scholarly journals Agonist-induced down-regulation of platelet-activating factor receptor gene expression in U937 cells

1994 ◽  
Vol 301 (3) ◽  
pp. 911-916 ◽  
Author(s):  
L Y Chau ◽  
K Peck ◽  
H H Yen ◽  
J Y Wang
Keyword(s):  
Prolonged Exposure ◽  
Radioligand Binding ◽  
Receptor Gene ◽  
Platelet Activating Factor ◽  
State Level ◽  
Transcriptional Level ◽  
U937 Cells ◽  
Down Regulation ◽  
Receptor Mrna ◽  
Paf Receptor

Prolonged exposure (8-24 h) of human promonocytic U937 cells to 100 nM 1-O-hexadecyl-2-N-methylcarbamyl-sn-glycero-3-phosphocholine (carbarmyl-PAF), a non-metabolizable analogue of platelet-activating factor (PAF), reduced the numbers of PAF receptors by 50-75%, as determined by the radioligand-binding assay. To clarify whether the down-regulation of receptor numbers is due to decreased expression level of the PAF-receptor gene, the effect of carbamyl-PAF on the steady-state level of PAF-receptor mRNA was examined by a highly sensitive reverse-transcriptase PCR method. A 50% decline in the level of PAF-receptor mRNA was observed in U937 cells pretreated with 100 nM carbamyl-PAF for 24 h. The effect of carbamyl-PAF was dose-dependent, with an EC50 value around 10 nM. PAF-receptor antagonist, SRI-63675, was able to attenuate the effect of carbamyl-PAF. Furthermore lysoPAF, at 1 uM, was unable to induce a significant decrease in PAF-receptor mRNA after incubation for 24 h, indicating that the effect of carbamyl-PAF was specific. The half-life of the PAF-receptor mRNA measured in the presence of actinomycin D was unaffected by carbamyl-PAF treatment. In contrast, nuclear run-off experiments demonstrated that the transcription rate of the PAF-receptor gene in carbamyl-PAF-treated cells was about 65% of that in control cells. These results suggest that the PAF receptor in U937 cells is subject to down-regulation by agonist, at least partly, at the transcriptional level.

Oxidized low-density lipoprotein and peroxisome-proliferator-activated receptor α down-regulate platelet-activating-factor receptor expression in human macrophages

2001 ◽  
Vol 354 (1) ◽  
pp. 225-232 ◽  
Author(s):  
Delphine HOURTON ◽  
Philippe DELERIVE ◽  
Jana STANKOVA ◽  
Bart STAELS ◽  
M. John CHAPMAN ◽  
...  
Keyword(s):  
Low Density Lipoprotein ◽  
Platelet Activating Factor ◽  
Density Lipoprotein ◽  
Receptor Expression ◽  
Peroxisome Proliferator ◽  
Low Density ◽  
Peroxisome Proliferator Activated Receptor ◽  
Receptor Mrna ◽  
Paf Receptor ◽  
Pparα Agonist

Regulation of the expression of platelet-activating factor (PAF) receptor by atherogenic lipoproteins might contribute to atherogenesis. We show that progressive oxidation of low-density lipoprotein (LDL) gradually inhibits PAF receptor expression on the macrophage cell surface. We tested the effect of oxidized LDL (oxLDL) on PAF receptor expression in human monocytes that do not contain peroxisome-proliferator-activated receptor γ (PPARγ), a nuclear receptor activated by oxLDL. OxLDL decreased by 50% (P ⩽0.001) and by 29% (P⩽0.05) the binding of PAF and the expression of PAF receptor mRNA respectively. Next we demonstrated that progressive oxidation of LDLs significantly activated PPARα-dependent transcription in transfected mouse aortic endothelial cells. Finally we demonstrated, in mature macrophages, that fenofibrate (20µM), a specific PPARα agonist, but not the specific PPARγ agonist BRL49653 (20nM), significantly decreased both PAF binding and PAF receptor mRNA expression, by 65% and 40% (P⩽0.001) respectively. Additionally, another PPARα agonist, Wy14,643, decreased PAF receptor promoter activity by 70% (P⩽0.05) in transfected THP-1 cells, suggesting the involvement of the proximal promoter region (-980 to -500) containing a series of four nuclear factor (NF)-κB motifs. Thus PPARα might be involved in the down-regulation of PAF receptor gene expression by oxLDLs in human monocytes/macrophages. The oxidation of one or more lipid components of LDLs might result in the formation of natural activators of PPARα. It is hypothesized that such activators might modulate inflammation and apoptosis upon atherogenesis by decreasing the expression of PAF receptor.

scholarly journals Expression of human platelet-activating factor receptor gene in EoL-1 cells following butyrate-induced differentiation

1995 ◽  
Vol 305 (3) ◽  
pp. 829-835 ◽  
Author(s):  
T Izumi ◽  
S Kishimoto ◽  
T Takano ◽  
M Nakamura ◽  
Y Miyabe ◽  
...  
Keyword(s):  
Receptor Gene ◽  
Platelet Activating Factor ◽  
Allergic Inflammation ◽  
Genomic Analysis ◽  
Receptor Protein ◽  
Cell Surface Expression ◽  
Lipid Mediator ◽  
Mrna Accumulation ◽  
Leukaemia Cell Line ◽  
Paf Receptor

Platelet-activating factor (PAF) is a potent lipid mediator of allergic inflammation through its interaction with eosinophils. Expression of the PAF receptor is modulated by many agents, including those responsible for cell differentiation. We report here that differentiation of a human eosinophilic leukaemia cell line, EoL-1, by sodium n-butyrate is associated with induction of PAF receptor gene expression, as indicated by: PAF receptor mRNA accumulation; increases in the binding of [3H]WEB 2086, a PAF antagonist; analysis of cell-surface expression of PAF receptor protein using a monoclonal anti-(PAF receptor) antibody; and augmentation of PAF-induced increase in the intracellular concentration of calcium. Using cDNA cloning, the receptor expressed in EoL-1 cells was identified as ‘Transcript 1’, one of two transcripts which was previously reported from human genomic analysis (Mutoh, Bito, Minami, Nakamura, Honda, Izumi, Nakata, Kurachi, Terano and Shimizu (1993) FEBS Lett. 322, 129-134). The PAF-induced calcium response and phosphoinositide turnover were decreased by pertussis toxin (PTX) treatment, suggesting that these signals are coupled largely with PTX-sensitive G-protein(s) in EoL-1 cells. These systems may provide a useful experimental model with which to investigate the relationship between eosinophilic differentiation and PAF receptor induction, and the role of eosinophils in allergic responses.

Platelet-activating factor receptors in lamb lungs are downregulated immediately after birth

2000 ◽  
Vol 278 (4) ◽  
pp. H1168-H1176 ◽  
Author(s):  
Basil O. Ibe ◽  
Fred C. Sander ◽  
J. Usha Raj
Keyword(s):  
Receptor Binding ◽  
Dissociation Constants ◽  
Receptor Gene ◽  
Choline Chloride ◽  
Platelet Activating Factor ◽  
Scatchard Analysis ◽  
Newborn Period ◽  
Paf Receptor ◽  
G Protein Coupled ◽  
Receptor Gene Expression

Platelet-activating factor (PAF) is a phospholipid with diverse biological functions mediated by a G protein-coupled receptor. We determined PAF receptor binding in lung membranes of four groups of perinatal lambs. Membrane protein (100 μg/ml) was incubated for 60 min at 30°C with 0.5–24 nM of acetyl-[3H]PAF in 30 mM Tris buffer, pH 7.2, containing 0.25% BSA, 10 mM MgCl2, and 125 mM choline chloride. PAF bound to membrane was isolated and quantified by scintillation spectrometry, followed with Scatchard analysis for receptor density (Bmax). The Bmax (means ± SE, fmol/mg protein) were 445.8 ± 12.3, 244.2 ± 3.3, 250.6 ± 3.6, and 419.9 ± 8.6 for the fetal, 90-min-old, <1-day-old, and 6- to 12-day-old lambs, respectively. The Bmax for the 90-min-old and <1-day-old lambs were not different but were significantly lower than those of either the term fetal or 6- to 12-day-old lambs. These data show a significant decrease in PAF binding to its receptor and in PAF Bmax in lung membranes of immediate newborn lambs. The dissociation constants ( K D, nM) were 7.7 ± 0.52, 11.5 ± 0.34, 6.9 ± 0.48, and 5.0 ± 0.53 for fetal, 90-min-old, <1-day-old, and 6- to 12-day-old newborn lamb lungs, respectively. The K D of the 90-min-old lamb was the highest of all. PAF receptor gene measured by RT-PCR showed a significant downregulation of PAF receptor gene mRNA in lungs of lambs <1 day old, suggesting a transcriptional regulation of PAF receptor gene expression in the immediate newborn period. We speculate that decreased PAF receptor binding immediately after birth will facilitate the fall in pulmonary vascular resistance in the immediate newborn period.

Down-Regulation of Vasopressin VlA Receptor mRNA in Diabetes Mellitus in the Rat

1995 ◽  
Vol 88 (6) ◽  
pp. 671-674 ◽  
Author(s):  
Paddy A. Phillips ◽  
John Risvanis ◽  
Anne-Marie Hutchins ◽  
Louise M. Burrell ◽  
Duncan MacGregor ◽  
...  
Keyword(s):  
Diabetes Mellitus ◽  
Radioligand Binding ◽  
Inositol Phosphates ◽  
Density Ratio ◽  
Northern Blotting ◽  
Diabetic Rats ◽  
Down Regulation ◽  
Receptor Mrna ◽  
V1a Receptor ◽  
Streptozotocin Induced Diabetes

1. To investigate the mechanism of hepatic V1a vasopressin receptor down-regulation in streptozotocin-induced diabetes mellitus in the rat, we measured hepatic V1a receptor mRNA by in situ hybridization histochemistry using oligonucleotide probes to the V1a receptor and Northern blotting. 2. Diabetes mellitus caused hyperglycaemia, hyperosmolality and increased plasma vasopressin concentrations (P < 0.01). Hepatoycte V1a receptor mRNA was reduced by 76% in diabetic rats (P < 0.01) and by 53% in insulin-treated diabetic rats (P < 0.01) versus control rats, in parallel with reduced V1a radioligand binding and vasopressin-stimulated inositol phosphates production. There was a similar decrease in hepatic V1a/18S mRNA density ratio in the diabetic and diabetic + insulin groups (both P < 0.05 versus control). 3. These findings suggest that altered V1a mRNA transcription is responsible for the reduced hepatic V1a receptor density in diabetes mellitus.

scholarly journals Regulation of platelet-activating-factor receptors and the desensitization response in polymorphonuclear neutrophils

1992 ◽  
Vol 288 (1) ◽  
pp. 241-248 ◽  
Author(s):  
J T O'Flaherty ◽  
D P Jacobson ◽  
J F Redman
Keyword(s):  
Actinomycin D ◽  
Platelet Activating Factor ◽  
Receptor Expression ◽  
Polymorphonuclear Neutrophils ◽  
Target Cells ◽  
Surface Compound ◽  
Down Regulation ◽  
High Affinity ◽  
Kinase C ◽  
Paf Receptor

Platelet-activating factor (PAF) desensitizes as well as stimulates its various target cells, We find that human polymorphonuclear neutrophils (PMN) exposed to PAF became maximally unresponsive to a second PAF challenge within 15-90 s in assays of Ca2+ mobilization and degranulation. The cells regained full PAF-sensitivity over the ensuing 20-40 min. These effects correlated with changes in PAF receptor availability. PMN treated with PAF, washed in regular buffer and assayed for PAF binding exhibited falls (maximal in 15 s), followed by rises (reaching control levels by 60 min), in the number of high-affinity PAF receptors. However, tracking studies showed that [3H]PAF accumulated on the cell surface for approximately 2 min before being internalized. Regular-buffer washes did not remove this superficial PAF, whereas a washing regimen using excess albumin to adsorb PAF removed 99% of the surface compound. PMN washed by the latter regimen after PAF exposure lost PAF receptors relatively slowly (maximal at approximately 5 min), but the ultimate extent of this loss and the rate at which receptor expression normalized were similar to those of cells washed in regular buffer. Neither cycloheximide nor actinomycin D influenced the course of the receptor changes, but two protein kinase C (PKC) blockers, staurosporine and 1-(5-isoquinolinesulphonyl)piperazine, inhibited the receptor-receptor-depleting actions of PAF. Indeed, a phorbol diester activator of PKC also caused PMN to decrease high-affinity PAF receptor numbers, and the two PKC blockers antagonized this action at concentrations that inhibited PAF-induced PAF receptor losses. We conclude that: (a) PAF induces PMN to down-regulate and then to re-express PAF receptors independently of protein synthesis; (b) these changes are likely to underlie the later stages and reversal of desensitization; (c) the onset (t < or = 2 min) of desensitization, however, precedes receptor down-regulation and must be due to receptor uncoupling from transductional elements; and (d) down-regulation of receptors for PAF appears to be mediated by PKC and/or elements inhibited by PKC blockers.

scholarly journals Regulation of nerve growth factor receptor gene expression by nerve growth factor in the developing peripheral nervous system.

1991 ◽  
Vol 112 (2) ◽  
pp. 303-312 ◽  
Author(s):  
F D Miller ◽  
T C Mathew ◽  
J G Toma
Keyword(s):  
Gene Expression ◽  
Nerve Growth Factor ◽  
Growth Factor ◽  
Receptor Gene ◽  
Transcriptional Level ◽  
Mrna Levels ◽  
Nerve Growth ◽  
Receptor Mrna ◽  
Ngf Receptor ◽  
Receptor Gene Expression

Nerve growth factor (NGF) is a target-derived neurotrophic protein that promotes the survival and growth of developing sympathetic and sensory neurons. We have examined NGF receptor gene expression in these neurons after NGF administration. Northern blot and in situ hybridization analyses demonstrated that NGF given systemically to neonatal rats increased levels of NGF receptor mRNA in sympathetic neurons within the superior cervical ganglion. This increase was accompanied by a differential regulation of genes associated with neurotransmitter phenotype; tyrosine hydroxylase mRNA was increased, but neuropeptide Y mRNA was not. NGF receptor mRNA levels were also increased in L4-L5 dorsal root ganglia, although this mRNA was not expressed uniformly in sensory neurons of control or NGF-treated animals. Levels of T alpha 1 alpha-tubulin mRNA, a marker of neuronal growth, also increased. In contrast to developing neurons, systemic NGF did not increase NGF receptor mRNA in nonneuronal cells of the sciatic nerve. To determine if NGF regulated NGF receptor gene expression at the transcriptional level, we examined PC12 cells. NGF treatment for 6 h increased NGF receptor mRNA fourfold; this increase was inhibited by cycloheximide. Nuclear run-off transcription assays demonstrated that the increase in steady-state NGF receptor mRNA levels was mediated at the transcriptional level. In contrast, although NGF treatment increased steady-state tyrosine hydroxylase mRNA levels, this effect was not blocked by cycloheximide, and was not due to increased transcription. These data raise the possibility that transcriptional regulation of NGF receptor gene expression by target-derived NGF could be a molecular mechanism for potentiating NGF's effects on neurons during developmental periods of neuronal competition and cell death.

cAMP mediates homologous downregulation of PAF receptor mRNA expression in mesangial cells

1997 ◽  
Vol 273 (3) ◽  
pp. F445-F450 ◽  
Author(s):  
A. Nakao ◽  
T. Watanabe ◽  
H. Bitoh ◽  
H. Imaki ◽  
T. Suzuki ◽  
...  
Keyword(s):  
Mrna Expression ◽  
Northern Blot Analysis ◽  
Mesangial Cells ◽  
Platelet Activating Factor ◽  
Receptor Mrna ◽  
A Cell ◽  
Paf Receptor ◽  
Camp Formation ◽  
Homologous Desensitization ◽  
Paf Receptor Antagonist

To clarify the molecular mechanism and significance of homologous desensitization of platelet-activating factor (PAF) signaling, we examined the effect of PAF on PAF receptor mRNA expression in rat mesangial cells by Northern blot analysis. Treatment of the cells with PAF (10(-7)-10(-8) M) reduced the expression of PAF receptor mRNA in 1 h, and this reduction was recovered by pretreatment of mesangial cells with a specific PAF receptor antagonist, WEB-2086 (10(-6) M), or a cyclooxygenase inhibitor, indomethacin (10(-6) M), for 10 min. PAF-stimulated prostaglandin E2 (PGE2) and adenosine 3',5'-cyclic monophosphate (cAMP) formation was measured by radioimmunoassays specific for each substance. Reduction of PAF receptor mRNA expression was mimicked by treatment of the cells with PGE2 (10(-6) M) or dibutyryl-cAMP (10(-3) M), a cell-permeable analog of cAMP. These results, taken together, suggest that PAF receptor mRNA expression is downregulated by exposure to PAF in cultured mesangial cells. This homologous downregulation is mediated by cAMP production through PGE2 synthesis.

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