Arachidonic Acid, a Growth Signal in Murine P815 Mastocytoma Cells1 (original) (raw)
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Arachidonic acid, a growth signal in murine P815 mastocytoma cells
Cancer research, 1993
Evidence is presented that inducing P815 murine mastocytoma cells to grow with serum activates a Ca(2+)-stimulated phospholipase A2 and the rapid release of arachidonic acid by the cells. Slower growth was also maintained by arachidonic acid or its immediate precursors or by diacylglycerols when bovine serum albumin replaced the serum. Together, arachidonic acid and 1-oleoyl-2-acetylglycerol stimulated growth at the same rate as 10% serum consistent with a role for both arachidonic acid and protein kinase C in the response to serum. Arresting cell growth with N6,O2'-dibutyryladenosine 3',5'-cyclic phosphate and theophylline inhibited the release of arachidonic acid in response to serum, suggesting that cyclic AMP prevents phospholipase activation as one of its pleiotypic effects on growth. Attempts to demonstrate metabolism of [3H]arachidonic acid to eicosanoids in serum-treated P815 cells by high-performance liquid chromatography or thin layer chromatography were unsucc...
Biochemistry and Cell Biology, 1997
Our previous studies demonstrated that inhibitors of arachidonate-phospholipid remodeling [i.e. the enzyme CoAindependent transacylase (CoA-IT)] decrease cell proliferation and induce apoptosis in neoplastic cells. The goal of the current study was to elucidate the molecular events associated with arachidonate-phospholipid remodeling that influence cell proliferation and survival. Initial experiments revealed the essential nature of cellular arachidonate to the signaling process by demonstrating that HL-60 cells depleted of arachidonate were more resistant to apoptosis induced by CoA-IT inhibition. In cells treated with CoA-IT inhibitors a marked increase in free arachidonic acid and AA-containing triglycerides were measured. TG enrichment was likely due to acylation of arachidonic acid into diglycerides and triglycerides via de novo glycerolipid biosynthesis. To determine the potential of free fatty acids to affect cell proliferation, HL-60 cells were incubated with varying concentrations of free fatty acids; exogenously provided 20-carbon polyunsaturated fatty acids caused a dose-dependent inhibition of cell proliferation, whereas oleic acid was without effect. Blocking 5-lipoxygenase or cyclooxygenases had no effect on the inhibition of cell proliferation induced by arachidonic acid or CoA-IT inhibitors. An increase in cell-associated ceramides (mainly in the 16:0-ceramide fraction) was measured in cells exposed to free arachidonic acid or to CoA-IT inhibitors. This study, in conjunction with other recent studies, suggests that perturbations in the control of cellular arachidonic acid levels affect cell proliferation and survival.
Biochemical and Biophysical Research Communications, 1986
Arachidonic acid, linolenic acid and 14 different oxygenated fatty acid derivatives were tested as activators of human protein kinase C in vitro using histone as substrate. Lipoxin A (5,6,15L-trihydroxy-7,9,11,13-eicosatetraenoic activated the kinase in the presence of calcium at 30 fold lower concentration (1 microM) than did arachidonic acid or 1,3-dioleoylglycerol. The methyl ester of lipoxin A and the free acids of leukotriene B4 as well as two lipoxin B isomers were without effect. In contrast, linolenic acid, leukotriene C4, certain mono- and dihydroxylated eicosanoids and one lipoxin B isomer had stimulatory effects, albeit at higher concentrations. The substrate specificity of protein kinase C activated by lipoxin A proved to be different from that of the phosphatidylserine or phorbol ester activated kinase. Results of the present study suggest that arachidonic acid derived oxygenation products, in particular lipoxin A, may serve as intracellular activators of protein kinase C.
Journal of Biological Chemistry, 1996
Through analysis of the effects of defined serum components and growth supplements, we reveal here that the factors in serum responsible for inducing recovery of Ca 2؉ pools and growth in thapsigargin-arrested DDT 1 MF-2 cells are exactly mimicked by the three essential fatty acids, arachidonic, linoleic, and ␣-linolenic acids. The EC 50 values for arachidonic and linoleic acids on growth induction of thapsigargin-arrested cells were the same, approximately 5 M. Nonessential fatty acids, including myristic, palmitic, stearic, oleic, and arachidic acids, were without any effect. Although not proven to be the active component of serum, levels of arachidonic and linoleic acids in serum were sufficient to explain serum-induced growth recovery. Significantly, arachidonic or linoleic acids induced complete recovery of bradykinin-sensitive Ca 2؉ pools within 6 h of treatment of thapsigarginarrested cells. Protein synthesis inhibitors (cycloheximide or puromycin) completely blocked the appearance of serum-induced or arachidonic acid-induced agonistsensitive pools. The sensitivity and fatty acid specificity of Ca 2؉ pool recovery in thapsigargin-arrested cells were almost identical to that for growth recovery. No pool or growth recovery was observed with 5,8,11,14eicosatetraynoic acid, the nonmetabolizable analogue of arachidonic acid, suggesting that conversion to eicosanoids underlies the pool and growth recovery induced by essential fatty acids. The results provide not only further information on the link between Ca 2؉ pools and cell growth but also evidence for a potentially important signaling pathway involved in inducing transition from a stationary to a proliferative growth state.
Journal of Biological Chemistry, 1998
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.
Endocrinology, 1999
Prior studies have shown that 24,25-dihydroxyvitamin D 3 2 D 3 ] plays a major role in resting zone chondrocyte differentiation and that this vitamin D metabolite regulates both phospholipase A 2 and protein kinase C (PKC) specific activities. Arachidonic acid is the product of phospholipase A 2 action and has been shown in other systems to affect a variety of cellular functions, including PKC activity. The aim of the present study was to examine the interrelationship between arachidonic acid and 24,25-(OH) 2 D 3 on markers of proliferation, differentiation, and matrix production in resting zone chondrocytes and to characterize the mechanisms by which arachidonic acid regulates PKC, which was shown previously to mediate the rapid effects of 24,25-(OH) 2 D 3 and arachidonic acid on these cells. Confluent, fourth passage resting zone cells from rat costochondral cartilage were used to evaluate these mechanisms. The addition of arachidonic acid to resting zone cultures stimulated [ 3 H]thymidine incorporation and inhibited the activity of alkaline phosphatase and PKC, but had no effect on proteoglycan sulfation. In contrast, 24,25-(OH) 2 D 3 inhibited [ 3 H]thymidine incorporation and stimulated alkaline phosphatase, proteoglycan sulfation, and PKC activity. In cultures treated with both agents, the effects of 24,25-(OH) 2 D 3 were reversed by arachidonic acid. The PKC isoform affected by arachidonic acid was PKC␣; cytosolic levels were decreased, but membrane levels were unaffected, indicating that translocation did not occur. Arachidonic acid had a direct effect on PKC in isolated plasma membranes and matrix vesicles, indicating a nongenomic mechanism. Plasma membrane PKC␣ was inhibited, and matrix vesicle PKC was stimulated; these effects were blocked by 24,25-(OH) 2 D 3 . Studies using cyclooxygenase and lipoxygenase inhibitors indicate that the effects of arachidonic acid are due in part to PG production, but not to leukotriene production. This is supported by the fact that H8-dependent inhibition of protein kinase A, which mediates the effects of PGE 2 , had no effect on the direct action of arachidonic acid but did mediate the role of arachidonic acid in the cell response to 24,25-(OH) 2 D 3 . Diacylglycerol does not appear to be involved, indicating that phospholipase C and/or D do not play a role. ␥-Linolenic acid, an unsaturated precursor of arachidonic acid, elicited a similar response in matrix vesicles but not plasma membranes, whereas palmitic acid, a saturated fatty acid, had no effect. These data suggest that arachidonic acid may act as a negative regulator of 24,25-(OH) 2 D 3 action in resting zone chondrocytes. (Endocrinology 140: 2991(Endocrinology 140: -3002, 1999) )
International Immunopharmacology, 2009
Arachidonic acid (AA) and its metabolites have recently generated a heightened interest due to growing evidence of their significant role in cancer biology. Thus, inhibitors of the AA cascade, first and foremost COX inhibitors, which have originally been of interest in the treatment of inflammatory conditions and certain types of cardiovascular disease, are now attracting attention as an arsenal against cancer. An increasing number of investigations support their role in cancer chemoprevention, although the precise molecular mechanisms that link levels of AA, and its metabolites, with cancer progression have still to be elucidated. This article provides an overview of the AA cascade and focuses on the roles of its inhibitors and their implication in cancer treatment. In particular, emphasis is placed on the inhibition of cell proliferation and neo-angiogenesis through inhibition of the enzymes COX-2, 5-LOX and CYP450. Downstream effects of inhibition of AA metabolites are analysed and the molecular mechanisms of action of a selected number of inhibitors of catalytic pathways reviewed. Lastly, the benefits of dietary omega-3 fatty acids and their mechanisms of action leading to reduced cancer risk and impeded cancer cell growth are mentioned. Finally, a proposal is put forward, suggesting a novel and integrated approach in viewing the molecular mechanisms and complex interactions responsible for the involvement of AA metabolites in carcinogenesis and the protective effects of omega-3 fatty acids in inflammation and tumour prevention.