Real-time differential tracking of human neutrophil and eosinophil migration in vivo

Abstract

Background: Hitherto, in vivo studies of human granulocyte migration have been based on indiscriminate labeling of total granulocyte populations. We hypothesized that the kinetics of isolated human neutrophil and eosinophil migration through major organs in vivo are fundamentally different, with the corollary that studying unseparated populations distorts measurement of both.

Methods: Blood neutrophils and eosinophils were isolated on 2 separate occasions from human volunteers by using Current Good Manufacturing Practice CD16 CliniMACS isolation, labeled with technetium 99m-hexamethylpropyleneamine oxime, and then reinfused intravenously. The kinetics of cellular efflux were imaged over 4 hours.

Results: Neutrophils and eosinophils were isolated to a mean purity of greater than 97% and greater than 95%, respectively. Activation of neutrophils measured as an increase in their CD11b mean fluorescence intensity in whole blood and after isolation and radiolabeling was 25.98 ± 7.59 and 51.82 ± 17.44, respectively, and was not significant (P = .052), but the mean fluorescence intensity of CD69 increased significantly on eosinophils. Analysis of the scintigraphic profile of lung efflux revealed exponential clearance of eosinophils, with a mean half-life of 4.16 ± 0.11 minutes. Neutrophil efflux was at a significantly slower half-life of 13.72 ± 4.14 minutes (P = .009). The migration of neutrophils and eosinophils was significantly different in the spleen at all time points (P = .014), in the liver at 15 minutes (P = .001), and in the bone marrow at 4 hours (P = .003).

Conclusions: The kinetics of migration of neutrophils and eosinophils through the lung, spleen, and bone marrow of human volunteers are significantly different. Study of mixed populations might be misleading.

Keywords: (99m)Tc-HMPAO; FITC; Fluorescein-5-isothiocyante; Granulocytes; HSA; Human serum albumin; MFI; Mean fluorescence intensity; PE; Phycoerythrin; ROI; Region of interest; TAC; Technetium 99m hexamethylpropyleneamine oxime; Time activity curve; bone marrow; eosinophils; kinetics; liver; lung; neutrophils; radiolabeled; spleen; technetium 99m–hexamethylpropyleneamine oxime.