability of Laue results in general. A study of multiple exposures taken under differing conditions of beam intensity, crystal quality, and temperature revealed information about ways to control damage of specimens by the x-ray beam.

  5. Laue diffraction as a tool in dynamic studies: Hydrolysis of a transiently stable intermediate in catalysis by trypsin

    Energy Technology Data Exchange (ETDEWEB)

    Singer, P.T.; Berman, L.E.; Cai, Z.; Mangel, W.F.; Jones, K.W.; Sweet, R.M. [Brookhaven National Lab., Upton, NY (United States); Carty, R.P. [State Univ. of New York, Brooklyn, NY (United States). Dept. of Biochemistry; Schlichting, I. [Brandeis Univ., Waltham, MA (United States). Rosenstiel Basic Medical Science Center; Stock, A. [Center for Advanced Biotechnology and Medicine, Piscataway, NJ (United States); Smalas, A. [Univ. of Tromso (Norway). Inst. of Mathematics and Physical Science

    1992-11-01

    A transiently stable intermediate in trypsin catalysis, guanidinobenzyol-Ser-195 trypsin, can be trapped and then released by control of the pH in crystals of the enzyme. This effect has been investigated by static and dynamic white-beam Laue crystallography. Comparison of structures determined before and immediately after a pH jump reveals the nature of concerted changes that accompany activation of the enzyme. Careful analysis of the results of several structure determinations gives information about the reliability of Laue results in general. A study of multiple exposures taken under differing conditions of beam intensity, crystal quality, and temperature revealed information about ways to control damage of specimens by the x-ray beam.

  6. Laue diffraction as a tool in dynamic studies: Hydrolysis of a transiently stable intermediate in catalysis by trypsin

    International Nuclear Information System (INIS)

    Singer, P.T.; Berman, L.E.; Cai, Z.; Mangel, W.F.; Jones, K.W.; Sweet, R.M.; Carty, R.P.; Smalas, A.

    1992-01-01

    A transiently stable intermediate in trypsin catalysis, guanidinobenzyol-Ser-195 trypsin, can be trapped and then released by control of the pH in crystals of the enzyme. This effect has been investigated by static and dynamic white-beam Laue crystallography. Comparison of structures determined before and immediately after a pH jump reveals the nature of concerted changes that accompany activation of the enzyme. Careful analysis of the results of several structure determinations gives information about the reliability of Laue results in general. A study of multiple exposures taken under differing conditions of beam intensity, crystal quality, and temperature revealed information about ways to control damage of specimens by the x-ray beam

  7. Less is More: Membrane Protein Digestion Beyond Urea–Trypsin Solution for Next-level Proteomics*

    Science.gov (United States)

    Zhang, Xi

    2015-01-01

    The goal of next-level bottom-up membrane proteomics is protein function investigation, via high-coverage high-throughput peptide-centric quantitation of expression, modifications and dynamic structures at systems scale. Yet efficient digestion of mammalian membrane proteins presents a daunting barrier, and prevalent day-long urea–trypsin in-solution digestion proved insufficient to reach this goal. Many efforts contributed incremental advances over past years, but involved protein denaturation that disconnected measurement from functional states. Beyond denaturation, the recent discovery of structure/proteomics omni-compatible detergent n-dodecyl-β-d-maltopyranoside, combined with pepsin and PNGase F columns, enabled breakthroughs in membrane protein digestion: a 2010 DDM-low-TCEP (DLT) method for H/D-exchange (HDX) using human G protein-coupled receptor, and a 2015 flow/detergent-facilitated protease and de-PTM digestions (FDD) for integrative deep sequencing and quantitation using full-length human ion channel complex. Distinguishing protein solubilization from denaturation, protease digestion reliability from theoretical specificity, and reduction from alkylation, these methods shifted day(s)-long paradigms into minutes, and afforded fully automatable (HDX)-protein-peptide-(tandem mass tag)-HPLC pipelines to instantly measure functional proteins at deep coverage, high peptide reproducibility, low artifacts and minimal leakage. Promoting—not destroying—structures and activities harnessed membrane proteins for the next-level streamlined functional proteomics. This review analyzes recent advances in membrane protein digestion methods and highlights critical discoveries for future proteomics. PMID:26081834

  8. Fish skin gelatin hydrolysates produced by visceral peptidase and bovine trypsin: Bioactivity and stability.

    Science.gov (United States)

    Ketnawa, Sunantha; Benjakul, Soottawat; Martínez-Alvarez, Oscar; Rawdkuen, Saroat

    2017-01-15

    The peptidase from the viscera of farmed giant catfish was used for producing gelatin hydrolysates (HG) and compared with those produced from commercial bovine trypsin (HB). The degree of hydrolysis (DH) observed suggests that proteolytic cleavage rapidly occurred within the first 120min of incubation, and there was higher DH in HG than in HB. HG demonstrated the highest ACE-inhibitory activity, DPPH, ABTS radical scavenging activity, and FRAP. HB showed the highest FRAP activity. The DPPH radical scavenging activity of HG was quite stable over the pH range of 1-11, but it increased slightly when the heating duration time reached 240min at 100°C. The ACE-inhibitory activity of HG showed the highest stability at a pH of 7, and it remained very stable at 100°C for over 15-240min. The visceral peptidase from farmed giant catfish could be an alternative protease for generating protein hydrolysates with desirable bioactivities. The resulting hydrolysates showed good stability, making them potential functional ingredients for food formulations. Copyright © 2016 Elsevier Ltd. All rights reserved.

  9. Spleen Stiffness Correlates with the Presence of Ascites but Not Esophageal Varices in Chronic Hepatitis C Patients

    Directory of Open Access Journals (Sweden)

    Kazuyo Mori

    2013-01-01

    Full Text Available Although spleen stiffness has recently been identified as potential surrogate marker for portal hypertension, the relationship between spleen stiffness and portal hypertension has not been fully elucidated. We attempted to determine the relationship between the liver or spleen stiffness and the presence of ascites or esophageal varices by acoustic radiation force impulse (ARFI imaging. A total of 33 chronic hepatitis C (CHC patients (median age 68; range 51–84 were enrolled. We evaluated the relationship between the liver or spleen stiffness and indicators of portal hypertension as well as clinical and biochemical parameters. Fourteen healthy volunteers were used for validating the accuracy of AFRI imaging. The liver and spleen stiffness increased significantly with progression of liver disease. A significant positive correlation was observed between the liver and spleen stiffness. However, spleen stiffness, but not liver stiffness, was significantly associated with the presence of ascites (, while there was no significant association between the spleen stiffness and spleen index/presence of esophageal varices in CHC patients. The area under the receiver operating characteristic curve based on the spleen stiffness was 0.80. In conclusion, spleen stiffness significantly correlates with the presence of ascites but not esophageal varices in CHC patients.

  10. A minimally invasive approach to spleen histopathology in dogs: A new method for follow-up studies of spleen changes in the course of Leishmania infantum infection.

    Science.gov (United States)

    Santos, Silvana Ornelas; Fontes, Jonathan L M; Laranjeira, Daniela F; Vassallo, José; Barrouin-Melo, Stella Maria; Dos-Santos, Washington L C

    2016-10-01

    Severe forms of zoonotic visceral leishmaniosis (ZVL) are associated with disruption of the spleen structure. However, the study of spleen histology requires splenectomy or necropsy. In this work, we present a minimally invasive cell-block technique for studying spleen tissue histology in dogs with ZVL. We examined 13 dogs with and seven dogs without Leishmania infantum infection. The dogs with Leishmania infection had a lower frequency of lymphoid follicles (2/13, Fisher's test, Pdogs (5/7 exhibiting lymphoid follicles and a plasma cell score of 1). The dogs with Leishmania infection also presented with granulomas (8/13) and infected macrophages (5/13). These differences in the histological presentations of spleen tissue from infected and uninfected dogs corresponded to changes observed in conventional histology. Hence, the cell-block technique described here may be used in the follow-up care and study of dogs with ZVL and other diseases in both clinical practice and research. Copyright © 2016 Elsevier Ltd. All rights reserved.

  11. Amino-acid sequence of two trypsin isoinhibitors, ITD I and ITD III from squash seeds (Cucurbita maxima).

    Science.gov (United States)

    Wilusz, T; Wieczorek, M; Polanowski, A; Denton, A; Cook, J; Laskowski, M

    1983-01-01

    The amino-acid sequences of two trypsin isoinhibitors, ITD I and ITD III, from squash seeds (Cucurbita maxima) were determined. Both isoinhibitors contain 29 amino-acid residues, including 6 half cystine residues. They differ only by one amino acid. Lysine in position 9 of ITD III is substituted by glutamic acid in ITD I. Arginine in position 5 is present at the reactive site of both isoinhibitors. The previously published sequence of ITD III has been shown to be incorrect.

  12. Prolonged hypoxic culture and trypsinization increase the pro-angiogenic potential of human adipose tissue-derived stem cells

    DEFF Research Database (Denmark)

    Rasmussen, Jeppe Grøndahl; Frøbert, Ole; Pilgaard, Linda

    2011-01-01

    Transplantation of mesenchymal stromal cells (MSC), including adipose tissue-derived stem cells (ASC), is a promising option in the treatment of vascular disease. Short-term hypoxic culture of MSC augments secretion of anti-apoptotic and angiogenic cytokines. We hypothesized that prolonged hypoxic...... (1% and 5% oxygen) culture and trypsinization would augment ASC expression of anti-apoptotic and angiogenic cytokines and increase the angiogenic potential of ASC-conditioned media....

  13. Comparison of various molecular forms of bovine trypsin: Correlation of infrared spectra with X-ray crystal structures

    Energy Technology Data Exchange (ETDEWEB)

    Prestrelski, S.J. (Mount Sinai School of Medicine of the City Univ. of New York (USA)); Byler, D.M. (U.S. Department of Agriculture, Philadelphia, PA (USA)); Liebman, M.N. (AMOCO Technology Corporation, Naperville, IL (USA))

    1991-01-01

    Fourier-transform infrared spectroscopy is a valuable method for the study of protein conformation in solution primarily because of the sensitivity to conformation of the amide I band (1700-1620 cm{sup {minus}1}) which arises from the backbone C{double bond}O stretching vibration. Combined with resolution-enhancement techniques such as derivative spectroscopy and self-deconvolution, plus the application of iterative curve-fitting techniques, this method provides a wealth of information concerning protein secondary structure. Further extraction of conformational information from the amide I band is dependent upon discerning the correlations between specific conformation types and component bands in the amide I region. In this paper the authors report spectra-structure correlations derived from conformational perturbations in bovine trypsin which arise from autolytic processing, zymogen activation, and active-site inhibition. IR spectra were collected for the single-chain ({beta}-trypsin) and once-cleaved, double-chain ({alpha}-trypsin) forms as well as at various times during the course of autolysis and also for zymogen, trypsinogen, and {beta}-trypsin inhibited with diisopropyl fluorophosphate. Spectral differences among the various molecular forms were interpreted in light of previous biochemical studies of autolysis and the known three-dimensional structures of the zymogen, the active enzyme, and the DIP-inhibited form. The spectroscopic results from these proteins in D{sub 2}O imply that certain loop structures may absorb in the region of 1655 cm{sup {minus}1}. They estimate that this approach to data analysis and interpretation is sensitive to changes of 0.01 unit or less in the relative integrated intensities of component bands in spectra whose peaks are well resolved.

  14. Extraction of gelatin from salmon (Salmo salar) fish skin using trypsin-aided process: optimization by Plackett-Burman and response surface methodological approaches.

    Science.gov (United States)

    Fan, HuiYin; Dumont, Marie-Josée; Simpson, Benjamin K

    2017-11-01

    Gelatin from salmon ( Salmo salar ) skin with high molecular weight protein chains ( α -chains) was extracted using trypsin-aided process. Response surface methodology was used to optimise the extraction parameters. Yield, hydroxyproline content and protein electrophoretic profile via sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis of gelatin were used as responses in the optimization study. The optimum conditions were determined as: trypsin concentration at 1.49 U/g; extraction temperature at 45 °C; and extraction time at 6 h 16 min. This response surface optimized model was significant and produced an experimental value (202.04 ± 8.64%) in good agreement with the predicted value (204.19%). Twofold higher yields of gelatin with high molecular weight protein chains were achieved in the optimized process with trypsin treatment when compared to the process without trypsin.

  15. Potentiometric determination of trypsin using a polymeric membrane polycation-sensitive electrode based on current-controlled reagent delivery.

    Science.gov (United States)

    Chen, Yan; Ding, Jiawang; Qin, Wei

    2012-12-01

    A potentiometric biosensor for the determination of trypsin is described based on current-controlled reagent delivery. A polymeric membrane protamine-sensitive electrode with dinonylnaphthalene sulfonate as cation exchanger is used for in situ generation of protamine. Diffusion of protamine across the polymeric membrane can be controlled precisely by applying an external current. The hydrolysis catalyzed with trypsin in sample solution decreases the concentration of free protamine released at the sample-membrane interface and facilitates the stripping of protamine out of the membrane surface via the ion-exchange process with sodium ions from the sample solution, thus decreasing the membrane potential, by which the protease can be sensed potentiometrically. The influences of anodic current amplitude, current pulse duration and protamine concentration in the inner filling solution on the membrane potential response have been studied. Under optimum conditions, the proposed protamine-sensitive electrode is useful for continuous and reversible detection of trypsin over the concentration range of 0.5-5UmL(-1) with a detection limit of 0.3UmL(-1). The proposed detection strategy provides a rapid and reagentless way for the detection of protease activities and offers great potential in the homogeneous immunoassays using proteases as labels. Copyright © 2012 Elsevier B.V. All rights reserved.

  16. Systematic and quantitative comparison of digest efficiency and specificity reveals the impact of trypsin quality on MS-based proteomics.

    Science.gov (United States)

    Burkhart, Julia Maria; Schumbrutzki, Cornelia; Wortelkamp, Stefanie; Sickmann, Albert; Zahedi, René Peiman

    2012-02-02

    Trypsin is the most frequently used proteolytic enzyme in mass spectrometry-based proteomics. Beside its good availability, it also offers some major advantages such as an optimal average peptide length of ~14 amino acids, and typically the presence of at least two defined positive charges at the N-terminus as well as the C-terminal Arg/Lys, rendering tryptic peptides well suited for CID-based LC-MS/MS. Here, we conducted a systematic study of different types of commercially available trypsin in order to qualitatively and quantitatively compare cleavage specificity, efficiency as well as reproducibility and the potential impact on quantitation and proteome coverage. We present a straightforward strategy applied to complex digests of human platelets, comprising (1) digest controls using a monolithic column HPLC-setup, (2) SCX enrichment of semitryptic/nonspecific peptides, (3) targeted MRM analysis of corresponding full cleavage/missed cleavage peptide pairs as well as (4) LC-MS analyses of complete digests with a three-step data interpretation. Thus, differences in digest performance can be readily assessed, rendering these procedures extremely beneficial to quality control not only the trypsin of choice, but also to effectively compare as well as optimize different digestion conditions and to evaluate the reproducibility of a dedicated digest protocol for all kinds of quantitative proteome studies. Copyright © 2011 Elsevier B.V. All rights reserved.

  17. Interspecific differences between D. pulex and D. magna in tolerance to cyanobacteria with protease inhibitors.

    Directory of Open Access Journals (Sweden)

    Christian J Kuster

    Full Text Available It is known that cyanobacteria negatively affect herbivores due to their production of toxins such as protease inhibitors. In the present study we investigated potential interspecific differences between two major herbivores, Daphnia magna and Daphnia pulex, in terms of their tolerance to cyanobacteria with protease inhibitors. Seven clones each of D. magna and of D. pulex were isolated from different habitats in Europe and North America. To test for interspecific differences in the daphnids' tolerance to cyanobacteria, their somatic and population growth rates were determined for each D. magna and D. pulex clone after exposure to varying concentrations of two Microcystis aeruginosa strains. The M. aeruginosa strains NIVA and PCC(- contained either chymotrypsin or trypsin inhibitors, but no microcystins. Mean somatic and population growth rates on a diet with 20% NIVA were significantly more reduced in D. pulex than in D. magna. On a diet with 10% PCC(-, the population growth of D. pulex was significantly more reduced than that of D. magna. This indicates that D. magna is more tolerant to cyanobacteria with protease inhibitors than D. pulex. The reduction of growth rates was possibly caused by an interference of cyanobacterial inhibitors with proteases in the gut of Daphnia, as many other conceivable factors, which might have been able to explain the reduced growth, could be excluded as causal factors. Protease assays revealed that the sensitivities of chymotrypsins and trypsins to cyanobacterial protease inhibitors did not differ between D. magna and D. pulex. However, D. magna exhibited a 2.3-fold higher specific chymotrypsin activity than D. pulex, which explains the observed higher tolerance to cyanobacterial protease inhibitors of D. magna. The present study suggests that D. magna may control the development of cyanobacterial blooms more efficiently than D. pulex due to differences in their tolerance to cyanobacteria with protease

  18. Studies on terrein as a new class of proteasome inhibitors

    International Nuclear Information System (INIS)

    Demasi, M.; Felicio, A.L.; Lima, C.; Pacheco, A.O.; Leite, H.G.; Andrade, L.H.

    2010-01-01

    The proteasome is an intracellular multicatalytic protease involved in the cell cycle regulation, signaling response, antigen presentation and apoptosis. Since proteasome inhibitors promote cell death by apoptosis, they have been proposed as new anti-tumoral drugs. Terrein, a secondary metabolite secreted by the fungus Aspergillus terreus, was firstly described in 1935. In the present work we report that terrein isolated through the screening for inhibitors of the 20S proteasome showed inhibitory effect upon both chymotrypsin- and trypsin-like activities of the multicatalytic core particle, the 20S proteasome. Despite of the high inhibitory concentration determined in vitro, that verified by incubating cells (fibroblasts and a pulmonary tumor cell line) in the presence of terrein was 4-fold lower indicating the proteasome as a selective intracellular target. Moreover, terrein promoted apoptotic cell death on both fibroblasts and pulmonary tumor cell line tested. Although terrein concentrations (mM range) necessary to elicit apoptosis in the cellular models herein tried were high when compared to those (muM and nM range) of other inhibitors recently described, its chemical structure is not correlated to any other inhibitor reported thus far. Therefore, the present results point out for the possibility of exploring terrein as a new molecular fragment for the development of synthetic proteasome inhibitors. (author)

  19. Isolation, cloning and structural characterisation of boophilin, a multifunctional Kunitz-type proteinase inhibitor from the cattle tick.

    Directory of Open Access Journals (Sweden)

    Sandra Macedo-Ribeiro

    Full Text Available Inhibitors of coagulation factors from blood-feeding animals display a wide variety of structural motifs and inhibition mechanisms. We have isolated a novel inhibitor from the cattle tick Boophilus microplus, one of the most widespread parasites of farm animals. The inhibitor, which we have termed boophilin, has been cloned and overexpressed in Escherichia coli. Mature boophilin is composed of two canonical Kunitz-type domains, and inhibits not only the major procoagulant enzyme, thrombin, but in addition, and by contrast to all other previously characterised natural thrombin inhibitors, significantly interferes with the proteolytic activity of other serine proteinases such as trypsin and plasmin. The crystal structure of the bovine alpha-thrombin.boophilin complex, refined at 2.35 A resolution reveals a non-canonical binding mode to the proteinase. The N-terminal region of the mature inhibitor, Q16-R17-N18, binds in a parallel manner across the active site of the proteinase, with the guanidinium group of R17 anchored in the S(1 pocket, while the C-terminal Kunitz domain is negatively charged and docks into the basic exosite I of thrombin. This binding mode resembles the previously characterised thrombin inhibitor, ornithodorin which, unlike boophilin, is composed of two distorted Kunitz modules. Unexpectedly, both boophilin domains adopt markedly different orientations when compared to those of ornithodorin, in its complex with thrombin. The N-terminal boophilin domain rotates 9 degrees and is displaced by 6 A, while the C-terminal domain rotates almost 6 degrees accompanied by a 3 A displacement. The reactive-site loop of the N-terminal Kunitz domain of boophilin with its P(1 residue, K31, is fully solvent exposed and could thus bind a second trypsin-like proteinase without sterical restraints. This finding explains the formation of a ternary thrombin.boophilin.trypsin complex, and suggests a mechanism for prothrombinase inhibition in vivo.

  20. Engineering of Yersinia Phytases to Improve Pepsin and Trypsin Resistance and Thermostability and Application Potential in the Food and Feed Industry.

    Science.gov (United States)

    Niu, Canfang; Yang, Peilong; Luo, Huiying; Huang, Huoqing; Wang, Yaru; Yao, Bin

    2017-08-30

    Susceptibility to proteases usually limits the application of phytase. We sought to improve the pepsin and trypsin resistance of YeAPPA from Yersinia enterocolitica and YkAPPA from Y. kristensenii by optimizing amino acid polarity and charge. The predicted pepsin/trypsin cleavage sites F89/K226 in pepsin/trypsin-sensitive YeAPPA and the corresponding sites (F89/E226) in pepsin-sensitive but trypsin-resistant YkAPPA were substituted with S and H, respectively. Six variants were produced in Pichia pastoris for catalytic and biochemical characterization. F89S, E226H, and F89S/E226H elevated pepsin resistance and thermostability and K226H and F89S/K226H improved pepsin and trypsin resistance and stability at 60 °C and low pH. All the variants increased the ability of the proteins to hydrolyze phytate in corn meal by 2.6-14.9-fold in the presence of pepsin at 37 °C and low pH. This study developed a genetic manipulation strategy specific for pepsin/trypsin-sensitive phytases that can improve enzyme tolerance against proteases and heat and benefit the food and feed industry in a cost-effective way.

  1. Ginsenoside Rg1 improves bone marrow haematopoietic activity via extramedullary haematopoiesis of the spleen.

    Science.gov (United States)

    Liu, Hua-Hsing; Chen, Fei-Peng; Liu, Rong-Kai; Lin, Chun-Lin; Chang, Ko-Tung

    2015-11-01

    Cyclophosphamide (CY) is a chemotherapeutic agent used for cancer and immunological diseases. It induces cytotoxicity of bone marrow and causes myelosuppression and extramedullary haematopoiesis (EMH) in treated patients. EMH is characterized with the emergence of multipotent haematopoietic progenitors most likely in the spleen and liver. Previous studies indicated that a Chinese medicine, ginsenoside Rg1, confers a significant effect to elevate the number of lineage (Lin(-) ) Sca-1(+) c-Kit(+) haematopoietic stem and progenitor cells (HSPCs) and restore the function of bone marrow in CY-treated myelosuppressed mice. However, whether the amelioration of bone marrow by Rg1 accompanies an alleviation of EMH in the spleen was still unknown. In our study, the cellularity and weight of the spleen were significantly reduced after Rg1 treatment in CY-treated mice. Moreover, the number of c-Kit(+) HSPCs was significantly decreased but not as a result of apoptosis, indicating that Rg1 alleviated EMH of the spleen induced by CY. Unexpectedly, the proliferation activity of c-Kit(+) HSPCs was only up-regulated in the spleen, but not in the bone marrow, after Rg1 treatment in CY-treated mice. We also found that a fraction of c-Kit(+) /CD45(+) HSPCs was simultaneously increased in the circulation after Rg1 treatment. Interestingly, the effects of Rg1 on the elevation of HSPCs in bone marrow and in the peripheral blood were suppressed in CY-treated splenectomized mice. These results demonstrated that Rg1 improves myelosuppression induced by CY through its action on the proliferation of HSPCs in EMH of the spleen and migration of HSPCs from the spleen to the bone marrow. © 2015 The Authors. Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and Molecular Medicine.

  2. Obesity-related chronic kidney disease is associated with spleen-derived IL-10.

    Science.gov (United States)

    Gotoh, Koro; Inoue, Megumi; Masaki, Takayuki; Chiba, Seiichi; Shiraishi, Kentaro; Shimasaki, Takanobu; Matsuoka, Kazue; Ando, Hisae; Fujiwara, Kansuke; Fukunaga, Naoya; Aoki, Kohei; Nawata, Tomoko; Katsuragi, Isao; Kakuma, Tetsuya; Seike, Masataka; Yoshimatsu, Hironobu

    2013-05-01

    Obesity is associated with systemic low-grade inflammation and is a risk factor for chronic kidney disease (CKD), but the molecular mechanism remains uncertain. We noticed spleen-derived interleukin (IL)-10 because it is observed that obesity reduces several cytokines in the spleen. We examined whether spleen-derived IL-10 regulates CKD caused by a high-fat diet (HF)-induced obesity as follows: (i) male mice were fed with HF (60% fat) during 8 weeks and IL-10 induction from the spleen was examined, (ii) glomerular hypertrophy, fibrosis, inflammatory responses in the kidney and systolic blood pressure (SBP) were evaluated in splenectomy (SPX)-treated mice fed HF, (iii) exogenous IL-10 was systemically administered to HF-induced obese mice and the alteration of obesity-induced pathogenesis caused by IL-10 treatment was assessed. (iv) IL-10 knockout (IL-10KO) mice were treated with SPX and glomerular hypertrophy, fibrosis and the inflammatory condition in the kidney and SBP were also investigated. Obesity decreased serum levels of only IL-10, an anti-inflammatory cytokine even though pro- and anti-inflammatory cytokine expression in the spleen was significantly lower in the obese group. SPX aggravated HF-induced inflammatory responses in the kidney and hypertension. These HF-induced alterations were inhibited by systemically administered IL-10. Moreover, SPX had little effect on inflammatory responses and SBP in the kidney of IL-10KO mice. We suggest that obesity reduces IL-10 induction from the spleen, and spleen-derived IL-10 may protect against the development of CKD induced by obesity.

  3. Estimation of the volume and percent uptake of the liver and spleen by SPECT

    International Nuclear Information System (INIS)

    Yamagata, Atushi

    1988-01-01

    The volume and percent uptake of the liver and spleen were estimated with single photon emission computed tomography (SPECT) using 99m Tc-phytate. Clinical usefulness of these parameters was evaluated by comparison with other liver function tests in 87 patients including 25 normal controls, 24 liver cirrhosis and 16 other chronic liver diseases. SPECT images were obtained by Maxi Camera 400T. Cut-off level for reconstruction of images and relationship between counts and activity (mCi) were obtained from phantom studies. Volumes estimated using SPECT and computed tomography were compared in 16 patients. Results obtained were as follows. 1) Optimal cut-off level for measurement of volumes for the liver was 37 % and for the spleen was 42 %. 2) Correlation between organ volumes estimated with CT and SPECT was good (r = 0.92 for the liver and r = 0.96 for the spleen), although volumes measured with SPECT were larger than those with CT. 3) Significant differences of percent uptake were observed between normal controls and liver cirrhosis. 4) Better correlation between spleen volumes and uptake was recognized in cases without liver cirrhosis than in cases with liver cirrhosis. The spleen uptake in liver cirrhosis was higher than those in others in comparison with the volume. 5) The liver/spleen ratio of 99m Tc-phytae uptake could most clearly differentiate liver cirrhosis from others. 6) Negative correlation was observed between liver volume or uptake and ICG (R 15 ). Estimation of volume and uptake of the liver and spleen could be a useful procedure to assess liver function, probably related with effective hepatic blood flow in liver cirrhosis. (author)

  4. The effect of the cytoskeletal inhibitors on the splenic lymphocyte traffic and homing in rats

    International Nuclear Information System (INIS)

    Yang Huibin

    1989-01-01

    The rat splenic lymphocyte traffic and homing in vivo and the effect of cytoskeletal inhibitors on this process were investigated using the technique of γ-counting of 51 Cr-labelled lymphocytes. The results suggests that:(1) After 2 of intravenous injection, the 51 Cr-labelled lymphocytes from donor rat spleen mainly home to recipient rat spleen, liver, lungs, mesenteric lymph modes (MLN) and gut-associated lymphoid tissues. (2) A significant inhibiting effect on the ability of preferential homing of splenic lymphocytes treated with sodium azide, cytochalasin B or colchicine shows that microtubles and microfilaments play an important role in the lymphocyte traffic and homing

  5. Hydroxyl and water molecule orientations in trypsin: Comparison to molecular dynamics structures

    Energy Technology Data Exchange (ETDEWEB)

    McDowell, R.S.; Kossiakoff, A.A. [Genentech, Inc., South San Francisco, CA (United States)

    1994-12-31

    A comparison is presented of experimentally observed hydroxyl and water hydrogens in trypsin determined from neutron density maps with the results of a 140ps molecular dynamics (MD) simulation. Experimental determination of hydrogen and deuterium atom positions in molecules as large as proteins is a unique capability of neutron diffraction. The comparison addresses the degree to which a standard force-field approach can adequately describe the local electrostatic and van der Waals forces that determine the orientations of these hydrogens. Neutron densities, derived from 2.1{Angstrom} D{sub 2}O-H{sub 2}O difference Fourier maps, provide a database of 27 well-ordered hydroxyl hydrogens. Most of the simulated hydroxyl orientations are within a standard deviation of the experimentally-observed positions, including several examples in which both the simulation and the neutron density indicate that a hydroxyl group is shifted from a {open_quote}standard{close_quote} rotamer. For the most highly ordered water molecules, the hydrogen distributions calculated from the trajectory were in good agreement with neutron density; simulated water molecules that displayed multiple hydrogen bonding networks had correspondingly broadened neutron density profiles. This comparison was facilitated by development of a method to construct a pseudo 2{Angstrom} density map based on the hydrogen atom distributions from the simulation. The degree of disorder of internal water molecules is shown to result primarily from the electrostatic environment surrounding that water molecule as opposed to the cavity size available to the molecule. A method is presented for comparing the discrete observations sampled in a dynamics trajectory with the time- averaged data obtained from X-ray or neutron diffraction studies. This method is particularly useful for statically-disordered water molecules, in which the average location assigned from a trajectory may represent a site of relatively low occupancy.

  6. Less is More: Membrane Protein Digestion Beyond Urea-Trypsin Solution for Next-level Proteomics.

    Science.gov (United States)

    Zhang, Xi

    2015-09-01

    The goal of next-level bottom-up membrane proteomics is protein function investigation, via high-coverage high-throughput peptide-centric quantitation of expression, modifications and dynamic structures at systems scale. Yet efficient digestion of mammalian membrane proteins presents a daunting barrier, and prevalent day-long urea-trypsin in-solution digestion proved insufficient to reach this goal. Many efforts contributed incremental advances over past years, but involved protein denaturation that disconnected measurement from functional states. Beyond denaturation, the recent discovery of structure/proteomics omni-compatible detergent n-dodecyl-β-d-maltopyranoside, combined with pepsin and PNGase F columns, enabled breakthroughs in membrane protein digestion: a 2010 DDM-low-TCEP (DLT) method for H/D-exchange (HDX) using human G protein-coupled receptor, and a 2015 flow/detergent-facilitated protease and de-PTM digestions (FDD) for integrative deep sequencing and quantitation using full-length human ion channel complex. Distinguishing protein solubilization from denaturation, protease digestion reliability from theoretical specificity, and reduction from alkylation, these methods shifted day(s)-long paradigms into minutes, and afforded fully automatable (HDX)-protein-peptide-(tandem mass tag)-HPLC pipelines to instantly measure functional proteins at deep coverage, high peptide reproducibility, low artifacts and minimal leakage. Promoting-not destroying-structures and activities harnessed membrane proteins for the next-level streamlined functional proteomics. This review analyzes recent advances in membrane protein digestion methods and highlights critical discoveries for future proteomics. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

  7. Acute cadmium administration to rats exerts both immunosuppressive and proinflammatory effects in spleen

    International Nuclear Information System (INIS)

    Demenesku, Jelena; Mirkov, Ivana; Ninkov, Marina; Popov Aleksandrov, Aleksandra; Zolotarevski, Lidija; Kataranovski, Dragan; Kataranovski, Milena

    2014-01-01

    Highlights: • The effect of cadmium on splenic T and innate immune cells in rats is explored. • Differential effects of 1 mg/kg on T cell and innate immune activities were shown. • Lower Cd dose (0.5 mg/kg) cause less pronounced immunosuppressive effects. • Proinflammatory effects on innate immune activities were seen at that dose. • Presented data depict complexity of immunomodulatory potential of this metal. - Abstract: Conflicting data (both suppression and augmentation as well as lack of the effect) exist in respect to cadmium (Cd) and splenic T cell-based immune cell activity. Spleen is also the site of innate immune responses but impact of Cd on this type of immunity has been less explored. In the present study the effects of acute Cd administration on basic aspects of both T cell-based and innate immune spleen cell activity were examined in rats. Intraperitoneal injection of 1 mg of Cd/kg resulted in decrease in concanavalin A (ConA) induced proliferation which seems to be more related to altered spleen cells responsiveness to IL-2 than to apoptosis. Differential effects on proinflammatory T cell derived cytokines were observed (decreases of IFN-γ gene expression and ConA-stimulated production, but increases in IL-17 mRNA levels with no effect on concentrations of protein product). Reduction of IFN-γ production seemed not to rely on IL-4 and IL-10, but at least partly on nitric oxide (NO). Increased activity relevant for innate immunity (granulocyte and CD11b + cell accumulation in the spleen, inducible nitric oxide synthase/iNOS expression and NO production by spleen cells) was observed, but there was a decrease in respiratory burst (dihydrorhodamine/DHR oxidation and nitroblue tetrazolium/NBT reduction). Increases of TNF-α and IL-1β gene expression and IL-1β protein product were noted as well. Administration of 0.5 mg Cd/kg resulted in less pronounced (ConA-induced proliferation) or lack of the effect (IFN-γ production) on spleen T cell

  8. Trauma center variation in splenic artery embolization and spleen salvage: a multicenter analysis.

    Science.gov (United States)

    Banerjee, Aman; Duane, Therese M; Wilson, Sean P; Haney, Starre; O'Neill, Patrick J; Evans, Heather L; Como, John J; Claridge, Jeffrey A

    2013-07-01

    This study aimed to evaluate if variation in management of blunt splenic injury (BSI) among Level I trauma centers is associated with different outcomes related to the use of splenic artery embolization (SAE). All adult patients admitted for BSI from 2008 to 2010 at 4 Level I trauma centers were reviewed. Use of SAE was determined, and outcomes of spleen salvage and nonoperative management (NOM) failure were evaluated. A priori, a 10% SAE rate was used to group centers into high- or low-use groups. There were 1,275 BSI patients. There were intercenter differences in age, injury severity, and grade of spleen injury (Spleen Injury Scale [SIS]). Mortality was similar by center; however, BSI treatment varied significantly by center. Overall, SAE use was highest at center A compared with B, C, and D (19%, 11%, 1%, and 4%, respectively; p trauma centers. Centers with higher rates of SAE use have higher spleen salvage and less NOM failure. SAE was shown to be an independent predictor of spleen salvage. Therapeutic study, level IV.

  9. Atheroprotector role of the spleen based on the teaching of Avicenna (Ibn Sina).

    Science.gov (United States)

    Emtiazy, Majid; Choopani, Rasool; Khodadoost, Mahmood; Tansaz, Mojgan; Nazem, Esmaiel

    2013-07-15

    Many studies have proven atherosclerosis is an inflammatory immune disease. The spleen plays an important immune role in the human body. Splenectomy is often used in several clinical disorders; but recent studies have shown that splenectomy may be effective in the development of atheroma lesions. Ibn Sina or Avicenna was known as one of the greatest philosopher and physician in Islam and in Medicine. He is remembered for his masterpiece, The "Al-Qanun fi al-Tibb" or "Qanun of medicine". According to the "Al-Qanun fi al-Tibb", spleen as storage organ plays an important role in absorption and secretion of the black bile in the human body. Therefore any disruption in the function of the spleen can lead to various diseases such as atherosclerosis. Based on his description, it is clear that Ibn Sina first described the role of spleen in prevention of atherosclerosis. In this review, we discuss the Avicenna (Ibn Sina) aspect of atheroprotector role of the spleen. Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.

  10. Trivial trauma and delayed rupture of a normal spleen: a case report

    Directory of Open Access Journals (Sweden)

    Sowers Nicholas

    2011-12-01

    Full Text Available Abstract Introduction Although a majority of splenic ruptures present acutely with a known mechanism of injury, a minority of patients present days to weeks following trauma with a delayed rupture. Also uncommon is the atraumatic rupture, the vast majority of which occur in patients with underlying splenic pathology. A handful of cases of apparently spontaneous rupture of a normal spleen are reported; however, there is debate about whether these actually represent delayed ruptures following a history of trauma that is not elicited. Although a few cases of delayed rupture of the spleen following trivial trauma have been reported, the majority of these present evidence of an underlying disease process. We found only two such cases that documented a normal spleen and three cases where underlying splenic pathology was not reported. We review the literature and discuss the phenomenon of delayed rupture of the normal spleen following trivial trauma. Case presentation A 27-year-old Caucasian man with no underlying splenic pathology presented with splenic rupture one week after playfully wrestling with his partner. The patient did not present at the time of the injury and only recalled it upon repeated questioning after computed tomography diagnosis. Conclusions This case lends support to the theory that the normal spleen can rupture some time after trivial trauma, which seems like a more plausible explanation than rupture without cause. However, given the dearth of similar reports in the literature, the possibility remains that the association we have observed is not causational.

  11. Comparative histology in the liver and spleen of three species of freshwater teleost

    Directory of Open Access Journals (Sweden)

    Camila F. Sales

    2017-03-01

    Full Text Available ABSTRACT We describe and compare the histology of liver and spleen ofGeophagus brasiliensis (Perciformes, Hypostomus francisci (Siluriformes and Hoplias aff. malabaricus (Characiformes, tropical freshwater fishes. InG. brasiliensisandH. aff. malabaricusthe hepatocytes were arranged in tubular form whereas in H. franciscithey cord-like. In all species, hepatocytes presented glycogen, but in G. brasiliensis and H. aff. malabaricus they showed strong stained for hemossiderin in the cytoplasm. InG. brasiliensis and H. aff. malabaricus, melanomacrophage centres (MMCs were associated to hepatic structures and only in G. brasiliensis was observed intrahepatic exocrine pancreas. The spleen, in all species, was characterized by red and white pulp without boundary between the two regions, but only in H. francisci was recorded nodular organization in splenic parenchyma. The G. brasiliensisandH. aff. malabaricuspresented in the white pulp MMCs linked mainly to ellipsoids. Besides, we observed large MMCs in the spleen in relation to liver of G. brasiliensis and H. aff. malabaricus. In liver, highest values of reticular fibers and collagen were observed inG. brasiliensis. In spleen, highest values of reticular fibers and collagen were recorded inH. aff. malabaricusandH. francisci, respectively. Histological differences confirm the hypothesis that the phylogenetic distance is reflected in liver and spleen.

  12. Pancreatic islet allograft in spleen with immunosuppression with cyclosporine. Experimental model in dogs.

    Science.gov (United States)

    Waisberg, Jaques; Neff, Charles Benjamin; Waisberg, Daniel Reis; Germini, Demetrius; Gonçalves, José Eduardo; Zanotto, Arnaldo; Speranzini, Manlio Basilio

    2011-01-01

    To study the functional behavior of the allograft with immunosuppression of pancreatic islets in the spleen. Five groups of 10 Mongrel dogs were used: Group A (control) underwent biochemical tests; Group B underwent total pancreatectomy; Group C underwent total pancreatectomy and pancreatic islet autotransplant in the spleen; Group D underwent pancreatic islet allograft in the spleen without immunosuppressive therapy; Group E underwent pancreatic islet allograft in the spleen and immunosuppression with cyclosporine. All of the animals with grafts received pancreatic islets prepared by the mechanical-enzymatic method - stationary collagenase digestion and purification with dextran discontinuous density gradient, implanted in the spleen. The animals with autotransplant and those with allografts with immunosuppression that became normoglycemic showed altered results of intravenous tolerance glucose (p < 0.001) and peripheral and splenic vein plasmatic insulin levels were significantly lower (p < 0.001) in animals that had allografts with immunosuppression than in those with just autotransplants. In the animals with immunosuppression with cyclosporine subjected to allograft of pancreatic islets prepared with the mechanical-enzymatic preparation method (stationary collagenase digestion and purification with dextran discontinuous density gradient), the production of insulin is decreased and the response to intravenous glucose is altered.

  13. The Spleen as an Optimal Site for Islet Transplantation and a Source of Mesenchymal Stem Cells

    Directory of Open Access Journals (Sweden)

    Naoaki Sakata

    2018-05-01

    Full Text Available This review demonstrates the unique potential of the spleen as an optimal site for islet transplantation and as a source of mesenchymal stem cells. Islet transplantation is a cellular replacement therapy used to treat severe diabetes mellitus; however, its clinical outcome is currently unsatisfactory. Selection of the most appropriate transplantation site is a major factor affecting the clinical success of this therapy. The spleen has long been studied as a candidate site for islet transplantation. Its advantages include physiological insulin drainage and regulation of immunity, and it has recently also been shown to contribute to the regeneration of transplanted islets. However, the efficacy of transplantation in the spleen is lower than that of intraportal transplantation, which is the current representative method of clinical islet transplantation. Safer and more effective methods of islet transplantation need to be established to allow the spleen to be used for clinical transplantation. The spleen is also of interest as a mesenchymal stem cell reservoir. Splenic mesenchymal stem cells contribute to the repair of damaged tissue, and their infusion may thus be a promising therapy for autoimmune diseases, including type 1 diabetes mellitus and Sjogren’s syndrome.

  14. Ames hypopituitary dwarf mice demonstrate imbalanced myelopoiesis between bone marrow and spleen.

    Science.gov (United States)

    Capitano, Maegan L; Chitteti, Brahmananda R; Cooper, Scott; Srour, Edward F; Bartke, Andrzej; Broxmeyer, Hal E

    2015-06-01

    Ames hypopituitary dwarf mice are deficient in growth hormone, thyroid-stimulating hormone, and prolactin. The phenotype of these mice demonstrates irregularities in the immune system with skewing of the normal cytokine milieu towards a more anti-inflammatory environment. However, the hematopoietic stem and progenitor cell composition of the bone marrow (BM) and spleen in Ames dwarf mice has not been well characterized. We found that there was a significant decrease in overall cell count when comparing the BM and spleen of 4-5 month old dwarf mice to their littermate controls. Upon adjusting counts to differences in body weight between the dwarf and control mice, the number of granulocyte-macrophage progenitors, confirmed by immunophenotyping and colony-formation assay was increased in the BM. In contrast, the numbers of all myeloid progenitor populations in the spleen were greatly reduced, as confirmed by colony-formation assays. This suggests that there is a shift of myelopoiesis from the spleen to the BM of Ames dwarf mice; however, this shift does not appear to involve erythropoiesis. The reasons for this unusual shift in spleen to marrow hematopoiesis in Ames dwarf mice are yet to be determined but may relate to the decreased hormone levels in these mice. Copyright © 2015. Published by Elsevier Inc.

  15. Transcription factor TLX1 controls retinoic acid signaling to ensure spleen development

    Science.gov (United States)

    Lenti, Elisa; Farinello, Diego; Penkov, Dmitry; Castagnaro, Laura; Lavorgna, Giovanni; Wuputra, Kenly; Tjaden, Naomi E. Butler; Bernassola, Francesca; Caridi, Nicoletta; Wagner, Michael; Kozinc, Katja; Niederreither, Karen; Blasi, Francesco; Pasini, Diego; Trainor, Paul A.

    2016-01-01

    The molecular mechanisms that underlie spleen development and congenital asplenia, a condition linked to increased risk of overwhelming infections, remain largely unknown. The transcription factor TLX1 controls cell fate specification and organ expansion during spleen development, and Tlx1 deletion causes asplenia in mice. Deregulation of TLX1 expression has recently been proposed in the pathogenesis of congenital asplenia in patients carrying mutations of the gene-encoding transcription factor SF-1. Herein, we have shown that TLX1-dependent regulation of retinoic acid (RA) metabolism is critical for spleen organogenesis. In a murine model, loss of Tlx1 during formation of the splenic anlage increased RA signaling by regulating several genes involved in RA metabolism. Uncontrolled RA activity resulted in premature differentiation of mesenchymal cells and reduced vasculogenesis of the splenic primordium. Pharmacological inhibition of RA signaling in Tlx1-deficient animals partially rescued the spleen defect. Finally, spleen growth was impaired in mice lacking either cytochrome P450 26B1 (Cyp26b1), which results in excess RA, or retinol dehydrogenase 10 (Rdh10), which results in RA deficiency. Together, these findings establish TLX1 as a critical regulator of RA metabolism and provide mechanistic insights into the molecular determinants of human congenital asplenia. PMID:27214556

  16. Spleen and thymic sonography as estimator of the immune response in children with repeated infections

    International Nuclear Information System (INIS)

    Rabaza, Jesus; Fundora, Hermes; Rodriguez, Alexis; Hernandez, Maria de los Angeles

    2010-01-01

    The spleen is a secondary lymphoid organ. The spleen has many functions: IgM class antibody production and phagocytosis of encapsulated bacteria. We propose to measure the dimensions of the spleen in children with repeated infections, in order to describe them as a diagnosis marker and to explore the probable correlation between the thymic characteristics and the dimensions of the spleen. We study 81 patients that attended the consultation of Immunology from Aballi hospital with repeated infections and they were from 8 months to 9 years old. Children with nutritional value up to the third percentile and those taking steroids 45 days before the study were excluded. We measured the thymic and splenic areas by mediastinal and abdominal echography. The length, the splenic index and the weight are less in patients with repeated infections. These data were significant in 12 to 23 month of age group. The length and the splenic index were less in patients with depletion of thymic area. The sonographic evaluation of the spleen is very important in patients from 1 to 2 years old and in patients with thymic depletion, because they are on the age of vaccine administration according to the Cuban schedule and having affected a main organ for immune response might be a reason for them to show a deficient response and need special schedules

  17. The Spleen as an Optimal Site for Islet Transplantation and a Source of Mesenchymal Stem Cells.

    Science.gov (United States)

    Sakata, Naoaki; Yoshimatsu, Gumpei; Kodama, Shohta

    2018-05-07

    This review demonstrates the unique potential of the spleen as an optimal site for islet transplantation and as a source of mesenchymal stem cells. Islet transplantation is a cellular replacement therapy used to treat severe diabetes mellitus; however, its clinical outcome is currently unsatisfactory. Selection of the most appropriate transplantation site is a major factor affecting the clinical success of this therapy. The spleen has long been studied as a candidate site for islet transplantation. Its advantages include physiological insulin drainage and regulation of immunity, and it has recently also been shown to contribute to the regeneration of transplanted islets. However, the efficacy of transplantation in the spleen is lower than that of intraportal transplantation, which is the current representative method of clinical islet transplantation. Safer and more effective methods of islet transplantation need to be established to allow the spleen to be used for clinical transplantation. The spleen is also of interest as a mesenchymal stem cell reservoir. Splenic mesenchymal stem cells contribute to the repair of damaged tissue, and their infusion may thus be a promising therapy for autoimmune diseases, including type 1 diabetes mellitus and Sjogren’s syndrome.

  18. The Spleen Is an Ideal Site for Inducing Transplanted Islet Graft Expansion in Mice.

    Directory of Open Access Journals (Sweden)

    Takeshi Itoh

    Full Text Available Alternative islet transplantation sites have the potential to reduce the marginal number of islets required to ameliorate hyperglycemia in recipients with diabetes. Previously, we reported that T cell leukemia homeobox 1 (Tlx1+ stem cells in the spleen effectively regenerated into insulin-producing cells in the pancreas of non-obese diabetic mice with end-stage disease. Thus, we investigated the spleen as a potential alternative islet transplantation site. Streptozotocin-induced diabetic C57BL/6 mice received syngeneic islets into the portal vein (PV, beneath the kidney capsule (KC, or into the spleen (SP. The marginal number of islets by PV, KC, or SP was 200, 100, and 50, respectively. Some plasma inflammatory cytokine levels in the SP group were significantly lower than those of the PV group after receiving a marginal number of islets, indicating reduced inflammation in the SP group. Insulin contents were increased 280 days after islet transplantation compared with those immediately following transplantation (p<0.05. Additionally, Tlx1-related genes, including Rrm2b and Pla2g2d, were up-regulated, which indicates that islet grafts expanded in the spleen. The spleen is an ideal candidate for an alternative islet transplantation site because of the resulting reduced inflammation and expansion of the islet graft.

  19. Factors influencing liver and spleen volume changes after donor hepatectomy for living donor liver transplantation

    International Nuclear Information System (INIS)

    Bae, Ji Hee; Ryeom, Hunku; Song, Jung Hup

    2013-01-01

    To define the changes in liver and spleen volumes in the early postoperative period after partial liver donation for living-donor liver transplantation (LDLT) and to determine factors that influence liver and spleen volume changes. 27 donors who underwent partial hepatectomy for LDLT were included in this study. The rates of liver and spleen volume change, measured with CT volumetry, were correlated with several factors. The analyzed factors included the indocyanine green (ICG) retention rate at 15 minutes after ICG administration, preoperative platelet count, preoperative liver and splenic volumes, resected liver volume, resected-to-whole liver volume ratio (LV R /LV W ), resected liver volume to the sum of whole liver and spleen volume ratio [LV R /(LV W + SV 0 )], and pre and post hepatectomy portal venous pressures. In all hepatectomy donors, the volumes of the remnant liver and spleen were increased (increased rates, 59.5 ± 50.5%, 47.9 ± 22.6%). The increment rate of the remnant liver volume revealed a positive correlation with LV R /LV W (r = 0.759, p R /LV W influences the increment rate of the remnant liver volume.

  20. The effect of lysate of spleen cells after low dose radiation (LDR) on NK activity

    International Nuclear Information System (INIS)

    Lu Duicai; Su Liaoyuan

    2003-01-01

    To find effect of lysate of spleen cells after LDR on NK activity of CD 57 cells or non-CD 57 cells, lysate of spleen cells after LDR were extracted. McAb (anti CD 57 cells) was used to separate CD 57 cells from human peripheral blood by Panning direct method. The CD 57 cells and non-CD 57 cells were used as effective cells. K 562 cells labelled by 3 H-TdR were used as target cells. The ratio of effect cells to target cells was 10:1. NK activity of CD 57 cells or non-CD 5 -7 cells with the lysate of spleen cells after LDR was reflected by the efficiency of anti tumor cells. The 3 H-TdR incorporation in K 562 cells cultured with non-CD 57 cells was significantly lower than that with CD 57 cells. After use of the lysate of spleen cells after LDR, NK activities of CD 57 cells and non-CD 57 cells were 1.24 and 1.58 respectively. They were both increased obviously compared with control groups. The effect of anti K 562 cells of non-CD 57 cells is even greater than that of CD 57 cells. The lysate of spleen cells after LDR can enhance the effect of both non-CD 57 cells and CD 57 cells

  1. Purification, characterization and cloning of an aspartic proteinase inhibitor from squash phloem exudate.

    Science.gov (United States)

    Christeller, J T; Farley, P C; Ramsay, R J; Sullivan, P A; Laing, W A

    1998-05-15

    Phloem exudate from squash fruit contains heat-inactivated material which inhibits pepsin activity. This inhibitory activity was purified by mild acid treatment, chromatography on trypsin-agarose, Sephadex G-75 and reverse-phase HPLC, resulting in the elution of three peaks with pepsin-inhibitory activity. N-terminal sequencing indicated a common sequence of MGPGPAIGEVIG and the presence of minor species with seven- or two-amino-acid N-terminal extensions beyond this point. Microheterogeneity in this end sequence was exhibited within and between two preparations. Internal sequencing of a major peak after a trypsin digestion gave the sequence FYNVVVLEK. The common N-terminal sequence was used to design a degenerate primer for 3' rapid amplification of cDNA ends and cDNA clones encoding two isoforms of the inhibitor were obtained. The open reading frames of both cDNAs encoded proteins (96% identical) which contained the experimentally determined internal sequence. The amino acid content calculated from the predicted amino acid sequence was very similar to that measured by amino acid analysis of the purified inhibitor. The two predicted amino acid sequences (96 residues) had neither similarity to any other aspartic proteinase inhibitor nor similarity to any other protein. The inhibitors have a molecular mass of 10,552 Da, measured by matrix-assisted laser-desorption ionisation time-of-flight mass spectrometry and approximately 10,000 Da by SDS/PAGE, and behave as dimers of approximately 21,000 Da during chromatography on Superdex G-75 gel-filtration medium. The calculated molecular masses from the predicted amino acid sequences were 10,551 Da and 10,527 Da. The inhibitor was capable of inhibiting pepsin (Ki = 2 nM) and a secreted aspartic proteinase from the fungus Glomerella cingulata (Ki = 20 nM). The inhibitor, which is stable over acid and neutral pH, has been named squash aspartic proteinase inhibitor (SQAPI).

  2. Sedimentation of nucleoids from thymus and spleen cells of rats after X-irradiation in vitro and in vivo

    International Nuclear Information System (INIS)

    Heinzelmann, R.

    1987-01-01

    The reaction to irradiation of thymocytes was tested immediately and 6 hours after whole body X-irradiation of rats with doses from 190 cGy up to 1520 cGy by nucleoid sedimentation. For comparison, examinations of thymus and spleen cells after X-irradiation in vitro were done. Preliminary analyses should find a possible coergism between X-rays on one side and hyperthermia and inhibitors of DNA-synthesis or DNA-repair (cytosinearabinoside, dideoxythymidine, 3-amino-benzamide, ethidiumbromide, and novobiocine) on the other side. From the results the following conclusions may be drawn: 1) With respect to the detection of in vivo effects of X-irradiation, the nucleoid sedimentation is less sensitive than biochemical methods. 2) Some hours after sublethal X-irradiation in vivo, free DNA and/or polydesoxyribonucleotides appear. At the same time cross-links can be detected in the chromatin fraction. 3) The reduction of the nucleoid sedimentation immediately after high doses of whole-body irradiation is the result of primary DNA lesions. The changes detectable some hours after are due to the secondary enzymatic changes, that are connected with the interphase death of thymocytes, and coincide with the present opinions about the irradiation induced apoptosis of cells. (orig./ECB) [de

  3. Changes of red blood cell aggregation parameters in a long-term follow-up of splenectomy, spleen-autotransplantation and partial or subtotal spleen resections in a canine model.

    Science.gov (United States)

    Miko, Iren; Nemeth, Norbert; Peto, Katalin; Furka, Andrea; Toth, Laszlo; Furka, Istvan

    2017-01-01

    Decrease or loss in splenic filtration function may influence the hemorheological state. To follow-up the long-term effects of splenectomy, spleen autotransplantation and spleen resections on red blood cell aggregation in a canine model. Beagle dogs were subjected to control (n = 6), splenectomy (SE, n = 4), spleen autotransplantation (AU, Furka's spleen-chip method, n = 8) or partial and subtotal spleen resection (n = 4/each) groups, and followed-up for 18 postoperative (p.o.) months. Erythrocyte aggregation was determined in parallel by light-transmittance aggregometry (Myrenne MA-1 aggregometer) and syllectometry (LoRRca). Erythrocyte aggregation decreased three months after splenectomy, with lower aggregation index and elongated aggregation time. It was more or less associated with relatively lower hematocrit and fibrinogen concentration. However, in autotransplantated animals a relatively higher fibrinogen did not increase the aggregation markedly. Spleen resection resulted in the most controversial red blood cell aggregation findings, and it seems, that the degree of the resection is an influencing factor. Splenectomy alters erythrocyte aggregation, spleen autotransplantation can be useful to preserve filtration function. However, the degree of restoration shows individual differences with a kind of 'functional periodicity'. Spleen resection controversially influences erythrocyte aggregation parameters. The subtotal resection is supposed to be worse than spleen autotransplantation.

  4. Multidetector computed tomography findings of mesenteroaxial gastric volvulus combined with torsion of wandering spleen: A case report and literature review

    International Nuclear Information System (INIS)

    Youn, In Kyung; Ku, Young Mi; Lee, Su Lim

    2016-01-01

    Gastric volvulus, defined as an abnormal rotation of stomach, may be idiopathic or secondary to abnormal fixation of intraperitoneal visceral ligaments. Wandering spleen is a movable spleen resulting from absence or underdevelopment of the splenic supporting ligaments that suspend the spleen to its normal position in the left part of the supramesocolic compartment of the abdomen. Wandering spleen increases the risk of splenic torsion. Both gastric volvulus and splenic torsion are potentially life-threatening if not urgently managed with surgery. Prompt and accurate diagnosis based on multidetector computed tomography (MDCT) is crucial to prevent unforeseen complications. Gastric volvulus and coexistent torsion of wandering spleen is a very rare condition. Herein, we described a case of gastric volvulus associated with wandering spleen and intestinal non-rotation in a 15-year-old girl focusing on MDCT findings

  5. Multidetector computed tomography findings of mesenteroaxial gastric volvulus combined with torsion of wandering spleen: A case report and literature review

    Energy Technology Data Exchange (ETDEWEB)

    Youn, In Kyung; Ku, Young Mi; Lee, Su Lim [Dept. of Radiology, Uijeongbu St. Mary' s Hospital, College of Medicine, The Catholic University of Korea, Uijeongbu (Korea, Republic of)

    2016-05-15

    Gastric volvulus, defined as an abnormal rotation of stomach, may be idiopathic or secondary to abnormal fixation of intraperitoneal visceral ligaments. Wandering spleen is a movable spleen resulting from absence or underdevelopment of the splenic supporting ligaments that suspend the spleen to its normal position in the left part of the supramesocolic compartment of the abdomen. Wandering spleen increases the risk of splenic torsion. Both gastric volvulus and splenic torsion are potentially life-threatening if not urgently managed with surgery. Prompt and accurate diagnosis based on multidetector computed tomography (MDCT) is crucial to prevent unforeseen complications. Gastric volvulus and coexistent torsion of wandering spleen is a very rare condition. Herein, we described a case of gastric volvulus associated with wandering spleen and intestinal non-rotation in a 15-year-old girl focusing on MDCT findings.

  6. Three sorghum serpin recombinant proteins inhibit midgut trypsin activity and growth of corn earworm

    Science.gov (United States)

    The sorghum (Sorghum bicolor) genome contains at least 17 putative serpin (serine protease inhibitor) open reading frames, some of which are induced by pathogens. Recent transcriptome studies found that most of the putative serpins are expressed but their roles are unknown. Four sorghum serpins were...

  7. Purification of a 6.5 kDa protease inhibitor from Amazon Inga umbratica seeds effective against serine proteases of the boll weevil Anthonomus grandis.

    Science.gov (United States)

    Calderon, L A; Teles, R C L; Leite, J R S A; Franco, O L; Grossi-de-Sá, M F; Medrano, F J; Bloch, C; Freitas, S M

    2005-08-01

    A 6.5 kDa serine protease inhibitor was purified by anion-exchange chromatography from the crude extract of the Inga umbratica seeds, containing inhibitor isoforms ranging from 6.3 to 6.7 kDa and protease inhibitors of approximately 19 kDa. The purified protein was characterized as a potent inhibitor against trypsin and chymotrypsin and it was named I. umbratica trypsin and chymotrypsin inhibitor (IUTCI). MALDI-TOF spectra of the IUTCI, in the presence of DTT, showed six disulfide bonds content, suggesting that this inhibitor belongs to Bowman-Birk family. The circular dichroism spectroscopy indicates that IUTCI is predominantly formed by unordered and beta-sheet secondary structure. It was also characterized, by fluorescence spectroscopy, as a stable protein at range of pH from 5.0 to 7.0. Moreover, this inhibitor at concentration of 75 microM presented a remarkable inhibitory activity (60%) against digestive serine proteases from boll weevil Anthonomus grandis, an important economical cotton pest.

  8. A new procedure for imaging liver and spleen with water soluble contrast media in liposomes

    International Nuclear Information System (INIS)

    Zherbin, E.A.; Davidenkova, E.F.; Khanson, K.P.; Gubareva, A.V.; Zhdanova, N.V.; Aliyakparov, M.T.; Loshakova, L.V.; Fomina, Eh.V.; Rozenberg, O.A.

    1983-01-01

    The problems of long-term, reversible, and safe contrast investigation of liver and spleen and reduction of the irritating action of water-soluble contrast media on the wall of blood vessels are unresolved. The production and experimental application of contrast media encapsulated in liposomes are described. It is possible to produce a liposome preparation with 10-20 % Verografin content. After intravenous injection it leads to a quick (after 16-30 min), persisting (10-12 h) and reversible (24-30 h) contrast imaging of liver and spleen in rodents. The contrast medium has no pathological effects on heart, blood and circulatory system and on the morphology of liver, spleen, heart, lungs, kidneys and urinary bladder. The perspectives of clinical application of such contrast media are discussed. (author)

  9. Usefulness of liposomes carrying losefamate for CT opacification of liver and spleen

    International Nuclear Information System (INIS)

    Seltzer, S.E.; Shulkin, P.M.; Adams, D.F.; Davis, M.A.; Hoey, G.B.; Hopkins, R.M.; Bosworth, M.E.

    1984-01-01

    Iosefamate, a hepatobiliary contrast agent, was encapsulated into liposomes to increase its ability to opacify the liver and spleen on computed tomographic (CT) images. Multilamellar lipid vesicles containing iosefamate in their aqueous phase were prepared. Seven dogs received intravenous injections of 100-300 mg l/kg in one of three forms; encapsulated, unencapsulated, or a mixture of the two in equal parts. Animals that received the opaque vesicles had marked opacification of their livers, bile ducts, gallbladders, spleens and gastrointestinal tracts. At the high-dose level, liver upake of the encapsulated materials was also greater. Liposome-encapsulated hepatobiliary contrast agents are effective liver and spleen opacifiers for CT imaging in the dog

  10. Extra-adrenal myelolipoma presenting in the spleen: A report of two cases

    Directory of Open Access Journals (Sweden)

    N.S. Aguilera

    2016-12-01

    Full Text Available Myelolipoma is a rare neoplasm composed of mature fat and bone marrow occurring most frequently in the adrenal gland with rare occurrences in extra adrenal locations including lung, liver, retroperitoneum, mediastinum and testes. Splenic myelolipomas are seen most commonly in non-human species including cat and dog. Only rare cases of splenic myelolipoma in humans have been reported previously. We present two cases of myelolipoma in the spleen. The first is a 62 year old female presenting with abdominal pain and a splenic mass. The second is a 44 year old male presenting with hematuria and a mass in the spleen. Both cases showed trilineage bone marrow elements with mature fat. These cases demonstrate that myelolipoma do rarely occur in human spleen and we highlight the distinction from extramedullary hematopoiesis, mature extramedullary myeloid tumor (myeloid sarcoma, lipoma and well differentiated liposarcoma.

  11. Effect of adeturone on the concentration of endogenous sulfhydryl groups in mouse spleen and liver

    International Nuclear Information System (INIS)

    Pantev, T.; Bychvarova, K.

    1981-01-01

    Levels of endogenous sulfhydryl groups (total, protein, and non-protein) in mouse liver and spleen were studied for response to the radioprotective drug Adeturone (AET adenosine triphosphate) as recorded at various time intervals (5 - 90 min) following administration of a 300 mg/kg b.w. dose. Spleen sulfhydryl concentration levels tended to elevation, with the peak effect noted at 45 min post-treatment. In the liver, augmentation was observed only for non-protein sylfhydryl groups, at 10 and 15 min post-treatment (time intervals when Adeturone affords maximum protection against radiation); at the 60 min, however, there was a statistically reliable drop. The findings indicate that Adeturone treatment produces response patterns of opposite directions in liver and spleen endogenous thiols. (A.B.)

  12. The Spleen Is an HIV-1 Sanctuary During Combined Antiretroviral Therapy.

    Science.gov (United States)

    Nolan, David J; Rose, Rebecca; Rodriguez, Patricia H; Salemi, Marco; Singer, Elyse J; Lamers, Susanna L; McGrath, Michael S

    2018-01-01

    Combined antiretroviral therapy (cART) does not eradicate HIV, which persists for years and can re-establish replication if treatment is stopped. The current challenge is identifying those tissues harboring virus through cART. Here, we used HIV env-nef single genome sequencing and HIV gag droplet digital PCR (ddPCR) to survey 50 tissues from five subjects on cART with no detectable plasma viral load at death. The spleen most consistently contained multiple proviral and expressed sequences (4/5 participants). Spleen-derived HIV demonstrated two distinct phylogenetic patterns: multiple identical sequences, often from different tissues, as well as diverse viral sequences on long terminal branches. Our results suggested that ddPCR may overestimate the size of the tissue-based viral reservoir. The spleen, a lymphatic organ at the intersection of the immune and circulatory systems, may play a key role in viral persistence.

  13. Measurement of spleen size using gamma camera scintigraphy in essential thrombocythaemia

    Energy Technology Data Exchange (ETDEWEB)

    Revesz, P. (Department of Medicine, Oestra Hospital, University of Goeteborg, Goeteborg (Sweden)); Carneskog, J.; Wadenvik, H.; Kutti, J. (Haematology Section, Department of Medicine, Sahlgrenska Hospital, University of Goeteborg, Goeteborg (Sweden)); Jarneborn, L. (Department of Clinical Physiology, Oestra Hospital, University of Goeteborg, Goeteborg (Sweden))

    1993-09-01

    By using gamma camera imaging the spleen size was determined in 33 consecutive patients with essential thrombocythaemia (ET) and in 33 consecutive patients with reactive thrombocytosis (RT). All ET patients were newly diagnosed and had not received myelosuppressive treatment prior to study; they all fulfilled the criteria for ET as established by the Polycythemia Vera Study Group. In both posterior and lateral projections, the spleen area in the group of ET patients was significantly larger than in the RT patients. The present study has shown that 39% of ET patients at diagnosis have splenic enlargement. Evaluation of Spleen size is therefore a useful diagnostic test in patients presenting with unexplained thrombocytosis. (au) (15 refs.).

  14. Yersinia enterocolitica-associated generalized microinfarctions of bone and spleen in a child

    International Nuclear Information System (INIS)

    Reiss-Zimmermann, Martin; Sorge, Ina; Hirsch, Wolfgang; Schille, Regine; Beer, Joerg

    2007-01-01

    We report a case of unusual extraintestinal yersiniosis in a 16-year-old girl with generalized microinfarctions of the bone and spleen. For the past 2 years she had been repeatedly admitted to our hospital with reactive arthritis, erythema nodosum and iridocyclitis of unknown aetiology. Ultrasound showed multiple round hypoechoic lesions in the spleen that were shown to have low T2 signal on MRI. MRI also showed disseminated nodular lesions of the skeleton that were low T1 and high T2 signal and demonstrated inhomogeneous contrast enhancement. The patient is currently in good health on low-dose nonsteroidal immunosuppressive therapy. This is a unique case of microinfarctions of the skeleton and spleen caused by a severe postinfectious autoimmune reaction following extraintestinal Yersinia enterocolitica infection. (orig.)

  15. Yersinia enterocolitica-associated generalized microinfarctions of bone and spleen in a child

    Energy Technology Data Exchange (ETDEWEB)

    Reiss-Zimmermann, Martin; Sorge, Ina; Hirsch, Wolfgang [University Hospital Leipzig, Paediatric Radiology, Department of Diagnostic and Interventional Radiology, Leipzig (Germany); Schille, Regine [University Hospital Leipzig, Paediatric Department, Leipzig (Germany); Beer, Joerg [University of Leipzig, Institute for Medical Microbiology and Epidemiology of Infectious Diseases, Leipzig (Germany)

    2007-12-15

    We report a case of unusual extraintestinal yersiniosis in a 16-year-old girl with generalized microinfarctions of the bone and spleen. For the past 2 years she had been repeatedly admitted to our hospital with reactive arthritis, erythema nodosum and iridocyclitis of unknown aetiology. Ultrasound showed multiple round hypoechoic lesions in the spleen that were shown to have low T2 signal on MRI. MRI also showed disseminated nodular lesions of the skeleton that were low T1 and high T2 signal and demonstrated inhomogeneous contrast enhancement. The patient is currently in good health on low-dose nonsteroidal immunosuppressive therapy. This is a unique case of microinfarctions of the skeleton and spleen caused by a severe postinfectious autoimmune reaction following extraintestinal Yersinia enterocolitica infection. (orig.)

  16. Dietary low or excess levels of lipids reduced growth performance, and impaired immune function and structure of head kidney, spleen and skin in young grass carp (Ctenopharyngodon idella) under the infection of Aeromonas hydrophila.

    Science.gov (United States)

    Ni, Pei-Jun; Jiang, Wei-Dan; Wu, Pei; Liu, Yang; Kuang, Sheng-Yao; Tang, Ling; Tang, Wu-Neng; Zhang, Yong-An; Zhou, Xiao-Qiu; Feng, Lin

    2016-08-01

    Our study explored the effect of dietary lipids on growth and immunity and structure (head kidney, spleen and skin) of young grass carp (Ctenopharyngodon idella). A total of 540 young grass carp with an average initial weight of 261.41 ± 0.53 g were fed diets containing six graded levels of lipids at 5.9-80.1 g/kg diet for 8 weeks. After that, a challenge trial was conducted by injection of Aeromonas hydrophila over 2 weeks. The results indicated that compared with optimal lipids supplementation, low and excess levels of lipids down-regulated the mRNA levels of antimicrobial peptides, anti-inflammatory cytokines, inhibitor of κBα (IκBα) and ribosomal p70S6 kinase (S6K1), and up-regulated pro-inflammatory cytokines, nuclear factor κB p65 (NF-κB p65), NF-κB c-Rel (not p52), IκB kinase α (IKKα), IKKβ, IKKγ, and eIF4E-binding protein (4EBP) mRNA levels in the head kidney and spleen of young grass carp (P lipids also increased reactive oxygen species (ROS) production and malondialdehyde (MDA) and protein carbonyl (PC) contents, reduced the activities of antioxidant enzymes (P lipids down-regulated the mRNA levels of B-cell lymphoma-2 (Bcl-2) and inhibitor of apoptosis protein (IAP) in the head kidney and spleen, whereas up-regulated the mRNA levels of apoptotic protease activating factor-1 (Apaf-1), caspase 3, 7, 8 and 9 mRNA levels in the head kidney and spleen and Fas ligand (FasL) mRNA levels in the spleen of young grass carp, suggesting that low or excess levels of lipids could decrease the head kidney and spleen immune function, induce oxidative damage and apoptosis and impair antioxidant system of young grass carp. At last, low or excess levels of lipids also impaired the immune function and structure in the skin of young grass carp. Based on the quadratic regression analysis for PWG, skin haemorrhage and lesions morbidity and IgM content, the dietary lipids requirements for young grass carp were estimated to be 43.7, 60.2, 55.0 and 52.1

  17. Natural history of nonoperative management for grade 4 and 5 liver and spleen injuries in children.

    Science.gov (United States)

    Yang, Jeannie C; Sharp, Susan W; Ostlie, Daniel J; Holcomb, George W; St Peter, Shawn D

    2008-12-01

    Nonoperative management is standard treatment of blunt liver or spleen injuries. However, there are few reports outlining the natural history and outcomes of severe blunt hepatic and splenic trauma. Therefore, we reviewed our experience with nonoperative management of grade 4 or 5 liver and spleen injuries. A retrospective analysis was performed on patients with grade 4 or 5 (high-grade) blunt liver and/or spleen injuries from April 1997 to July 2007 at our children's hospital. Demographics, hospital course data, and follow-up data were analyzed. There were 74 high-grade injuries in 72 patients. There were 30 high-grade liver and 44 high-grade spleen injuries. Two patients had both a liver and splenic injury. High-grade liver injuries had a significantly longer length of intensive care and hospital stay compared to high-grade spleen injuries. There were also a significantly higher number of transfusions, radiographs, and total charges in the high-grade liver injuries when compared to the high-grade splenic injuries. The only mortality from solid organ injury was a grade 4 liver injury with portal vein disruption. In contrast, there was only one complication from a high-grade splenic injury-a pleural effusion treated with thoracentesis. There were 5 patients with complications from their liver injury requiring 18 therapeutic procedures. Three patients (10%) with liver injury required readmission as follows: one 5 times, one 3 times, and another one time. Patients with high-grade liver injuries have a longer recovery, more complications, and greater use of resources than in patients with similar injuries to the spleen.

  18. Parasitic load and histological aspects in different regions of the spleen of dogs with visceral leishmaniasis.

    Science.gov (United States)

    Bagues, Naiara Carvalho Teixeira; Pinheiro, Cristiane Garboggini Melo de; Bastos, Leila Andrade; Fraga, Deborah Bittencourt Mothé; Veras, Patrícia Sampaio Tavares; Pontes-de-Carvalho, Lain Carlos; Dos-Santos, Washington L C; Oliveira, Geraldo Gileno de Sá

    2018-02-01

    Leishmania infantum causes from subclinical infection to severe disease in humans and dogs. The spleen is one of the organs most affected by the infection. Although evidence exists that the parasitic load distribution and histological alterations may not be homogeneous in the affected organs of naturally infected individuals, it has not been formally demonstrated using the current techniques used for studying the disease. In six dogs naturally infected with Leishmania, parasitic load and histological changes were compared in samples collected from the lower, middle and upper third of the spleen. Parasitic load in the spleen of the group of dogs was variable, revealing a difference of 61 times between animals with the lowest and the highest parasitism. The set of parasitic load values of each dog showed a cluster trend, when compared to the other animals. Nevertheless, the parasitic load values of each dog showed a variation ranging from 3.2 to 34.7 times between lowest and highest value. Histological changes showed recognizable variation in frequency (granulomas) or intensity (perisplenitis) in the spleen of 2 out of the 6 dogs. The agreement of histological findings between samples collected from the different thirds of the spleen was good (kappa coeficient, 0.61-0.80) very good (0.81-0.99) or perfect (1.00), for most of the parameters analyzed. Variability of parasitic load and, to a lesser extent, histological changes in spleen of dogs with visceral leishmaniasis is observed. Such variability may be taken in account in the design of studies on pathogenesis, vaccine and therapeutic drug development. Copyright © 2017 Elsevier Ltd. All rights reserved.

  19. Shear-wave elastography of the liver and spleen identifies clinically significant portal hypertension

    DEFF Research Database (Denmark)

    Jansen, Christian; Bogs, Christopher; Verlinden, Wim

    2017-01-01

    BACKGROUND & AIMS: Clinically significant portal hypertension (CSPH) is associated with severe complications and decompensation of cirrhosis. Liver stiffness measured either by transient elastography (TE) or Shear-wave elastography (SWE) and spleen stiffness by TE might be helpful in the diagnosis...... correlate with portal pressure and can both be used as a non-invasive method to investigate CSPH. Even though external validation is still missing, these algorithms to rule-out and rule-in CSPH using sequential SWE of liver and spleen might change the clinical practice....

  20. Locally advanced leiomyosarcoma of the spleen. A case report and review of the literature

    Directory of Open Access Journals (Sweden)

    Recchia Franco

    2007-11-01

    Full Text Available Abstract Background Leiomyosarcomas are rare tumours, predominantly localized in the stomach, small intestine and retroperitoneum. Only one case of primary leiomyosarcoma of the spleen is described in human beings in literature. Case presentation We report a case of locally advanced primary leiomyosarcoma of the spleen in a 54 year-old woman, diagnosed only after splenectomy, performed with the suspicion of splenic haematoma. Conclusion Due to the lack of cases, no specific chemotherapy regimen has been tested to provide a longer survival.