Stimulation of p38 Phosphorylation and Activity by Arachidonic Acid in HeLa Cells, HL60 Promyelocytic Leukemic Cells, and Human Neutrophils

Abstract

Although it is well appreciated that arachidonic acid, a second messenger molecule that is released by ligandstimulated phospholipase A 2 , stimulates a wide range of cell types, the mechanisms that mediate the actions of arachidonic acid are still poorly understood. We now report that arachidonic acid stimulated the appearance of dual-phosphorylated (active) p38 mitogen-activated protein kinase as detected by Western blotting in HeLa cells, HL60 cells, human neutrophils, and human umbilical vein endothelial cells but not Jurkat cells. An increase in p38 kinase activity caused by arachidonic acid was also observed. Further studies with neutrophils show that the stimulation of p38 dual phosphorylation by arachidonic acid was transient, peaking at 5 min, and was concentration-dependent. The effect of arachidonic acid was not affected by either nordihydroguaiaretic acid, an inhibitor of the 5-, 12-, and 15-lipoxygenases or by indomethacin, an inhibitor of cyclooxygenase. Arachidonic acid also stimulated the phosphorylation and/or activity of the extracellular signal-regulated protein kinase and of c-jun N-terminal kinase in a cell-typespecific manner. An examination of the mechanisms through which arachidonic acid stimulated the phosphorylation/activity of p38 and extracellular signal-regulated protein kinase in neutrophils revealed an involvement of protein kinase C. Thus, arachidonic acid stimulated the translocation of protein kinase C ␣, ␤I, and ␤II to a particulate fraction, and the effects of arachidonic acid on mitogen-activated protein kinase phosphorylation/activity were partially inhibited by GF109203X, an inhibitor of protein kinase C. This study is the first to demonstrate that a polyunsaturated fatty acid causes the dual phosphorylation and activation of p38.

2,2′,4,4′-Tetrachlorobiphenyl upregulates cyclooxygenase-2 in HL-60 cells via p38 mitogen-activated protein kinase and NF-κB

“…Furthermore, free AA activates a variety of proteins involved in the regulation of COX-2 expression, including p38 MAP kinase (Hii et al 1998), peroxisome proliferator-activated receptor γ (Pawliczak et al 2002(Pawliczak et al , 2004 and phosphatidyl inositol-3-kinase (Monick et al 2002). Interestingly, expression of COX-2 was decreased in brains of PLA 2 -deficient mice relative to wild type mice (Bosetti and Weerasinghe, 2003).…”

Examination of the signal transduction pathways leading to activation of extracellular signal‐regulated kinase by formyl‐methionyl‐leucyl‐phenylalanine in rat neutrophils

“…The endogenous AA released is subject to conversion to the metabolites of cyclo-oxygenase (COX) and lipoxygenase (LO). A recent report indicates that AA, acting as a second messenger, is able to stimulate the phosphorylation and activity of ERK in a variety of cell types including human neutrophils [20]. The activation of ERK by AA through an LO-dependent but COX-independent pathway has been reported in vascular smooth muscle cells [21] and in human neutrophils [22].…”

A Novel Long Chain Polyunsaturated Fatty Acid, β-Oxa 21:3n-3, Inhibits T Lymphocyte Proliferation, Cytokine Production, Delayed-Type Hypersensitivity, and Carrageenan-Induced Paw Reaction and Selectively Targets Intracellular Signals

“…The Ag-Ab complexes were precipitated by the addition of protein A-Sepharose (20 l/sample) and then washed. p38 activity was determined by measuring the incorporation of 32 P into myelin basic protein (19). Phosphorylated myelin basic protein was resolved by 16% SDS-PAGE and detected, and the amount of incorporated radioactivity was quantitated as described above.…”

“…A solid-phase assay was used to assay JNK activity, as described previously (19). Briefly, T lymphocytes (1 ϫ 10 7 ; 1 ϫ 10 6 /ml) were pretreated with ␤-oxa 21:3n-3 (20 M) or an equivalent amount of DPC and then stimulated with PMA (10 ng/ml) and A23187 (1 M) for 30 min.…”