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Microfluidic Platform for Measuring Neutrophil Chemotaxis from Unprocessed Whole Blood
Published on: June 3, 2014
Hydroxyethyl Starch 130/0.4 Binds to Neutrophils Impairing Their Chemotaxis through a Mac-1 Dependent Interaction
Alessandro Trentini1, Francesca Murganti2,3, Valentina Rosta4
1Section of Medical Biochemistry, Molecular Biology and Genetics, Department of Biomedical and Specialist Surgical Sciences, University of Ferrara, 44121 Ferrara, Italy. alessandro.trentini@unife.it.
Abstract:
Several studies showed that hydroxyethyl starch (HES), a synthetic colloid used in volume replacement therapies, interferes with leukocyte-endothelium interactions. Although still unclear, the mechanism seems to involve the inhibition of neutrophils' integrin. With the aim to provide direct evidence of the binding of HES to neutrophils and to investigate the influence of HES on neutrophil chemotaxis, we isolated and treated the cells with different concentrations of fluorescein-conjugated HES (HES-FITC), with or without different stimuli (N-Formylmethionine-leucyl-phenylalanine, fMLP, or IL-8). HES internalization was evaluated by trypan blue quenching and ammonium chloride treatment. Chemotaxis was evaluated by under-agarose assay after pretreatment of the cells with HES or a balanced saline solution. The integrin interacting with HES was identified by using specific blocking antibodies. Our results showed that HES-FITC binds to the plasma membrane of neutrophils without being internalized. Additionally, the cell-associated fluorescence increased after stimulation of neutrophils with fMLP (p < 0.01) but not IL-8. HES treatment impaired the chemotaxis only towards fMLP, event mainly ascribed to the inhibition of CD-11b (Mac-1 integrin) activity. Therefore, the observed effect mediated by HES should be taken into account during volume replacement therapies. Thus, HES treatment could be advantageous in clinical conditions where a low activation/recruitment of neutrophils may be beneficial, but may be harmful when unimpaired immune functions are mandatory.
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