Secretion Of Platelet-activating Factor Acetylhydrolase Research Articles

ROBO4 deletion ameliorates PAF-mediated skin inflammation via regulating the mRNA translation efficiency of LPCAT1/LPCAT2 and the expression of PAF receptor.

The diminished level of platelet-activating factor acetylhydrolase (PAFAH) in milk causes an enhanced level of platelet activating factor (PAF) in the skin, leading to a severe hair loss phenotype during neonatal pup's lactation. The deletion of very-low-density-lipoprotein receptor (VLDLR) prevents the expression and secretion of PAFAH. Here we revealed that deletion of Roundabout 4 (ROBO4) in mice ameliorated hair loss phenotype via reducing PAF concentration in skin. As a consequence, the neonatal pups with ROBO4 deletion lactated by mother with VLDLR deletion showed normal hair phenotype during lactation. In details,ROBO4 deletion reduced the protein but not mRNA expression of two PAF synthetic enzymes LPCAT1/LPCAT2 in macrophage as well as the expression of PAF receptor in both macrophage and ocular tissue, but increased PAFAH protein in serum. On the other hand, RNA expression profile analysis in macrophages revealed that the genes involving in oxidative phosphorylation and ribosome obviously decreased their expression in response to ROBO4 deletion. Moreover, through High Performance Liquid Chromatography (HPLC) analysis, we found that ATP concentration also reduced in ROBO4 deletion macrophages. Because ribosome and energy are very important factors for the mRNA translation, we then tested whether ROBO4 deletion affects LPCAT1/LPCAT2 mRNA translation using polyribosome assay. As expected, the mRNA level of LPCAT1/LPCAT2 significantly decreased in polyribosome in ROBO4 deletion macrophage comparing to that of wild type. Additionally, mice with ROBO4 deletion suppressed LPS-induced IL-6 expression as well as the phosphorylation of p44/42 and p65, but enhanced the AKT phosphorylation. Collectively, ROBO4 deletion alleviates PAF- and LPS-mediated inflammation. And above results also indicate PAF signal might be a crosstalk point of ROBO4- and VLDLR-activated pathways.

PAF-acetylhydrolase activity in plasma of patients with chronic kidney disease. Effect of long-term therapy with erythropoietin

Platelet activating factor acetylhydrolase (PAF-AH) is a Ca2+-independent phospholipase A2 that is secreted mainly from monocytes/macrophages. In human plasma, PAF-AH is associated primarily with low-density lipoprotein (LDL), while a small proportion of enzyme is associated with high-density lipoprotein (HDL). The ratio of HDL-PAF-AH to total plasma enzyme activity may represent a potential marker of atherogenicity. We evaluated possible alterations of lipoprotein-associated enzyme activity in Chronic Kidney Disease (CKD) patients, stages 3-4, and further investigated whether long-term therapy with recombinant human erythropoietin (epoetin) has any influence on the plasma PAF-AH activity in vivo or on the enzyme activity secreted from peripheral blood monocytes (PBMs), in vitro. Forty-eight patients, 28 men and 20 women, with CKD (stages 3-4) participated in the study. Patients were randomized into groups I and II. Patients of group I (n = 28) were administered subcutaneously epoetin, 50 units/kg once per week. The Hb target was 13 g/dl. In group II (n = 20), epoetin was initiated only when the Hb levels decreased during follow-up to less than 9 g/dl. All patients were seen on an outpatient basis at 2, 4 and 6 months. Twenty-two normolipidemic age- and sex-matched healthy volunteers also participated in the study and were used as controls. The PAF-AH activity in plasma of both patient groups at baseline was higher compared to controls, whereas no difference in the HDL-PAF-AH activity was observed among the studied groups. Thus, the ratio of HDL-PAF-AH to the plasma enzyme activity was significantly lower in both patient groups compared to controls. Epoetin administration in the patients of group I was associated with a significant increase in the plasma PAF-AH and in HDL-PAF-AH activities 2 months after treatment, which remained stable for up to 6 months of therapy, a phenomenon not observed in untreated patients of group II. Thus, the ratio of HDL-PAF-AH to the plasma enzyme activity was significantly increased in patients of group I compared to the baseline values, a phenomenon not observed in patients of group II. In vitro treatment with epoetin of PBMs from patients of group I (undergoing therapy with epoetin) resulted in a dose-dependent increase in total and secreted enzyme activity, a phenomenon not observed in patients of group II who did not receive therapy with epoetin. This suggests that the in vivo increase in lipoprotein-associated PAF-AH observed in patients treated with epoetin may be attributed to the drug-induced enhanced secretion of PAF-AH from PBMs of these patients. CKD patients of stages 3-4 are characterized by an increase in plasma PAF-AH activity and a low ratio of HDL-PAF-AH to total plasma enzyme activity. Long-term therapy with epoetin may improve this atherogenic ratio thus this drug may play an important antiatherogenic role in CKD.

Activation of protein kinase C inhibits the secretion of platelet-activating factor acetylhydrolase by human decidual macrophages.

Background and Aims: Platelet activating factor (PAF), a potent phospholipid mediator, has been implicated in a number of reproductive processes through ovulation to parturition. To clarify the regulatory mechanism of PAF metabolism in the decidua, we have investigated the effect of activation of protein kinase C (PKC) on the secretion of PAF-acetylhydrolase (PAF-AH), a PAF-inactivating enzyme, by human decidual macrophages. Methods: Decidual macrophage populations were isolated from human decidua by using enzymic digestion, Ficoll-Paque centrifugation, or flow cytometric sorting. The cells were treated with a PKC activator (TPA), H-7, dibutyryl cyclic adenosine monophosphate (AMP), Bis-(o-aminophenoxy)-ethane-N,N,N',N'-tetraacetic acid, tetra (acetoxymethyl)-ester (BAPTA/AM) and/or nifedipine. The activity of PAF-AH secreted in the culture medium was assayed. Results: The PKC activator, TPA, inhibited the PAF-AH secretion by decidual cells in a dose-dependent manner. The TPA also decreased the enzyme secretion by flow cytometrically purified macrophages. The inhibitory effect of TPA was blocked by a PKC inhibitor, H-7. Protein kinase A (PKA) activation by dibutyryl cyclic AMP was without effect on the enzyme secretion. Calcium channel blockers, BAPTA/AM and nifedipine had no effect on the PAF-AH secretion. Conclusion: It is suggested that the TPA-induced inhibition of PAF-AH secretion may be mediated, in part, by a PKC-dependent signal transduction, and that activation of PKC may result in the increase in the local concentration of PAF in the decidua because of its inhibitory effect on the PAF-AH secretion by decidual macrophages. (Reprod Med Biol 2003; 2: 121-126).

Effects of colony-stimulating factors on the secretion of platelet-activating factor acetylhydrolase by human decidual macrophages

Objective: The aim is to clarify the role of platelet-activating factor and colony-stimulating factors in term and preterm parturition. Study Design: Decidual macrophage populations were obtained by an enzymic digestion, Ficoll-Paque centrifugation, or flow cytometric sorting. The effects of colony-stimulating factors on the platelet-activating factor acetylhydrolase secretion by these cells were examined. Results: Granulocyte-macrophage colony-stimulating factor inhibited the platelet-activating factor acetylhydrolase secretion by decidual macrophages. Granulocyte colony-stimulating factor also decreased the enzyme secretion but at higher concentrations than those required for granulocyte colony-stimulating factor. In contrast, macrophage colony-stimulating factor increased the enzyme secretion. These colony-stimulating factor-induced effects were specifically neutralized by the corresponding antibodies. Conclusion: Colony-stimulating factors may modulate the local concentration of platelet-activating factor in the decidua via their inhibitory or stimulatory effect on the secretion of platelet-activating factor acetylhydrolase, contributing to the regulation of term and preterm parturition at the maternal-fetal interface. (Am J Obstet Gynecol 2003;188:157-61.)

N-linked glycosylation of macrophage-derived PAF-AH is a major determinant of enzyme association with plasma HDL

Human plasma PAF-AH (platelet-activating factor-acetylhydrolase) is a Ca2+-independent phospholipase A2 of hematopoietic origin associated with LDL and HDL; it degrades PAF and oxidizes phospholipids. We show that human macrophages synthesize PAF-AH as a premedial Golgi precursor containing high mannose N-linked glycans. Secreted PAF-AH possesses a molecular mass of ~55 kDa and contains mature N-linked glycans. Secreted PAF-AH activity (90 ± 4% of the total) bound to a wheat germ lectin column and could be eluted with N-acetylglucosamine, whereas digestion with N-acetylneuraminidase II completely abolished enzyme absorption. Tunicamycin significantly reduced cell-associated PAF-AH activity and inhibited enzyme secretion; but it did not alter the ratio of secreted to cell-associated enzyme (1.8 at 6 h and 3.1 at 24 h), suggesting that glycosylation is not essential for PAF-AH secretion. Digestion of cell-associated PAF-AH or secreted PAF-AH with peptide N-glycosidase F affected neither catalytic activity nor its resistance to proteolysis with trypsin or proteinase K; in addition, it did not affect PAF-AH association with LDL, but significantly increased its association with HDL.We suggest that macrophage-derived PAF-AH contains heterogeneous asparagine-conjugated sugar chain(s) involving sialic acid, which hinders its association with HDL but does not influence the secretion, catalytic activity, or resistance of PAF-AH to proteases.

Inhibitory effect of interleukin-8 on the secretion of platelet-activating factor acetylhydrolase by human decidual macrophages

To determine whether interleukin-8 (IL-8) regulates platelet-activating factor acetylhydrolase (PAF-AH) secretion by decidual cells in vitro. Decidual macrophages were obtained from human term decidual tissue by enzymic digestion and Ficoll-Paque centrifugation. The effect of IL-8 and phorbol esters on the secretion of PAF-acetylhydrolase by these cells was examined. IL-8 inhibited PAF-AH secretion by decidual macrophages in a dose-dependent manner. The inhibition was reversed partially by antibodies against tumor necrosis factor-alpha and IL-1 beta. IL-8-induced inhibition was blocked partially by the protein kinase C (PKC) inhibitors, sphingosine, and H-7 1-(5-isoquinoline sulfonyl)-2-methylpiperazine. A PKC activator, 12-O-tetradecanoyl phorbol 13-acetate, decreased PAF-AH secretion. IL-8 may increase the PAF concentration in the decidua via its inhibitory effect on PAF-AH secretion by decidual macrophages. IL-8-induced inhibition of enzyme secretion may have been mediated in part by PKC-dependent signal transduction.

Inhibitory Effect of Interleukin-8 on the Secretion of Platelet-Activating Factor Acetylhydrolase by Human Decidual Macrophages

Inhibitory Effect of Interleukin-8 on the Secretion of Platelet-Activating Factor Acetylhydrolase by Human Decidual Macrophages

Hepatic regulation of platelet-activating factor acetylhydrolase and lecithin:cholesterol acyltransferase biliary and plasma output in rats exposed to bacterial lipopolysaccharide.

Normal rat bile contains secretory platelet-activating factor acetylhydrolase (PAF-AH), the enzyme capable of hydrolyzing the inflammatory mediator platelet-activating factor (PAF), and phospholipids containing oxidized truncated fatty acids. Because lecithin:cholesterol acyltransferase (LCAT) possesses intrinsic PAF-AH-like activity, it also may represent a potential anti-inflammatory enzyme. The behavior of PAF-AH and LCAT in hepatobiliary inflammatory responses in vivo has not been characterized. We therefore investigated the biliary and plasma secretion and pharmacological characteristics of these enzymes in rats subjected to intraportal bacterial endotoxin exposure (lipopolysaccharide [LPS], Escherichia coli, 055:B5). Portal vein LPS infusion (1 mg/kg, bolus) resulted in a maximal 4- to 5-fold increase in bile PAF-AH-specific activity with a gradual decline to baseline by 18 hours. Biliary PAF-AH hydrolyzed also the truncated sn-2-succinoyl and sn-2-glutaroyl analogs of PAF, indicating a broader activity of PAF-AH in bile toward byproducts of glycerophospholipid peroxidation. Plasma PAF-AH activity was not altered 5 hours after LPS injection compared with saline injection, but it was significantly elevated 18 hours after endotoxin exposure. The levels of LCAT in bile were low and declined to nearly undetectable values by 5 hours after cannulation in both control and LPS-exposed rats. Plasma LCAT activity was significantly increased after 5 hours and decreased 18 hours after LPS injection. In summary, hepatic exposure to endotoxin results in a rapid increase in biliary secretion of PAF-AH followed by elevation of LCAT and PAF-AH levels in plasma. We propose that biliary secretion of PAF-AH may be involved in the hepatic response to endotoxic insult by counteracting potential inflammatory damage in the biliary tree and gastrointestinal tract, whereas plasma increases in LCAT and PAF-AH may promote elimination of excess PAF and oxidized phospholipids in the circulation.

The inhibitory effect of protein kinase C activation on the secretion of platelet-activating factor-acetylhydrolase by human decidual macrophages

The inhibitory effect of protein kinase C activation on the secretion of platelet-activating factor-acetylhydrolase by human decidual macrophages

The inhibitory effect of 1,25-dihydroxyvitamin D3 on the secretion of platelet-activating factor acetylhydrolase by human decidual macrophages.

The effects of 1,25-dihydroxyvitamin D3 [1,25-(OH)2D3] and 25-hydroxyvitamin D3 on the secretion of a platelet-activating factor (PAF)-inactivating enzyme, PAF-acetylhydrolase (PAF-AH), by decidual macrophage populations were examined. 1,25-(OH)2D3 inhibited PAF-AH secretion by either decidual cells or flow cytometrically purified decidual macrophages. 25-Hydroxyvitamin D3 also decreased the enzyme secretion but at higher concentrations than those required for 1,25-(OH)2D3. The 1,25-(OH)2D3-induced inhibition was partially blocked by protein kinase C (PKC) inhibitors, sphingosine and 1-(5-isoquinolinesulfonyl)-2-metylpiperazine (H-7). An intracellular calcium channel blocker, bis-(o-aminophenoxy)-ethane-N,N,N',N'-tetraacetic acid, tetra(acetoxymethyl)-ester (BAPTA/AM), also partially blocked the inhibition by 1,25-(OH)2D3, whereas extracellular calcium channel blockers, verapamil and nifedipine, failed to prevent the inhibition. H-7 and BAPTA/AM additively blocked the 1,25-(OH)2D3-induced inhibition. A PKC activator, 12-O-tetradecanoylphorbol 13-acetate, also decreased PAF-AH secretion. The decrease was abolished by sphingosine or H-7. It is suggested that 1,25-(OH)2D3 may increase the concentration of PAF in the decidua via its inhibitory effect on PAF-AH secretion by decidual macrophages and that the inhibitory action may be mediated by intracellular calcium and PKC-dependent signal transduction. PAF and 1,25-(OH)2D3 may, therefore, play a cooperative role in the regulation of PAF metabolism at the maternal-fetal interface.

The inhibitory effect of 1,25-dihydroxyvitamin D3 on the secretion of platelet-activating factor acetylhydrolase by human decidual macrophages

The inhibitory effect of 1,25-dihydroxyvitamin D3 on the secretion of platelet-activating factor acetylhydrolase by human decidual macrophages

Smoking and preterm labor: Effect of a cigarette smoke extract on the secretion of platelet-activating factor—acetylhydrolase by human decidual macrophages

Maternal smoking in pregnancy is associated with a significant increase in the incidence of preterm labor, premature rupture of membranes, and premature delivery. Our aim was to clarify the cause underlying this association. The effect of cigarette smoke extract on the secretion of platelet-activating factor-acetylhydrolase by both decidual macrophages and peripheral blood monocytes and macrophages was investigated. The cigarette smoke extract inhibited the platelet-activating factor-acetylhydrolase secretion by these cells. The inhibitory effect of cigarette smoke extract on the secretion was a hundred times more potent compared with its direct effect on the plasma enzyme. Glutathione and dithiothreitol blocked the inhibition, whereas catalase or superoxide dismutase did not. Nicotine and cotinine have no effect on the secretion. The presence in cigarette smoke extract of a potent inhibitor(s) of platelet-activating factor-acetylhydrolase secretion by decidual macrophages may provide an insight into the pathogenesis of preterm labor, premature rupture of membranes, and premature delivery in women who smoke during pregnancy.

Secretion of platelet-activating factor acetylhydrolase by human decidual macrophages.

Platelet-activating factor (PAF) metabolism in the maternal-fetal decidual interface has been investigated. Human decidua was obtained from patients at term and not in labor after cesarean section. The cells were isolated by enzymic digestion, followed by Ficoll-Paque centrifugation, or were purified further by discontinuous Percoll density gradient centrifugation. Cell populations were analyzed by flow cytometry after labeling with macrophage-specific antibodies. Twenty-seven percent of the cells obtained after enzymic digestion and Ficoll-Paque density gradient centrifugation had macrophage surface markers. The decidual cell population secreted PAF-acetylhydrolase (PAF-AH) activity. The secreted PAF-AH was the plasma-type isozyme. Synthesis and secretion was inhibited by actinomycin-D or cycloheximide. The PAF-AH activity secreted into the culture medium correlated positively with the number of macrophages. Flow cytometric purification yielded a 96% macrophage marker-positive population. The macrophages were shown to be the only cell types of decidual tissue that secreted PAF-AH. Treatment with anti-CD14 monoclonal antibody and complement specifically blocked PAF-AH secretion by collagenase-dispersed cells. It is concluded that decidual macrophages produce and secrete PAF-AH of the plasma type, and it is suggested that these cells may play an important role in PAF metabolism during parturition.

Secretion of platelet-activating factor acetylhydrolase by human decidual macrophages

Secretion of platelet-activating factor acetylhydrolase by human decidual macrophages

Effects of endotoxins and cytokines on the secretion of platelet-activating factor-acetylhydrolase by human decidual macrophages

The aim was to clarify the role of platelet-activating factor in parturition, preterm labor, and premature rupture of membranes. Decidual macrophage populations were obtained by enzymic digestion, Ficoll-Paque centrifugation, or flow cytometric sorting. The effects of endotoxins and cytokines on platelet-activating factor-acetylhydrolase secretion by these cells were examined. Lipopolysaccharide inhibited the platelet-activating factor-acetylhydrolase secretion by decidual macrophages. The inhibition was partially reversed by interleukin-1 receptor antagonist or by neutralizing antibodies against interleukin-1 alpha, interleukin-1 beta, or tumor necrosis factor-alpha. Tumor necrosis factor-alpha, interleukin-1 alpha, and interleukin-1 beta also decreased the enzyme secretion. The inhibitory actions of tumor necrosis factor-alpha and interleukin-1 beta were specifically neutralized by the corresponding antibodies. The effect of interleukin-1 alpha or interleukin-1 beta on the secretion was abolished by interleukin-1 receptor antagonist. It is suggested that platelet-activating factor is involved in the pathogenesis of preterm labor or premature rupture of membranes caused by endotoxins and the subsequent activation of cytokine network.

Effect of 17β-estradiol on secretion of platelet-activating factor acetylhydrolase by HepG2 cells

Estrogen has been shown to decrease plasma platelet-activating factor (PAF) acetylhydrolase activity, but the precise mechanisms are not known. We examined the effect of estradiol on the secretion of PAF acetylhydrolase by HepG2 cells. In our previous study, we demonstrated the production of this enzyme by HepG2 cells, which we used as an experimental model of normal hepatocytes. 17β-Estradiol mildly but consistently inhibited the secretion of PAF acetylhydrolase by HepG2 cells in a concentration-dependent manner. Under basal conditions, HepG2 cells secreted 42.3 pmol/mg cell protein/min PAF acetylhydrolase in 24 hours (mean of 8 dishes), and the presence of 10 −7 mol/L 17β-estradiol decreased the secretion to 77% ± 10.3% of control values (mean ± SD, n = 8, P < .02). 17β-Estradiol treatment affected neither the secretion of apolipoprotein (apo) A-I nor cell-associated PAF acetylhydrolase activity. Electrophoretic separation of [ 35S]methionine-labeled PAF acetylhydrolase revealed a single band whose molecular weight was approximately 43,000 d. We conclude that estrogen decreases the secretion of PAF acetylhydrolase by the liver, and it may explain, at least in part, the effect of estrogen on plasma PAF acetylhydrolase.

Secretion of Platelet-Activating Factor Acetylhydrolase Following Phorbol Ester-Stimulated Differentiation of HL-60 Cells

Platelet-activating factor (PAF) plays an important role in a number of biological processes ranging from inflammation to reproductive biology. We have reported that the enzyme that inactivates this potent autacoid, PAF-acetylhydrolase (PAF-AH), is decreased in maternal plasma during the latter stages of pregnancy. This enzyme is associated with the plasma lipoprotein fraction and therefore its tissue origin was thought to be the liver. Prescott and colleagues ( J. Biol. Chem. 265, 17381, 1990) have reported that both a rat liver cell line (HepG2 cells) and human peripheral macrophages secrete PAFAH of the plasma type. We have shown previously that the injection of rats with dexamethasone or medroxyprogesterone causes an increase and estrogen a decrease in the plasma PAF-AH activity. To clarify the mechanism of hormonal regulation of PAF-AH production, we employed a monocyte-macrophage model system to investigate the secretion of PAF-AH during differentiation. In the present study, we have demonstrated that a myelocytic leukemic cell line (HL-60) produces and secretes PAF-AH into a defined medium when the cells are differentiated into macrophages following stimulation by 12- O-tetradecanoylphorbol-13-acetate (TPA). The medium obtained from unstimulated HL-60 cells did not contain detectable amounts of PAF-AH activity. Stimulation with TPA caused a dose- and time-dependent increase in PAF-AH activity in the media. No increase in cell number was observed in the HL-60 cells during the culture period after the cells were treated with TPA. Cell lysis was excluded by the demonstration that the TPAinduced adherent cells excluded trypan blue and did not release lactate dehydrogenase activity into the medium. The increase in PAF-AH activity was inhibited by actinomycin D and cycloheximide. Dexamethasone and medroxyprogesterone markedly increased the secretion of PAF-AH by these cells, while estrogen was without effect. Bacterial endotoxin (lipopolysaccaride, LPS) inhibited the production of PAF-AH by these cells in a dose-dependent manner. The stimulation of PAF-AH secretion during differentiation of HL-60 cells and its modulation by LPS and steroid hormones may provide a useful model system for studying PAF metabolism during the inflammatory response and pregnancy.

Liver cells secrete the plasma form of platelet-activating factor acetylhydrolase.

Platelet-activating factor (PAF) is a phospholipid (1-O-alkyl-2-acetyl-sn-glycero-3-phosphocholine) with diverse physiological effects. It has been implicated as a mediator of inflammation, allergy, shock, and thrombosis. Plasma contains an enzyme, PAF acetylhydrolase, that catalyzes the degradation of PAF, and the level of this enzyme may regulate the concentration of PAF in the blood and extracellular spaces under some conditions. Thus, the cellular source(s) of this enzyme and the factors that regulate its synthesis and secretion are issues that may have important physiological and pathological implications. We found that cultures of Hep G2, a human hepatocarcinoma line, secreted PAF acetylhydrolase activity. Optimal secretion occurred in medium that contained serum, and the newly secreted PAF acetylhydrolase was associated with high density and low density lipoproteins (LDL and HDL, respectively), just as the enzyme is in plasma. In the absence of serum. PAF acetylhydrolase was secreted with a particle that had a density similar to HDL. Apolipoproteins B and E were found in the same fractions. We tested the effects of a variety of hormones on the secretion of PAF acetylhydrolase and found that secretion was inhibited by 17 alpha-ethynylestradiol with a maximal effect at 30 microM. This may account for the observation of others that estrogens reduce the activity of PAF acetylhydrolase in the plasma. The PAF acetylhydrolase secreted by Hep G2 cells appeared to be identical to the enzyme in human plasma based on substrate specificity, association with LDL and HDL, response to inhibitors, and reactivity with antibodies against the plasma PAF acetylhydrolase. In conclusion, we have demonstrated that hepatocytes in culture secrete a PAF acetylhydrolase that is apparently identical to the plasma form. The secretion is constitutive but may also be regulated in response to hormonal stimulation.