PubMed Health⌕ Search

Biomedical subjects

I Kudo

Publications and source records attributed to I Kudo.

At least 19 recordsLinked to original sources
PubMed1995

Detection and purification of two 14 kDa phospholipase A2 isoforms in rat kidney: their role in eicosanoid synthesis.

Phospholipase A2 (PLA2) activity in the soluble fraction of rat kidney yielded three peaks on DEAE cellulose column chromatography. From these three, we purified two PLA2 isoforms to near-homogeneity. Both had a molecular weight of approx. 14,000 on SDS-PAGE, and immunochemical and enzymological studies indicated that one is a 14 kDa type I PLA2 and the other a 14 kDa type II PLA2. RNA blot analysis confirmed that rat kidney contains both types of PLA2 and that administration of lipopolysaccharides and mercury chloride into rats increased type II PLA2 mRNA levels in kidney. When cultured rat mesangial cells were incubated with purified type I or type II PLA2 in combination with the calcium ionophore A23187 at suboptimal condition, augmentation of prostaglandin E2 production was observed. Type I and type II forms of PLA2 may play a role in arachidonate metabolism in rat kidney.

Animals↗
PubMed1995

Lipoxygenase-catalyzed oxygenation of arachidonylethanolamide, a cannabinoid receptor agonist.

Various purified lipoxygenases were incubated with [14C]arachidonylethanolamide which is an endogenous ligand for cannabinoid receptors. When radioactive products were analyzed by thin-layer chromatography, porcine leukocyte 12-lipoxygenase and rabbit reticulocyte and soybean 15-lipoxygenases produced polar compounds at about the same reaction rates as that of oxygenation of free arachidonic acid. In contrast, the reaction of human platelet 12-lipoxygenase proceeded at a much lower rate, and porcine leukocyte 5-lipoxygenase was totally inactive. The result indicated that the lipoxygenases, which had been shown previously to be capable of oxygenating esterified polyunsaturated fatty acids, were also active with the arachidonylethanolamide. High-performance liquid chromatography, ultraviolet and mass spectrometry and nuclear magnetic resonance spectroscopy identified the major product by leukocyte 12-lipoxygenase as 12-hydroperoxy-5,8,10,14-eicosatetraenoylethanolamide and that by 15-lipoxygenases as 15-hydroperoxy-5,8,11,13-eicosatetraenoylethanolamide. The 15-hydroxy derivative inhibited electrically-evoked contraction of mouse vas deferens with an IC50 of 0.63 microM as well as arachidonylethanolamide (0.17 microM), but the 12-hydroxy derivative was much less effective.

Animals↗
PubMed1995

Phospholipid degradation in rat calcium ionophore-activated platelets is catalyzed mainly by two discrete secretory phospholipase As.

An "A1 type" phospholipase activity with serine-phospholipid preference was released by rat activated platelets. It was distinct from the secretory type II phospholipase A2 [Horigome, K., Hayakawa, M., Inoue, K., and Nojima, S. (1987) J. Biochem. 101, 625-631] and co-purified with the secretory lysophosphatidylserine-selective lysophospholipase activity [Higashi, S., Kobayashi, T., Kudo, I., and Inoue, K. (1988) J. Biochem. 103, 442-447]. Several lines of evidence indicated that a single protein was responsible for the phospholipase A1 and lysophospholipase activities. Marked accumulation of lysophospholipids was observed in rat calcium ionophore-activated washed platelets and both phospholipase A1/lysophospholipase and type II phospholipase A2 were shown to contribute to this phospholipid degradation. A selective inhibitor of type II phospholipase A2 reduced the phospholipid degradation and enhanced the clotting time and prothrombinase activity. These results indicate that secretory platelet phospholipases may play a role in regulation of blood clotting.

Animals↗
PubMed1995

A case of osteoporosis with bilateral defects in the mandibular processes.

We carried out a detailed total body examination of a 62-year-old woman with osteoporosis who had bilateral defects in the mandibular processes. It was inferred that the defects in both articular heads were caused by resorption of small bone fragments following fracture. The quantity of bone salt was determined by microdensitometry, and a diagnosis of osteoporosis was then established. An improved bite was obtained by treatment consisting of tooth extraction and the preparation of partial dentures.

Absorptiometry, Photon↗
PubMed1994

Dual regulation of cytosolic phospholipase A2 in mast cells after cross-linking of FC epsilon-receptor.

We have reported previously that cultured mast cells (MC) express three discrete phospholipases A2 (PLA2s), one of which corresponds to arachidonoyl-preferential cytosolic PLA2 (cPLA2). In the present study, we investigated the possible role of cPLA2 in eicosanoid synthesis by activating mouse bone marrow-derived mast cells (BMMC) through cross-linking of the high affinity IgE receptor (Fc epsilon RI) with a specific Ag. BMMC released arachidonic acid within 2 min after Fc epsilon RI cross-linking. A rapid, transient phosphorylation of cPLA2 was observed after Fc epsilon RI cross-linking, reaching the maximum within 2 min, and accompanied by an increase of cPLA2 activity in the cell lysate. Exposure of BMMC to the IgE-Ag for longer periods resulted in a time-dependent increase of the cPLA2 protein. The increase was detected within 10 h after stimulation and reached the maximum within 30 h. Dexamethasone inhibited the Ag-stimulated cPLA2 induction significantly. cPLA2 activity in cells stimulated for 24 h was increased significantly, and suppressed in cells treated with dexamethasone. When the cells were exposed to IgE-Ag for 36 h and then challenged with a secondary agonist, thrombin, arachidonate release was augmented significantly in comparison with cells without the Ag pretreatment. Thus, cPLA2 activation in BMMC by short term exposure to the Ag might be regulated by post-Fc epsilon RI modification (phosphorylation) of pre-existing enzyme, whereas that observed after long term exposure might be explained by the increase in cPLA2 protein.

Animals↗
PubMed1994

A possible role for extracellular bicarbonate in U-46619-induced rat platelet aggregation.

U-46619, a thromboxane A2 agonist, has been believed not to induce aggregation of rat platelets. However, we have found that U-46619 evoked rat platelet aggregation when the cells were suspended in HCO3-containing medium, whereas it was inactive in medium lacking HCO3-. 4,4'-Diisothiocyanostilbene-2,2'-disulfonate (DIDS), selective inhibitor of the HCO3/Cl- exchanger, inhibited the aggregation of rat platelets induced by U-46619 as well as by collagen and ADP. Thrombin-induced aggregation was also inhibited by DIDS, although to a much lesser extent. Several inhibitors of anion transporters, such as phloretin, pyridoxal phosphate and ethacrynic acid, suppressed U-46619-induced aggregation. These observations indicate that HCO3-influx via an anion exchanger may be involved in the signal transduction pathway of U-46619 in rat platelets.

15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5↗
PubMed1994

Complete discrimination of docosahexaenoate from arachidonate by 85 kDa cytosolic phospholipase A2 during the hydrolysis of diacyl- and alkenylacylglycerophosphoethanolamine.

In our previous report (Shikano, M., Masuzawa, Y. and Yazawa, K. (1993) J. Immunol. 150, 3525-3533), we described that the enrichment of docosahexaenoic acid (DHA, 22:6(n - 3)) reduces both arachidonic acid (AA, 20:4(n - 6)) release and platelet-activating factor (PAF) synthesis in human eosinophilic leukemia cells, Eol-1. Since no DHA release was observed in response to Ca-ionophore stimulation, we presumed that the phospholipase A2 (PLA2) responsible for AA release and PAF synthesis can not hydrolyze the DHA moiety of phospholipids. In the present paper, we examined whether DHA-containing diacyl- and alkenylacylglycerophosphoethanolamine (DHA-diacylGPE and DHA-alkenylacyGPE) are susceptible to the action of AA-preferential 85 kDa cytosolic phospholipase A2 (cPLA2) from rabbit platelets in comparison with AA and eicosapentaenoic acid (EPA, 20:5(n - 3)) derivatives. When diacylGPE was used as a substrate, DHA release was almost negligible under the assay condition that allowed AA and EPA to be liberated at the rates of 4.3 mumol/min per mg protein and 2.5 mumol/min per mg protein, respectively. On the other hand, 14 kDa type II PLA2 hydrolyzed DHA-diacylGPE as well as AA-diacylGPE and EPA-diacylGPE. When DHA-diacylGPE and AA-diacylGPE were mixed at equimolar concentrations, DHA release by cPLA2 was not observed and AA release was reduced to 32% in the case without DHA-diacylGPE. This indicated that DHA-diacylGPE is a poor substrate but possesses the inhibitory activity for cPLA2. cPLA2 does not clearly discriminate between AA-alkenylacylGPE and AA-diacylGPE. As in the case using diacylGPE as a substrate, DHA-alkenylacylGPE was completely discriminated from AA-alkenylacylGPE by cPLA2. The roles of DHA and cPLA2 in the synthesis of lipid mediators will be discussed in relation to the new aspects of the substrate specificity of cPLA2 provided here.

Animals↗
PubMed1994

Role of type II phospholipase A2 in the inflammatory process of carrageenan-induced pleurisy in rats.

In order to investigate the role of type II phospholipase A2 (PLA2) in the inflammatory process, the effect of a monoclonal antibody specific to type II PLA2 on carrageenan-induced pleurisy was studied in rats. Intravenous injection of the antibody (MB5.2), which inhibits the catalytic activity of type II PLA2, significantly reduced both the pleural exudate volume and the intrapleural leukocyte count, while a control antibody did not have any appreciable effect. MB5.2 caused no change in the level of type II PLA2 in the pleural fluid. These results suggest that type II PLA2 generated at inflamed sites, at least in part, have a crucial role in the pathogenesis of acute inflammation.

Animals↗
PubMed1994

Suppressive effects of the anti-allergic drugs, tranilast and azelastine, on the lysophosphatidylserine-dependent activation of rat mast cells.

Anti-allergic drugs, tranilast and azelastine, were examined for their effects on lysophosphatidylserine (lysoPS)-dependent histamine release from rat mast cells. Although both compounds suppressed the histamine release in a dose-dependent manner, the inhibition was affected by lysoPS concentration differently. In the presence of an increasing concentration of lysoPS, the suppressive effect of tranilast decreased. The inhibition by azelastine, however, was independent of the concentration of lysoPS. The findings suggest that these two drugs inhibit lysoPS-depedent histamine release through essentially different routes.

Animals↗
PubMed1994

Characterization of cytosolic phospholipase A2 in rat mastocytoma RBL-2H3.

We previously reported that cultured mast cells expressed three discrete phospholipases A2 (PLA2S), one of which showed a remarkable preference for phospholipids bearing an arachidonoyl residue at the sn-2 position [M. Murakami, et al., J. Biochem., 111, 175 (1992)]. In the present study, we have purified and characterized this enzyme using rat mastocytoma RBL-2H3 as an enzyme source. The elution profiles of the arachidonoyl-preferential PLA2 activity of rat mastocytoma RBL-2H3 cells on several column chromatographies were indistinguishable from those of 85-kDa cytosolic PLA2 (cPLA2) characterized so far. The molecular mass of the partially purified PLA2 was estimated to be about 90 kDa by gel filtration and it hydrolyzed arachidonate-containing phospholipids preferentially in the presence of submicromolar Ca2+ concentrations. Furthermore, it was immunoprecipitated with an anti-rabbit cPLA2 antibody almost completely. From these observations, we have concluded that the arachidonoyl-preferential PLA2 in mast cells belongs to the "cPLA2" family.

Animals↗
PubMed1993

Participation in cellular prostaglandin synthesis of type-II phospholipase A2 secreted and anchored on cell-surface heparan sulfate proteoglycan.

Rat-liver-derived BRL-3A cells, which express both type-II phospholipase A2 (PLA2) and cytosolic PLA2 (cPLA2), generated prostaglandin E2 (PGE2) in the presence of fetal calf serum. When the cells were treated with tumor necrosis factor (TNF), PGE2 generation was greatly stimulated. The production of PGE2 observed in both cases was suppressed by a type-II PLA2-specific inhibitor, thielocin A1. Appreciable amounts of type-II PLA2 were released into the medium from the TNF-stimulated cells when heparin was added extracellularly. The release of type-II PLA2 from TNF-stimulated cells was also found in the presence of heparan sulfate or dextran sulfate, whereas other glycosaminoglycans showed no effects under the same conditions. These findings suggest that type-II PLA2 expressed in BRL-3A cells mostly associates with the cell surface by binding to cellular heparan sulfate proteoglycan. Removal of cell-surface-associated type-II PLA2, by either extracellular addition of heparin or by prior treatment of the BRL-3A cells with heparitinases, resulted in marked reduction of PGE2 synthesis in the cells. Exposure of BRL-3A cells to thrombin also induced the apparent secretion of type-II PLA2, and thrombin-stimulated PGE2 generation was suppressed by heparin effectively. Type-II PLA2 secreted and attached to heparan sulfate on the cell surface may therefore play an essential role in PGE2 synthesis by BRL-3A cells.

Animals↗
PubMed1993

Characteristics of lysophospholipase activity expressed by cytosolic phospholipase A2.

Evidence has accumulated to suggest that a wide variety of mammalian cells and tissues express a cytosolic phospholipase A2 with arachidonoyl preference (cPLA2). Purified rabbit platelet-derived cPLA2, as well as the human recombinant enzyme originally identified in the monocytic leukemic cell line U937, exhibit significant lysophospholipase activity. Several series of experiments indicated that a single protein mediated both activities. Treatment of the purified enzyme with p-bromophenacylbromide or an anti-(rabbit platelet cPLA2) monoclonal antibody, RHY-5, suppressed the activity of phospholipase A2 without any appreciable effect on lysophospholipase activity, suggesting that the domain(s) required for phospholipase A2 activity may be located separately from that for lysophospholipase activity. Lysophospholipase activity was appreciably detected above the critical micellar concentration of the substrate. Lysophosphatidylcholine was also hydrolyzed efficiently when it was incorporated into liposomes made of dialkylphosphatidylcholine. The hydrolysis of lysophospholipid was dependent on the fatty acid bound at the sn1 position; the relative rates of hydrolysis of 1-oleoyllysophosphatidylcholine, 1-palmitoyllysophosphatidylcholine, and 1-stearoyllysophosphatidylcholine were 23, 8, and 1, respectively. A similar order of reactivity was observed with lysophospholipid incorporated into dialkylphosphatidylcholine liposomes. cPLA2 may function not only as an arachidonate liberation enzyme but also as an enzyme responsible for degradation of certain molecular species of lysophospholipids formed in membranes.

Animals↗
PubMed1993

Triggering of degranulation in mast cells by exogenous type II phospholipase A2.

We have previously shown the possibility that endogenous type II phospholipase A2 (PLA2) might participate in degranulation in mast cells (MC) (Murakami, M., et al. 1992. Eur. J. Biochem. 209:257). Now we have examined whether or not exogenously added type II PLA2 triggers MC degranulation. When rat peritoneal connective tissue MC (CTMC) were exposed to purified rat type II PLA2 at concentrations of more than 10 micrograms/ml, significant release of histamine was observed, whereas PGD2 was not generated under the same conditions. Mouse peritoneal CTMC as well as bone marrow-derived immature MC also responded to PLA2. Preincubation of CTMC with tyrosine kinase inhibitors, genistein, and herbimycin A, but not with pertussis toxin, resulted in abolition of the sensitivity to PLA2. The ability of type II PLA2 to induce histamine release was inhibited by an antibody or chemicals, both of which blocked the catalytic activity of type II PLA2. Heparin or an antibody recognizing the heparin-binding domain of type II PLA2 also suppressed the MC-degranulating activity, probably due to inhibition of binding of PLA2 to the cells. The interaction between heparan sulfate on cell surface and the heparin-binding domain of type II PLA2 may be important for the induction of exocytosis. The catalytic domain of the enzyme is also crucially important for the degranulation induction. Furthermore, we found that nerve growth factor, one of the potent regulators of MC function, significantly potentiated type II PLA2-induced histamine release from rat CTMC. These results suggest the possible role of extracellular type II PLA2 in activation of CTMC primed with nerve growth factor at inflamed sites.

Animals↗
PubMed1993

Possible role of mammalian secretory group II phospholipase A2 in T-lymphocyte activation: implication in propagation of inflammatory reaction.

Both 2-lysophosphatidylcholine and cis-unsaturated fatty acids were previously shown to intensify agonist-induced cellular responses by enhancing the diacylglycerol-dependent activation of protein kinase C. Consistent with these observations, extracellular, secretory group II phospholipase A2, when added directly to human resting T lymphocytes, greatly potentiates their activation that was induced by a membrane-permeant diacylglycerol and ionomycin, as determined by the expression of the alpha subunit of the interleukin 2 receptor and thymidine incorporation into DNA. Diacylglycerol and ionomycin were essential for this cellular response, and phospholipase A2 alone showed no effect. The amount of 2-lysophosphatidylcholine produced in these cells by the exogenous phospholipase A2 was greatly increased in the presence of diacylglycerol and ionomycin, suggesting that the membrane phospholipids became susceptible to the phospholipase A2 when protein kinase C was activated. The results suggest that phospholipase A2 secreted into inflammatory sites plays a role in the propagation of cellular responses. Protein kinase C may function in the hydrolysis of membrane phospholipids by the exogenous phospholipase A2, and the products of this phospholipid hydrolysis enhance agonist-induced protein kinase C activation, thereby intensifying cellular responses.

Cell Membrane Permeability↗