Cell surface distribution of Fc receptors II and III on living human neutrophils before and during antibody dependent cellular cytotoxicity

Abstract

Microscopic techniques have been employed to study the cell surface distributions of the immunoglobulin Fc receptors (FcR) II and III on living human neutrophils. Fluorescein-or rhodamine-conjugated monoclonal IgG or Fab fragments directed against FcRII (CDw32) and FcRIII (CD16) were employed to label receptors. FcRII and III were found to be uniformly distributed at neutrophil surfaces during resting conditions. During neutrophil polarization and migration FcRII but not FcRIII preferentially accumulated at the uropod. Sheep erythrocytes (SRBCs) were opsonized with IgG and then incubated with neutrophils. When neutrophils were labeled prior to target addition, FcRII but not FcRIII were found to cluster at the target-effector interface. Little or no clustering of FcRs was observed if labeling was performed after target binding. SRBC oxidation was observed using Soret band illumination during transmitted light microscopy. Time-lapse studies of FcRII distribution and target oxidation were performed. FcRII formed clusters at target effector interfaces prior to target oxidation. Three lines of evidence suggest that clustering is not a general plasma membrane response. Firstly, FcRIII do not cluster lannic acid-modified erythrocytes avidly bound to neutrophils but did not trigger clustering of FcRII. Furthermore, irrelevant neutrophil membrane labels were unaffected by the presence of IgG-opsonized erythrocytes. We suggest that FcRII clustering is one important component leading to the oxidative destruction of target cells.

Shape, F‐actin, and surface morphology changes during chemotactic peptide‐induced polarity in human neutrophils

“…Some authors have suggested that the differences are caused by a n increase in the density of the molecules a t the anterior pole (Haston and Maggs, 1990;McKay et al, 1991;Sullivan et al, 1984;Walter et al, 1980;Weinbaum et al, 1980). Alternatively, they may result from the accumulation of plasma membrane in the front of polar cells (Petty et al, 1989;Pytowski et al, 1990). We hope that our findings will make it possible to analyze the expression, spatial distribution, and mobilization of different cell surface molecules linked to r the F-actin based cytoskeleton (e.g., integrins and selectins) during polarization and cellular motility, and will shed light on the role of these molecules during neutrophil-mediated cell functions.…”

The NADPH oxidase complex of phagocytic leukocytes: a biochemical and cytochemical view

“…Petty and his co-workers (Francis et al 1988;Petty et al 1989 have employed other microscopic techniques to monitor ROS production and delivery to phagocytic targets. They have utilized Soret band (absorption contrast) transmitted light microscopy to monitor changes in hemoglobin due to the entry of ROS into target erythrocytes within the phagosomes of nentrophils.…”

Imaging neutrophil activation: Analysis of the translocation and utilization of NAD(P)H‐associated autofluorescence during antibody‐dependent target oxidation

“…Previous studies in this laboratory (Francis et al, 1988;Petty et al, 1989) have shown that the entry of oxidative molecules into target erythrocytes can be conveniently monitored by imaging the Soret band at its 430 nm edge using transmitted light microscopy. We have combined these two methods to spatially and temporally resolve the metabolic changes in neutrophils that accompany the oxidative destruction of targets.…”