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Biomedical subjects

K Waku

Publications and source records attributed to K Waku.

At least 37 records · Page 2Linked to original sources
PubMed1999

Evidence that the cannabinoid CB1 receptor is a 2-arachidonoylglycerol receptor. Structure-activity relationship of 2-arachidonoylglycerol, ether-linked analogues, and related compounds.

An endogenous cannabimimetic molecule, 2-arachidonoylglycerol, induces a rapid, transient increase in intracellular free Ca2+ concentrations in NG108-15 cells through a cannabinoid CB1 receptor-dependent mechanism. We examined the activities of 24 relevant compounds (2-arachidonoylglycerol, its structural analogues, and several synthetic cannabinoids). We found that 2-arachidonoylglycerol is the most potent compound examined so far: its activity was detectable from as low as 0.3 nM, and the maximal response induced by 2-arachidonoylglycerol exceeded the responses induced by others. Activities of HU-210 and CP55940, potent cannabinoid receptor agonists, were also detectable from as low as 0.3 nM, whereas the maximal responses induced by these compounds were low compared with 2-arachidonoylglycerol. Anandamide was also found to act as a partial agonist in this assay system. We confirmed that free arachidonic acid failed to elicit a response. Furthermore, we found that a metabolically stable ether-linked analogue of 2-arachidonoylglycerol possesses appreciable agonistic activity, although its activity was apparently lower than that of 2-arachidonoylglycerol. We also confirmed that pretreating cells with various cannabinoid receptor agonists nullified the response induced by 2-arachidonoylglycerol, whereas pretreating cells with other neurotransmitters or neuromodulators did not affect the response. These results strongly suggested that the cannabinoid CB1 receptor is originally a 2-arachidonoylglycerol receptor, and 2-arachidonoylglycerol is the intrinsic physiological ligand for the cannabinoid CB1 receptor.

Animals↗
PubMed1999

Giant hydronephrosis due to a ureteral stone, and elevated serum levels of CA 19-9.

CA 19-9 is a widely used tumor marker. However, an elevation in serum CA 19-9 can occur in some patients with benign disorders such as cholecystolithiasis in the absence of tumor. We treated a case of acquired ureteral stone-induced giant hydronephrosis with markedly elevated serum CA 19-9 values. After nephrectomy, the serum CA 19-9 level returned to normal. No malignant cells were found in the tissues of the resected kidney. Localization of CA 19-9 was confirmed by immunohistochemical staining of the renal pelvic mucosa. A detailed case report is presented with a review of the literature.

Adult↗
PubMed1998

Accumulation of various N-acylethanolamines including N-arachidonoylethanolamine (anandamide) in cadmium chloride-administered rat testis.

Changes in the levels of various molecular species of N-acylethanolamine in CdCl2-administered rat testis were examined. We found that the levels of various N-acylethanolamines including anandamide (N-arachidonoylethanolamine), an endogenous cannabinoid receptor ligand, were dramatically increased in CdCl2-admin-istered rat testis. Such changes were particularlyprominent for saturated and monoenoic species such as N-palmitoyl species (39-fold at 9 h) and N-stearoyl species (21-fold at 9 h), compared with unsaturated fatty acid-containing species such as anandamide (5-fold at 9 h). Noticeably, increased levels were observed of not only N-acylethanolamines but also several species of N-acylphosphatidylethanolamine, potential precursors for N-acylethanolamines. We confirmed that the rat testis microsomal fraction contains phosphodiesterase activity catalyzing the release of N-acylethanolamine from N-acylphosphatidylethanolamine and transacylase activity catalyzing the formation of N-acylphosphatidylethanolamine from phosphatidylethanolamine and phosphatidylcholine. These enzyme activities were not dramatically different in the microsomal fraction obtained from CdCl2-administered rat testis compared with that in the case of control rat testis, at least when estimated in cell-free assay systems, suggesting that the accessibility of the substrates to the enzymes may be increased in CdCl2-administered rat testis to generate a large amount of N-acylethanolamine. Possible pathophysiological implications of the augmented generation of N-acylethanolamine including anandamide in CdCl2-administered rat testis were discussed.

Animals↗
PubMed1998

2-Arachidonoylglycerol, an endogenous cannabinoid receptor agonist: identification as one of the major species of monoacylglycerols in various rat tissues, and evidence for its generation through CA2+-dependent and -independent mechanisms.

The molecular species compositions of monoacylglycerols obtained from various rat tissues were examined by reverse-phase high-performance liquid chromatography (HPLC) and gas chromatography-mass spectrometry (GC-MS) analyses. We confirmed that 2-arachidonoylglycerol, an endogenous cannabinoid receptor agonist, is one of the most abundant molecular species of monoacylglycerols in the brain. Substantial amounts of 2-arachidonoylglycerol were also found in the liver, spleen, lung and kidney, but the levels were considerably lower than that in the brain. We found that a small amount of 2-arachidonoylglycerol was generated in a brain homogenate during incubation in the absence of Ca2+. Importantly, the generation of 2-arachidonoylglycerol was markedly augmented in the presence of Ca2+, suggesting that Ca2+ plays a key role in regulation of the generation of 2-arachidonoylglycerol in this tissue.

Animals↗
PubMed1998

Detection of an endogenous cannabimimetic molecule, 2-arachidonoylglycerol, and cannabinoid CB1 receptor mRNA in human vascular cells: is 2-arachidonoylglycerol a possible vasomodulator?

Human vascular endothelial cells were found to generate and release 2-arachidonoylglycerol, an endogenous cannabinoid receptor ligand, upon stimulation with thrombin or A23187. We confirmed that vascular smooth muscle cells as well as endothelial cells possess cannabinoid CB1 receptor mRNA. 2-Arachidonoylglycerol, generated in vascular tissues, may play an important role in modulating vascular tone through acting on the cannabinoid CB1 receptor expressed on vascular smooth muscle cells, endothelial cells as well as peripheral nerve terminals.

Aorta↗
PubMed1997

Inhibition by 2-arachidonoylglycerol, a novel type of possible neuromodulator, of the depolarization-induced increase in intracellular free calcium in neuroblastoma x glioma hybrid NG108-15 cells.

2-Arachidonoylglycerol was found to inhibit the depolarization-induced increase in [Ca2+]i in NG108-15 cells differentiated with prostaglandin E1 and theophylline in a dose-dependent manner. Such an effect appears to be rather specific to polyunsaturated fatty acid-containing monoacylglycerols such as 2-arachidonoylglycerol. Neither 2-palmitoylglycerol nor free arachidonic acid exhibited appreciable inhibitory activity. These observations raise the possibility that 2-arachidonoylglycerol attenuates the increase in [Ca2+]i, thereby modulating several neural functions in this type of cell.

Arachidonic Acids↗
PubMed1997

Inhibition of UDP-glucuronosyltransferase activity by fatty acyl-CoA. Kinetic studies and structure-activity relationship.

We previously identified and purified UDP-glucuronosyltransferase (UGT) isoforms as targets of protein acylation from rat liver microsomes (Yamashita et al., Biochem J 312: 301-308, 1995). The acylation of UGT isoforms occurred upon incubation with acyl-CoA without another protein acyltransferase, suggesting that it was autoacylation. The study revealed the interaction of UGT isoforms with acyl-CoA. In the present study, the effects of fatty acyl-CoA on UGT activities were examined thoroughly, using a rat liver microsomal and purified enzyme fractions. The UGT activities of both fractions were inhibited by acyl-CoA in a concentration-dependent manner. The effect of acyl-CoA was observed on the activities toward various substrates, suggesting that the effect shows the wide spectrum of the isoforms of UGT. To assess the mechanism underlying the inhibition of UGT activity by acyl-CoA, the relationship of the inhibition, acyl-CoA binding to the proteins, and changes in the tertiary structure of the enzyme were examined. The kinetics of these phenomena were related closely with each other. Furthermore, the inhibition of UGT activity was specified for acyl-CoA, though a structurally related compound, acyl-3-dephosphoCoA, had no inhibitory effect. The results suggested that the specific binding of acyl-CoA to UGT isoforms induced conformational changes of the enzymes and resultant inhibition of UGT activity.

Acyl Coenzyme A↗
PubMed1997

Acyltransferases and transacylases involved in fatty acid remodeling of phospholipids and metabolism of bioactive lipids in mammalian cells.

Over 100 different phospholipid molecular species are known to be present in mammalian cells and tissues. Fatty acid remodeling systems for phospholipids including acyl-CoA: lysophospholipid acyltransferases, CoA-dependent and CoA-independent transacylation systems and lysophospholipase/transacylase are involved in the biosynthesis of these molecular species. Acyl-CoA:1-acyl-2-lysophospholipid acyltransferase prefers polyunsaturated fatty acyl-CoAs as acyl donors while acyl-CoA:2-acyl-1-lysophospholipid acyltransferase prefers saturated fatty acyl-CoAs. Therefore, the acyl-CoA:lysophospholipid acyltransferase system is involved in the synthesis of the phospholipid molecular species containing sn-1 saturated and sn-2 unsaturated fatty acids. The CoA-dependent transacylation system catalyzes the transfer of fatty acids esterified in phospholipids to lysophospholipids in the presence of CoA without the generation of free fatty acids. The CoA-dependent transacylation reaction in rat liver exhibits strict fatty acid specificity, i.e., three types of fatty acids (20:4, 18:2, and 18:0) are transferred. On the other hand, the CoA-independent transacylase catalyzes the transfer of C20 and C22 polyunsaturated fatty acids from diacyl phospholipids to various lysophospholipids, in particular, ether-containing lysophospholipids, in the absence of any cofactors. The CoA-independent transacylase is assumed to be involved in the accumulation of polyunsaturated fatty acids in ether-containing phospholipids and in the removal of deleterious ether-containing lysophospholipids. These acyltransferases and transacylases are involved in not only the remodeling of fatty acids but also the synthesis and degradation of some bioactive lipids and their precursors. In this review, the properties of these fatty acid remodeling systems and their possible roles in the biosynthesis of bioactive lipids are described.

1-Acylglycerophosphocholine O-Acyltransferase↗
PubMed1997

Is the cannabinoid CB1 receptor a 2-arachidonoylglycerol receptor? Structural requirements for triggering a Ca2+ transient in NG108-15 cells.

The effects of delta9-tetrahydrocannabinol and 2-arachidonoylglycerol on the intracellular free Ca2+ concentration ([Ca2+]i) in NG108-15 cells were examined in detail. We found that delta9-tetrahydrocannabinol induces a rapid, modest increase in [Ca2+]i. The response was detectable with 3 nM delta9-tetrahydrocannabinol. We also found that very low concentrations of 2-arachidonoylglycerol elicit a rapid, more prominent increase in [Ca2+]i. Such a response was observed not only in NG108-15 cells but also in N18TG2 cells. The response induced by 2-arachidonoylglycerol in either NG108-15 cells or N18TG2 cells was abolished by pretreatment of the cells with a cannabinoid CB1 receptor specific antagonist, SR141716A, suggesting that 2-arachidonoylglycerol interacts with the CB1 receptor to induce the response. The results of an experiment involving a phospholipase C inhibitor suggested that phospholipase C is involved in the rapid increase in [Ca2+]i induced by 2-arachidonoylglycerol. We also found that 1(3)-arachidonoylglycerol exhibits similar activity to that of 2-arachidonoylglycerol, although its activity at low concentrations was somewhat weak compared with that of 2-arachidonoylglycerol. We further confirmed that several structural analogues of 2-arachidonoylglycerol were less active compared with 2-arachidonoylglycerol. These results suggest that the structure of 2-arachidonoylglycerol is strictly recognized by the CB1 receptor, which raises the possibility that the CB1 receptor is originally a 2-arachidonoylglycerol receptor.

Calcium↗
PubMed1997

Comparison of platelet-activating factor receptor mRNA levels in peripheral blood eosinophils from normal subjects and atopic asthmatic patients.

Platelet-activating factor (PAF) modulates the functions of eosinophils, which play an important role in allergic inflammation, through a specific receptor(s) expressed on the cell surface. We compared expression levels of mRNA for the PAF receptor (PAF-R) in peripheral blood eosinophils from atopic asthmatic patients with those from normal healthy subjects using a relative quantification method based on the reverse transcription-polymerase chain reaction (RT-PCR). We found that the levels of PAF-R mRNA in eosinophils from the patients were significantly higher than those in normal subjects. These results suggest that the increased expression of PAF-R in eosinophils may be relevant to the pathogenesis of atopic asthma.

Adult↗
PubMed1996

2-Arachidonoylglycerol, a putative endogenous cannabinoid receptor ligand, induces rapid, transient elevation of intracellular free Ca2+ in neuroblastoma x glioma hybrid NG108-15 cells.

Low concentrations of 2-arachidonoylglycerol were found to induce rapid, transient elevation of intracellular free Ca2+ in NG108-15 cells (EC50 was 150 nM). Free arachidonic acid, 2-palmitoylglycerol, 2-oleoylglycerol, 2-linoleoylglycerol and 2-docosahexaenoylglycerol were inactive. Anandamide acted as a partial agonist. Importantly, desensitization was observed upon sequential challenge with 2-arachidonoylglycerol. Furthermore, cross-desensitization was observed between 2-arachidonoylglycerol and WIN 55212-2, a cannabinoid receptor agonist. Pretreatment of the cells with SR141716A, a cannabinoid receptor antagonist, abolished the activities of both 2-arachidonoylglycerol and WIN 55212-2. These results strongly suggest that 2-arachidonoylglycerol and WIN 55212-2 bind to a common cannabinoid receptor to elicit cellular responses and that 2-arachidonoylglycerol has some physiological role in nervous tissues.

Arachidonic Acids↗
PubMed1996

Cadmium suppresses delta 9 desaturase activity in rat hepatocytes.

The effects of cadmium (Cd) on the fatty acid composition and delta 9 desaturation activity were studied in hepatocytes which had been cultured in serum-free medium. The aim of this study was to determine whether the inhibition of delta 9 desaturase seen in Cd-treated rat liver (Kudo et al. (1991) Toxicology, 68, 133-142) is the direct effect of Cd or not. When hepatocytes were cultured in serum-free medium, increases in the proportions of monounsaturated fatty acids such as oleic acid (18:1) were observed in the phospholipid (PL) and triacylglycerol (TG) fractions of hepatocytes, suggesting that delta 9 desaturase was induced. By contrast, essential fatty acids such as linoleic acid (18:2) and arachidonic acid (20:4) decreased. These changes were similar to those seen in the livers of essential fatty acid (EFA)-deficient rats. When hepatocytes were cultured in the presence of CdCl2, the changes in the fatty acid composition were suppressed. The effect of CdCl2 was concentration-dependent. The rate of the conversion of 18:0 to 18:1, which is catalyzed by delta 9 desaturase, was reduced by treatment with Cd, whereas the rate of conversion of 18:2 to 20:4, which is catalyzed by delta 6 desaturase, was slightly increased. Cd did not inhibit liver microsomal delta 9 desaturase activity in vitro. We concluded that Cd suppressed the induction of delta 9 desaturase in cultured hepatocytes. These phenomena can explain the selective inhibition of delta 9 desaturase in the livers of Cd-treated rats.

Animals↗
PubMed1996

Regulation by IL-5 of expression of functional platelet-activating factor receptors on human eosinophils.

Platelet-activating factor (PAF) and IL-5 are both important mediators of various allergic reactions, although the relationship between these mediators in allergic responses has not yet been fully elucidated. In this study, we investigated the effects of human rIL-5 on the expression of the PAF receptor (PAF-R) on eosinophils from healthy human volunteers. The specific binding of [3H]WEB 2086, a specific ligand for PAF-R, to eosinophils treated with 5 ng/ml of IL-5 for 12 h was significantly higher than that to untreated cells. The Bmax value for IL-5-treated eosinophils was 1.8-fold higher than that for untreated cells, while the Kd values remained unchanged. This clearly indicates that surface expression of PAF-R on eosinophils is enhanced by IL-5. The enhancement was first observed at 6 h and reached a plateau at 12 to 18 h. The effect of IL-5 was abolished in the presence of cycloheximide or actinomycin D. When the relative amount of PAF-R mRNA was determined by reverse transcription PCR, the message was found to be increased through activation of transcription of transcript 1 on exposure to IL-5. Furthermore, we found that the PAF-induced increase in intracellular calcium ion concentration in eosinophils was markedly augmented by exposure to IL-5 for 12 h. Several lines of data suggest that the enhanced expression of PAF-R on eosinophils due to IL-5 is the cause of the augmented response to PAF. These findings will be of value for understanding the mechanisms of selective infiltration and activation of eosinophils in allergic diseases in which PAF and IL-5 are crucially involved.

Azepines↗
PubMed1996

Potential biosynthetic connections between the two cannabimimetic eicosanoids, anandamide and 2-arachidonoyl-glycerol, in mouse neuroblastoma cells.

Anandamide (arachidonoyl-ethanolamide, AnNH) and 2-arachidonoyl-glycerol (2-AG) have been suggested to act as endogenous agonists at the brain cannabinoid receptor, and their biosynthetic and degradative mechanisms in nervous tissues and cells have also been partially elucidated. Here we present evidence for the presence, in mouse N18TG2 neuroblastoma cells, of enzymatic activities potentially responsible for the biosynthesis of AnNH and 2-AG from a common phospholipid precursor. Cell homogenates were shown to catalyze: (a) the transfer of an arachidonoyl moiety from the sn-1 position of sn-1,2-di-arachidonoyl-phosphatidylcholine (AAPC) to phosphatidyl-ethanolamine (PE) to form N-arachidonoyl-PE (N-ArPE) and sn-1-lyso-2-arachidonoyl-PC (lyso-APC), (b) the hydrolysis of N-AtPE to AnNH, (c) the hydrolysis of lyso-APC to 2-AG, (d) the hydrolysis of AAPC to sn-1,2-di-arachidonoyl-glycerol (AAG), and (e) the hydrolysis of AAG to 2-AG. From these findings it is possible to suggest that AAPC may serve as precursor for both AnNH and 2-AG biosynthesis through three different pathways.

Animals↗
PubMed1996

Transacylase-mediated and phosphodiesterase-mediated synthesis of N-arachidonoylethanolamine, an endogenous cannabinoid-receptor ligand, in rat brain microsomes. Comparison with synthesis from free arachidonic acid and ethanolamine.

The levels of N-arachidonoylethanolamine (anandamide), an endogenous cannabinoid-receptor ligand, and a relevant molecule, N-arachidonoylphosphatidylethanolamine (N-arachidonoylPtdEtn), in rat brain were investigated using a newly developed sensitive analytical method. We found that rat brain contains small but significant amounts of these two types of N-arachidonoyl lipids (4.3 pmol/g tissue and 50.2 pmol/g tissue, respectively). Then, we investigated how N-arachidonoylethanolamine (anandamide) is produced in the brain. We found that anandamide can be formed enzymatically via two separate synthetic pathways in the brain: enzymatic condensation of free arachidonic acid and ethanolamine; and formation of N-arachidonoylPtdEtn from PtdEtn and arachidonic acid esterified at the 1-position of phosphatidyl-choline (PtdCho), and subsequent release of anandamide from N-arachidonoylPtdEtn through the action of a phosphodiesterase. We confirmed that rat brain contains both the enzyme activities and lipid substrates involved in these reactions. Several lines of evidence strongly suggest that the second pathway, rather than the first one, meets the requirements and conditions for the synthesis of various species of N-acylethanolamine including anandamide in the brain.

Acyltransferases↗
PubMed1996

Enzymatic synthesis of anandamide, an endogenous cannabinoid receptor ligand, through N-acylphosphatidylethanolamine pathway in testis: involvement of Ca(2+)-dependent transacylase and phosphodiesterase activities.

Rat testis was shown to contain significant amounts of both N-acylethanolamine, including N-arachidonoylethanolamine (anandamide), and N-acylphosphatidylethanolamine (N-acylPE), including N-arachidonoylPE. The fatty acid profiles of the N-acyl moieties of the two classes resembled each other. We confirmed that testis microsomes contain a phosphodiesterase activity catalyzing the release of anandamide from N-arachidonoylPE. They also contain an enzyme activity catalyzing the transfer of arachidonic acid from the 1-position of diacylphospholipids to PE to form N-arachidonoylPE. These results suggest that the N-acylPE pathway is important in the synthesis of anandamide in this tissue.

Acyltransferases↗
PubMed1996

Biphasic effect of cadmium ions on the secretion of leukotriene B4 in rabbit alveolar macrophages.

One major role of alveolar macrophages is the production of eicosanoids, which modulate immune and inflammatory processes in the lung. In this study, the effects were investigated of cadmium ions on the secretion of leukotriene (LT)B4 and prostaglandin (PG)E2, predominant products of lipoxygenase and cyclooxygenase, respectively. Cd2+ had an inhibitory effect on the secretion of LTB4 and PGE2 in response to A23187 stimulation at concentrations > 3 x 10(-5)M. This effect can be explained by the inhibition of arachidonic acid (20:4) liberation from membrane phospholipids by Cd2+, because Cd2+ inhibits both [3H] arachidonic acid (20:4) liberation from [3H]20:4-prelabeled macrophages and the cytosolic phospholipase A2 activity. At concentrations < 3 x 10(-5)M, Cd2+ had no effect on PGE2 secretion but showed an augmentation of LTB4 secretion. In vitro study using macrophage lysate showed enhanced LTB4 synthesis from arachidonic acid by Cd2+, which could be responsible for the augmentation of LTB4 secretion in cells. These results indicate that Cd2+ may increase inflammation by increasing LTB4 production in lung.

Analysis of Variance↗