Crystal‐induced neutrophil activation. II. evidence for the activation of a phosphatidylcholine‐specific phospholipase D

Abstract

Objective. To investigate the involvement of phospholipase D in the signaling pathways activated by 2 pathologically relevant inflammatory microcrystals, monosodium urate (MSU) and calcium pyrophosphate dihydrate (CPPD).Methods. Human peripheral blood neutrophils were used throughout. Phospholipase D activity was monitored by measuring 3 separate indices: 1) the mass of phosphatidic acid, 2) the levels of alkyl-phosphatidic acid, and 3) the levels of formation, in the presence of ethanol, of phosphatidylethanol. The latter 2 parameters were measured in cells labeled with 1-0-3H-alkyl-2-acetyl-sn-glycero-3-phosphocholine. The cells were stimulated with microcrystals of triclinic morphology.Results. Both MSU and CPPD crystals induced a time-and concentration-dependent accumulation of phosphatidic acid mass and elevation in levels of alkylphosphatidic acid and phosphatidylethanol in prelabeled cells. The activation of phospholipase D by the microcrystals was partially sensitive to colchicine and largely resistant to pertussis toxin. Inhibition of phosphatidic acid formation by wortmannin or ethanol reduced the microcrystal-stimulated production of superoxide anions.Conclusion. These results indicate that microcrystals stimulate phospholipase D in human neutrophils and that at least some of the functional consequences of neutrophil-microcrystal interactions may be dependent on this biochemical pathway.A significant body of evidence indicates that an interaction of monosodium urate (MSU) or of calcium pyrophosphate dihydrate (CPPD) crystals with neutrophils is involved in the etiology of gouty arthritis and of articular chondrocalcinosis, respectively ( 1,2). This conclusion is supported, inter alia, by the presence of microcrystals in the synovial environment including the synovial fluid, and by the ability to reproduce the major symptoms of gout and articular chondrocalcinosis by introduction of these crystals into normal joints (3-6).Microcrystals interact with all of the major synovial cell types, including neutrophils, monocyte/

Inflammatory Microcrystals Alter the Functional Phenotype of Human Osteoblast-Like Cells In Vitro: Synergism with IL-1 to Overexpress Cyclooxygenase-2

“…The present study indicated that MSUM or CPPD microcrystals were capable of stimulating hOB to express a significant activity of COX-2 at concentrations lower than those previously reported for activation of neutrophils in vitro (28,(41)(42)(43)(44)(45). Compared with CPPD microcrystals, the stronger activity of MSUM microcrystals to stimulate the production of PGE 2 in hOB was also demonstrated for calcium mobilization, superoxide production, and phospholipase D activity in human neutrophils (28,41,46), suggesting that MSUM microcrystals could be more potent activators of cellular functions than CPPD microcrystals. Also, similar to our data on aggressiveness differences between MSUM and CPPD microcrystals on hOB ( Fig.…”

Cholesterol-modulating Agents Selectively Inhibit Calcium Influx Induced by Chemoattractants in Human Neutrophils

“…3 H]alkyl-2-lyso-phosphatidylcholine (2 Ci/10 7 cells) for 90 min as described previously (25)(26)(27). For the last 30 min, 10 mM M␤CD were added to the cells.…”

A Phosphatidic Acid-activated Protein Kinase and Conventional Protein Kinase C Isoforms Phosphorylate p22 , an NADPH Oxidase Component

“…1) that includes activation of phospholipases, generation of lipid second messengers, and the activation of protein kinases. One of the phospholipases activated in response to many physiological agonists of neutrophils is phospholipase D (13)(14)(15)(16)(17)(18)(19)(20)(21)(22)(23)(24)(25)(26)(27). Phospholipase D cleaves neutrophil phospholipids to form phosphatidic acid (PA), 1 which can then be converted to diacylglycerol by PA phosphohydrolase.…”