The involvement of phosphatidylinositol 3-kinase in crystal induced human neutrophil activation.
OBJECTIVE
We investigated whether phosphatidylinositol (PI) 3-kinase is involved in the signal transduction pathway leading to neutrophil activation by inflammatory microcrystals.


METHODS
Neutrophil chemiluminescence and degranulation responses to opsonized crystals were measured in the presence of selective inhibitors known to inhibit PI 3-kinase activity in neutrophils.


RESULTS
Wortmannin and LY 294002, 2 selective inhibitors of PI 3-kinase, were shown to inhibit neutrophil activation induced by plasma opsonized crystals of calcium pyrophosphate dihydrate (CPPD) [both monoclinic (M) and triclinic (T) forms] and monosodium urate monohydrate (MSUM). IC50 for wortmannin or LY 294002 inhibition of crystal induced respiratory burst (measured by chemiluminescence) was about 3 nM and 0.3 microM, respectively, proving the pivotal role of PI 3-kinase in neutrophil respiratory burst activation by all 3 crystals. Degranulation responses of neutrophils to CPPD(M) and CPPD(T) crystals were also inhibited by about 50% by wortmannin in the 10 to 20 nM concentration range, supporting the direct involvement of PI 3-kinase in signal transduction pathways leading to crystal induced neutrophil degranulation. All 3 crystals induced the activation of PI 3-kinase in neutrophils as measured by the increased PI 3-kinase activity associated with immunoprecipitated tyrosine phosphorylated proteins from 1 min crystal-neutrophil incubations. Neutrophils pretreated with wortmannin at 10 nM showed sub-basal levels of PI 3-kinase activity at all time points measured.


CONCLUSIONS
PI 3-kinase plays a central role in the signal transduction pathways leading to respiratory burst and degranulation responses in neutrophils activated by inflammatory microcrystals.
Jackson
J K
JK
Faculty of Pharmaceutical Sciences, University of British Columbia, Vancouver, Canada.
Lauener
R
R
Duronio
V
V
Burt
H M
HM
D000730:Androstadienes / Q000494:pharmacology; D002131:Calcium Pyrophosphate / Q000494:pharmacology*; D002478:Cells, Cultured; D002867:Chromones / Q000494:pharmacology; D003460:Crystallization; D003594:Cytoplasmic Granules / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D008163:Luminescent Measurements; D009025:Morpholines / Q000494:pharmacology; D018375:Neutrophil Activation / Q000187:drug effects; D009504:Neutrophils / Q000187:drug effects / Q000201:enzymology*; D019869:Phosphatidylinositol 3-Kinases; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000187:drug effects / Q000378:metabolism*; D016897:Respiratory Burst / Q000187:drug effects; D015398:Signal Transduction; D013481:Superoxides / Q000378:metabolism; D014527:Uric Acid / Q000494:pharmacology*; D000077191:Wortmannin
D000730:Androstadienes; D002867:Chromones; D004791:Enzyme Inhibitors; D009025:Morpholines; D013481:Superoxides; D014527:Uric Acid; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; D019869:Phosphatidylinositol 3-Kinases; D017853:Phosphotransferases (Alcohol Group Acceptor); D002131:Calcium Pyrophosphate; D000077191:Wortmannin