Introduction To date, our knowledge of in vivo migration of neutrophils and eosinophils in homeostasis and disease states is based on granulocytes. Here we present a pilot study using purified human eosinophils or neutrophils and demonstrate their differential in vivo kinetics in asthmatic and healthy volunteers. Methods: On two separate occasions 100 ml of blood was obtained from eight human volunteers (4 mild stable asthmatics and 4 non asthmatic, healthy volunteers) Granulocytes were separated using gradient Ficoll-Paque PLUS 1.084 centrifugation. Superparamagnetic particles coupled to a monoclonal antibody against CD16, a surface marker present in neutrophils, were incubated with the granulocytes (containing eosinophils and neutrophils). CliniMACS system (Miltenyi biotec, Bergisch-Gladbach, Germany; and Becton-Dickinson, Oxford, UK) was used to obtain highly purified (>93% pure) human blood eosinophils or neutrophils (>; 97%). Purified cells were labelled with Tc-99m HMPAO (Ceretec, GE Healthcare) under aseptic cGMP conditions and 75–100 MBq of labelled cells were administered intravenously. Dynamic lung images were acquired for the first 30 minutes. Further static scans of 5 minutes each were acquired at 1; 2 and 4 hours. Results: We were able to obtain highly purified neutrophils (positive selection) or eosinophils (negative selection). Kinetics of eosinophils in lung, liver and spleen differed significantly from kinetics of neutrophils. Initial dynamic lung images revealed a significant difference in the time activity curves for eosinophils and neutrophils. Migration of eosinophils from the lungs followed a monexponential clearance (t1/2) of 4.16 min. While neutrophil had significantly different clearance half-lives of 13.72 min (p=0.0019). There were significant differences in eosinophil and neutrophil migration and distribution in the liver and spleen (p<0.0018 and p<0.0325). There was a trend towards faster neutrophil migration in the asthmatics. This was not statistically significant.
Conclusions For the first time it has been possible to identify distinct patterns of neutrophil and eosinophil migration through lung, liver and spleen in both healthy volunteers and stable asthmatics. This technique provides the opportunity for rapid throughput screening of novel therapeutic agents designed to alter leukocyte migration in disease conditions, or to further phenotype disease such as asthma.
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