ANCA-induced neutrophil F-actin polymerization: Implications for microvascular inflammation

doi:doi:10.1111/j.1523-1755.2005.00063.x

Abstract

ANCA-induced neutrophil F-actin polymerization: Implications for microvascular inflammation.

Background

The antineutrophil cytoplasmic antibody (ANCA)-positive vasculitides are characterized by a necrotizing vasculitis of small vessels with neutrophil infiltration. The reasons behind the selectivity for small vessels remain unclear, but may relate to the necessity for neutrophils to deform in order to pass through capillaries. The resistance to deformation of neutrophils largely arises from their actin cytoskeleton. It is hypothesized that ANCA, by inducing actin polymerization, increases neutrophil rigidity and contributes to their sequestration in capillaries.

Methods

To test this hypothesis, neutrophils were treated with IgG-ANCA and the following characterizations: formation of filamentous F-actin (by flow cytometry); changes in morphology (by fluorescence and electron microscopy); and the potential to obstruct microvessels (by measuring entry times into micropipettes with comparable diameters to capillaries). The neutrophil signaling mechanisms activated by IgG-ANCA were investigated using blocking antibodies to Fc gamma receptors and inhibitors of tyrosine phosphorylation. Protein tyrosine phosphorylation was examined by immunoblotting of cell lysates, and calcium fluxes were measured by spectrofluorimetry of Fura-2 pentakis (acetoxymethyl) ester (Fura 2-AM) labeled neutrophils.

Results

IgG-ANCA led to a significant dose-dependent actin polymerization over about 10 minutes. Over the same period, neutrophils became distorted in shape and more resistant to micropipette aspiration. Treatment with normal IgG caused less marked and delayed changes in these parameters. Actin polymerization required engagement of Fc gamma RIIa receptor, tyrosine phosphorylation, and calcium fluxes.

Conclusion

These novel findings reveal signaling mechanisms that underlie ANCA-induced actin polymerization and might explain the predilection for small vessels in IgG-ANCA–associated vasculitis.

Keywords:

neutrophils, recirculation, recruitment, signal transduction, Fc gamma receptors

Cell Biology – Immunology – Pathology