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T W Kuypers

Publications and source records attributed to T W Kuypers.

4 recordsLinked to original sources
PubMed1993

Down modulation of L-Selectin expression on eosinophils recovered from bronchoalveolar lavage fluid after allergen provocation.

In allergic asthma eosinophils infiltrate into the lung after allergen challenge. The mechanism of this cellular infiltration is not fully understood. L-Selectin is involved in leucocyte-endothelial cell recognition and participates in homing of leucocytes into sites of inflammation. To find indications for a role of L-Selectin in the migration of eosinophils to the bronchoalveolar space we measured L-Selectin expression on eosinophils in peripheral blood and bronchoalveolar lavage fluid (BAL) 4 hr after the early allergic reaction after allergen challenge. Nine patients with allergic asthma participated in the study. An eosinophil specific high depolarization signal enabled us to measure L-Selectin expression on eosinophils in a FACS analysis without isolation of these cells. Eosinophils recovered from BAL showed a strong decrease of L-Selectin expression compared to blood eosinophils. This decrease in L-Selectin expression can be induced in vitro by activation of eosinophils with PMA or FMLP whereas priming of eosinophils during several hours with GM-CSF did not influence L-Selectin expression. Our results are a first indication that L-Selectin may play a role during homing of eosinophils in the lung in asthma after allergen challenge. Moreover, the low expression of L-Selectin on eosinophils in the lung is a further indication that these cells exhibit an activated phenotype.

Adult↗
PubMed1990

Continuous cell activation is necessary for stable interaction of complement receptor type 3 with its counter-structure in the aggregation response of human neutrophils.

Human neutrophils aggregate after stimulation with various stimuli; this response is completely absent in neutrophils from patients with leukocyte adhesion deficiency (LAD). To investigate the cellular requirements of this process a method was used in which neutrophils are separately loaded with hydroethidine (HE) and sulfofluorescein (SFDA), to give them either red fluorescence or green fluorescence. After mixing HE- and SFDA-labeled cells in a ratio of 1:1, the number of double-colored aggregates formed after activation was determined by analysis on a fluorescence-activated cell sorter. In this way, essential information is obtained when cells of different origin are used. The formation of aggregates between neutrophils of an LAD patient and control neutrophils was thus quantified. Because neutrophil aggregation is dependent mostly on the presence of complement receptor type 3 (CR3), which is not present on LAD neutrophils, this result revealed the presence of a counter-structure for CR3 on LAD neutrophils (and hence on normal human neutrophils). In addition to the presence of these proteins on the cell surface, aggregation required continuous cell triggering as indicated by the transient aggregation induced by short-term activation of protein kinase C. This phenomenon was substantiated by the fact that energy depletion caused profound disaggregation. The present study reveals that neutrophil aggregation is a well-controlled process, which needs constant activation of CR3 for a stable interaction with a constitutively expressed counter-structure.

Adenosine Triphosphate↗
PubMed1989

A rapid turbidimetric assay of phagocytosis and serum opsonizing capacity.

An in vitro assay has been developed to measure the opsonizing capacity of serum and the extent of bacterial uptake by phagocytes. Various micro-organisms were preopsonized for 10 min with a serum concentration previously determined to be optimal for the respective types of micro-organism. Subsequently, neutrophils from a healthy donor were added to the preopsonized bacteria in a cuvette of a spectrophotometer. The decrease in turbidity at 400 nm, resulting from the uptake of the micro-organisms by the neutrophils, was measured for 20-30 min and the area under the curves was taken as a measure of the opsonizing capacity of the serum or the phagocytic capacity of the neutrophils. The results correlated well with standard opsonophagocytic assays. By excluding Ca2+ from the buffer of the assay, phagocytosis was distinguished from the combined response of phagocytosis and aggregation. In the presence of Ca2+ ions, both phagocytosis and aggregation contributed to the decrease in turbidity. In the absence of Ca2+, phagocytosis was normal, but aggregation was completely inhibited. Phagocytosis in the absence of Ca2+ was also observed using microscopic and radiometric methods of evaluation. Neutrophils from a patient with a deficiency of leukocyte adhesion molecules, ingested as many bacteria as did normal neutrophils without Ca2+. Experiments with NaF, to inhibit phagocytosis, indicated that the change in turbidity measured in the absence of Ca2+ was mainly caused by phagocytosis, not by attachment of bacteria to the neutrophils. The opsonizing capacity of sera, as determined in our assay, depended both on antibodies and on an intact complement system and the inter-assay variance was less than 5%. We found a close correlation between turbidity changes measured in the presence or absence of Ca2+, suggesting that both phagocytosis and aggregation are opsonin-dependent. This assay is applicable to a variety of opsonizing fluids and micro-organisms, and can be used for assessing the phagocytic capacity of patients' neutrophils as well as for assessing the opsonizing capacity of patients' sera.

Blood Bactericidal Activity↗