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Biomedical subjects

C A Owen

Publications and source records attributed to C A Owen.

At least 19 recordsLinked to original sources
PubMed1995

Inducible binding of bioactive cathepsin G to the cell surface of neutrophils. A novel mechanism for mediating extracellular catalytic activity of cathepsin G.

Catalytically active cathepsin G that is bound to the cell surface of human neutrophils may play a variety of roles in normal neutrophil biology and in pathobiology associated with inflammation. In this study, we describe expression of neutrophil cell surface-bound cathepsin G in response to TNF-alpha and platelet-activating factor (PAF) under conditions in which minimal free release of cathepsin G is detected. TNF-alpha and PAF alone induced modest (two- to threefold) increases in cell surface-bound cathepsin G, but exhibited a marked dose- and time-dependent priming effect for subsequent chemoattractant-induced responses (up to 15- to 25-fold increases in cell surface expression). When optimally primed (TNF-alpha, 100 U/ml, or PAF, 10(-9) M), neutrophils expressed five- to sixfold more cell surface-bound cathepsin G, in comparison with cells exposed to FMLP alone. Priming responses were more rapid with PAF (15 s to 5 min) than with TNF-alpha (1 to 60 min). Optimally primed and FMLP-stimulated neutrophils express approximately 160 ng of catalytically active cathepsin G per 10(6) cells, which represents approximately 11% of the cellular content of unstimulated cells. Cathepsin G binds to the cell surface by a charge-dependent mechanism since: 1) incubation of cells with highly positively charged molecules abrogated agonist-induced up-regulation of the cell surface expression of cathepsin G and 2) cathepsin G was eluted from the cell surface by high concentrations of NaCl. These data indicate that interactions between biologically relevant pro-inflammatory cytokines and chemoattractants serve to markedly up-regulate cell surface-bound cathepsin G. The focused catalytic activity of cell surface-bound cathepsin G may alter endothelial and epithelial barriers, promote thrombogenesis, injure extracellular matrix, and/or facilitate directed migration of neutrophils during inflammation.

Cathepsin G↗
PubMed1995

Multiple genes code for high-molecular-mass rhoptry proteins of Plasmodium yoelii.

We have examined the number of genes coding for a group of high-molecular-mass rhoptry protein(s) in the malaria parasite Plasmodium yoelii, and studied variation in the gene family within the parasite's genome. A region of the genes was amplified using oligonucleotides based on conserved DNA sequences and the products cloned. The sequences could be divided into 7 groups by restriction-fragment-length polymorphism. Further variation was detected by sequence analysis; 11 different sequences were detected in the 16 clones analyzed. The genes in the family were distributed on 6 chromosomes probably at 9 or more loci.

Animals↗
PubMed1995

Cell surface-bound elastase and cathepsin G on human neutrophils: a novel, non-oxidative mechanism by which neutrophils focus and preserve catalytic activity of serine proteinases.

Serine proteinases of human polymorphonuclear neutrophils play an important role in neutrophil-mediated proteolytic events; however, the non-oxidative mechanisms by which the cells can degrade extracellular matrix in the presence of proteinase inhibitors have not been elucidated. Herein, we provide the first report that human neutrophils express persistently active cell surface-bound human leukocyte elastase and cathepsin G on their cell surface. Unstimulated neutrophils have minimal cell surface expression of these enzymes; however, phorbol ester induces a 30-fold increase. While exposure of neutrophils to chemoattractants (fMLP and C5a) stimulates modest (two- to threefold) increases in cell surface expression of serine proteinases, priming with concentrations of lipopolysaccharide as low as 100 fg/ml leads to striking (up to 10-fold) increase in chemoattractant-induced cell surface expression, even in the presence of serum proteins. LPS-primed and fMLP-stimulated neutrophils have approximately 100 ng of cell surface human leukocyte elastase activity per 10(6) cells. Cell surface-bound human leukocyte elastase is catalytically active, yet is remarkably resistant to inhibition by naturally occurring proteinase inhibitors. These data indicate that binding of serine proteinases to the cell surface focuses and preserves their catalytic activity, even in the presence of proteinase inhibitors. Upregulated expression of persistently active cell surface-bound serine proteinases on activated neutrophils provides a novel mechanism to facilitate their egress from the vasculature, penetration of tissue barriers, and recruitment into sites of inflammation. Dysregulation of the cell surface expression of these enzymes has the potential to cause tissue destruction during inflammation.

Cathepsin G↗
PubMed1994

A discrete subpopulation of human monocytes expresses a neutrophil-like proinflammatory (P) phenotype.

We have demonstrated that a discrete and naturally occurring subpopulation of human monocytes expresses a neutrophil-like proinflammatory (P) phenotype. P monocytes constitute 20-30% of the circulating monocyte pool and are characterized by 1) avid adherence to extracellular matrix through high-level cell-surface expression of alpha 5-, beta 1-, and beta 2-integrins; 2) high capacity to produce reactive oxygen species; 3) high content of serine proteinases and alpha 1-proteinase inhibitor; and 4) proteolytic activity against a soluble peptide human leukocyte elastase substrate, [3H]elastin, and solid-phase fibronectin, even in the presence of proteinase inhibitors. However, P monocytes express little or no cell-surface HLA-DR antigen, suggesting that they are unable to participate in specific immune responses. In contrast, the remainder of circulating monocytes have a low proinflammatory potential but contain the population of monocytes with high-level expression of HLA-DR antigen. P monocytes can readily be separated from the remainder of monocytes on the basis of 1) their capacity to adhere to fibronectin; and 2) their absent expression of HLA-DR antigen when flow cytometry or immunomagnetic beads are used. Our data indicate that, when recruited to sites of inflammation, P monocytes can either promote resolution of inflammation or contribute to tissue injury.

Cell Separation↗
PubMed1994

Monocytes recruited to sites of inflammation express a distinctive proinflammatory (P) phenotype.

Only a minor proportion of monocytes responds to chemoattractants. To test the possibility that chemoattractant-responsive monocytes have distinctive functional characteristics, we enriched or depleted monocyte preparations for cells having a proinflammatory (P) phenotype and tested their responses to biologically relevant chemoattractants. We prepared monocyte subpopulations by one of three independent techniques to minimize the chances of artifacts: 1) depletion of P monocytes by adherence to fibronectin; 2) enrichment for P monocytes by negative selection for HLA-DR antigen; and 3) flow cytometric sorting. We measured responsiveness of monocyte subpopulations to N-formyl-Met-Leu-Phe, C5a, zymosan-activated serum, and monocyte chemoattractant protein-1 by three parameters: 1) polarization, 2) actin polymerization, and 3) directed migration. With each chemoattractant and each parameter, there was a striking direct relationship between the responsiveness of the monocyte preparations and their content of P monocytes. Our data indicate that the capacity of monocytes to be recruited rapidly from the vasculature into sites of inflammation is a property of a subpopulation of monocytes with a distinctive, neutrophil-like proinflammatory phenotype.

Actins↗
PubMed1992

Monocyte adherence to fibronectin: role of CD11/CD18 integrins and relationship to other monocyte functions.

Adherence of monocytes to extracellular matrix components is critical for their accumulation at sites of infection. To gain insight into the factors that regulate monocyte recruitment, we have studied monocyte adherence with regard to the regulatory effects of bacterial lipopolysaccharide (LPS) and the mechanisms involved; moreover, we have contrasted the phenotypes of adherent and nonadherent cells. Our results show that only a minor subpopulation of monocytes (20-25%) adhere spontaneously to fibronectin and that LPS stimulated a threefold increase in the proportion of adherent cells. Basal adherence and LPS-stimulated adherence of monocytes to fibronectin were substantially mediated by CD11/CD18 integrins. Further studies revealed that spontaneously adherent monocytes were 14-fold more actively phagocytic, released 1.6-fold more superoxide anion, and contained 20-fold more peroxidase activity than nonadherent cells, whereas LPS-adherent cells had an intermediate phenotype. These results indicate that LPS may enhance the accumulation of monocytes with an antimicrobial phenotype and thereby promote resolution of tissue infection.

Antigens, CD↗
PubMed1992

Increased adherence of monocytes to fibronectin in bronchiectasis. Regulatory effects of bacterial lipopolysaccharide and role of CD11/CD18 integrins.

Regulated adherence of monocytes to extracellular matrix is a prerequisite for accumulation of mononuclear phagocytes during pulmonary infection and inflammation. We have obtained monocytes from patients with an inflammatory lung disease (bronchiectasis) and from control subjects and have compared their adherence to fibronectin. Spontaneous adherence of monocytes from the control subjects was 20 +/- 2%, whereas that of patients' cells was markedly higher and correlated with the severity of airway inflammation: 65 +/- 5% and 40 +/- 8% in patients with purulent and mucoid sputum, respectively. Endotoxin and cytokines from areas of airway disease are likely to be responsible for the observed monocyte activation, since: (1) endotoxin was detectable in all of the patients but in none of the control subjects; (2) LPS produced a dose-related increase in adherence of normal monocytes in vitro (maximal 65 +/- 2% adherence at 1 microgram/ml of LPS); (3) recombinant cytokines and LPS produced additive effects on monocyte adherence in vitro. The adherence of the patients' monocytes to fibronectin was substantially mediated by CD11/CD18 integrins, via both RGD-dependent and RGD-independent mechanisms. These data indicate that signals arising from foci of infection and inflammation can influence the adherence of monocytes, and they are likely to be determinants of the accumulation of mononuclear phagocytes in the lungs of patients with bronchiectasis.

Adult↗
PubMed1991

Structure of Sm25, an antigenic integral membrane glycoprotein of adult Schistosoma mansoni.

Sm25 is the principal antigen recognised by antibodies from mice protectively vaccinated with isolated tegumental membranes of adult Schistosoma mansoni. The full-length amino acid sequence of this protein has been deduced from the sequence of two cDNAs, one isolated by screening a cDNA library and the other, including the 5' end of the gene, amplified directly from adult worm RNA using the polymerase chain reaction. The predicted sequence represents a nascent polypeptide of Mr 21,500. Following cleavage of a predicted signal sequence, the Mr of the resulting polypeptide is 17,600. The polypeptide contains 2 potential sites for N-linked glycosylation and a hydrophobic domain at the C-terminus that could facilitate membrane association. Analysis of the mature gene product confirmed that Sm25 is an N-glycosylated integral membrane protein and that the Mr of the deglycosylated polypeptide is between 15,000 and 20,000.

Amino Acid Sequence↗
PubMed1990

Historical account of tests of hemostasis.

Although the fact that blood clotted when it was released from the body was well known to Hippocrates, Plato, Aristotle, Celsus, and Galen, it was not believed to have any physiologic or pathologic significance. Petit, a surgeon, recognized in the 1730s that clotting was hemostatically important in amputations. Finally, the mechanism of clotting began to be studied by Buchanan (1838), who recognized thrombin; Hammarsten (1875), who purified fibrinogen; and Arthus (1890), who discovered the need for calcium. The fact that platelets existed and had a hemostatic function was developed in the 1800s. Not until the late 1940s did the explosion in the discovery of new clotting factors begin; they now number up to Factor XIII, plus many more that have no Roman numeral designation. Discovery of clotting factors led to their assays. The use of whole blood clotting times was improved by measuring the clotting times of plasma. This was followed by the partial thromboplastin time (PTT) and the activated partial thromboplastin time (APTT). The prothrombin time became an important laboratory test. Specific factors were assayed by progressively more specific tests. These included elements of the fibrinolytic and inhibitory systems. Platelets were counted and their functions measured by bleeding times, clot retraction, adhesion, and aggregation. The diagnosis of hemostatic disorders has improved in parallel with the discoveries of new factors and the development of their precise assays.

Blood Coagulation Tests↗
PubMed1989

Kappa Delta Award paper. Tissue fluid pressures: from basic research tools to clinical applications.

The two basic research tools developed to measure tissue fluid pressure (wick catheter) and osmotic pressure (colloid osmometer) have undergone extensive validation and refinement over the past 20 years. Using these techniques, basic science investigations were undertaken of edema in Amazon reptiles, pressure-volume relations in animals and plants, adaptive physiology of Antarctic penguins and fishes, edema in spawning salmon, tissue fluid balance in humans under normal conditions and during simulated weightlessness, and orthostatic adaptation in a mammal with high and variable blood pressures--the giraffe. Following and sometimes paralleling this basic research have been several clinical applications related to use of our colloid osmometer and wick technique. Applications of the osmometer have included insights into (a) reduced osmotic pressure of sickle-cell hemoglobin with deoxygenation and (b) reduced swelling pressure of human nucleus pulposus with hydration or certain enzymes. Clinical uses of the wick technique have included (a) improvement of diagnosis and treatment of acute and chronic compartment syndromes, (b) elucidation of tissue pressure thresholds for neuromuscular dysfunction, and (c) development of a better tourniquet design for orthopaedics. This article demonstrates that basic research tools open up areas of basic, applied, and clinical research.

Animals↗
PubMed1989

Evaluation of preoperative hematology-coagulation screening in liver transplantation.

We retrospectively reviewed the results of preoperative hematology-coagulation studies in 66 patients who underwent orthotopic liver transplantation-24 with the primary diagnosis of chronic active hepatitis (CAH), 22 with primary sclerosing cholangitis (PSC), and 20 with primary biliary cirrhosis (PBC). The mean prothrombin time was above normal in all three diagnostic groups, patients with CAH having the highest values. The mean activated partial thromboplastin time was normal in patients with PSC or PBC but elevated in those with CAH. Fibrinogen levels were above normal in patients with PBC but decreased in 1 patient (5%) with PSC and 10 (42%) with CAH. Mean platelet counts were below normal in 68% and 55% of patients with PSC and PBC, respectively, but in 96% of those with CAH. The mean Ivy bleeding time was normal in patients with PSC or PBC but prolonged in those with CAH. Patients with PSC or PBC had normal mean activity levels of factors II, V, VII, IX, and X, whereas those with CAH had below normal mean values for factors II and VII. The antithrombin III activity level was normal in patients with PSC or PBC but reduced in those with CAH. Thus, patients with CAH have a greater derangement in results of clotting studies in comparison with those who have PSC or PBC, but the use of blood did not differ among the three diagnostic groups.

Antithrombin III↗
PubMed1987

Hemostatic evaluation of patients undergoing liver transplantation.

A detailed coagulation and thromboelastographic study was done on the first 50 liver transplantation procedures performed at the Mayo Clinic between March 1985 and June 1986. Most of the patients suffered from primary sclerosing cholangitis, primary biliary cirrhosis, or chronic active hepatitis. Seven patients required a second liver transplantation, and six patients died, none intraoperatively. Most of the patients had distorted hemostatic mechanisms preoperatively, as would be expected because the liver generates most of the clotting factors. The outstanding exception was factor VIII, which was usually in the high-normal range or even more elevated. Substantial deterioration of coagulation factors occurred regularly during reperfusion of the donor liver. In some instances, this trend was corrected within 1 hour, but platelet counts continued to decrease, and some coagulation factors rebounded only partially. Because thromboelastographic tracings are quickly available to the liver transplant team and because they tend to forewarn of impending hemostatic problems, we believe that thromboelastography is a reasonably effective procedure for monitoring coagulation during liver transplantation.

Blood Coagulation Factors↗
PubMed1985

A clinical evaluation of automated chromogenic tests as substitutes for conventional prothrombin time and activated partial thromboplastin time tests.

Automated procedures involving a chromogenic substrate sensitive to thrombin-sarcosine-Pro-Arg p-nitroanilide were compared with conventional tests for prothrombin times and activated partial thromboplastin times (APTT) and with specific assays for factors V, VII, VIII, IX, X, XI, and XII. The reproducibility and sensitivity of the chromogenic tests were compared with those of the clotting tests. Further, we have confirmed that the chromogenic test for APTT is sensitive to factor VII deficiency, unlike the clotting test for APTT. This might be an advantage in monitoring orally anticoagulated patients. The ready availability of the automated equipment for performing the chromogenic tests suggests their potential for routine use. However, some discrepant results in certain patients with liver disease and in others with factor VIII inhibitors warrant caution.

Autoanalysis↗