Published online before print January 29, 2007

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© by The Society for Leukocyte Biology
Journal of Leukocyte Biology, doi:10.1189/jlb.0406248

Received for publication April 5, 2006.
Revised December 15, 2006.
Accepted for publication December 15, 2006.

Article

Potent inhibition of store-operated Ca²⁺ influx and superoxide production in HL60 cells and polymorphonuclear neutrophils by the pyrazole derivative BTP2

Natacha Steinckwich ^*@, Jean-Pol Frippiat ^*, Marie-José Stasia , Marie Erard , Rachel Boxio ^*, Christiane Tankosic ^*, Isabelle Doignon ^||, and Oliver Nüße ^||

*University Henri Poincaré Nancy 1, EA3442, Vandoeuvre-les-Nancy, France; University Joseph Fourier, GREPI EA2938, Grenoble, France; CNRS, UMR8000, Orsay, France; University Paris-Sud, Orsay, France; and ^||INSERM, U757, Orsay, France

^@ To whom correspondence should be addressed. E-mail: natacha.steinckwich{at}scbiol.uhp-nancy.fr.

Abstract

Store-operated calcium entry (SOCE) is a key regulator in the activation of leukocytes. 3,5-Bistrifluoromethyl pyrazole (BTP) derivatives have been identified recently as inhibitors of T lymphocyte activation. The inhibitory effect of one of these compounds, N-(4-[3,5-bis(trifluoromethyl)-1H-pyrazol-1-yl]phenyl)-4-methyl-1,2,3-thiadiazole-5-carboxamide (BTP2), appears to be a result of inhibition of SOC influx. Polymorphonuclear neutrophils provide effective protection against bacterial infection, but they are also involved in tissue damage during chronic inflammation. As for T lymphocytes, their activation relies on SOCE. We therefore investigated the effect of BTP2 on calcium homeostasis and functional responses of human neutrophils. BTP2 significantly inhibited the calcium influx after stimulation with thapsigargin or fMLF. This inhibition was seen after 5 min of incubation with 10 µM BTP2 and after 24 h with lower concentrations. With 24 h incubation, the effect appeared irreversible, as the removal of BTP2 3 h before the experiment did not reduce this inhibition in granulocyte-differentiated HL60 cells. In human neutrophils, BTP2 reduced superoxide anion production by 82% after 24 h of incubation. On the contrary, phagocytosis, intraphagosomal radical production, and bacterial killing by neutrophils were not reduced significantly, even after 24 h treatment with 10 µM BTP2. This work suggests that BTP2 could become an important tool to characterize calcium signaling in neutrophils. Furthermore, BTP2 or related compounds could constitute a new approach to the down-regulation of neutrophils in chronic inflammatory disease without compromising antibacterial host defense.

Key Words: human cells • signal transduction • calcium channel inhibitor • phagocytosis • bacterial killing

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