Influence of iron- and zinc-chelating agents on neutrophil extracellular trap formation

Release of neutrophil extracellular traps (NETs) is one of the neutrophils' mechanisms involved in the response to infection. NETs are released from the cell in response to a biological or synthetic stimulus to entrap, immobilize and kill pathogens. Metal ions and metal binding proteins were identified in the structure of NETs, but their role in NET release remains unclear. The aim of this study was to assess how lack of iron and zinc generated by ion sequestration using chelators affects NET release. Neutrophils were isolated from whole blood or buffy coats of healthy blood donors by density gradient centrifugation and incubated with zinc chelators: 20 mu M N,N,N',N'-Tetrakis(2-pyridylmethyl)ethylenediamine (TPEN), 40 mu M diethylenetriaminepentaacetic acid (DTPA) or iron chelators: 400 mu M deferoxamine mesylate salt (DFO) and 50 mu M iminodiacetic acid (IDA). Next, 100 nM phorbol 12-myristate 13-acetate (PMA) was added to stimulate release of NETs. The amount of released DNA was measured by fluorometry and NETs were visualized by immunofluorescence microscopy. This study demonstrates that iron and zinc chelators are able to modulate NET release. Here we show that preincubation of neutrophils with TPEN and IDA inhibits NET release in cells stimulated with PMA. On the other hand, DFO stimulates NET release. Incubation of cells with DTPA does not affect release of NETs.

Automated summary

This study demonstrates that iron and zinc chelators can modulate the release of NETs. Preincubation of neutrophils with TPEN and IDA inhibits the release of NETs in cells stimulated with PMA, while DFO stimulates the release of NETs. Incubation with DTPA has no effect on the release of NETs.