arachidonyltrifluoromethane and Inflammation

TCDD (dioxin) induces a rapid inflammatory response from 3T3-L1 adipocytes as judged by prominent induction of the mRNA expression of prostaglandin-endperoxide synthase 2 (Cox-2) along with other inflammation markers within 1 h. This action of TCDD is clearly antagonized by cell pretreatment with AACOCF3 (an inhibitor of cPLA2), nifedipine (a Ca(2+) channel blocker), or 3'-methyl-4'-nitroflavone (MNF), an antagonist of the Ah receptor (AhR), suggesting the possible involvement of the nongenomic pathway of action of TCDD as shown previously in MCF10A cells [Dong, B., and Matsumura, F. (2008) Mol. Pharmacol. 74 (1), 255-263]. This early inflammatory action of TCDD is clearly different from that mediated by its classical action pathway in that the former is mediated by protein kinases such as PKC, PKA, and tyrosine kinases, but not by ARNT. Furthermore, the former is not blocked by two "DRE-decoy" treatments. Such an inflammatory effect of TCDD on 3T3-L1 adipocyes persists at least for 5 days, when the affected adipocytes exhibit significant reduction in their adipocyte characteristics. To assess the cause for the long-lasting influence of this nongenomic action of TCDD, we tested the effects of AACOCF3, exogenous arachidonic acid (AA), and H89 (an inhibitor of PKA) on the 5 day action of TCDD. These agents clearly antagonized all the long-term actions of TCDD except that on CYP1A1 induction, indicating that the influence of the nongenomic action of TCDD lasts a long time in this cell material. One of the major factors mediating its long-lasting effects has been identified to be PKA.

Topics: 3T3-L1 Cells; Adipocytes; Animals; Arachidonic Acid; Arachidonic Acids; Cyclic AMP-Dependent Protein Kinases; Inflammation; Isoquinolines; Mice; Polychlorinated Dibenzodioxins; Sulfonamides; Time Factors

Lysophosphatidylcholine (LPC) is the major bioactive lipid component of oxidized LDL, thought to be responsible for many of the inflammatory effects of oxidized LDL described in both inflammatory and endothelial cells. Inflammation-induced transformation of vascular smooth muscle cells from a contractile phenotype to a proliferative/secretory phenotype is a hallmark of the vascular remodeling that is characteristic of atherogenesis; however, the role of LPC in this process has not been fully described. The present study tested the hypothesis that LPC is an inflammatory stimulus in coronary artery smooth muscle cells (CASMCs). In cultured human CASMCs, LPC stimulated time- and concentration-dependent release of arachidonic acid that was sensitive to phospholipase A2 and C inhibition. LPC stimulated the release of arachidonic acid metabolites leukotriene-B4 and 6-keto-prostaglandin F1alpha, within the same time course. LPC was also found to stimulate basic fibroblast growth factor release as well as stimulating the release of the cytokines GM-CSF, IL-6, and IL-8. Optimal stimulation of these signals was obtained via palmitic acid-substituted LPC species. Stimulation of arachidonic acid, inflammatory cytokines and growth factor release, implies that LPC might play a multifactorial role in the progression of atherosclerosis, by affecting inflammatory processes.

Topics: 6-Ketoprostaglandin F1 alpha; Arachidonic Acid; Arachidonic Acids; Cells, Cultured; Coronary Vessels; Cytokines; Fibroblast Growth Factor 2; Humans; Inflammation; Leukotriene B4; Lipids; Lysophosphatidylcholines; Myocytes, Smooth Muscle; Phospholipases; Tritium

Group IIA secretory phospholipase A(2) (sPLA(2)-IIA) is a prototypic sPLA(2) enzyme that may play roles in modification of eicosanoid biosynthesis as well as antibacterial defense. In several cell types, inducible expression of sPLA(2) by pro-inflammatory stimuli is attenuated by group IVA cytosolic PLA(2) (cPLA(2)alpha) inhibitors such as arachidonyl trifluoromethyl ketone, leading to the proposal that prior activation of cPLA(2)alpha is required for de novo induction of sPLA(2). However, because of the broad specificity of several cPLA(2)alpha inhibitors used so far, a more comprehensive approach is needed to evaluate the relevance of this ambiguous pathway. Here, we provide evidence that the induction of sPLA(2)-IIA by pro-inflammatory stimuli requires group VIB calcium-independent PLA(2) (iPLA(2)gamma), rather than cPLA(2)alpha, in rat fibroblastic 3Y1 cells. Results with small interfering RNA unexpectedly showed that the cytokine induction of sPLA(2)-IIA in cPLA(2)alpha knockdown cells, in which cPLA(2)alpha protein was undetectable, was similar to that in replicate control cells. By contrast, knockdown of iPLA(2)gamma, another arachidonyl trifluoromethyl ketone-sensitive intracellular PLA(2), markedly reduced the cytokine-induced expression of sPLA(2)-IIA. Supporting this finding, the R-enantiomer of bromoenol lactone, an iPLA(2)gamma inhibitor, suppressed the cytokine-induced sPLA(2)-IIA expression, whereas (S)-bromoenol lactone, an iPLA(2)beta inhibitor, failed to do so. Moreover, lipopolysaccharide-stimulated sPLA(2)-IIA expression was also abolished by knockdown of iPLA(2)gamma. These findings open new insight into a novel regulatory role of iPLA(2)gamma in stimulus-coupled sPLA(2)-IIA expression.

Topics: Animals; Arachidonic Acids; Calcium; Cell Line; Cytokines; Eicosanoids; Enzyme Induction; Fibroblasts; Group II Phospholipases A2; Group VI Phospholipases A2; Inflammation; Lipopolysaccharides; Naphthalenes; Phosphodiesterase Inhibitors; Phospholipases A; Phospholipases A2; Pyrones; Rats

Cytosolic phospholipase A(2) (cPLA(2)) plays a pivotal role in inflammation by catalyzing the release of arachidonic acid, a substrate for lipoxygenase and cyclooxygenase enzymes, from membrane phospholipids. In the present study we examined the role of cPLA(2) in inflammatory responses through the use of a specific inhibitor of the enzyme, cPLA(2), arachidonyl trifluoromethyl ketone (AACOCF3). Interestingly, we observed that AACOCF3 is an inhibitor of chronic but not acute inflammatory responses. Specifically, AACOCF3 inhibited phorbol 12-myristate 13-acetate (PMA)-induced chronic ear edema in mice. Additionally, oral treatment of ovalbumin-sensitized/ovalbumin-challenged BALB/c mice with 20 mg/kg AACOCF3 prevented the development of airway hyper-responsiveness in a model of asthma. Furthermore, AACOCF3 decreased cellular recruitment in the airway lumen and airway inflammation after the ovalbumin challenge. Taken together, these results suggest that a potent and specific chemical inhibitor of cPLA(2) may be useful for the treatment of chronic inflammatory diseases including rheumatoid arthritis, inflammatory bowel disease, psoriasis, and asthma.

Topics: Allergens; Animals; Arachidonic Acids; Asthma; Cell Movement; Chronic Disease; Cytosol; Dose-Response Relationship, Drug; Drug Delivery Systems; Edema; Enzyme Inhibitors; Female; Group IV Phospholipases A2; Inflammation; Leukocytes; Male; Mice; Mice, Inbred BALB C; Phospholipases A; Phospholipases A2

Multiple sclerosis (MS) is an inflammatory demyelinating disease of the central nervous system (CNS) that results in motor and sensory deficits. Although MS and its animal model, experimental autoimmune encephalomyelitis (EAE), are thought to be T cell-mediated diseases, the mechanisms underlying the lesions in the CNS are not fully understood. We propose that a strong candidate as a central mediator in evoking the complex pathological changes seen in MS and EAE is the enzyme cytosolic phospholipase A2 (cPLA2). One of the metabolic products of this enzyme is pro-inflammatory, while the other induces myelin breakdown, demyelination, and chemokine/cytokine expression. We provide evidence that cPLA2 is highly expressed in EAE lesions and show that blocking this enzyme leads to a remarkable reduction in the onset and progression of EAE.

Topics: Animals; Arachidonic Acids; Axons; Blotting, Western; CD4-Positive T-Lymphocytes; Cell Count; Chemokines; Cyclooxygenase 2; Cytokinins; Cytosol; Disease Models, Animal; Endothelial Cells; Enzyme Inhibitors; Female; Gene Expression; Immunohistochemistry; Inflammation; Isoenzymes; Macrophages; Mice; Mice, Inbred C57BL; Microscopy, Electron; Multiple Sclerosis; Oligonucleotide Array Sequence Analysis; Phospholipases A; Phospholipases A2; Prostaglandin-Endoperoxide Synthases; Random Allocation; Receptors, Chemokine; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Spinal Cord; Time Factors

Monocyte chemoattractant protein 1 (MCP-1) has an important influence on monocyte migration into sites of inflammation. Our understanding of the signal transduction pathways involved in the response of monocytes to MCP-1 is quite limited yet potentially significant for understanding and manipulating the inflammatory response. Prior studies have demonstrated a crucial regulatory role for cytosolic phospholipase A(2) (cPLA(2)) in monocyte chemotaxis to MCP-1. In these studies we investigated the role for another PLA(2), calcium-independent PLA(2) (iPLA(2)) in comparison to cPLA(2). Pharmacological inhibitors of PLA(2) were found to substantially inhibit chemotaxis. Using antisense oligodeoxyribonucleotide treatment we found that iPLA(2) expression is required for monocyte migration to MCP-1. Complete blocking of the chemotactic response was observed with inhibition of either iPLA(2) or cPLA(2) expression by their respective antisense oligodeoxyribonucleotide. In reconstitution experiments, lysophosphatidic acid completely restored MCP-1-stimulated migration in iPLA(2)-deficient monocytes, whereas lysophosphatidic acid was without effect in restoring migration in cPLA(2)-deficient monocytes. To the contrary, arachidonic acid fully restored migration of cPLA(2)-deficient monocytes while having no effect on the iPLA(2)-deficient monocytes. Additional studies revealed that neither enzyme appears to be upstream of the other indicating that iPLA(2) and cPLA(2) represent parallel regulatory pathways. These data demonstrate novel and distinct roles for these two phospholipases in this critical step in inflammation.

Topics: Aminobenzoates; Arachidonic Acid; Arachidonic Acids; Aristolochic Acids; Chemokine CCL2; Chemotaxis, Leukocyte; Chlorobenzoates; Cinnamates; Enzyme Inhibitors; Fatty Acids; Group IV Phospholipases A2; Group VI Phospholipases A2; Humans; Inflammation; Lysophospholipids; Monocytes; Naphthalenes; Oligodeoxyribonucleotides, Antisense; ortho-Aminobenzoates; Phenanthrenes; Phospholipases A; Pyrones; Signal Transduction