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Biomedical subjects

K Ohlsson

Publications and source records attributed to K Ohlsson.

At least 253 records · Page 14Linked to original sources
PubMed1977

Localization of chymotrypsin-like cationic protein, collagenase and elastase in azurophil granules of human neutrophilic polymorphonuclear leukocytes.

The subcellular localization of granulocyte collagenase, elastase and chymotrypsin-like cationic protein was determined using velocity centrifugation of cytoplasmic granules of human polymorphonuclear leukocytes. The proteases were assayed by immunochemical and enzymatic methods. Measurements of lactoferrin and myeloperoxidase distinguish exactly between constituents of specific and azurophil granules. Collagenase, elastase and chymotrypsin-like cationic proteins showed an almost identical sharp and unimodal distribution. They co-sedimented with myeloperoxidase demonstrating that these enzymes are localized exclusively in the azurophil granules.

Adult↗
PubMed1977

Localization of a low molecular weight protease inhibitor to tracheal and mixillary sinus mucosa.

Normal human bronchial secretion contains protease inhibitors from plasma and an acid-stable, low molecular weight inhibitor which strongly inactivates granulocyte elastase and chymotrypsin-like cationic proteins. The distribution of this bronchial protease inhibitor in tracheal mucosa was analyzed by an indirect immunoperoxidase method. Strong peroxidase staining was obtained in the columnar ciliated epithelium of the trachea and in the sero-mucus glands. The findings support a local mucosal production of the inhibitor. Maxillary sinus mucosa was also positively stained for the inhibitor.

Animals↗
PubMed1977

Elastases from human and canine granulocytes, II. Interaction with protease inhibitors of animal, plant, and microbial origin.

Inhibitors of animal, plant, and microbial origin were tested against human and canine granulocytic elastases. The trypsin-chymotrypsin inhibitors from dog submandibular glands, from soybeans (Bowman-Birk) and from chickpeas show strong interaction with these proteases (Ki = 10(-8) - 10(-9)M). The trypsin-kallikrein inactivator of bovine organs (Trasylol) is not active against granulocytic elastases or against human granulocytic cathepsin G. Elastatinal, a specific inhibitor of elastases, isolated from actinomycetes (Streptomyces griseoruber), forms stable complexes with elastase from human (Ki = 6.2 X 10(-6)M) and canine granulocytes (Ki = 1.1 X 10(-6)M). A possible therapeutic application of these inhibitors for the inactivation of granulocytic proteases, which are able to degrade connective tissue in different pathological states, is discussed.

Animals↗
PubMed1977

Isolation and partial characterization of a low molecular weight acid stable protease inhibitor from human bronchial secretion.

An acid stable protease inhibitor was isolated from human bronchial secretion. Two important stages of the purification procedure were affinity chromatography on trypsin bound to Affi-Gel 10 and ion-exchange chromatography on SP-Sephadex C-50. The isolated inhibitor appeared as a single band on analytical disc electrophoresis and eluted as a homogeneous protein peak on gel filtration on Sephadex G-75 corresponding to a molecular weight of about 10500. Amino acid analyses showed no tryptophan or histidine and as N-terminal amino acid tyrosine. No glucosamine or galactosamine was detected. The results of the analyses suggest that the purified inhibitor is identical to the low molecular weight trypsin-chymotrypsin inhibitor of human seminal plasma (HUSI-I).

Bronchi↗
PubMed1977

Isolation and characterisation of a low molecular weight inhibitor (of chymotrypsin and human granulocytic elastase and cathepsin G) from leeches.

Two protein proteinase inhibitors were isolated and purified from the leech Hirudo medicinalis by means of gel filtration and ion-exchange chromatography. They inhibit chymotrypsin, subtilisin and the granulocytic neutral proteases elastase and cathepsin G. They proved to be homogeneous in polyacrylamide and dodecylsulfate gel electrophoresis and by end group analysis; only threonine was found as N-terminal amino acid residue using the dansylation technique. These inhibitors, which we call eglins, are stable in neutral and weakly acid (pH 3) solutions and resist non-specific proteolysis. From the amino acid compositions, a molecular weight of 6 600 - 6 800 is calculated for both inhibitory proteins, which is in good agreement with a value of about 6000 estimated by dodecylsulfate electrophoresis. The eglins contain an unusually large amount of hydrophobic amino acid residues but no methionine, isoleucine or--a rarity--cysteine residues or disulfide bridges. To our knowledge, the eglins are the first examples of proteinase inhibitors of the protein type which are not stabilized by disulfide bridges.

Amino Acids↗
PubMed1977

Comparison of the reactions of neutral granulocyte proteases with the major plasma protease inhibitors and with antiplasmin.

Reaction mixtures of human serum and increasing amounts of granulocyte collagenase, elastase and chymotrypsin-like enzyme were studied by crossed immunoelectrophoresis utilizing antibodies against alpha1-antitrypsin, alpha1-antichymotrypsin, and antiplasmin. The increasing complex formation of alpha1-antitrypsin and alpha 1-antichymotrypsin with the different granulocyte proteases was not accompanied by any changes in the electrophoretic mobility or precipitate pattern of antiplasmin until the protease binding capacity of serum was saturated. The antiplasmin component in the reaction mixtures of human serum and granulocyte collagenase or elastase was not precipitated by antibodies against the proteases. The results indicate that none of the granulocyte proteases are bound by antiplasmin and that these enzymes do not activate plasminogen in serum.

Antibodies↗
PubMed1977

Elevated fecal levels of endogenous pancreatic endopeptidases after antibiotic treatment.

Feces from normal and antibiotic-treated persons were analyzed for the content of immunoreactive trypsin and elastase. In the control group the mean concentration of immunoreactive trypsin was 13 microgram per g feces as compared to 147 in the antibiotic-treated group. Elastase was demonstrable in only 3 of 30 samples in the control group but in 20 of 26 in the antibiotic-treated group. The decreased inactivation of pancreatic proteases must depend on an altered intestinal microflora. The results suggest that reestablishment of a normal enteric flora may take months after the short time oral administration of antibiotics.

Anti-Bacterial Agents↗
PubMed1977

Immunochemical quanitation of pancreatic endopeptidases in the intestinal contents of germfree and conventional rats.

Two electrophoretically distinct trypsins and chymotrypsins and an elastolytic enzyme were isolated from rat pancreatic juice. Rabbit antisera against these enzymes were produced, and with an immunochemical technique the trypsins, chymotrypsins, and elastase were studied in the intestinal contents of conventional and germfree rats. In both types of rat the anionic trypsin and chymotrypsin were the most abundant and found in higher concentrations in the distal than in the proximal small intestine. The cecal and fecal concentrations of anionic trypsin were markedly higher in the germfree rat when compared to the conventional rat. Cymotrypsin was undetectable in the large intestine of either the conventional or germfree rat when this technique was used. Immunoreactive elastase was found in greater amounts in the distal small intestine, and high concentrations were demonstrated in the cecal contents and feces of the germfree rat. In contrast, no immunoreactive elastase was detected in the large intestine of the conventional rat. Gel filtration indicated that the immunoreactive anionic trypsin and elastase found in fecal extracts were of about the same molecular size as the native enzymes. The findings suggest that the intestinal microflora is instrumental in the inactivation and degradation of pancreatic trypsin and elastase but not chymotrypsin.

Animals↗
PubMed1977

Neutral proteases of human granulocytes. IV. Interaction between human granulocyte collagenase and plasma protease inhibitors.

Purified human granulocyte collagenase was inactivated by serum through the formation of complexes with alpha 1-antitrypsin and alpha 2-macroglobulin. A molar combining ratio of 1:1 was observed for each inhibitor. The affinity of alpha 2-macroglobulin was about 10 times that of alpha 1-antitrypsin for granulocyte collagenase. The molar concentration of alpha 1-antitrypsin in the blood exceeds that of alpha 2-macroglobulin by about 12 times, so that the inhibitors may be equally important for defence against granulocyte collagenase.

Granulocytes↗
PubMed1976

alpha1-Antichymotrypsin interaction with cationic proteins from granulocytes.

Human granulocytes contain cationic proteins with chymotrypsin-like activity. These proteases showed a higher relative affinity for alpha1-antichymotrypsin than for alpha1-antitrypsin but the highest affinity for alpha2-macroglobulin. The complexes between cationic protein and alpha1-antichymotrypsin migrate as beta-globulin on agarose gel electrophoresis.

Chymotrypsin↗
PubMed1976

Isolation and partial characterization of elastase from dog granulocytes.

1. An elastolytic enzyme has been isolated from dog granulocyte leukocytes. The purification procedure included preparation of the granula fraction, chromatography on Sephadex G-75 and ion-exchange chromatography on SP-Sephadex C-50 at pH 6.0. 2. The elastase isolated was homogeneous in analytical disc electrophoresis and showed in sodium dodecylsulfate electrophoresis a single protein component with the molecular weight of 24800. The enzyme lacked tyrosine and lysine and the N-terminal amino acid was phenylalanine. No carbohydrate or sialic acid were detected. 3. The dog granulocyte elastase showed similar activities as human granulocyte elastase on elastin and fibrin. The Km value for 3-carboxypropionyl-L-alanyl-L-alanyl-L-alanyl-p-nitroanilide was 2.50 mM and the pH optimum 8.5. The elastase preparation obtained was 99.5% active as judged from active-site titration. 4. The enzyme is a cationic protein and shows pronounced trailing on agarose gel electrophoresis. 5. A monospecific antiserum against the purified enzyme was produced in rabbits.

Amino Acids↗
PubMed1976

Demonstration and semiquantitative determination of complexes between various proteases and human alpha2-macroglobulin.

Complex formation in vitro between human alpha2-macroglobulin and the human proteases cationic trypsin, chymotrypsin, plasmin and granulocyte elastase and collagenase was clearly visualized by the use of thin-layer electrofocusing in polyacrylamide gel followed by electrophoresis in agarose gel containing antibodies against human alpha2-macroglobulin. The technique permits semi-quantitative determination of the amount of complex and can demonstrate the formation of complexes between alpha2-macroglobulin and protease in vivo in ascitic fluid in peritonitis.

Binding Sites↗
PubMed1976

The disappearance of enzyme-inhibitor complexes from the circulation of man.

1. Complexes of human trypsin and human granulocyte elastase with alpha1-anti-trypsin and alpha2-macroglobulin were isolated and injected intravenously into human volunteers. 2. The elimination of alpha2-macroglobulin complexes with trypsin and elastase followed single-exponential functions with half-lives of 9 and 12 min respectively. The complexes showed no tendency to dissociate. 3. Complexes of alpha1-anti-trypsin with trypsin persisted in the circulation much longer, with a half-life of 3-5 h; complexes of alpha1-anti-trypsin with elastase had an intermediate half-life of 1 h. 4. Dissociation was observed of alpha1-anti-trypsin complexes with transfer of trypsin and elastase to alpha2-macroglobulin. 5. Dialysable radioactivity appeared in the urine soon after the injection of alpha2-macroglobulin complexes, which suggested a breakdown of complexes by cells in the reticuloendothelial system. Radioactivity over the liver achieved maximum values within 30-40 min after the injection of alpha2-macroglobulin complexes but not until 50-70 min after the injection of alpha1-anti-trypsin comlexes. 6. These results support the concept of a key position for alpha2-macroglobulin in the protective mechanisms against endogenous proteases.

Binding Sites↗
PubMed1976

Effects of granulocyte neutral proteases on complement components.

The proteolytic effects of collagenase and elastase from human granulocytes were investigated on the human complement components C3 and C5 in serum and with purified components. The conversion of C3 was analyzed with crossed immunoelectrophoresis as described by Ganrot, and the conversion of C5 was detected with immunoelectrophoresis according to Scheidegger's method. Collagenase converted C3 to C3b but had no detectable effect on C5. Elastase converted C3 to C3b and converted C5 to a C5b-like fragment. The proteolysis by collagenase and elastase in serum was not detectable until the molar ratio for enzyme to the protease inhibitors alpha1-antitrypsin and alpha2-macroglobulin was greater than 1.

Complement C3↗
PubMed1976

Purification and partial characterization of human pancreatic elastase.

1) An elastolytic enzyme has been isolated from human pancreatic juice. The concentration of the enzyme was about 0.6 mg/ml corresponding to about 10% of the protein concentration of the juice. 2) The purification procedure included chromatography on Sephadex G-25 followed by ion-exchange chromatography on SP-Sephadex C-50 at pH 6.0, affinity chromatography on Trasylol-Sepharose 4-B and a final ion-exchange chromatography on SP-Sephadex at pH 7.6. The yield was about 50%. 3) The elastase isolated was homogeneous in analytical disc electrophoresis and showed a single protein component with a molecular weight of 26 300 in dodecylsulfate-electrophoresis. There are similarities in the amino acid compositions of human and porcine pancreatic elastases. 4) The human enzyme has a lower activity on elastin than porcine elastase but similar activities on casein and fibrin. The Km value for Boc-Ala-ONp was 5.13 X 10(-4) M. The elastase isolated was 98% active, as judged from active site titration. 5) The results of immunodiffusion studies of activated pancreatic juice and duodenal juice with specific elastase-directed antibodies indicate that the purified enzyme was the single elastolytic enzyme present. 6) It is a cationic protein without any carbohydrate. The activation of the pro-elastase resulted in the formation of an active enzyme with a higher isoelectric point than the zymogen.

Amino Acids↗
PubMed1976

Elastases from human and canine granulocytes, I. Some proteolytic and esterolytic properties.

The lysosome-like granules of human and canine granulocytes contain an enzyme with elastinolytic activity. The enzymatic behaviour of these elastases was further characterized using the protein substrates elastin-orcein and azocasein and the synthetic substrates tert.-butyloxycarbonyl-alanine p-nitrophenylester (Boc-Ala-ONp) and 3-carboxypropionyl-L-alanyl-L-alanyl-L-alanine p-nitroanilide (Suc-Ala3-NHNp) in photometric assays. The affinities of the granulocyte elastases and of porcine pancreatic elastase to these substrates are very similar, e.g. human granulocyte elastase: KM (Boc-Ala-ONp) = 0.35mM, KM (Suc-Ala3-NHNp) = 1.25mM, porcine pancreatic elastase: KM (Boc-Ala-ONp) = 0.3mM, KM (Suc-Ala3-NHNp) - 1.15mM. The most convenient substrate for the assay of human and dog granulocyte elastases and for kinetic measurements with these enzymes is Suc-Ala3-NHNp. Using this substrate, the dissociation constant of the complex of human granulocyte elastase with human alpha1-antitrypsin could be determined (Ki = 3.5 x 10(-10)M).

Alanine↗