Sodium Fluoride Mimics the Effect of Prostaglandin E2 on Catecholamine Release from Bovine Adrenal Chromaffin Cells

We have reported recently that prostaglandin E2 (PGE2) stimulated phosphoinositide metabolism in bovine adrenal chromaffin cells and that PGE2 and ouabain, an inhibitor of Na+, K(+)-ATPase, synergistically induced a gradual secretion of catecholamines from the cells. Here we examined the involvement of a GTP-binding protein(s) in PGE receptor-induced responses by using NaF. In the presence of Ca2+ in the medium, NaF stimulated the formation of all three inositol phosphates, i.e., inositol monophosphate, bisphosphate, and trisphosphate, linearly over 30 min in a dose-dependent manner (15-30 mM). This effect on phosphoinositide metabolism was accompanied by an increase in cytosolic free Ca2+. NaF also induced catecholamine release from chromaffin cells, and the dependency of stimulation of the release on NaF concentration was well correlated with those of NaF-enhanced inositol phosphate formation and increase in cytosolic free Ca2+. Although the effect of NaF on PGE2-induced catecholamine release in the presence of ouabain was additive at concentrations below 20 mM, there was no additive effect at 25 mM NaF. Furthermore, the time course of catecholamine release stimulated by 20 mM NaF in the presence of ouabain was quite similar to that by 1 microM PGE2, and both stimulations were markedly inhibited by amiloride, with half-maximal inhibition at 10 microM. Pretreatment of the cells with pertussis toxin did not prevent, but rather enhanced, PGE2-induced catecholamine release over the range of concentrations examined. These results demonstrate that NaF mimics the effect of PGE2 on catecholamine release from chromaffin cells and suggest that PGE2-evoked catecholamine release may be mediated by the stimulation of phosphoinositide metabolism through a putative GTP-binding protein insensitive to pertussis toxin.     

Elicitor-induced hydroxycinnamoyl-CoA:tyramine hydroxycinnamoyltransferase in plant cell suspension cultures

Treatment of Nicotiana glutinosa L. cell suspension cultures with chitosan results in the co-induction of phenylalanine ammonia lyase, 4-eoumarate:CoA ligase, tyrosine decarboxylase and tyramine hydroxycinnamoyltransferase, all involved in the biosynthesis of hydroxycinnamoyltyramines. The highest enzyme activities were observed around 12 h after addition of 0.8 to 1 mg chitosan per g fresh weight of cells. No hydroxycinnamoyltyramines could be detected by TLC or HPLC of extracts made from non-treated or elicited cells. [¹⁴C]-Tyramine was incorporated into insoluble polymeric material at a higher rate in elicitor-treated than in non-treated cells of N. glutinosa. Tyramine hydroxycinnamoyltransferase could be induced in suspension cultured cells of Eschscholtzia califortnca Cham, but not in cells of Phaseolus vulgaris L. or Catharanthus roseus (L.) G. Don by addition of a yeast elicitor to the growth medium.     

Adipose triglyceride lipase regulates eicosanoid production in activated human mast cells

Human mast cells (MCs) contain TG-rich cytoplasmic lipid droplets (LDs) with high arachidonic acid (AA) content. Here, we investigated the functional role of adipose TG lipase (ATGL) in TG hydrolysis and the ensuing release of AA as substrate for eicosanoid generation by activated human primary MCs in culture. Silencing of ATGL in MCs by siRNAs induced the accumulation of neutral lipids in LDs. IgE-dependent activation of MCs triggered the secretion of the two major eicosanoids, prostaglandin D2 (PGD2) and leukotriene C4 (LTC4). The immediate release of PGD2 from the activated MCs was solely dependent on cyclooxygenase (COX) 1, while during the delayed phase of lipid mediator production, the inducible COX-2 also contributed to its release. Importantly, when ATGL-silenced MCs were activated, the secretion of both PGD2 and LTC4 was significantly reduced. Interestingly, the inhibitory effect on the release of LTC4 was even more pronounced in ATGL-silenced MCs than in cytosolic phospholipase A₂-silenced MCs. These data show that ATGL hydrolyzes AA-containing TGs present in human MC LDs and define ATGL as a novel regulator of the substrate availability of AA for eicosanoid generation upon MC activation.     

Activation of OR1A1 suppresses PPAR-γ expression by inducing HES-1 in cultured hepatocytes

Olfactory receptors (ORs) comprise the largest G protein-coupled receptor gene superfamily. Recent studies indicate that ORs are also expressed in non-olfactory organs, including metabolically active tissues, although their biological functions in these tissues are largely unknown. In this study, OR1A1 expression was detected in HepG2 liver cells. OR1A1 activation by (−)-carvone, a known OR1A1 ligand, increased the cyclic adenosine monophosphate (cAMP), but not intracellular Ca²⁺ concentration, thereby inducing protein kinase A (PKA) activity with subsequent phosphorylation of cAMP response element-binding protein (CREB) and upregulation of the CREB-responsive gene hairy and enhancer of split (HES)-1, a corepressor of peroxisome proliferator-activated receptor-γ (PPAR-γ) in hepatocytes. In (−)-carvone-stimulated cells, the repression of PPAR-γ reduced the expression of the target gene, mitochondrial glycerol-3-phosphate acyltransferase, which encodes a key enzyme involved in triglyceride synthesis. Intracellular triglyceride level and lipid accumulation were reduced in cells stimulated with (−)-carvone, effects that were diminished following the loss of OR1A1 function. These results indicate that OR1A1 may function as a non-redundant receptor in hepatocytes that regulates the PKA-CREB-HES-1 signaling axis and thereby modulates hepatic triglyceride metabolism.     

Chrotacumines G–J,chromone alkaloids from Dysoxylum acutangulum with osteoclast differentiation inhibitory activity

Four new chromone alkaloids, chrotacumines G–J (1–4), have been isolated from the barks of Dysoxylum acutangulum. Their structures and absolute configurations were elucidated on the basis of NMR and CD data. Chrotacumines G and J (1 and 4) showed osteoclast differentiation inhibitory activity in a dose dependent manner.     

Quantitative proteomic profiling identifies global protein network dynamics in murine embryonic heart development

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Regulation of the phosphate (Pi) concentration in UMR 106 osteoblast-like cells: Effect of Pi,Na+ and K+

Osteoblast-like cells possess Na-dependent transporters which accumulate orthophosphate (Pi) from the extracellular medium. This may be important in bone formation. Here we describe parallel measurements of Pi uptake and cellular [Pi] in such cells from the rat (UMR 106–01 and UMR 106–06) and human (OB), and in non-osteoblastic human fibroblasts (Detroit 532 (DET)). In UMR 106–01, cellular [Pi] was weakly dependent on extracellular [Pi] and higher than expected from passive transport alone. [³²Pi]-uptake was inhibited by Na deprivation, but paradoxically increased on K deprivation. With Na, 87 per cent of cellular ³²P was found in organic phosphorus pools after only 5 min. Na deprivation also decreased cellular [Pi], in both UMR 106–01 and DET, but the decrease was smaller than that in [³²Pi]-uptake. Ouabain decreased [³²Pi]-uptake and cellular [Pi] in DET, but not in UMR 106–01. Regulation of cellular [Pi] is therefore at least partly dependent on Na/Pi co-transport, but this does not seem to be an exclusive property of osteoblasts.