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Biomedical subjects

D Roos

Publications and source records attributed to D Roos.

At least 289 records · Page 16Linked to original sources
PubMed1982

Characterization of the peroxidase in human eosinophils.

Human eosinophil peroxidase is a cationic protein with a higher content of arginine, the enzyme being poorly soluble in water. The purified enzyme is able to carry out the peroxidative chlorination of monochlorodimedon. Like myeloperoxidase the position of the pH optimum of this reaction depends on the ration of the concentrations of chloride and H2O2. Compared to myeloperoxidase the pH optimum is shifted by 0.8 pH unit to more acid pH values. The physiological consequences of the properties of the eosinophil peroxidase are discussed.

Eosinophils↗
PubMed1982

Characterization of hereditary partial myeloperoxidase deficiency.

We studied a family with a partial myeloperoxidase deficiency. The myeloperoxidase in the neutrophils and monocytes of the parents and their two sons had normal spectral properties (determined optically and by EPR). Enzymic characteristics (oxidation of iodide) were indistinguishable from those of normal myeloperoxidase; moreover, immunological identity between the myeloperoxidase in the leukocytes of the family members and normal myeloperoxidase was found. No differences in heat stability were observed. The neutrophils and monocytes of the sons contained 9% to 18% of the myeloperoxidase content of normal cells; the neutrophils and monocytes of the parents contained 45% to 58%. These data suggest either that the parents are heterozygous and the sons homozygous for hereditary partial myeloperoxidase deficiency or that each parent is heterozygous for a different type of myeloperoxidase deficiency and the sons combine both deficiencies. The oxidative metabolism of the neutrophils during phagocytosis was not affected by the myeloperoxidase deficiency. The killing of Staphylococcus aureus was apparently normal. The perforation of Escherichia coli by the neutrophils of the sons, however, was retarded in comparison with normal neutrophils.

Adult↗
PubMed1982

Isolation and partial characterization of a novel subset of human T lymphocytes defined by monoclonal antibodies.

A novel subset of human blood lymphocytes was isolated by means of labelling with monoclonal antibodies and fluorescence-activated cell sorting. In normal individuals, the new subset accounts for about 2% of the blood T lymphocytes. The cells of this subset bind monoclonal antibodies specific for T lymphocytes in general [e.g. OKT3, Hu-Lyt 3(9 . 6) and Leu-22] and they also form E rosettes. However, no binding is seen with monoclonal antibodies to T-lymphocyte subsets (OKT4, OKT8, Leu-2A and Leu-3A). Moreover, the lymphocytes of this new subset express neither Ia antigens nor membrane immunoglobulins. They do not bind OKM1, an antibody against cells of the myelomonocytic lineage that also reacts with natural killer cells, nor do they bind OKT6 or OKT10, specific for thymocyte antigens. The cells have a high specific gravity, a thymocyte-like pattern of lactate dehydrogenase isoenzymes and do not contain terminal deoxynucleotidyl transferase. Although these lymphocytes are viable, also after culture in vitro, and can be stored in liquid nitrogen, they are inert in all functional systems tested: they neither proliferate upon stimulation with mitogens or allogeneic cells, nor do they display suppressor or natural killer cell activity. A patient who was successfully reconstituted by bone marrow transplantation for severe combined immunodeficiency, was found to contain an abnormally high (25%--30%) fraction of these OKT3 positive, OKT4 and OKT8 negative cells among his circulating T lymphocytes.

Antibodies, Monoclonal↗
PubMed1982

Characterization of two subsets of human T gamma cells.

Normal human E rosette-forming, Fc-IgG receptor-bearing cells (so-called T gamma cells) were separated into two functionally different subpopulations. Both subpopulations bind the monoclonal antibody OKM1 (directed against an antigen present also on monocytes and granulocytes). The first subpopulation accounts for about 70% of the total T gamma cell population, does not bind OKT3 (a monoclonal antibody directed against an antigen present on most T lymphocytes), and displays strong killer (K) cell and natural killer (NK) cell activity. The second subpopulation accounts for about 30% of the total T gamma population, binds both OKT3 and OKM1 (confirmed with a double-labeling assay), and displays low K and NK cell activity. Each subset contained less than 10% null cells. Comparison of T gamma cell populations obtained by adherence to a monolayer of IgG-coated human erythrocytes or by rosette formation with these cells revealed that pure T gamma cells with normal killer cell and natural killer cell activity were best obtained with the monolayer technique. Comparison of the enzymic and functional profile of T gamma cells, monocytes and granulocytes, as well as changes during culture of these cells in vitro, failed to indicate a relationship between T gamma cells and cells of the myelomonocytic lineage.

Antibodies, Monoclonal↗
PubMed1982

Phagocytosis and degradation of DNA-anti-DNA complexes by human phagocytes: influence of the antibody class.

DNA-anti-DNA complexes were prepared with igG anti-DNA or IgM anti-DNA and incubated with monocytes and neutrophils from human blood. The capacity of the complexes to induce phagocytosis and degradation of the antigen by the phagocytes varied with the composition of the complexes. Complexes prepared with IgG anti-DNA were readily processed (phagocytosed and degraded) by the cells, provided the complexes were large (precipitating at 3000 x g in 15 minutes). Processing of the complex required the Fc part of the IgG antibody. Complexes prepared with IgM anti-DNA were hardly processed at all, although they were as large as the IgG anti-DNA-containing complexes that were processed. IgM anti-DNA enhanced the processing of IgG anti-DNA-DNA complexes by increasing the size of the complexes.

Antigen-Antibody Complex↗
PubMed1981

Phagocytosis and degradation of DNA-anti-DNA complexes by human phagocytes. I. Assay conditions, quantitative aspects and differences between human blood monocytes and neutrophils.

The uptake in vitro was studied of 3H-labeled DNA-anti-DNA complexes by neutrophils and monocytes from human blood. Complexes were prepared from 3H-labeled circular double-stranded (dS) DNA of bacteriophage PM2 and anti-dsDNA-containing sera from patients with systemic lupus erythematosus. After phagocytosis, cells and medium were separated. The cells were treated with DNase to remove adherent and noningested complexes before the cell-associated radioactivity was counted. Thus, only complexes inside the cells were measured. The medium was analyzed for acid-precipitable radioactivity. In this way, we found that neutrophils only phagocytose the complexes, whereas monocytes phagocytose the complexes and degrade the antigen. In contrast, both types of phagocyte degraded the antigen in tetanus-anti-tetanus complexes. The degradation took place after phagocytosis, inside the cells. The difference in DNA degradation between neutrophils and monocytes correlated with the difference in acid DNase activity of the lysosomal fractions: monocytes contained DNase activity, neutrophils did not. With complexes made from DNA with 131I-labeled anti-DNA, we found that both cell types degraded the antibody. Uptake of complexes and degradation of antigen increased with incubation time and cell concentration and was saturable with respect to complex concentration. The processes were inhibited by 5 mM mono-iodoacetic acid or by low temperatures.

Antigen-Antibody Complex↗
PubMed1981

Phagocytosis and degradation of DNA-anti-DNA complexes by human phagocytes II. Influence of the size of the complexes.

The influence was studied of the size of DNA-anti-DNA complexes on their capacity to induce phagocytosis and degradation by monocytes and neutrophils from human blood. An estimate of the size of the complexes was obtained by precipitation analysis, sucrose-gradient velocity sedimentation and filtration over Nucleopore filters. The results show that only those complexes are processes that precipitate in 15 min at 3000 x g. These complexes have an S value of more than 25000 and a diameter of more than 0.4 micrometer. Likewise, tetanus-anti-tetanus complexes also had to be precipitable in 15 min at 3000 x g to be processed by the phagocytes.

Antigen-Antibody Complex↗
PubMed1981

Large-scale purification and cryopreservation of human monocytes.

Human blood monocytes were purified by a new method capable of handling 3 X 10(9) mononuclear leukocytes, which does not involve adherence of the cells and takes about 3 h to perform. The yield of monocytes is 70%, the purity about 90% and the viability 99%. Monocytes purified by this method were cryopreserved at -196 degrees C. The function of the cells was tested before freezing and after thawing. We found that the capacity of cryo-preserved monocytes to move to the source of a chemotactic gradient, to ingest particles, to mount a respiratory burst during phagocytosis, to kill intracellular bacteria, to lyse anti-D sensitized erythrocytes and to help autologous lymphocytes in a proliferative response to mitogens or antigens, was preserved by 70% or more as compared with non-frozen cells. Thus, cryopreservation of human blood monocytes is possible with maintenance of functional capacities.

Blood Bactericidal Activity↗
PubMed1981

Some enzymatic characteristics of eosinophil peroxidase from patients with eosinophilia and from healthy donors.

Some enzymatic characteristics of human eosinophil peroxidase were compared with those of human myeloperoxidase. Both enzymes catalyzed the oxidation of iodide by hydrogen peroxide. This assay proved to be very sensitive; the activity of 100 eosinophils/ml could be measured. The position of the pH optimum of this reaction was linearly dependent on the logarithm of the iodide/H2O2 ratio. At the same substrate ratio, this optimum was about 1 pH unit higher for eosinophil peroxidase than for myeloperoxidase. This difference may be related to the action of myeloperoxidase inside an acidified phagolysosome as opposed to the extracellular action of eosinophil peroxidase on the surface of certain parasites. Under defined conditions (KI, 1.4 mM; H2O2, 0.18 mM; cetyltrimethylammonium bromide, 0.008% [wt/vol]; pH 6), the activity of eosinophil peroxidase could be measured in a mixed granulocyte suspension independently of myeloperoxidase. Eosinophils from patients with eosinophilia were found to contain as much peroxidase activity as did eosinophils from healthy donors. No enzymatic differences in eosinophil peroxidase were found between the two types of donors.

Cetrimonium↗
PubMed1981

Phagocytosing human neutrophils inactivate their own granular enzymes.

During phagocytosis, neutrophils generate reactive oxygen metabolites and release lysosomal enzymes into the extracellular medium. We have investigated the possibility that these enzyme are inactivated by the oxygen compounds. Phagocytosing neutrophils from 12 patients with chronic granulomatous disease, which do not generate these oxygen metabolites, released two to three times more activity of lysozyme and beta-glucuronidase than did normal neutrophils. This difference proved to be due to a decrease of approximately 20% of the total activity of these enzymes in normal neutrophils, but not in neutrophils of patients with chronic granulomatous disease. This inactivation of enzymes took place during phagocytosis of opsonized zymosan particles as well as during stimulation of normal cells with phorbol myristate acetate. The inactivation was not due to formation of inhibitors. The lysosomal enzymes were not activated when the neutrophils were stimulated under anaerobic conditions. Addition of catalase, superoxide dismutase, or albumin gave no protection against the oxidative damage; reduced glutathione gave partial protection. The oxidative inactivation was more pronounced in the presence of azide. Measurement of the activity and the amount of protein of acid alpha-glucosidase in the cells showed that the specific activity of this enzyme decreased by approximately 50% during 30 min of phagocytosis. This indicates that the inactivation of the lysosomal enzymes takes place in the phagolysosomes, before the enzymes have leaked into the extracellular medium.

Extracellular Space↗
PubMed1981

Extracellular proton release by stimulated neutrophils.

We have tried to elucidate the mechanism of phagosome acidification in human neutrophils. Assuming that phenomena occurring at the plasma membrane reflect reactions in the phagocytic vacuoles, we have stimulated human neutrophils with agents that induce a "respiratory burst," and we have measured the release of protons into the extracellular medium. Phorbol myristate acetate, N-formyl-methionyl-leucyl-phenylalanine and serum-opsonized zymosan particles each caused a rapid release of protons, concomitant with the increase in oxygen consumption. The stimulated release of protons was strictly coupled to the increase respiration of the cells, because inhibition of the respiration of either anaerobiosis, chlorpromazine, or glycolytic inhibitors also inhibited the release of protons. Also, in the presence of the above-mentioned stimulating agents, neutrophils from three patients with chronic granulomatous disease enhanced neither respiration not proton release. In normal cells, the ratio of deltaH+/-deltaO2 was 1.04 +/- 0.19 (mean +/ SD, n = 13). The mechanism of this proton release is not clear. The amount of lactic and carbonic acid produced by stimulated neutrophils was inadequate to explain the amount of protons released. Perhydroxyl radicals were also ruled out as the source of the protons. Because the cells did not release measurable amounts of phosphate ions, a phosphate-hydroxyl-ion antiport was also excluded. Finally, the lack of any effect of uncouplers renders it unlikely that a respiration-driven proton gradient is built up across the plasma membrane.

Granulomatous Disease, Chronic↗
PubMed1981

Effect of ascorbate on abnormal neutrophil, platelet and lymphocytic function in a patient with the Chediak-Higashi syndrome.

A diminished chemotactic response was observed with the neutrophils of a patient with the Chediak-Higashi syndrome, who was not in the accelerated phase of the disease. An abnormally low release of myeloperoxidase from these cells during phagocytosis was also noted; this resulted in a decreased iodination capacity and probably also caused the defect in the intracellular killing of bacteria by the neutrophils. The level of cyclic AMP in these cells was elevated, but decreased after treatment with ascorbate either in vitro or in vivo. During ascorbate therapy, the bactericidal activity of the neutrophils normalized, whereas the chemotactic response remained low. Nevertheless, the patient had significantly less infections during ascorbate therapy. The bleeding tendency, due to a storage-pool disorder of the Chediak-Higashi platelets, was unaffected by treatment with ascorbate. The patient's lymphocytes did not display any activity in antibody-dependent lymphocytotoxicity. This defect was not affected by treatment with ascorbate either.

Adenosine Diphosphate↗
PubMed1981

Reactivity differences among human T cells from blood and lymphoid organs, analysed by limiting dilution: correlation with specific gravity and binding of peanut lectin.

Human T cells from peripheral blood, cord blood, thymus, spleen and lymph node were analysed for their proliferative response capacity to mitogens, for their specific gravity and size, and for their binding capacity of peanut agglutinin. A close correlation was found between these parameters: thymocytes and T cells from spleen were consistently heavier and smaller, and showed a lower proliferative response capacity, than T cells from blood or lymph node. Similarly, within each cell population, the small, heavy cells were least reactive. The limiting-dilution analysis revealed that heavy T cells from peripheral blood contain a lower number of reactive cells than the light peripheral T lymphocytes. Because heavy T cells from the thymus bound more peanut lectin than did light cells, it is speculated that the differences between T cells of high and low specific gravity might reflect differences in maturation level.

Adult↗