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Satoshi Ishii

Publications and source records attributed to Satoshi Ishii.

At least 37 records · Page 2Linked to original sources
PubMed2004

Platelet-activating factor receptor develops airway hyperresponsiveness independently of airway inflammation in a murine asthma model.

Lipid mediators play an important role in modulating inflammatory responses. Platelet-activating factor (PAF) is a potent proinflammatory phospholipid with eosinophil chemotactic activity in vitro and in vivo. We show in this study that mice deficient in PAF receptor exhibited significantly reduced airway hyperresponsiveness to muscarinic cholinergic stimulation in an asthma model. However, PAF receptor-deficient mice developed an eosinophilic inflammatory response at a comparable level to that of wild-type mice. These results indicate an important role for PAF receptor, downstream of the eosinophilic inflammatory cascade, in regulating airway responsiveness after sensitization and aeroallergen challenge.

Animals↗
PubMed2004

Dyslipidemia associated with atherosclerotic disease systemically alters dendritic cell mobilization.

High LDL and/or low HDL are risk factors for atherosclerosis and are also a common clinical feature in systemic lupus erythematosus, rheumatoid arthritis, and psoriasis. Here, we show that changes in lipid profiles that reflect atherosclerotic disease led to activation of skin murine dendritic cells (DCs) locally, promoted dermal inflammation, and induced lymph node hypertrophy. Paradoxically, DC migration to lymph nodes was impaired, suppressing immunologic priming. Impaired migration resulted from inhibitory signals generated by platelet-activating factor (PAF) or oxidized LDL that acts as a PAF mimetic. Normal DC migration and priming was restored by HDL or HDL-associated PAF acetylhydrolase (PAFAH), which mediates inactivation of PAF and oxidized LDL. Thus, atherosclerotic changes can sequester activated DCs in the periphery where they may aggravate local inflammation even as they poorly carry out functions that require their migration to lymph nodes. In this context, HDL and PAFAH maintain a normally functional DC compartment.

1-Alkyl-2-acetylglycerophosphocholine Esterase↗
PubMed2004

Efficient and rapid purification of recombinant human alpha-galactosidase A by affinity column chromatography.

The lysosomal enzyme alpha-galactosidase A (alpha-Gal A) metabolizes neutral glycosphingolipids that possess alpha-galactoside residues at the non-reducing terminus, and inherited defects in the activity of alpha-Gal A lead to Fabry disease. We describe here an efficient and rapid purification procedure for recombinant alpha-Gal A by sequential Concanavalin A (Con A)-Sepharose and immobilized thio-alpha-galactoside (thio-Gal) agarose column chromatography. Optimal elution conditions for both columns were obtained using overexpressed human alpha-Gal A. We recommend the use of a mixture of 0.9 M methyl alpha-mannoside and 0.9 M methyl alpha-glucoside in 0.1 M acetate buffer (pH 6.0) with 0.1 M NaCl for the maximum recovery of glycoproteins with multiple high-mannose type sugar chains from Con A column chromatography, and that the Con A column should not be reused for the purification of glycoproteins that are used for structural studies. Binding of the enzyme to the thio-Gal column requires acidic condition at pH 4.8. A galactose-containing buffer (25 mM citrate-phosphate buffer, pH 5.5, with 0.1 M galactose, and 0.1 M NaCl) was used to elute alpha-Gal A. This procedure is especially useful for the purification of mutant forms of alpha-Gal A, which are not stable under conventional purification techniques. A protocol that purifies an intracellular mutant alpha-Gal A (M279I) expressed in COS-7 cells within 6h at 62% overall yield is presented.

Animals↗
PubMed2004

Platelet-activating factor receptor-deficient mice show an unaltered clearance of nontypeable Haemophilus influenzae from their respiratory tract.

Platelet-activating factor (PAF), a glycerophospholipid with proinflammatory properties, exerts its biological effects by interacting with the PAF receptor (PAFR) expressed on many different cell types. The PAFR specifically binds phosphorylcholine, the biologically active component of PAF. However, phosphorylcholine is also a component of the cell wall of nontypeable Haemophilus influenzae (NTHi). In recently published in vitro experiments, the invasion of respiratory epithelial cells by NTHi was mediated by the PAFR. To determine the role of the PAFR in host defense against pneumonia induced by NTHi, PAFR-deficient (PAFR-/-) and normal wild-type mice were intranasally inoculated with NTHi. The absence of a functional PAFR was associated with a normal innate immune response as indicated by similar bacterial counts, myeloperoxidase activity, and inflammation within the pulmonary compartment of PAFR-/- and wild-type mice. These data indicate that the PAFR does not interfere with the clearance of NTHi from the respiratory tract.

Animals↗
PubMed2004

Absence of platelet-activating factor receptor protects mice from osteoporosis following ovariectomy.

While platelet-activating factor (PAF) is produced in various diseases associated with bone resorption, its functions in bone metabolism remain unknown. Using PAF receptor-deficient mice, we evaluated the role of PAF in the development of bone resorption following ovariectomy, a model of postmenopausal osteoporosis. Through observations of bone mineral density and histomorphometric parameters, it was found that bone resorption was markedly attenuated in PAF receptor-deficient mice, indicating that PAF links estrogen depletion and osteoporosis in vivo. Osteoclasts expressed higher amounts of the enzymes required for PAF biosynthesis than osteoblasts. TNF-alpha and IL-1beta increased the acetyl-coenzyme A:lyso-PAF acetyltransferase activity in osteoclasts. Osteoclasts, but not osteoblasts, expressed the functional PAF receptor. PAF receptor stimulation prolonged the survival of osteoclasts in vitro. Furthermore, osteoclasts treated with a PAF receptor antagonist, and also those from PAF receptor-deficient mice, showed reductions in survival rate and Ca resorption activity. Consistently, in organ cultures, bone resorption was significantly suppressed by a PAF receptor antagonist treatment or genetic PAF receptor deficiency. Thus, these results suggest that, through the inflammatory cytokines, estrogen depletion enhances PAF production as a unique autocrine factor for osteoclast functions. Inhibition of PAF function might pave the way for a new strategy to prevent postmenopausal bone loss without disturbing osteoblast functions.

Animals↗
PubMed2003

Identification of p2y9/GPR23 as a novel G protein-coupled receptor for lysophosphatidic acid, structurally distant from the Edg family.

Lysophosphatidic acid (LPA) is a bioactive lipid mediator with diverse physiological and pathological actions on many types of cells. LPA has been widely considered to elicit its biological functions through three types of G protein-coupled receptors, Edg-2 (endothelial cell differentiation gene-2)/LPA1/vzg-1 (ventricular zone gene-1), Edg-4/LPA2, and Edg-7/LPA3. We identified an orphan G protein-coupled receptor, p2y9/GPR23, as the fourth LPA receptor (LPA4). Membrane fractions of RH7777 cells transiently expressing p2y9/GPR23 displayed a specific binding for 1-oleoyl-LPA with a Kd value of around 45 nm. Competition binding and reporter gene assays showed that p2y9/GPR23 preferred structural analogs of LPA with a rank order of 1-oleoyl- > 1-stearoyl- > 1-palmitoyl- > 1-myristoyl- > 1-alkyl- > 1-alkenyl-LPA. In Chinese hamster ovary cells expressing p2y9/GPR23, 1-oleoyl-LPA induced an increase in intracellular Ca2+ concentration and stimulated adenylyl cyclase activity. Quantitative real-time PCR demonstrated that mRNA of p2y9/GPR23 was significantly abundant in ovary compared with other tissues. Interestingly, p2y9/GPR23 shares only 20-24% amino acid identities with Edg-2/LPA1, Edg-4/LPA2, and Edg-7/LPA3, and phylogenetic analysis also shows that p2y9/GPR23 is far distant from the Edg family. These facts suggest that p2y9/GPR23 has evolved from different ancestor sequences from the Edg family.

Animals↗
PubMed2003

Preparation of 7-substituted ginkgolide derivatives: potent platelet activating factor (PAF) receptor antagonists.

Ginkgolides are structurally unique constituents of Ginkgo biloba extracts and are known antagonists of the platelet-activating factor (PAF) receptor. Ginkgolide C is 25-fold less potent than ginkgolide B as a PAF receptor antagonist, due to the presence of the 7beta-OH. Recently, we found that 7alpha-fluoro ginkgolide B was equipotent to ginkgolide B underlining the critical importance of the 7-position of ginkgolides for PAF receptor activity. Herein we describe the synthesis of a series of ginkgolide B derivatives with modifications at the 7-position and the pharmacological evaluation of these derivatives as assayed by cloned PAF receptors. In two cases nucleophilic attack on a 7beta-O-triflate ginkgolide B did not lead to the expected products, but gave rise to two unprecedented ginkgolide derivatives, one with a novel rearranged skeleton. Furthermore, standard reduction of 7alpha-azido ginkgolide B did not give the expected primary amine, but instead yielded alkylated amines depending on the solvent employed. Pharmacological testing with cloned PAF receptors showed that ginkgolides with 7alpha-substitutents had increased affinity compared to 7beta-substituents, in particular 7alpha-chloro ginkgolide B, the most potent nonaromatic ginkgolide derivative described to date with a K(i) value of 110 nM.

Animals↗
PubMed2003

Experimental Trypanosoma cruzi infection in platelet-activating factor receptor-deficient mice.

The generation of an inflammatory response driven by Trypanosoma cruzi or its subproducts appears to be essential for tissue injury and disease pathogenesis. However, this inflammatory response is also relevant in the control of T. cruzi replication. The lipid mediator platelet-activating factor (PAF) has been implicated in a number of pathological conditions characterized by tissue inflammation. In the present study, we aimed at evaluating the role of PAF during T. cruzi infection by using mice that were genetically deficient in the PAF receptor. We observed that infected hearts of PAFR(-/-) mice had an increased number of parasite nests, associated with a more intense inflammatory infiltrate. This was associated with greater parasitemia and lethality. When wild-type and PAFR(-/-) mice were compared, there were no marked changes in the kinetics of the expression of MCP-1, RANTES, IFN-gamma and TNF-alpha in heart tissue of infected animals. Moreover, serum concentrations of TNF-alpha, nitrate and parasite-specific IgM were similar in both groups of mice. In vitro, macrophages from PAFR(-/-) animals did not phagocytose trypomastigote forms when activated with PAF, leukotriene B(4) or MCP-1 and produced less nitric oxide when infected and activated with IFN-gamma. These results are consistent with the hypothesis that endogenous synthesis of PAF and activation of PAF receptors control T. cruzi replication in mice in great part via facilitation of the uptake of the parasite and consequent activation of macrophages.

Animals↗
PubMed2003

Role of PAF receptors during intestinal ischemia and reperfusion injury. A comparative study between PAF receptor-deficient mice and PAF receptor antagonist treatment.

1 The reperfusion of ischemic tissues may be associated with local and systemic inflammation that prevents the full benefit of blood flow restoration. The present study aimed to confirm a role for platelet-activating factor receptor(s) (PAFR) during ischemia and reperfusion injury by using genetically modified mice deficient in the PAFR (PAFR(-/-) mice) and to evaluate comparatively the effectiveness of pharmacological treatment using the PAFR antagonist UK-74,505 (modipafant). 2 The reperfusion of the ischemic superior mesenteric artery (SMA) induced marked local (intestine) and remote (lungs) tissue injury, as assessed by the increase in vascular permeability, neutrophil influx and intestinal hemorrhage and in the production of TNF-alpha. There was also a systemic inflammatory response, as shown by the increase in serum TNF-alpha concentrations and marked reperfusion-associated lethality. 3 After reperfusion of the ischemic SMA, PAFR(-/-) mice had little tissue or systemic inflammation and lethality was delayed, but not prevented, in these mice. Interestingly, the reperfusion-associated increases in tissue concentrations of IL-10 were significantly greater in PAFR(-/-) than wild-type mice. 4 Pretreatment with PAFR antagonist UK-74,505 (1 mg kg(-1)) markedly prevented tissue injury, as assessed by the increase in vascular permeability, neutrophil accumulation, hemorrhage and TNF-alpha concentrations in the intestine and lungs. In contrast, UK-74,505 failed to affect reperfusion-associated lethality and increases in serum TNF-alpha when used at 1 mg kg(-1). 5 Reperfusion-associated lethality and increase in serum TNF-alpha were only affected when a supra-maximal dose of the antagonist was used (10 mg kg(-1)). At this dose, UK-74,505 also induced a marked enhancement of reperfusion-associated increases in tissue concentrations of IL-10. However, at the same dose, UK-74,505 failed to prevent reperfusion-associated lethality in PAFR(-/-) mice any further. 6 The present studies using genetically modified animals and a receptor antagonist firmly establish a role of PAFR activation for the local, remote and systemic inflammatory injury and lethality which follows reperfusion of the ischemic SMA in mice. Moreover, it is suggested that high doses of PAFR antagonists need to be used if the real efficacy of these compounds is to be tested clinically.

Animals↗
PubMed2003

Host response of platelet-activating factor receptor-deficient mice during pulmonary tuberculosis.

Platelet-activating factor (PAF) is a phospholipid with potent, diverse actions, which has been implicated as an important mediator in host defence against several intracellular pathogens. To determine the role of PAF in host defence in pulmonary tuberculosis, PAF receptor-deficient (PAFR-/-) and wild-type (PAFR+/+) mice were infected intranasally with a virulent strain of Mycobacterium tuberculosis. Mycobacterial outgrowth in lungs and liver did not differ significantly between PAFR-/- and PAFR+/+ mice at 2 or 6 weeks postinfection. After 28 weeks, 86% of PAFR-/- mice and 79% of PAFR+/+ mice had died (non-significant). In addition, both mouse strains were indistinguishable with respect to histopathology, the recruitment and activation of lymphocytes, and cytokine concentrations in the lung. These data suggest that PAF is not involved in the protective immune response to tuberculosis.

Animals↗
PubMed2003

Involvement of platelet-activating factor and LIS1 in neuronal migration.

Platelet-activating factor (PAF, 1-O-alkyl-2-acetyl-sn-glycero-3-phosphocholine) is a biologically active lipid mediator. We have previously shown the expression of PAF receptor in neurons and microglia. PAF is produced in the brain from its precursor, and degraded by the enzyme PAF acetylhydrolase. LIS1 is a regulatory subunit of PAF acetylhydrolase, and is identical to a gene whose deletion causes the human neuronal migration disorder, type I lissencephaly. Indeed, Lis1 mutant mice display defects in neuronal migration and layering in vivo, and also in cerebellar granule cell migration in vitro. However, the roles of PAF and the PAF receptor in the neuronal migration remain to be determined. Here, we show that PAF receptor-deficient mice exhibited histological abnormalities in the embryonic cerebellum. PAF receptor-deficient cerebellar granule neurons migrated more slowly in vitro than wild-type neurons, consistent with the observation that a PAF receptor antagonist reduced the migration of wild-type neurons in vitro. Synergistic reduction of neuronal migration was observed in a double mutant of PAF receptor and LIS1. Unexpectedly, PAF affected the migration of PAF receptor-deficient neurons, suggesting a receptor-independent pathway for PAF action. The PAF receptor-independent response to PAF was abolished in granule neurons derived from the double mutant mice. Thus, our results suggest that the migration of cerebellar granule cells is regulated by PAF through receptor-dependent and receptor-independent pathways, and that LIS1 is a pivotal molecule that links PAF action and neuronal cell migration both in vivo and in vitro.

1-Alkyl-2-acetylglycerophosphocholine Esterase↗
PubMed2002

A potent inhibitor of cytosolic phospholipase A2, arachidonyl trifluoromethyl ketone, attenuates LPS-induced lung injury in mice.

Acute respiratory distress syndrome (ARDS) is an acute lung injury of high mortality rate, and sepsis syndrome is one of the most frequent causes of ARDS. Metabolites of arachidonic acid, including thromboxanes and leukotrienes, are proinflammatory mediators and potentially involved in the development of ARDS. A key enzyme for the production of these inflammatory mediators is cytosolic phospholipase A(2) (cPLA(2)). Recently, it has been reported that arachidonyl trifluoromethyl ketone (ATK) is a potent inhibitor of cPLA(2). In the present study, we hypothesized that pharmacological intervention of cPLA(2) could affect acute lung injury. To test this hypothesis, we examined the effects of ATK in a murine model of acute lung injury induced by septic syndrome. The treatment with ATK significantly attenuated lung injury, polymorphonuclear neutrophil sequestration, and deterioration of gas exchange caused by lipopolysaccharide and zymosan administration. The current observations suggest that pharmacological intervention of cPLA(2) could be a novel therapeutic approach to acute lung injury caused by sepsis syndrome.

Animals↗
PubMed2002

Ginkgolide derivatives for photolabeling studies: preparation and pharmacological evaluation.

The terpene trilactones (TTLs), ginkgolides and bilobalide, are structurally unique constituents of Ginkgo biloba extracts, which exhibit various neuromodulatory properties. Although the TTLs are believed to be responsible for some of these effects, the specific interactions with targets in the central nervous system remain to be elucidated on a molecular level. Ginkgolides are known antagonists of the platelet-activating factor (PAF) receptor. Herein, we describe the first examination of the binding of native TTLs and their derivatives to the cloned PAF receptor, confirming that of the TTLs, ginkgolide B is the most potent PAF receptor antagonist. Ginkgolide derivatives carrying photoactivatable and fluorescent groups for the purpose of performing photolabeling have been prepared and evaluated using the cloned PAF receptor. These studies have shown that ginkgolide derivatives with aromatic photoactivatable substituents are potent PAF receptor antagonists with K(i) values of 0.09-0.79 microM and hence excellent ligands for clarifying the binding of ginkgolides to PAF receptor by photolabeling studies. Ginkgolide derivatives incorporating both fluorescent and photoactivatable groups still retained binding affinity to the PAF receptor and hence should be promising ligands for photolabeling and subsequent sequencing studies.

Animals↗
PubMed2002

Characterization of mouse cysteinyl leukotriene receptors mCysLT1 and mCysLT2: differential pharmacological properties and tissue distribution.

Cysteinyl leukotrienes (LTs) are important proinflammatory mediators. Their precise roles in mice need to be elucidated to interpret mouse models of inflammatory diseases. For this purpose, we cloned and characterized mouse receptors for cysteinyl LTs, mCysLT(1) and mCysLT(2). mCysLT(1) and mCysLT(2) were composed of 339 amino acids with 87.3% identity and 309 amino acids with 73.4% identity to human orthologues, respectively. A pharmacological difference was noted between mouse and human CysLT(2). Pranlukast, a specific inhibitor for human CysLT(1), antagonized mCysLT(2) responses as determined by Ca(2+) elevation and receptor-induced promoter activation. The mRNA expressions of both mCysLTs were higher in C57BL/6 mice than in 129 mice. mCysLT(1) mRNA was expressed mainly in skin, lung, and small intestine. mCysLT(2) was seen more ubiquitously with high expressions in spleen, lung, and small intestine. By in situ hybridization we demonstrated for the first time that mCysLT(1) and mCysLT(2) were expressed in subcutaneous fibroblasts. The different pharmacological characteristics of CysLT(2) between human and mouse and the different distributions of CysLTs between mouse strains suggest that careful choice and interpretation are necessary for a study of CysLTs using animal models.

Amino Acid Sequence↗
PubMed2002

Alternative splicing in the alpha-galactosidase A gene: increased exon inclusion results in the Fabry cardiac phenotype.

Fabry disease is an inborn error of glycosphingolipid catabolism, resulting from deficient activity of lysosomal alpha-galactosidase A (alpha-Gal A). A rare alternative splicing that introduces a 57-nucleotide (nt) intronic sequence to the alpha-Gal A transcript from intron 4 of the gene has been identified. In addition, a novel midintronic base substitution that results in substantially increased alternative splicing has been identified in a patient with Fabry disease who has the cardiac variant phenotype. The sequence of the patient's intron 4 contains a single G-->A transversion at genomic nt 9331 (IVS4+919 G-->A ), located at the minus sign4 position of the 3' end of the intronic insertion (nts 9278--9334 in the genomic sequence). Minigene constructs containing the entire intron 4 sequence with G, A, C, or T at nt 9331 within an alpha-Gal A complementary DNA expression vector were prepared and expressed in COS-1 cells. Whereas transfection of the G or T minigenes transcribed predominantly normal-sized transcripts, the transfection of the A or C minigenes produced a large amount of the alternatively spliced transcript. These results suggest that the G-->A mutation, within an A/C-rich domain, results in increased recognition of the alternative splicing by an A/C-rich enhancer-type exonic splicing enhancer. The intronic mutation was not observed in 100 unrelated unaffected men but was present in 6 unrelated patients with cardiac Fabry disease. Reverse-transcriptase polymerase chain reaction of total RNA of various normal human tissues revealed that the alternatively spliced transcript was present in all of the samples, and especially at a higher ratio in the lung and muscle. The normal transcript was present in the patients' lymphoblasts and resulted in approximately 10% residual enzyme activity, leading to a cardiac phenotype of Fabry disease.

Alternative Splicing↗
PubMed2002

Receptor-dependent metabolism of platelet-activating factor in murine macrophages.

Degradation of platelet-activating factor (PAF) was examined by incubating PAF with macrophages from PAF receptor-deficient mice. The degradation rate was halved as compared with wild-type mice. The reduction of the rate was comparable with the presence of a PAF antagonist WEB 2086 in wild-type cells. PAF was internalized rapidly (t(12) approximately 1 min) into wild-type macrophages. The PAF internalization was inhibited by the treatment of 0.45 m sucrose but was not affected by phorbol 12-myristate 13-acetate, suggesting that PAF internalizes into macrophages with its receptor in a clathrin-dependent manner. Internalized PAF was degraded into lyso-PAF with a half-life of 20 min. Treatment of concanavalin A inhibited the conversion of PAF into lyso-PAF, suggesting that uptake of PAF enhances PAF degradation. Lyso-PAF was subsequently metabolized into 1-alkyl-2-acyl-phosphatidylcholine. In addition, release of PAF acetylhydrolase from macrophages was enhanced when wild-type macrophages were stimulated with PAF but not from macrophages of PAF receptor-deficient mice. Thus, the PAF stimulation of macrophages leads to its degradation through both intracellular and extracellular mechanisms.

1-Alkyl-2-acetylglycerophosphocholine Esterase↗
PubMed2002

Platelet-activating factor receptor.

Platelet-activating factor (PAF, 1-O-alkyl-2-acetyl-sn-glycero-3-phosphocholine) is a biologically active phospholipid mediator. Although PAF was named for its potential to induce platelet aggregation, intense investigations have elucidated potent biological actions of PAF in a broad range of cell types and tissues. PAF acts by binding to a unique G-protein-coupled seven transmembrane receptor, and activates multiple intracellular signaling pathways. In the last decade, we have identified the PAF receptor structures, intracellular signaling mechanisms, and genomic organizations. Recently, we found a single nucleotide polymorphism of the human PAF receptor (A224D) with an allele frequency of 7.8% in Japanese. Cells expressing this receptor exhibited the reduced cellular signaling, although the binding parameters remain unchanged. We have established two different types of genetically altered mice, i.e. PAF receptor-overexpressing mouse and PAF receptor-deficient mouse. These mutant mice provide a novel and specific approach for identifying the pathophysiological and physiological functions of PAF in vivo. This review focuses on phenotypes of these mutant mice and summarizes the previous reports regarding PAF and PAF receptor.

Animals↗