Involvement of ion channels in human eosinophil respiratory burst☆☆☆

Abstract 

Background: Human eosinophils possess a variety of ion channels that play a crucial role in the regulation of cellular activity. During eosinophil respiratory burst, efflux of H⁺ ions through H⁺ channels provides an efficient mechanism of H⁺ extrusion and charge compensation. Interestingly, recent studies suggest that other ion channels may also be involved in this process. Objective: We sought to investigate the role of ion channels in phorbol 12-myristate 13-acetate–induced superoxide (O₂^•–) generation by human eosinophils. Methods: O₂^•– production was measured by using the superoxide dismutase–inhibitable reduction of cytochrome c. Ion channel expression and function were studied by using RT-PCR and the patch clamp technique, respectively. Results: O₂^•– generation was affected by several ion channel blockers, especially 4,4-diisothio-cyanostilbene-2,2′-disulfonic acid. The involvement of Cl^– channels in this process was confirmed by replacement of Cl^– with gluconate or other anions. The halide dependence of O₂^•– production could be described by the sequence Cl^–≥Br^–>I^–, which is similar to the selectivity sequence of several members of the chloride channel (ClC) family. RT-PCR studies performed with primers for ClC-2, ClC-3, ClC-4, ClC-5, ClC-6, and the cystic fibrosis transmembrane conductance regulator showed only the expression of ClC-3. The presence of phorbol 12-myristate 13-acetate–sensitive Cl^– channels in human eosinophils with biophysical properties similar to the ClC-3 channel has been studied. Conclusion: Cl^– channels play an important role in the regulation of O₂^•– production by human eosinophils. (J Allergy Clin Immunol 2000;106:272-9.)

Keywords:  Eosinophils, Cl^– channels, superoxide, patch clamp, phorbol 12-myristate 13-acetate

Abbreviations:  CFTR: , Cystic fibrosis transmembrane conductance regulator, ClC: , Chloride channel, DIDS: , 4,4-Diisothiocyanostilbene-2,2'-disulfonic acid, DNDS: , 4,4-Dinitrostilbene-2,2'-disulfonic acid, DPC: , Diphenylamine-2-carboxylate, NADPH: , Reduced nicotinamide adenine dinucleotide phosphate, O₂^•–: , Superoxide, PMA: , Phorbol 12-myristate 13-acetate, SOD: , Superoxide dismutase