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Biomedical subjects

J Hed

Publications and source records attributed to J Hed.

At least 73 records · Page 4Linked to original sources
PubMed1989

Characteristics of alveolar cells and soluble components in bronchoalveolar lavage fluid from non-smoking aluminium potroom workers.

Aluminium potroom workers have been reported to develop severe pneumoconiosis and bronchial hyperreactivity. The influence of inhalation of aluminium oxide and fluorides on the alveolar milieu was studied by bronchoalveolar lavage (BAL) in 14 male non-smoking potroom workers; 28 non-smoking healthy volunteers served as controls. The total numbers, concentrations, and proportions of various alveolar cells did not differ between the groups. The concentrations of albumin and fibronectin in BAL fluid were significantly higher (p less than 0.01 for both) in the exposed workers, reflecting an increased alveolar capillary permeability and an activation of alveolar macrophages (AMs). The concentration of angiotensin converting enzyme, another AM marker, was, however, decreased (p less than 0.01) in the workers. The concentration of hyaluronan, a fibroblast marker, did not differ between the groups. AMs from workers had a decreased capacity (p less than 0.05) to interact with yeast C3b particles but not to ingest them. The expression of HLA-DR and OKM1 on the cell surfaces of AMs were equal in the two groups. The BAL findings were not accompanied by restrictive lung disease in the workers. The fact that only a discrete alveolitis was found in the potroom workers may be due to a low grade of exposure to alumina and fluorides and to frequent use of respiratory protection equipment.

Adult↗
PubMed1989

Quantitative rather than qualitative differences between monocytes with respect to IgE Fc receptor expression as studied by flow cytofluorometry.

In this study we have used aggregated human IgE, conjugated with fluoroisothiocyanate, as a ligand in flow cytofluorometry to detect the low-affinity Fc receptor for IgE on human leukocytes. The percentage of IgE-binding monocytes increased up to 44%, in a dose-related manner, as the concentration of aggregated IgE in the fluid phase increased to the highest concentration (300 micrograms/ml) used in this study. No such dose-related increase was shown for granulocytes (1.8% IgE-binding cells) or lymphocytes (2.1% IgE-binding cells). The IgE-binding was inhibited by IgE but not by IgG, whereas the opposite effect was obtained for IgG binding. We could not verify a distinct IgE-binding subpopulation of monocytes in peripheral blood. The results rather indicate that most monocytes bind IgE to a variable degree depending on the concentration of the ligand in the fluid phase.

Antigens, Differentiation, B-Lymphocyte↗
PubMed1989

Autofluorescence in human alveolar macrophages from smokers: relation to cell surface markers and phagocytosis.

Flow cytometry was used to study the influence of smoking histories on autofluorescence, expression of surface markers, and phagocytic ability in alveolar macrophages (AM) recruited by bronchoalveolar lavage (BAL) from healthy smokers (n = 13) and nonsmokers (n = 13). Alveolar macrophages have an autofluorescence that can be quenched by a recently developed technique. In the present study, this technique was used in combination with flow cytofluorometry. Alveolar macrophages from smokers (mean 10.6 +/- 7.6 pack-years) showed a significantly (p less than .001) increased autofluorescence compared to nonsmokers. This autofluorescence was associated with an increased complexity of the cells but not with altered cell volumes. No correlation was seen between the mean fluorescence intensity and the cigarette consumption among smokers. Despite the difference in autofluorescence, no altered expression of surface markers known as markers of cell activation (HLA-DR, CR3) was detected in AMs from smokers compared to nonsmokers. The functional ability of AMs to ingest C3b-coated particles analyzed with a fluorescence quenching assay did not differ between the groups. The lack of correlation between the cigarette consumption and the autofluorescence suggests a maximal fluorescence intensity in the present population of smokers. The biological mechanism behind this autofluorescence needs to be further investigated.

Adult↗
PubMed1988

Enumeration of IFN-gamma-producing cells by flow cytometry. Comparison with fluorescence microscopy.

A new intracytoplasmic immunofluorescence staining to detect and quantify human interferon-gamma (IFN-gamma)-producing cells by means of flow cytometry is described. Mononuclear leukocytes, stimulated in vitro to produce IFN-gamma, were fixed and made permeable to antibodies by sequential exposure to paraformaldehyde and the detergent n-octyl-glucoside. Cytoplasmic IFN-gamma was demonstrated by indirect immunofluorescence using IFN-gamma-specific mouse monoclonal antibodies. The staining exhibited a very characteristic morphology and was localized in the Golgi apparatus. An excellent agreement between the enumeration of cytoplasmic IFN-gamma-positive cells by immunofluorescence microscopy and flow cytometry was noted. However, the latter has the advantage of a standardized control, is less labor consuming and is observer independent.

Adult↗
PubMed1988

The expression of CR1 and CR3 on non-modulated and modulated granulocytes of healthy blood donors as measured by flow cytofluorometry.

Complement receptors, CR1 and CR3, on neutrophils increase their cellular spontaneously at 37 degrees C or after mechanical stress during the cell preparation. We have established a cell preparation procedure and a cytofluorometric immunoassay method to evaluate the receptor expression in vivo in this study. The expression of CR1 and CR3 was studied after haemolysis in NH4Cl at different temperatures and incubation intervals. It was shown that cell preparation and receptor analysis must be performed at 15 degrees C or lower to avoid up-regulation of the receptor structures. Two minutes' incubation at 20 degrees C was sufficient to modulate the cells in this regard. Granulocytes from healthy blood donors were analysed and the mean fluorescence intensity (MFI), which reflects the number of receptors on the cell surface, showed a normal distribution for the CR1 (n = 158) and CR3 (n = 76) expression in the healthy population. The MFI of the two receptors showed a correlation (r = 0.71). Granulocytes from all donors could be modulated at 37 degrees C to a similar degree for both receptors (r = 0.76), despite the fact that they are supposed to be mobilized from different intracellular pools. A group of patients with localized inflammatory processes, such as sinusitis, differed from the healthy controls. In this group, the CR1 expression was not normally distributed, but 15 out of 26 patients (58%) had granulocytes with a CR1 expression exceeding the mean +/- SD of the normal population.

Adolescent↗
PubMed1987

The use of fluorescence quenching in flow cytofluorometry to measure the attachment and ingestion phases in phagocytosis in peripheral blood without prior cell separation.

Flow cytofluorimetry identifies and quantifies cell markers of different leukocyte subpopulations by combining cytofluorimetry with the differences in the light scattering properties of the leukocytes in mixed populations. In the phagocytic assay, reported in this paper, the experimental conditions were selected in such a way that it was possible to analyse the phagocytic function of granulocytes in peripheral blood without time-consuming cell separation. The percentage of phagocytosing granulocytes was not dependent on the concentration of granulocytes at the selected incubation time and particle (yeast-C3b) concentration. Furthermore, it was possible to adapt a previously described fluorescence quenching technique (FQ method) to differentiate between attachment and ingestion. Crystal violet, originally used in the FQ method, could not be used in this assay due to its lysomotropic effect. Trypan blue at a concentration of 0.25 mg/ml or higher at pH 4.5 showed a plateau effect in fluorescence quenching indicating an effect on attached but not ingested particles. This assay offers a simple technique to screen the functional properties of phagocytic cells in peripheral blood.

Cell Membrane↗
PubMed1987

Hydrocortisone both decreases the up-regulation of complement receptors CR1 and CR3 and the ingestion process of human granulocytes.

We have studied the effect of hydrocortisone on the complement receptor expression (CR1 and CR3) of human granulocytes during the up-regulation phase and the following stable period when exposed to N-formyl-methionyl-leucyl-phenylalanine (fMLP) or the medium alone at 37 degrees C. The receptor expression was also correlated to the C3bi-mediated phagocytosis with special reference to attachment and ingestion. The results showed that after incubation with fMLP the increased expression of CR1 and CR3 is accompanied by an increased ingestion, but not attachment, of complement-opsonized yeast particles. This increase was significantly lower with regard both to attachment and ingestion, as well as receptor expression if hydrocortisone was present during the up-regulation phase. However, the addition of hydrocortisone after the up-regulation of fMLP-treated granulocytes decreased the ingestion of particles but not the CR1 and CR3 expression. The results indicate a membrane effect of hydrocortisone that affects both the receptor mobilization and ingestion mechanism of the phagocytes.

Granulocytes↗
PubMed1985

The role of negative charge in the complement-dependent dissociation of IgG-mediated aggregation.

The IgG-mediated aggregation of yeast particles was used as a model to study the dissociating effect of human serum complement on immune aggregates. A 92% decrease in non-aggregated particles was obtained by coating particles with IgG. After incubation of the aggregates in normal human serum (NHS) at 37 degrees C the number of free particles increased almost 10 times. The effect could be obtained with ethyleneglycol-bis-(beta-aminoethylether)-N,N'-tetraacetic acid-containing serum but not ethylenediaminetetraacetic acid-containing serum, indicating the importance of the alternate pathway of complement activation. Treatment with NHS did not release anti-yeast IgG from the particles, indicating that disruption of antigen-antibody bonds did not explain the phenomenon. The negative charge of the different particles was studied by measuring the interaction to diethylaminoethyl-Sephacel beads. Compared with non-coated and IgG-coated particles, the NHS-treated particles were not released until the ionic strength increased to 200 mM NaCl or the pH decreased to 4.4, indicating a strong negative charge of NHS-treated particles. We propose that complement increases the negative charge of immune complexes, thereby inducing repulsive forces that counteract Fc-Fc interactions and increase the solubility of the complexes.

Antibodies, Fungal↗
PubMed1985

Erythrocyte enhancement of C3b-mediated phagocytosis by human neutrophils in vitro: a combined effect of the erythrocyte complement receptors CR1 and erythrocyte scavengers to reactive oxygen metabolites (ROM).

Human erythrocyte CR1 receptors have been shown to bind complement-fixing immune complexes and, thus, facilitate their elimination from the circulation. The autotoxic effect of free radicals released from phagocytes during phagocytosis can be alleviated by scavengers like catalase and superoxide dismutase. Erythrocytes are known to contain these antioxidants. This study showed that 74% of opsonized yeast particles adhered to human erythrocytes. No difference was seen between yeast opsonized with C3b and yeast opsonized with both IgG and C3b. This adherence was due to the C3b receptor (CR1), as monoclonal antibodies against the CR1 receptor could abrogate the adherence. The yeast phagocytosis by neutrophils was increased by 15% when yeast-C3b was used, and by 34% when yeast-IgG/C3b was used in the presence of human red blood cells. The increase of phagocytosis was not seen when rat erythrocytes (lacking CR1) were present. The cytochrome c reduction decreased with the presence of human erythrocytes during phagocytosis, indicating a scavenging effect on the superoxide anions. The addition of scavengers or erythrocyte lysate, instead of erythrocytes, enhanced phagocytosis of yeast-IgG/C3b to at least the same extent as the erythrocytes. These observations suggest that human erythrocytes primarily enhance phagocytosis through the scavenging effect of those erythrocytes which are concurrently attached with the prey through its CR1 receptor, and then attached to the PMN.

Complement C3b↗
PubMed1984

Myeloperoxidase reduces the opsonizing activity of immunoglobulin G and complement component C3b.

The effect of myeloperoxidase, hydrogen peroxide (H2O2) and a halide (Cl) on the opsonizing molecules in immunoglobulin G (IgG) and complement factor C3b was assayed. At concentrations of the enzyme (1 microgram/ml) that can be found in the extracellular fluid during inflammation, the myeloperoxidase-H2O2-Cl system inhibited the opsonizing effect of IgG and C3b measured as phagocytic uptake and superoxide generation. The effect was related to the enzymatic peroxidative activity of the protein. The presence of albumin (10 mg/ml) reduced the effect of myeloperoxidase with 10-20%. Taurine, which in the presence of myeloperoxidase-H2O2-Cl forms hydrophilic chloramines, and D-penicillamine, which scavenges HOCl, neutralize the inhibitory effect of myeloperoxidase. This suggests that either hypochlorous acid or lipophilic chloramines may exert its effect by oxidizing free sulphydryl groups exposed on the opsonizing ligands. Since the myeloperoxidase-H2O2-halide system also affects chemotactic factors, leukotrienes, proteinases and membrane receptors, the system may in several ways affect the development of the inflammatory response.

Complement C3b↗
PubMed1984

Chemotaxis of polymorphonuclear leukocytes in response to surface-bound complement-derived chemoattractants generated in situ.

Polymorphonuclear leukocytes (PMNLs) were allowed to migrate on slides with fixed yeast particles dotted about on the surface. Locomotion was quantified by counting the number of yeast particles in association with a PMNL. Yeast particles that differed in their ability to consume hemolytic complement differed also in their ability to generate chemoattractants. Addition of a complement source to yeast particles able to activate the complement system resulted in a chemotactic response, as well as when fluid-phase attractants were removed prior to the measurement of PMNL chemotaxis, indicating that the chemoattractants generated were absorbed to the surface. Using an immunofluorescence technique, it was found that complement factor 5 coated a circular area around each yeast particle, provided that the particles were able to activate the complement system.

Chemotactic Factors↗
PubMed1984

Complement activation according to the alternate pathway by glass and plastic surfaces and its role in neutrophil adhesion.

C3-deposition, generated by complement activation according to the alternate pathway, was detected on borosilicate glass slides and polystyrene Petri dishes. The C3-depositions grew peripherally until the entire surface was covered. The deposits were also visualized with scanning electron microscopy and could not be washed away with low-pH (3.5) or high-pH (9.6) buffers. No consumption of complement function was detected indicating a phenomenon restricted to the glass and plastic surfaces. The C3-deposits could mediate an adherence of human neutrophils.

Cell Adhesion↗
PubMed1984

Metabolic activation of human neutrophils by fluid-phase and particle-bound ligands.

The ability of particle-bound and fluid-phase ligands (IgG, C3b, and ConA) to induce metabolic activation in cytochalasin B-treated human neutrophils was studied. Different activation mechanisms of the neutrophils are proposed, since the activation of cytochalasin B-treated neutrophils is not dependent on whether or not the ligands are particle-bound but not on the nature of ligand per se.

Complement C3b↗
PubMed1984

The effect of reactive oxygen metabolites (ROM) on the attachment and ingestion phases of C3b- and IgG-mediated phagocytosis by macrophages.

The effects of enzymatically produced reactive oxygen metabolites (ROM) on the attachment and ingestion phases of C3b- and IgG-mediated phagocytosis by cultured mouse peritoneal macrophages (MPM) was investigated using a hypoxanthine-xanthine oxidase ROM-generating system. ROM-exposure at a dose which did not affect cell viability caused a slight decrease in the percentage of phagocytosing cells. The total number of cell-associated (attached and ingested) C3b- and IgG-coated particles initially decreased in relation to controls. After 120 min the number of attached and ingested C3b-particles had returned to the level of controls, while the corresponding value for IgG-particles lagged behind. The number of ingested particles decreased in both C3b- and IgG-groups at each time-point studied (30-150 min). A linear increase in the formation of lipid peroxidation products was measured during the period of observation, while transmission electron microscopical studies showed largely intact morphology. These results indicate that ROM species may induce membrane-related changes in inflammatory cells such as macrophages, probably due to lipid peroxidation; affecting the binding functions of the C3b- and Fc-receptors, without any obvious alteration in cellular fine structure.

Animals↗