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Biomedical subjects

O Cromwell

Publications and source records attributed to O Cromwell.

At least 73 records · Page 4Linked to original sources
PubMed1988

Antigen-induced neutrophil chemotactic factor from cloned human T lymphocytes.

Here we report the presence of a low molecular weight (10,000 neutrophil chemotactic factor (NCF) in the supernatants of activated human T lymphocytes. Appreciable amounts of the 10,000 MW NCF were generated by peripheral blood mononuclear cells (PBMC) stimulated with anti-CD3 antibody, and comparable NCF was secreted by both long-term human T-helper (CD4+) cell lines reactive with house dust mite (Dermatophagoides farinae) and influenza A virus-immune T-cell clones. When the cloned T cells were stimulated with specific antigen in the presence of irradiated accessory cells (AC) or insolubilized anti-CD3 antibody the 10,000 MW NCF was readily identifiable in 24-hr culture supernatants. Cultures of AC and antigen alone produced negligible neutrophil chemotactic activity, as did control cultures using an irrelevant allergen (mixed grass pollen). These findings indicate that the 10,000 MW NCF may be T-lymphocyte derived and that formation and release are dependent upon stimulation via the antigen receptor.

Antigens↗
PubMed1988

The identification and partial characterization of a human mononuclear cell-derived neutrophil chemotactic factor apparently distinct from IL-1, IL-2, GM-CSF, TNF and IFN-gamma.

We have identified a neutrophil chemotactic factor (NCF) in supernatants from human blood mononuclear cells (MNC) cultured in the presence of phytohaemagglutinin (PHA). Maximal activity was observed 48 hr after culture. Following gel filtration, NCF eluted as a single major peak, together with proteins, having a molecular size of approximately 10,000 MW. The material gave a single band on SDS-PAGE but was heterogeneous following chromatofocusing (pIs approximately 6.8-7.0, 5.5-6.0 and 5.0). The biological activity of the partially purified material was abolished by trypsin and chymotrypsin treatment. NCF was heat stable (70 degrees, 60 min) and promoted both directional migration (chemotaxis) of neutrophils and, to a lesser extent, stimulated random locomotion (chemokinesis). The factor was not associated with detectable amounts of IL-1, IL-2 or interferon-gamma (IFN-gamma). MNC-derived NCF had a molecular size lower than recombinant granulocyte-monocyte colony-stimulating factor (rGM-CSF) and recombinant tumour necrosis factor (rTNF), and was considerably more active in chemotaxis. Optimal chemotactic concentrations of partially purified MNC-derived NCF were of comparable potency to FMLP and LTB4 and had about 60% of the activity of optimal concentrations of C5a, C5a-des-Arg and platelet-activating factor (PAF). These experiments indicate that the human MNC-derived NCF is a potent chemo-attractant distinct from other cytokines previously reported to promote neutrophil locomotion.

Cell Movement↗
PubMed1987

Allergen-induced recruitment of bronchoalveolar helper (OKT4) and suppressor (OKT8) T-cells in asthma. Relative increases in OKT8 cells in single early responders compared with those in late-phase responders.

We measured the numbers of helper (OKT4) and suppressor (OKT8) T-cells in blood and bronchoalveolar lavage (BAL) in 6 patients who developed single early reactions after allergen inhalational challenge and in 6 subjects who developed dual (early- and late-phase) responses. The asthmatics were lavaged twice. On the first occasion, they inhaled the diluent control (Dil) solution, and after an interval of at least 7 days, allergen (Ag) inhalational challenge was undertaken. Significant differences were observed in the single early responders (SER) only. For instance, when the results obtained on the Dil day were compared with those obtained on the Ag day, there was a significant (p less than 0.05) increase in the percentage of OKT4 cells in the blood of SER. Also in the BAL of SER there was a significant (p less than 0.05) decrease in the percentage of OKT4 and an increase (p less than 0.05) in the percentage and absolute numbers of OKT8 cells. Furthermore, the percentage of OKT4 was highly significantly (p less than 0.005) lower on the Ag day in the SER than on the Ag day in the late-phase responders (LPR). The OKT4/OKT8 ratio was significantly (p less than 0.05) lower on the Ag day than on the Dil day in BAL from SER and highly significantly decreased (p less than 0.001) when results on the Ag day in the SER were compared with those on the Ag day in the LPR. These findings raise the possibility that mobilization of suppressor T-cells into the lung after allergen-induced single early reactions in asthma might be associated with the prevention of a subsequent late-phase response.

Adult↗
PubMed1987

Neutrophil chemotactic activity in acute severe asthma (status asthmaticus).

Serum neutrophil chemotactic activity (NCA) was measured in patients with acute severe asthma (status asthmaticus) and compared with that in control subjects (mild asthma, stable chronic irreversible air-flow obstruction, allergic rhinitis, noninfective lung conditions, or asymptomatic). There were 9 subjects in each group. Statistically significant elevations (p less than 0.002) in NCA were detected in acute severe asthma when compared with each control group. Serial measurements of NCA were subsequently undertaken in 12 patients with acute asthma, at the time of admission to hospital, after 3 days of treatment, and on discharge after approximately 7 days. A highly significant (p less than 0.001) reduction in serum NCA activity on Day 7 compared with that on Day 0 was observed, and this correlated inversely with the improvement in lung function (PEFR). Gel filtration by fast protein liquid chromatography (FPLC) using Superose 6 prep grade (6PG) indicated that NCA in acute severe asthma was heterogeneous and consisted of at least 4 peaks of activity associated with proteins with molecular weights of approximately 800, 600, 150, and less than 20 kD. The 800- and 150-kD peaks were also observed in control subjects, but to a lesser degree. The 600- and less than 20-kD activities were virtually confined to the patients with acute severe asthma. FPLC chromatofocusing of the 600-MW peak from the acute asthmatics, using a Mono-P column and a pH gradient from 8.3 to 5.0, revealed considerable activity in fractions eluting between pH 6.0 and 7.0, which was not observed in the normal control subjects.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1987

Leukotriene B4 generation by human neutrophils following IgG-dependent stimulation.

It has previously been shown that human neutrophils generate substantial quantities of LTB4 when stimulated with the calcium ionophore, A23187, or with unopsonized zymosan. We now report that normal human neutrophils produced substantial quantities of LTB4 (measured by radioimmunoassay and validated by RP-HPLC) when incubated with large non-phagocytosable IgG-coated beads (Sepharose 4B). LTB4 was identified in both the extra and intracellular compartments. The production of LTB4 was dependent upon the number of IgG-coated particles and the concentration of IgG bound to the beads. Release was maximal after a 15-30 min incubation time and was enhanced by prior activation of the neutrophils with the synthetic bacterial product f-met-leu-phe. Comparable LTB4 production was also observed when neutrophils were incubated with antigen (Aspergillus fumigatus)-coated beads sensitized with purified IgG obtained from the sera of patients with allergic bronchopulmonary aspergillosis. These results suggest a further mechanism by which neutrophils may be activated to produce inflammatory mediators in the tissues.

Antigens, Fungal↗
PubMed1986

Enteral and systemic release of leukotrienes during anaphylaxis of Nippostrongylus brasiliensis-primed rats.

Rats with acquired immunity to the intestinal nematode Nippostrongylus brasiliensis develop anaphylaxis after i.v. challenge with an extract of worm antigen, with the small intestine being the primary shock organ. In the present study we have shown that these events were associated with significant elevations in intestinal and plasma concentrations of leukotrienes LTB4 and LTC4. The changes were observed in immune rats over 10-, 30-, and 60-min intervals after antigen challenge but were absent in control animals. These lipid mediators were identified both in the perfusate of the gut lumen, which contained large quantities of mucus, and in homogenates of intestinal tissue. In addition, significant elevations in the concentrations of plasma LTB4 and LTC4 were detected in immune challenged rats but not in controls. Leukotrienes were identified by radioimmunoassay and validated by reverse-phase high-performance liquid chromatography (RP-HPLC). RP-HPLC analysis of SRS-A leukotrienes in immune challenged rats indicated that LTC4 was the predominant sulfidopeptide leukotriene at 10 min, with almost complete biodegradation to LTD4 and LTE4 within 30 min. Infected rats also had significant increases in the numbers of intestinal mucosal mast cells (MMC) and eosinophils. Evidence of MMC activation during anaphylaxis was obtained by showing significant elevations of intestinal and systemic concentrations of their exclusive serine enzyme, rat mast cell proteinase II (RMCPII). Thus, the release of substantial amounts of leukotrienes in the gut and plasma of N. brasiliensis-primed rats after interaction with worm antigens suggests that these potent mediators may play an important role in allergic-type hypersensitivity known to occur during immune reactions against parasitic helminths.

Anaphylaxis↗
PubMed1986

Morphological and secretory properties of bronchoalveolar lavage mast cells in respiratory diseases.

We have studied various functional and morphological characteristics of mast cells obtained in bronchoalveolar lavage from fifty-two patients with several lung diseases. The percentage of mast cells ranged from 0.04 to 0.6% (bronchial carcinoma), 0.05-0.3% (sarcoidosis), 0.06-0.25% (asthma), 0.04-1.8% (miscellaneous) and 0.02-0.04% (normals). There were no significant differences in the mast cell counts between the disease groups. Lung mast cells exhibited heterogeneity of size, shape and intensity of staining. Cells from thirty-seven subjects were further studied for total histamine content and histamine release using various secretagogues. There was a significant correlation (P less than 0.001) between the histamine content of the total lavage cell population and mast cell counts. The calculated mean histamine content per mast cell was 6.35 pg. Histamine was released in a dose-dependent fashion after stimulation with anti-IgE, calcium ionophore and phorbol myristate acetate with a time course of histamine release characteristic of the mast cell. Unlike peripheral blood basophils, no release was observed following incubation with f-met-leu-phe (10(-6)-10(-8) M) and neither cell type released histamine following incubation with 48/80 (10 micrograms/ml). Inhibition of anti-IgE-induced histamine release was obtained following pre-incubation with salbutamol (10(-4)-10(-6) M). These studies indicate that bronchoalveolar lavage is a suitable model for the study of human lung mast cells.

Adult↗
PubMed1986

Platelet-activating factor. A potent chemotactic and chemokinetic factor for human eosinophils.

Platelet-activating factor (PAF-acether), an inflammatory mediator with a wide range of biological activities including neutrophil aggregation and chemotaxis, was studied for its effect on human eosinophil locomotion (chemotaxis and chemokinesis). Human eosinophils (25-95% purity) were obtained from donors with a variety of diseases associated with hypereosinophilia. PAF-acether elicited directional locomotion of eosinophils, in a time- and dose-dependent fashion, at concentrations from 10(-5) to 10(-8) M; lyso-PAF had minimal activity over the same dose range. Compared with PAF-acether, the eosinophil locomotory responsiveness of leukotriene B4 (LTB4), histamine, and the valyl- and alanyl-eosinophil chemotactic factor of anaphylaxis (ECF-A) tetrapeptides was negligible. Conversely, neutrophil responsiveness to PAF-acether (optimum 10(-6) M) was comparable in effect to LTB4 (optimum dose 10(-8) M). It was shown that PAF-acether elicited both chemotaxis and chemokinesis of eosinophils. Comparison of normal density and light density eosinophils revealed no qualitative difference in the response to PAF-acether and the other chemoattractants, although the light density cells seemed to demonstrate a greater degree of locomotion to PAF-acether and LTB4. Thus, PAF-acether appears to be a potent eosinophilotactic agent which may play a role in inflammatory reactions characterized by eosinophil infiltration.

Cell Movement↗
PubMed1986

The effects of eosinophil activating factor on IgG-dependent sulphidopeptide leukotriene generation by human eosinophils.

Eosinophil activating factor (EAF) is a 40 kD protein released from cultured, unstimulated human monocytes which enhances the IgG-dependent eosinophil-mediated cytotoxicity of helminthic larvae. We have recently shown that eosinophils elaborate substantial quantities of leukotriene C4 (LTC4) during incubation with IgG-coated particles and now report that EAF, partially purified by sequential chromatography on Sephacryl S-200 and DEAE-cellulose, enhanced this IgG-dependent LTC4 production by human eosinophils in a dose- and time-dependent fashion. LTC4 production by normal density eosinophils, separated on discontinuous metrizamide gradients, was significantly increased after incubation with several dilutions of EAF (P less than 0.05), although an increase was not seen with low density cells. The enhancement was similar in degree to that seen when normal density eosinophils were activated with the bacterial analogue, f-met-leu-phe (fMLP). EAF produced a time-dependent increase in LTC4 which was significantly greater (P less than 0.01) than the control. Sulphidopeptide leukotriene (LT) generation was validated by reverse phase high pressure liquid chromatography (RP-HPLC). These results indicate that there is a firm association between monocytes, eosinophils and LTC4; an observation which may be of relevance to mechanisms in chronic asthma and related disorders, and in immune reactions against migrating helminthic larvae.

Dose-Response Relationship, Drug↗
PubMed1985

Inhibition of leukotriene C4 and B4 generation by human eosinophils and neutrophils with the lipoxygenase pathway inhibitors U60257 and BW755C.

Human eosinophils and neutrophils have the capacity to generate leukotriene C4 (LTC4) and leukotriene B4 (LTB4) respectively when stimulated by calcium ionophore A23187. Leukotriene production by mixtures of these cell types was measured by radioimmunoassay for LTC4 and LTB4, and the specificities of the assays determined by assessing cross-reactivities with a number of other arachidonic acid metabolites. The IC50S for LTC4 and LTB4 in their respective assays were 1.76 +/- 0.04 nmol and 3.00 +/- 0.08 nmol. Cross-reactivity for anti-LTC4 was shown by leukotriene D4 (LTD4) (70%) and leukotriene E4 (LTE4) (8%), when compared to LTC4, whereas in the radioimmunoassay for LTB4, only the 5(S), 12(R) 6-trans isomer of LTB4 showed appreciable interaction (12%). LTC4 production by eosinophil enriched cell fractions obtained from metrizamide gradients was inhibited in a dose-dependent fashion by the prostacyclin analogue, 6,9-deepoxy-6,9-phenylimino-delta 6,8-prostaglandin I, (U60257) and by 3-amino-1-(3-trifluoromethyl phenyl)-2-pyrazole (BW755C). The ID50 values for U60257 and BW755C were 2 X 10(-6) and 5 X 10(-6) M respectively. This demonstration of LTC4 production by human eosinophils, which are known to be important cells in clinical asthma, provides an in vitro model to assess 5-lipoxygenase inhibitors in human tissue.

4,5-Dihydro-1-(3-(trifluoromethyl)phenyl)-1H-pyraz↗
PubMed1985

Allergen-induced release of sulphidopeptide leukotrienes (SRS-A) and LTB4 in allergic rhinitis.

Leukotrienes are a recently discovered group of arachidonic acid-derived lipid mediators. Using radioimmunoassay and high pressure liquid chromatography (HPLC), we have identified the SRS-A sulphidopeptide leukotrienes (LTC4, LTD4 and LTE4) in nasal washings from patients with allergic rhinitis who underwent nasal challenge with specific allergen. Smaller, but significant, amounts of LTB4 were also detected. The concentrations of nasal leukotrienes were directly related to the dose of allergen, and were recovered in washings in a time-dependent fashion after challenge. When the patients were subjected to methacholine nasal challenge on a control day, we found only negligible amounts of either the sulphidopeptide leukotrienes or LTB4. These findings support the view that LTC4, LTD4 and LTE4 might contribute to the pathogenesis of allergic rhinitis as a result of their recognized effects on mucous hypersecretion and vasopermeability, and that the potent chemoattractant LTB4 might be involved in the subsequent infiltration of inflammatory cells.

Adult↗
PubMed1984

Immunologic studies in allergen-induced late-phase asthmatic reactions.

We have measured plasma histamine, serum neutrophil chemotactic activity (NCA) and complement (C3 and C4) over a 24-hour period in patients experiencing either early- and late-phase (dual) or single early asthmatic reactions to inhaled allergens. There was a significant biphasic elevation in plasma histamine, which paralleled the fall in forced expiratory volume in 1 sec in 10 patients with dual responses, whereas in seven subjects with single early reactions, only a single early increase in histamine concentrations was observed. In general, in the individual subjects, the changes in plasma histamine paralleled both the elevations in serum NCA and the decreases in forced expiratory volume in 1 sec. By gel filtration on Sephacryl S-400, anion exchange chromatography on DEAE Sephacel, and chromatofocusing with Polybuffer Exchanger 94, the major NCA of both the early and the late reactions was associated with proteins having an estimated molecular size of 600,000 daltons, an elution from DEAE Sephacel at 0.15M to 0.30M of NaCl (pH 8.1), and a pI of approximately 6.5. There were no appreciable changes in serum C3 and C4 up to 24 hr after challenge in subjects with late-phase responses. The patterns of asthmatic response were not related to either the total or allergen-specific serum IgE or IgG4 concentrations. These results support the view that mediators of hypersensitivity participate in late-phase as well as early asthmatic reactions.

Adolescent↗
PubMed1984

Mediators in exercise-induced asthma.

Circulating concentrations of the mast cell-associated mediators, histamine and neutrophil chemotactic factor (NCF) of high molecular weight, were measured in atopic and nonatopic asthmatics after treadmill exercise. Elevations in the concentrations of both mediators accompanied the development of exercise-induced asthma (EIA). Normal individuals did not release mediators or develop bronchoconstriction after an identical exercise. The elaboration of mediators was not due to the onset of airflow obstruction, the postexercise basophilia, or the exercise task per se. A treadmill exercise undertaken while inhaling fully conditioned air inhibited EIA and NCF release; in contrast the same exercise undertaken while breathing cold, dry air elicited EIA and the production of mediators. This suggests that the stimulus for EIA and mediator release may be identical. Late-phase asthmatic reactions occur 3 to 9 hr after exercise in some asthmatics and are accompanied by the appearance of circulating NCF, as previously reported in allergen-induced late responses. In addition to the contribution of mediators to the spasmogenic reaction in EIA, mediators may contribute to bronchial inflammation by activating circulating leukocytes. There was a kinetic increase in the expression of neutrophil C3b receptors in EIA (+) asthmatics for up to 60 min after treadmill exercise. The enhancement of C3b receptors, as evidence of neutrophil activation, was preceded by release of NCF and reductions in peak expiratory flow rates. The prior administration of cromolyn inhibited EIA, NCF release, and enhancement of C3b receptors. These changes were not observed in EIA (-) asthmatics after an identical exercise task. These findings support the view that mediators are released in EIA and may play an important role in its pathogenesis.

Asthma↗
PubMed1984

Partial characterization of allergens associated with hypersensitivity to the 'green nimitti' midge (Cladotanytarsus lewisi, Diptera: Chironomidae).

Allergens in extracts of the 'green nimitti' midge, Cladotanytarsus lewisi Freeman (Diptera: Chironomidae), a cause of widespread hypersensitivity in the Sudan, were isolated and partially characterized by assays which depend on the binding of 125I-anti-IgE to allergen-IgE complexes. These methods included RAST inhibition, crossed radioimmunoelectrophoresis (CRIE) and rocket radioimmunoelectrophoresis (RRIE). Following Sephadex G100 chromatography the 'major peak' of allergenicity as determined by RAST inhibition, RRIE and SDS-PAGE was associated with molecules of approximately 17 000 daltons. The peak eluting at Vo contained material of molecular weight 66 000 daltons which also bound 125I-anti-IgE, but had only 61% of the activity of the 'major peak' by RAST inhibition. By isoelectric focusing and RRIE of fractions obtained by chromatofocusing with polybuffer exchanger 94, the 'major peak' was associated with multiple bands with a pI range of 3.5-5.5. These results indicate that the major allergens from C. lewisi are a group of closely related acidic peptides.

Allergens↗
PubMed1984

Neutrophil chemotactic activity and histamine in atopic and nonatopic subjects after exercise-induced asthma.

Seven atopic and 6 nonatopic asthmatics with exercise-induced asthma were subjected to a treadmill exercise task. After this procedure, both groups had elevations in the concentrations of plasma histamine and serum high molecular weight neutrophil chemotactic activity (as assessed by Sephadex G-200 chromatography). These findings support the view that the release of mast-cell-associated mediators can be independent of the atopic state.

Adolescent↗
PubMed1984

Human granulocyte/pollen-binding protein. Recognition and identification as transferrin.

Normal human serum was found to contain a heat-stable protein which promoted the binding of granulocytes to timothy grass pollen (granulocyte/pollen-binding protein [GPBP]). GPBP was purified by gel filtration, anion exchange, and affinity chromatography. Virtually all of the granulocyte/pollen-binding activity was associated with a beta-1-protein having a molecular mass of approximately 77,000 D and an isoelectric point of between 5.5 and 6.1. By immunoelectrophoresis and sodium dodecyl sulfate-polyacrylamide gel electrophoresis, the protein was identified as transferrin. Monospecific antisera raised against either GPBP or transferrin removed biological activity from GPBP preparations, and GPBP and transferrin gave lines of identity with these two antisera. The apparent heterogeneity in the molecular size and charge of GPBP observed during progressive purification was minimal when GPBP was saturated with ferric ions before the separation procedures. These experiments indicate that granulocyte/pollen binding is a hitherto unrecognized property of transferrin which appears to be unrelated to iron transport and raises the possibility that transferrin might have a physiological role in the removal of certain organic matter.

Carrier Proteins↗