Mechanism of neutrophil activation by an unopsonized inflammatory particulate: Monosodium urate crystals induce pertussis toxin-insensitive hydrolysis of phosphatidylinositol 4,5-bisphosphate

Monosodium urate crystals are believed to trigger acute inflammation via the direct stimulation of leukocytes. Unopsonized urate crystals activate neutrophil (PMN) membrane G proteins in a pertussis toxin (PT)-sensitive manner, but induce PT-insensitive cytosolic [Ca²⁺]_i elevation. Thus, we have further defined the mechanism of PMN responsiveness to urate crystals in this study. Though urate crystals can increase membrane permeability by lytic effects, we observed elevation of PMN cytosolic [Ca²⁺]_i in the absence of extracellular [Ca²⁺]_i. In addition, the early, crystal-induced cytosolic [Ca²⁺]_i transient was buffered in cells loaded with a [Ca²⁺]_i-chelator. This suggested mobilization of internal [Ca²⁺]_i stores, which was supported by demonstrating rapid phosphatidylinositol bisphosphate (PIP2) hydrolysis, and the formation of inositol (1,4,5) trisphosphate (as well as phosphatidic acid) in a PT-insensitive manner. Importantly, PMN activation by urate crystals was discriminatory, as evidenced by the absence of phosphatidylinositol trisphosphate formation, a PT-sensitive event triggered by chemotactic factors. Urate crystal-induced PIP2 hydrolysis was not a nonspecific consequence of the early cytosolic [Ca²⁺]_i transient itself, and it did not require phagocytosis. However, crystal-induced O₂^- release was markedly inhibited by buffering of the early cytosolic [Ca²⁺]_i transient under conditions where crystal phagocytosis and PMA-induced O₂^- release were unaffected. We conclude that urate crystals activate PT-insensitive PIP2 hydrolysis, resulting in IP3 generation, and early urate crystal-induced mobilization of cytosolic [Ca²⁺]_i. This pathway appears to modulate crystal-induced O₂^- release.