DOCSLIB.ORG

Production of Secretory Leucocyte Protease Inhibitor

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
Production of Secretory Leucocyte Protease Inhibitor Research Paper Mediators of Inflammation, 8, 147–151 (1999) SECRETORY leucocyte protease in hibitor (SLPI) is a Production of secretory leucocyte potent in hibitor of granulocyte elastase and cath- epsin G, and also an inhibitor of pancreatic enzym es protease inhibitor (SLPI) in human like trypsin , chym otrypsin and pancreatic elastase. SLPI has also been show n to in hibit HIV-1 in fections pancreatic b -cells by blocking viral DNA synthesis. Since SLPI is an in hibitor of pancreatic proteases we wis hed to in ves- tigate w heth er SLPI was also actually produced in the Max Nystrom,È 1,2 Magnus Bergenfeldt,1,2 pancreas. M-RNA from human pancreatic tissue Irena Ljungcrantz,2 sa Lindeheim2 and show ed evidence of SLPI production usin g the reverse 2,C A transcriptase polym er chain reaction technique (RT- Kjell Ohlsson PCR). Using im m unohistochem ical m ethods SLPI was dem onstrated in the b -ce1ls of the islets of Langer- hans. Th e function could be local protease/anti- 1Department of Surgery, Lund University, Lund, and protease regulation or antiviral/antibacterial defence 2Department of Surgical Pathophysiology, MalmoÈ in the close vicin ity of the cell surface, or even in side University Hospital, SE-205 02 Malmo,È Sweden the b -cell itself. Key words: SLPI, Islets of Langerhans, Pancreas, b -Cell, CA Antiviral, Antibacterial Corresponding Author Tel: (+46) 40 33 31 50/51 Fax: (+46) 40 33 62 40 Introduction blocking DNA synthesis.22 SLPI is probably involved in inflammatory regulation in the intestines as w ell as in SLPI is an acid-resistant 11.8-kDa non-glycosylated the respiratory and urogenital tracts. In patients w ith serine antiprotease. Originally isolated from human ulcerative colitis, active pancreatic proteases have parotid gland secretions,1 it is found in cells on a been found in faeces. Since SLPI is an inhibitor of both variety of mucosal surfaces. For example, the seminal pancreatic and leukocytic proteases, it could be of vesicles, prostate, epididymis and cervix ,2 –7 in serous importance in the pathogenesis of ulcerative colitis. cells of the parotid and submandibular salivary Having recently demonstrated SLPI production in the glands,1,8,9 in the upper respiratory tract, in the intestinal mucosa,14 and know ing that SLPI produced lacrimal glands,10 as w ell as in the Clara cells and in the upper respiratory tract is rapidly degraded in goblet cells of the low er respiratory tract.11–13 SLPI gastric and duodenal juice,23 w e also w anted to has also been found in von Ebner’s glands at the base investigate other possibilities of origin. Earlier studies of the tongue and submucosal cells of the oesopha- failed to show production or presence of SLPI in the gus.10 Recently, SLPI w as found in Paneth cells in the pancreas.10 The aim of this study w as, therefore, to small intestine, and in scattered mucosa cells of reinvestigate the possible presence and production goblet-type in the epithelium of both the large and of SLPI in pancreatic tissue using more sensitive small bow el. In addition, SLPI has frequently been methods. found in colonic adenomas.14 SLPI is thought to be an inflammatory regulator w ith Methods and materials strong inhibitory capacity aimed tow ards proteases present in various body fluids. Thus, SLPI is a potent Xylene, ethanol, hydrochloric acid, hydrogen perox- inhibitor of leucocyte elastase and cathepsin G, and a idase, formaldehyde, haematoxylin and Tris-buffered moderate inhibitor of trypsin and chymotrypsin.1,1 5,1 6 saline (0.05 mol/l Tris in NaCl 0.15 mol/l, pH 7.6) Furthermore, it is a moderate inhibitor of pancreatic w ere standard chemicals for laboratory use. Recombi- elastase,17 and it is also know n to inhibit both mast cell nant human SLPI was a kind gift from Robert chymase activity and histamine release from mast Thompson at Synergen Inc. (Boulder, CO, USA). Goat cells.18 ,19 Its proteinase-inhibitory activity is know n to anti-SLPI antiserum was produced in the laboratory by be located in the second COOH-terminal domain. immunising Swedish landrace goats w ith recombinant Although no inhibitory capacity has been detected in SLPI. Normal goat and rabbit serum w as obtained the NH2-terminal domain, antimicrobial capacity has from Dakopatt AB (Copenhagen, Denmark). Bio- been reported.20,2 1 It has also been reported that SLPI tinylated rabbit anti-goat IgG antibodies and avidin- blocks the human immunodeficiency virus type 1 by biotinylated horseradish peroxidase complex (ABC ISSN 0962-9351 print/ISSN 1466-1861 online/99/030147-05 1999 Taylor & Francis Ltd 147 M. Ny stro¨ m et al. complex) were purchased from Vector Laboratories After hybridisation the membrane w as w ashed in 2 (Burlingame, CA, USA). Diaminobenzidine w as a 3 SSC containing 0.1% SDS, 1 3 15 min, 0.1 3 SSC product of Saveen Biotech AB (Malmo¨ , Sw eden). containing 0.1% SDS, 2 3 15 min, both at room Crystalline3 porcine pepsin w as obtained from Sigma temperature. Detection of the target DNA hybrid w as Chemicals (St. Louis, MO, USA). achieved as follow s. After a brief rinse in 100 mM Tris – HCl, 150 mM NaCl, pH 7.5, the membrane w as incubated in 0.5% (w /v) blocking reagent in the same RT-PCR and Southern blot buffer for 30 min. This w as follow ed by a brief w ash in Human pancreas Poly(A+ ) RNA w as obtained from buffer. Diluted anti-digoxigenin antibody–AP-conju- Clontec (Palo Alto, CA, USA). Specific primers (R&D gate (1:500) w as applied to the membrane for 30 min. Systems Europe Ltd, Abingdon, UK) w ere designed Rinses in buffer, 2 3 15 min follow ed. The staining according to the nucleic acid sequence of SLPI w as carried out by incubating the membrane in a cDNA.24 The upstream and dow nstream primers for solution containing 45 m l Nitroblue Tetrasodium salt amplification of the human SLPI cDNA fragments (NBT), 35 m l 5-bromo-4-chloro-33 indolyl phosphate w ere: 59 -TGT CCT GAC ACT TGT GGC AT-39 and (X-phosphate) in 10 ml buffer (100 mM Tris –HCl, 59 -CGA TCA ACT GGC ACT TCT TG-39 . The oligonu- 100 mM NaCl, 50 mM MgCl2 , pH 9.5). The experiment cleotide used for probing the Southern blot of the RT- w as repeated three times. PCR product had the sequence 59 -GGT TTG GGG TGT CAA CAG GAT CCA GGC AT-39 w ith the modification Immunohistochemical staining 39 +59 digoxigenin.24 The first strand of cDNA w as synthesised using Pancreas specimens from five different patients col- reverse transcriptase, isolated from avian myelo- lected during surgery for pancreatic adenocarcinoma blastosis virus (Appligene, Illkirch, France) in a 50-m l w ere investigated. The specimens were from micro- reaction mixture containing 2.5 m l 400 mM Tris –HCl, and macroscopically normal pancreatic tissue. The pH 8.3, 2.5 m l 400 mM KCl, 1 m l 300 mM MgCl2 , 5 m l patients were two men and three w omen betw een 41 100 mM dithiothretiol (DTT), 5 m l 5 mM 4dNTPmix, and 69 years of age. They had no earlier history of 2 m l 2 mg/ml actinomycin D, 1 m l 4 m M dow nstream pancreatic disease. Informed consent for the proce- primer and approx. 1–2 m g poly(A+ ) RNA (approx. dure w as obtained. The tissue samples w ere cut at 10 m l). The reaction mix ture w as incubated at 42°C for random and fixed in 4% formaldehyde. Paraffin- 60 mm, then diluted w ith 450 m l 10 mM Tris –HCl/ embedded tissue w as routinely stained w ith haema- 10 mM EDTA, pH 7.5. To 5 m l of this diluted mixture toxylin for light microscopic examination. The fixed w ere added 5 m l of each amplification primer, 20 mM specimens were subjected to pepsin digestion (pep- each, 4 m l 5 mM 4dNTPmix, 10 m l 103 amplification sin 4 mg/ml in 0.01 mol/l HCl) for 30 min at 37°C and buffer (500 mM KCl, 100 mM Tris –HCl, pH 8.4, and w ere transferred into Tris-buffered saline. To quench 1 mg/ml gelatine), 1.5 m l 100 mM MgCl2 and 69 m l H2O. endogenous tissue peroxidase activity, in the next The PCR reaction w as performed using the ‘hot start’ step, the sections were incubated w ith 0.3% H2O2 in method w ith paraffin pellets, heating at 94°C for 2 min methanol for 30 min at room temperature. To block follow ed by chilling on ice. Then 2.5 units (0.5 m l) of non-specific staining, the specimens w ere incubated Taq DNA polymerase (Appligene) w ere added to the for 10 min in 5% normal rabbit serum. In order to tubes. Amplification w as carried out in a thermo-cycler show specific staining in the b -cells, the specimens (Hybaid Omnigene, Teddington, UK) w ith the follow - w ere treated as follow s. ing cycling programme: 39 cycles of 2 min at 55°C; 2 min at 72°C and 1 min at 94°C; follow ed by one cycle (1) SLPI of 2 min at 55°C and 7 min at 72°C. The PCR product The slides were incubated w ith goat anti-human SLPI w as then analysed by agarose gel electrophoresis and (1:1000) for 30 min followed by the addition of visualised after ethidium bromide staining (0.8% biotinylated rabbit anti-goat IG antibodies (5 m g/ml agarose gel in 0.53 TBE buffer).
Load more

Recommended publications
  • Purification and Identification of a Binding Protein for Pancreatic
    Biochem. J. (2003) 372, 227–233 (Printed in Great Britain) 227 Purification and identification of a binding protein for pancreatic secretory trypsin inhibitor: a novel role of the inhibitor as an anti-granzyme A Satoshi TSUZUKI*1,2,Yoshimasa KOKADO*1, Shigeki SATOMI*, Yoshie YAMASAKI*, Hirofumi HIRAYASU*, Toshihiko IWANAGA† and Tohru FUSHIKI* *Laboratory of Nutrition Chemistry, Division of Food Science and Biotechnology, Graduate School of Agriculture, Kyoto University, Kitashirakawa Oiwake-cho, Sakyo-ku, Kyoto 606-8502, Japan, and †Laboratory of Anatomy, Graduate School of Veterinary Medicine, Hokkaido University, Kita 18-Nishi 9, Kita-ku, Sapporo 060-0818, Japan Pancreatic secretory trypsin inhibitor (PSTI) is a potent trypsin of GzmA-expressing intraepithelial lymphocytes in the rat small inhibitor that is mainly found in pancreatic juice. PSTI has been intestine. We concluded that the PSTI-binding protein isolated shown to bind specifically to a protein, distinct from trypsin, on from the dispersed cells is GzmA that is produced in the the surface of dispersed cells obtained from tissues such as small lymphocytes of the tissue. The rGzmA hydrolysed the N-α- intestine. In the present study, we affinity-purified the binding benzyloxycarbonyl-L-lysine thiobenzyl ester (BLT), and the BLT protein from the 2 % (w/v) Triton X-100-soluble fraction of hydrolysis was inhibited by PSTI. Sulphated glycosaminoglycans, dispersed rat small-intestinal cells using recombinant rat PSTI. such as fucoidan or heparin, showed almost no effect on the Partial N-terminal sequencing of the purified protein gave a inhibition of rGzmA by PSTI, whereas they decreased the inhi- sequence that was identical with the sequence of mouse granzyme bition by antithrombin III.
    [Show full text]
  • Serine Proteases with Altered Sensitivity to Activity-Modulating
    (19) & (11) EP 2 045 321 A2 (12) EUROPEAN PATENT APPLICATION (43) Date of publication: (51) Int Cl.: 08.04.2009 Bulletin 2009/15 C12N 9/00 (2006.01) C12N 15/00 (2006.01) C12Q 1/37 (2006.01) (21) Application number: 09150549.5 (22) Date of filing: 26.05.2006 (84) Designated Contracting States: • Haupts, Ulrich AT BE BG CH CY CZ DE DK EE ES FI FR GB GR 51519 Odenthal (DE) HU IE IS IT LI LT LU LV MC NL PL PT RO SE SI • Coco, Wayne SK TR 50737 Köln (DE) •Tebbe, Jan (30) Priority: 27.05.2005 EP 05104543 50733 Köln (DE) • Votsmeier, Christian (62) Document number(s) of the earlier application(s) in 50259 Pulheim (DE) accordance with Art. 76 EPC: • Scheidig, Andreas 06763303.2 / 1 883 696 50823 Köln (DE) (71) Applicant: Direvo Biotech AG (74) Representative: von Kreisler Selting Werner 50829 Köln (DE) Patentanwälte P.O. Box 10 22 41 (72) Inventors: 50462 Köln (DE) • Koltermann, André 82057 Icking (DE) Remarks: • Kettling, Ulrich This application was filed on 14-01-2009 as a 81477 München (DE) divisional application to the application mentioned under INID code 62. (54) Serine proteases with altered sensitivity to activity-modulating substances (57) The present invention provides variants of ser- screening of the library in the presence of one or several ine proteases of the S1 class with altered sensitivity to activity-modulating substances, selection of variants with one or more activity-modulating substances. A method altered sensitivity to one or several activity-modulating for the generation of such proteases is disclosed, com- substances and isolation of those polynucleotide se- prising the provision of a protease library encoding poly- quences that encode for the selected variants.
    [Show full text]
  • Human Elastase 1: Evidence for Expression in the Skin and the Identi®Cation of a Frequent Frameshift Polymorphism
    View metadata, citation and similar papers at core.ac.uk brought to you by CORE provided by Elsevier - Publisher Connector Human Elastase 1: Evidence for Expression in the Skin and the Identi®cation of a Frequent Frameshift Polymorphism Ulvi Talas, John Dunlop,1 Sahera Khalaf , Irene M. Leigh, and David P. Kelsell Center for Cutaneous Research, St. Bartholomew's and the Royal London School of Medicine and Dentistry, Whitechapel, London, U.K. Human pancreatic elastase 1 is a serine protease individuals with/without the keratoderma revealed a which maps to the chromosomal region 12q13 close sequence variant, which would result in a premature to a locus for an autosomal dominant skin disease, truncation of the protein. This sequence variant, diffuse nonepidermolytic palmoplantar keratoderma, however, did not segregate with the skin disease and, and was investigated as a possible candidate gene for indeed, was found to occur at a relatively high fre- this disorder. Expression of two elastase inhibitors, quency in the population. Individuals homozygous ela®n and SLPI, has been related to several hyper- for the variant do not have any obvious skin proliferative skin conditions. elastase 1 is functionally abnormalities. Based on the analysis of the secondary silent in the human pancreas but elastase 1 expres- structure of the translated putative protein, the sion at the mRNA level was detected in human truncation is unlikely to result in knock-out of the cultured primary keratinocytes. Antibody staining elastase, but may cause destabilization of the localized the protein to the basal cell layer of the enzyme±inhibitor complex. Key words: ela®n/keratino- human epidermis at a number of sites includingthe cyte/protein truncation/serine protease.
    [Show full text]
  • Design of Ultrasensitive Probes for Human Neutrophil Elastase Through Hybrid Combinatorial Substrate Library Profiling
    Design of ultrasensitive probes for human neutrophil elastase through hybrid combinatorial substrate library profiling Paulina Kasperkiewicza, Marcin Porebaa, Scott J. Snipasb, Heather Parkerc, Christine C. Winterbournc, Guy S. Salvesenb,c,1, and Marcin Draga,b,1 aDivision of Bioorganic Chemistry, Faculty of Chemistry, Wroclaw University of Technology, Wroclaw 50-370, Poland; bProgram in Cell Death and Survival Networks, Sanford-Burnham Medical Research Institute, La Jolla, CA 92024; and cCentre for Free Radical Research, Department of Pathology, University of Otago Christchurch, Christchurch 8140, New Zealand Edited* by Vishva M. Dixit, Genentech, San Francisco, CA, and approved January 15, 2014 (received for review October 1, 2013) The exploration of protease substrate specificity is generally possibilities. Here we demonstrate a general approach for the syn- restricted to naturally occurring amino acids, limiting the degree of thesis of combinatorial libraries containing unnatural amino acids, conformational space that can be surveyed. We substantially with subsequent screening and analysis of large sublibraries. We enhanced this by incorporating 102 unnatural amino acids to term this approach the Hybrid Combinatorial Substrate Library explore the S1–S4 pockets of human neutrophil elastase. This ap- (HyCoSuL). We demonstrate the utility of this approach in the proach provides hybrid natural and unnatural amino acid sequen- design of a highly selective substrate and activity-based probe. ces, and thus we termed it the Hybrid Combinatorial Substrate As a target protease, we selected human neutrophil elastase Library. Library results were validated by the synthesis of individ- (EC 3.4.21.37) (NE), a serine protease restricted to neutrophil ual tetrapeptide substrates, with the optimal substrate demon- azurophil granules (14).
    [Show full text]
  • Isolation and Characterization of Porcine Ott-Proteinase Inhibitor1),2) Leukocyte Elastase-Inhibitor Complexes in Porcine Blood, I
    Geiger, Leysath and Fritz: Porcine otrproteinase inhibitor 637 J. Clin. Chem. Clin. Biochem. Vol. 23, 1985, pp. 637-643 Isolation and Characterization of Porcine ott-Proteinase Inhibitor1),2) Leukocyte Elastase-Inhibitor Complexes in Porcine Blood, I. By R. Geiger, Gisela Leysath and H. Fritz Abteilung für Klinische Chemie und Klinische Biochemie (Leitung: Prof. Dr. H. Fritz) in der Chirurgischen Klinik Innenstadt der Universität München (Received March 18/June 20, 1985) Summary: arProteinase inhibitor was purified from procine blood by ammonium sulphate and Cibachron Blue-Sepharose fractionation, ion exchange chromatography on DEAE-Cellulose, gel filtration on Sephadex G-25, and zinc chelating chromatography. Thus, an inhibitor preparation with a specific activity of 1.62 lU/mgprotein (enzyme: trypsin; Substrate: BzArgNan) was obtained. In sodium dodecyl sulphate gel electroph- oresis one protein band corresponding to a molecular mass of 67.6 kDa was found. On isoelectric focusing 6 protein bands with isoelectric points of 3.80, 3.90, 4.05, 4.20, 4.25 and 4.45 were separated. The amino acid composition was determined. The association rate constants for the Inhibition of various serine proteinases were measured. Isolierung und Charakterisierung des on-Proteinaseinhibitors des Schweins Leukocyten-oLj-Proteinaseinhibitor-Komplexe in Schweineblut, L Zusammenfassung; arProteinaseinhibitor wurde aus Schweineblut mittels Ammoniumsulfatfallung und Frak- tionierung an Cibachron-Blau-Sepharose, lonenaustauschchromatographie an DEAE-Cellulose, Gelfiltration an Sephadex G-25 und Zink-Chelat-Chromatographie isoliert. Die erhaltene Inhibitor-Präparation hatte eine spezifische Aktivität von 1,62 ITJ/mg Protein (Enzym: Trypsin; Substrat: BzArgNan). In der Natriumdodecyl- sulfat-Elektrophorese wurde eine Proteinbande mit einer dazugehörigen Molekülmasse von 67,6 kDa erhalten.
    [Show full text]
  • A Proteinase 3 Contribution to Juvenile Idiopathic Arthritis-Associated Cartilage Damage
    Brief Report A Proteinase 3 Contribution to Juvenile Idiopathic Arthritis-Associated Cartilage Damage Eric K. Patterson 1 , Nicolas Vanin Moreno 1, Douglas D. Fraser 2,3,4 , Gediminas Cepinskas 1,5, Takaya Iida 1 and Roberta A. Berard 2,4,6,* 1 Centre for Critical Illness Research, Lawson Health Research Institute, London, ON N6A 5W9, Canada; [email protected] (E.K.P.); [email protected] (N.V.M.); [email protected] (G.C.); [email protected] (T.I.) 2 Lawson Health Research Institute, Children’s Health Research Institute, London, ON N6A 5W9, Canada; [email protected] 3 Department of Physiology and Pharmacology, Schulich School of Medicine and Dentistry, Western University, London, ON N6A 5C1, Canada 4 Department of Paediatrics, Schulich School of Medicine and Dentistry, Western University, London, ON N6A 5C1, Canada 5 Department of Medical Biophysics, Western University, London, ON N6A 5C1, Canada 6 Division of Rheumatology, London Health Sciences Centre, Children’s Hospital, London, ON N6A 5W9, Canada * Correspondence: [email protected] Abstract: A full understanding of the molecular mechanisms implicated in the etiopathogenesis of juvenile idiopathic arthritis (JIA) is lacking. A critical role for leukocyte proteolytic activity (e.g., elastase and cathepsin G) has been proposed. While leukocyte elastase’s (HLE) role has been documented, the potential contribution of proteinase 3 (PR3), a serine protease present in abundance Citation: Patterson, E.K.; Vanin in neutrophils, has not been evaluated. In this study we investigated: (1) PR3 concentrations in Moreno, N.; Fraser, D.D.; Cepinskas, the synovial fluid of JIA patients using ELISA and (2) the cartilage degradation potential of PR3 by G.; Iida, T.; Berard, R.A.
    [Show full text]
  • Fecal Elastase-1 Is Superior to Fecal Chymotrypsin in the Assessment of Pancreatic Involvement in Cystic Fibrosis
    Fecal Elastase-1 Is Superior to Fecal Chymotrypsin in the Assessment of Pancreatic Involvement in Cystic Fibrosis Jaroslaw Walkowiak, MD, PhD*; Karl-Heinz Herzig, MD, PhD§; Krystyna Strzykala, M ChemA, Dr Nat Sci*; Juliusz Przyslawski, M Phar, Dr Phar‡; and Marian Krawczynski, MD, PhD* ABSTRACT. Objective. Exocrine pancreatic function ystic fibrosis (CF) is the most common cause in patients with cystic fibrosis (CF) can be evaluated by of exocrine pancreatic insufficiency in child- direct and indirect tests. In pediatric patients, indirect hood. Approximately 85% of CF patients are tests are preferred because of their less invasive charac- C 1,2 pancreatic insufficient (PI). Thus, the assessment of ter, especially in CF patients with respiratory disease. exocrine pancreatic function in CF patients is of great Fecal tests are noninvasive and have been shown to have clinical importance. For the evaluation, both direct a high sensitivity and specificity. However, there is no 3,4 comparative study in CF patients. Therefore, the aim of and indirect tests are used. The gold standard is the present study was to compare the sensitivity and the the secretin-pancreozymin test (SPT) or one of its specificity of the fecal elastase-1 (E1) test with the fecal modifications. However, this test is invasive, time chymotrypsin (ChT) test in a large cohort of CF patients consuming, expensive, and not well standardized in and healthy subjects (HS). children. Therefore, its use is limited to qualified Design. One hundred twenty-three CF patients and gastroenterologic centers. 105 HS were evaluated. In all subjects, E1 concentration Several indirect tests, such as serum tests—amy- and ChT activity were measured.
    [Show full text]
  • Pancreatic Elastase EC 3.4.21.36
    Pancreatic Elastase EC 3.4.21.36 Structure: Elastase is a compact globular protein consisting of a single polypeptide chain of 240 amino acids cross- linked by 4 disulfide bridges. It has a hydrophobic core and exhibits extensive sequence homology with other serine proteinases, such as trypsin and chymotrypsin (1,2) . The tertiary structure of elastase has been elucidated (1) . The enzyme is synthesized in porcine pancreas as a pre-proelastase (3) . After processing to proelastase, it is stored in the zymogen granules and later activated to elastase in the duodenum by tryptic cleavage of a single peptide bond in the inactive precursor molecule (4) . This process is closely resembling those of trypsinogen and chymotrypsinogen activation which results in the removal from the N-terminal end of the molecule of a small activation peptide, enabling the enzyme to adopt its native conformation. The existence of two forms of elastase has been reported which differ in respect to their catalytic properties (5) . Elastase contains no prosthetic groups or metal ions and is not subject to any allosteric activatory or inhibitory control. Its enzymatic activity results solely from the specific three-dimensional conformation which its single polypeptide chain adopts. Therefore activity is lost by denaturation and conformational changes (6) . Specificity: Elastase is a serine proteinase with broad substrate specificity. It preferentially cleaves peptide bonds at the carbonyl end of amino acid residues with small hydrophobic side chains, such as glycine, valine, leucine, isoleucine, and particularly alanine. The wide specificity of elastase for non-aromatic uncharged side chains explains its unique ability to digest native elastin, a protein rich in aliphatic side chains.
    [Show full text]
  • Faecal Elastase 1: Not Helpful in Diagnosing Chronic Pancreatitis Associated with Mild to Gut: First Published As 10.1136/Gut.42.4.551 on 1 April 1998
    Gut 1998;42:551–554 551 Faecal elastase 1: not helpful in diagnosing chronic pancreatitis associated with mild to Gut: first published as 10.1136/gut.42.4.551 on 1 April 1998. Downloaded from moderate exocrine pancreatic insuYciency P G Lankisch, I Schmidt, H König, D Lehnick, R Knollmann, M Löhr, S Liebe Abstract tion. The former is particularly helpful Background/Aim—The suggestion that only in detecting severe EPI, but not the estimation of faecal elastase 1 is a valuable mild to moderate form, which poses the new tubeless pancreatic function test was more frequent and diYcult clinical prob- evaluated by comparing it with faecal chy- lem and does not correlate significantly motrypsin estimation in patients catego- with the severe morphological changes rised according to grades of exocrine seen in chronic pancreatitis. pancreatic insuYciency (EPI) based on (Gut 1998;42:551–554) the gold standard tests, the secretin- pancreozymin test (SPT) and faecal fat Keywords: faecal elastase 1; faecal chymotrypsin; analysis. secretin-pancreozymin test; faecal fat analysis; exocrine pancreatic insuYciency; diagnosis Methods—In 64 patients in whom EPI was suspected, the following tests were per- formed: SPT, faecal fat analysis, faecal The diagnosis of chronic pancreatitis is usually chymotrypsin estimation, faecal elastase 1 based on abnormal results from pancreatic estimation. EPI was graded according to function tests and morphological examin- the results of the SPT and faecal fat ation.1 For the evaluation of exocrine pancre- analysis as absent, mild, moderate, or atic function, there are both direct and indirect severe. The upper limit of normal for fae- tests.
    [Show full text]
  • Role of Chicken Pancreatic Trypsin, Chymotrypsin and Elastase in the Excystation Process of Eimeria Tenella Oocysts and Sporocysts
    View metadata, citation and similar papers at core.ac.uk brought to you by CORE provided by Obihiro University of Agriculture and Veterinary Medicine Academic Repository Role of Chicken Pancreatic Trypsin, Chymotrypsin and Elastase in the Excystation Process of Eimeria tenella Oocysts and Sporocysts 著者（英） Guyonnet Vincent, Johnson Joyce K., Long Peter L. journal or The journal of protozoology research publication title volume 1 page range 22-26 year 1991-10 URL http://id.nii.ac.jp/1588/00001722/ J. Protozool. Res., 1. 22-26(1991) Copyright © 1991 , Research Center for Protozoan Molecular Immunology Role of Chicken Pancreatic Trypsin, Chymotrypsin and Elastase in the Excystation Process of Eimeria tenella Oocysts and Sporocysts VINCENT GUYONNET, JOYCE K. JOHNSON, and PETER L. LONG Department of Poultry Science, The University of Georgia Athens, GA 30602, U.S.A. Recieved 2 September 1991 / Accepted October 5 1991 Key words: chymotrypsin, Eimeria tenella, elastase, excystation, trypsin ABSTRACT The role of pancreatic proteolytic enzymes in the excystation process of Eimeria tenella oocysts and sporocysts was studied in vitro. Intact sporulated oocysts were preincubated in phosphate buffer, NaCl 0.9% (PBS) added with 0.5% chicken bile extract in a 5% C02 atmosphere for 30 minutes prior to exposure to either 0.25% (w/v) chicken trypsin, chymotrypsin, pancreatic elastase, or a 1% (w/v) crude extract of unsporulated and sporulated oocysts of E. tenella (Expt.1). No excystation was observed under these conditions. Sporocysts were also incubated under the same conditions without pretreatment in C02. Excystation was observed for sporocysts incubated with either trypsin, chymotrypsin or pancreatic elastase, the best percentage of excystation being recorded for the latter after 5 hours (Expt.
    [Show full text]
  • A Genomic Analysis of Rat Proteases and Protease Inhibitors
    A genomic analysis of rat proteases and protease inhibitors Xose S. Puente and Carlos López-Otín Departamento de Bioquímica y Biología Molecular, Facultad de Medicina, Instituto Universitario de Oncología, Universidad de Oviedo, 33006-Oviedo, Spain Send correspondence to: Carlos López-Otín Departamento de Bioquímica y Biología Molecular Facultad de Medicina, Universidad de Oviedo 33006 Oviedo-SPAIN Tel. 34-985-104201; Fax: 34-985-103564 E-mail: [email protected] Proteases perform fundamental roles in multiple biological processes and are associated with a growing number of pathological conditions that involve abnormal or deficient functions of these enzymes. The availability of the rat genome sequence has opened the possibility to perform a global analysis of the complete protease repertoire or degradome of this model organism. The rat degradome consists of at least 626 proteases and homologs, which are distributed into five catalytic classes: 24 aspartic, 160 cysteine, 192 metallo, 221 serine, and 29 threonine proteases. Overall, this distribution is similar to that of the mouse degradome, but significatively more complex than that corresponding to the human degradome composed of 561 proteases and homologs. This increased complexity of the rat protease complement mainly derives from the expansion of several gene families including placental cathepsins, testases, kallikreins and hematopoietic serine proteases, involved in reproductive or immunological functions. These protease families have also evolved differently in the rat and mouse genomes and may contribute to explain some functional differences between these two closely related species. Likewise, genomic analysis of rat protease inhibitors has shown some differences with the mouse protease inhibitor complement and the marked expansion of families of cysteine and serine protease inhibitors in rat and mouse with respect to human.
    [Show full text]
  • Persistent Elastase/Proteinase Inhibitor Imbalance During
    003 1-3998/89/2604-035 1$02.00/0 PEDIATRIC RESEARCH Vol. 26, No. 4, 1989 Copyright O 1989 International Pediatric Research Foundation, Inc. Printed in U.S.A. Persistent Elastase/Proteinase Inhibitor Imbalance during Prolonged Ventilation of Infants with Bronchopulmonary Dysplasia: Evidence for the Role of Nosocomial Infections H. WALTI, C. TORDET, L. GERBAUT, P. SAUGIER, G. MORIETTE, AND J. P. RELIER Neonatal Intensive Care Unit, Port-Royal Maternity, Paris [H. W., G.M., J.P.R., P.S.]; CNRS, Cellular Biology Center, Ivry [C. T.]; and Department of Biochemistry, Hopital St Vincent-de-Paul, Paris [L.G.] France ABSTRACT. Acute imbalance between elastase and a-1- Premature infants with acute neonatal lung injury, usually proteinase inhibitor (a1Pi) may contribute to the develop- RDS, may develop neonatal chronic pulmonary disease, of which ment of bronchopulmonary dysplasia (BPD). The question the most common form is BPD. The exact pathogenesis of BPD of whether such an imbalance persists in BPD infants still is unresolved, however, many recent reports emphasize the im- requiring mechanical ventilation after 4 wk of life has not portance of inflammatory events (1, 2). Histologic and cytologic been previously addressed. We studied 14 infants still on studies of infants with RDS have shown an early and significant mechanical ventilation at 4 wk of age: nine had BPD and alveolitis which consists of inflammatory cells (PMN and AM) five did not. Weekly (4 to 9 wk) serum and bronchoalveolar (1-4). This alveolitis is prolonged in infants with RDS who lavage (BAL) specimens were taken. alPi and a-2-mac- develop BPD (3).
    [Show full text]