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K Hanasaki

Publications and source records attributed to K Hanasaki.

At least 37 records · Page 2Linked to original sources
PubMed1995

CD22-mediated cell adhesion to cytokine-activated human endothelial cells. Positive and negative regulation by alpha 2-6-sialylation of cellular glycoproteins.

We previously showed that cultured human umbilical vein endothelial cells (HEC) exposed to the inflammatory cytokines tumor necrosis factor-alpha or interleukin-1 display increased activity of beta-galactoside alpha 2,6-sialyltransferase. This is associated with enhanced expression of ligands for the B cell receptor CD22 beta, which recognizes alpha 2-6-linked sialic acids (Hanasaki, K., Varki, A., Stamenkovic, I., and Bevilacqua, M. P. (1994) J. Biol. Chem. 269, 10637-10643). Here we report that increased expression of CD22 ligands is a feature of dermal microvascular endothelial cells as well, and is also observed in response to the cytokine interleukin-4. Tumor necrosis factor-alpha stimulation of HEC causes no change in the profile of endothelial glycoproteins recognized by CD22, but doubles the proportion of total cellular N-linked oligosaccharides capable of binding tightly to CD22. This modest change is sufficient to cause a marked increase in alpha 2-6-linked sialic acid-dependent binding of Chinese hamster ovary (CHO) cells expressing recombinant human CD22. In contrast, B lymphoma cell lines expressing higher levels of cell surface CD22 do not show such sialic acid-dependent binding to activated HEC. Since B lymphoma cells themselves also express high levels of alpha 2-6-linked sialic acids, their CD22 molecules might be rendered nonfunctional by endogenous ligands. In support of this, the lectin function of CD22 can be directly detected on transfected CHO cells, but not on B lymphoma cells. Furthermore, coexpression of beta-galactoside alpha 2,6-sialyltransferase with CD22 in the CHO cells abrogates sialic acid-dependent binding to cytokine-activated HEC. However, such co-transfected cells can bind to B lymphoma cells in a manner apparently less dependent upon alpha 2-6-linked sialic acid, suggesting CD22-mediated interactions that may not be directly dependent on its lectin function. Thus, CD22-mediated interactions between B cells and activated vascular endothelium may be positively regulated by induction of alpha 2-6-linked sialic acid-bearing endothelial cell ligands, but negatively regulated by such ligands on the B cells expressing CD22. Since expression of both CD22 and beta-galactoside alpha 2,6-sialyltransferase are regulated during B cell ontogeny, these findings could be of importance in B cell function and/or trafficking.

Animals↗
PubMed1995

Binding of human plasma sialoglycoproteins by the B cell-specific lectin CD22. Selective recognition of immunoglobulin M and haptoglobin.

CD22 is a cell-surface receptor of resting mature B cells that recognizes sialic acid (Sia) in the natural structure Sia alpha 2-6Gal beta 1-4GlcNAc (Powell, L. D., Jain, R. K., Matta, K. L., Sabesan, S., and Varki, A. (1995) J. Biol. Chem. 270, 7523-7532). Human umbilical vein endothelial cells (HEC) treated with inflammatory cytokines such as tumor necrosis factor-alpha (TNF-alpha) display increases in cell-surface CD22 ligands, caused by increased expression of the enzyme beta-galactoside alpha 2,6-sialyltransferase (Hanasaki, K., Varki, A., Stamenkovic, I., and Bevilacqua, M. P. (1994) J. Biol. Chem. 269, 10637-10643; Hanasaki, K., Varki, A., and Powell, L. D. (1995) J. Biol Chem. 270, 7533-7542). Thus, CD22 could direct potential interactions between mature B cells and endothelial cells during inflammatory states. However, this would have to occur in the presence of blood plasma, which contains many sialoglycoproteins known to carry alpha 2-6-linked sialic acids. We show here that human plasma can indeed inhibit Sia-dependent binding of a recombinant soluble chimeric form of human CD22 (CD22Rg) to TNF-alpha activated HEC. Affinity adsorption of individual human plasma samples with immobilized CD22Rg showed that, of the numerous alpha 2-6-sialic acid containing glycoproteins in plasma, only three polypeptides with apparent molecular mass (under reducing conditions) of 74, 44, and 25 kDa bound, and were specifically eluted with alpha 2-6-sialyllactose. NH2-terminal amino acid sequencing of these high affinity CD22 ligands revealed that they are subunits of immunoglobulin M (IgM) and haptoglobin. Purified human IgM from pooled human plasma can be quantitatively bound by CD22Rg, and binding is blocked by alpha 2-6-sialyllactose, but not by alpha 2-3-sialyllactose. Pretreatment by sialidase or by mild periodate oxidation of sialic acid side chains abolishes these interactions. IgM at physiological concentrations also inhibits CD22Rg binding to TNF-alpha-activated HEC in a manner dependent not only upon its sialylation but also requiring its intact multimeric structure. These data show that CD22 is capable of highly selective recognition of certain multimeric plasma sialoglycoproteins that carry alpha 2-6-linked sialic acids. Notably, the two proteins that are selectively recognized are known to be involved in immune and inflammatory responses. Haptoglobin synthesis by the liver is markedly increased during the "acute phase response" to systemic inflammation, while IgM is the major product resulting from activation of resting CD22-positive B cells.

Antigens, CD↗
PubMed1994

Inositol polyanions. Noncarbohydrate inhibitors of L- and P-selectin that block inflammation.

Selectins are cell adhesion molecules known to support the initial attachment of leukocytes to inflamed vascular endothelium through their recognition of carbohydrate ligands such as the tetrasaccharide sialyl Lewisx (Neu5Ac alpha 2-3Gal beta 1-4(Fuc alpha 1-3)GlcNAc-). In the present study, we describe the inhibition of L- and P-selectin function by inositol polyanions, simple 6-carbon ring structures that have multiple ester-linked phosphate or sulfate groups. In a purified component competition assay, binding of L- and P-selectin-Ig fusion proteins to immobilized bovine serum albumin-sialyl Lewisx neoglycoprotein was inhibited by inositol hexakisphosphate (InsP6, IC50 = 2.1 +/- 1.4 microM and 160 +/- 40 microM), by inositol pentakisphosphate (InsP5, IC50 = 1.4 +/- 0.2 and 260 +/- 40 microM), and by inositol hexakissulfate (InsS6, IC50 = 210 +/- 80 microM and 2.8 +/- 0.9 mM); E-selectin-Ig binding was unaffected. Inositol polyanions diminished the adhesion of LS180 colon carcinoma cells to plates coated with L- and P-selectin-Ig but not with E-selectin-Ig. Inositol polyanions blocked polymorphonuclear leukocyte (PMN) adhesion to COS cells expressing recombinant transmembrane P-selectin but not to those expressing E-selectin. In addition, inositol polyanions diminished PMN adhesion to activated endothelial cells under rotation-induced shear stress, a process known to require L-selectin function. In vivo, the effects of inositol polyanions were studied in two murine models of acute inflammation. Intravenously administered InsP6 (two doses of 40 mumol/kg) inhibited PMN accumulation in thioglycolate-induced inflammation (55 +/- 10% inhibition) and in zymosan-induced inflammation (61 +/- 4% inhibition). InsP5 and InsS6 also inhibited inflammation in these models, although higher doses were required for InsS6. In conclusion, inositol polyanions are noncarbohydrate small molecules that inhibit L- and P-selectin function in vitro and inflammation in vivo.

Animals↗
PubMed1994

Cytokine-induced beta-galactoside alpha-2,6-sialyltransferase in human endothelial cells mediates alpha 2,6-sialylation of adhesion molecules and CD22 ligands.

Sialic acids decorating blood and cell surface proteins can play important roles in various biological processes. The inflammatory cytokines tumor necrosis factor-alpha (TNF-alpha) and interleukin-1, as well as bacterial lipopolysaccharide, can activate vascular endothelium, increasing expression of several surface glycoproteins. Here we show that treatment of cultured human endothelial cells (HEC) with TNF-alpha, interleukin-1, or lipopolysaccharide causes increased expression of the enzyme beta-galactoside alpha-2,6-sialytransferase (alpha 2-6STN). TNF-alpha was most effective, inducing a 3.5-fold enhancement of cell-associated sialytransferase activity by 72 h. In addition, activated HEC secreted a large portion of the induced sialyltransferase activity into the medium. Analysis of labeled HEC showed both a relative and an absolute increase of alpha 2,6-linked sialic acid on N-linked oligosaccharides after TNF-alpha stimulation. This coincided with increased expression of endothelial glycoproteins bearing N-linked glycans with alpha 2,6-linked sialic acid detected by the lectin Sambucus nigra agglutinin. The cytokine-inducible endothelial cell adhesion molecules E-selectin, ICAM-1, and VCAM-1 are among these glycoprotein substrates for alpha 2-6STN. These changes also correlated with a substantial increase in binding sites for CD22 beta, a mammalian lectin known to recognize oligosaccharides carrying multiple copies of alpha 2,6-linked sialic acid. Northern analysis revealed increased levels of mRNA encoding alpha 2-6STN. Thus, activation of endothelial cells during inflammatory and immunological processes may induce alpha 2-6STN, which can participate in sialylation of other activation-dependent molecules.

Antigens, CD↗
PubMed1994

Molecular cloning of pancreatic group I phospholipase A2 receptor.

We have recently reported that mammalian pancreatic group I phospholipase A2 (PLA2-I) has its specific receptor (PLA2 receptor) on a variety of mammalian cells and that various biological responses are elicited by PLA2-I via this receptor. In this study, we cloned cDNAs encoding a protein corresponding to the bovine PLA2 receptor purified from the corpora lutea on the basis of its partial amino acid sequences. The identity of a protein encoded by the cloned cDNA with the bovine PLA2 receptor was verified by a transient expression experiment using COS-7 cells. Interestingly, the deduced primary structure of the PLA2 receptor (1,463 amino acid residues) exhibits a close relatedness throughout the molecule to that of the macrophage mannose receptor, a unique member of Ca(2+)-dependent (C-type) animal lectin family, in spite of their functional diversity. Based on this sequence similarity between these two receptors, the domain organization of the PLA2 receptor could be tentatively assigned as follows; 10 extracellular domains including 8 tandem repeats homologous to C-type carbohydrate-recognition domains (CRDs) and a single transmembrane region followed by a short cytoplasmic tail. The results of transient expression experiments for mutant PLA2 receptors supported this assignment and furthermore suggested the region responsible for PLA2-I binding corresponds to CRDs in the mannose receptor.

Amino Acid Sequence↗
PubMed1992

Effect of pancreatic type phospholipase A2 on isolated porcine cerebral arteries via its specific binding sites.

The addition of porcine pancreatic group I phospholipase A2 (PLA2-I) produced a transient contraction followed by a relaxation in helical strips of porcine cerebral arteries. Its ED50 value (2.3 nM) was almost identical to the Kd value (3.9 nM) calculated from the specific binding of 125I-labeled porcine PLA2-I in cultured porcine cerebral arterial smooth muscle cells. Type-specific action of PLA2s and homologous desensitization strongly implicated the involvement of PLA2-I-specific sites in the response. The transient contraction was abolished by treatment with indomethacin as well as by the removal of endothelium, indicating the dependence of vasoconstrictor prostaglandins synthesized by PLA2-I in endothelium. The PLA2-I-induced relaxation response was also observed in bovine and cat cerebral arteries, thus providing a new aspect of PLA2-I as a vasoactive substance.

Animals↗
PubMed1992

Purification and characterization of a high-affinity binding protein for pancreatic-type phospholipase A2.

A high-affinity and specific binding site for mammalian group I phospholipase A2 (PLA2-I) was found on the membranes of bovine corpus luteum. Affinity labeling experiments revealed that PLA2-I binds to a single polypeptide with a mass of 190-200 kDa. The PLA2-I binding protein in the membranes was solubilized in an active form with n-octyl beta-D-thioglucoside, and then purified approx. 16,000-fold. The purification procedures consisted of diethylaminoethyl-Sephacel chromatography, PLA2-I-affinity gel chromatography and gel-filtration high-performance liquid chromatography on a TSKgel G3,000SWXL column. The final preparation migrated as a single molecular species of 190 kDa on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), and identification of the 190 kDa protein as the PLA2-I binding protein was demonstrated by ligand blotting analysis. The purified protein possessed a binding capacity with high affinity and specificity for a mammalian mature type of PLA2-I. Treatment of the purified material with N-glycosidase F resulted in increased mobility of the protein on SDS-PAGE as well as considerable abolition of the PLA2-I binding activity, thus suggesting the requirement of the carbohydrate moiety of the PLA2-I binding protein for receptor-ligand interactions.

Animals↗
PubMed1992

Migration of vascular smooth muscle cells by phospholipase A2 via specific binding sites.

Pancreatic-type group I phospholipase A2 (PLA2-I), EC 3.1.1.4, long thought to act as a digestive enzyme, has a specific binding site in several types of tissues and cells and these sites promote PLA2-I-stimulated DNA synthesis. In this study we report a PLA2-I action on the migration of rat embryonic thoracic aorta smooth muscle cells (A7r5). A7r5 cells had a single class of PLA2-I binding site with an equilibrium binding constant (Kd) value of 1.7 nM and a maximum binding capacity (Bmax) of 40,000 sites/cell. The migration activity of PLA2-I for A7r5 cells was examined using modified Boyden chambers. PLA2-I stimulated the migration dose-dependently, and the ED50 value was about 1 nM, which was almost the same as the Kd value for PLA2-I binding. Checkerboard analysis showed that the response of A7r5 cells to PLA2-I was chemokinetic, but not chemotactic. These findings reveal a new aspect of PLA2-I in the modulation of vascular function.

12-Hydroxy-5,8,10,14-eicosatetraenoic Acid↗
PubMed1992

Characterization of a high affinity binding site for pancreatic-type phospholipase A2 in the rat. Its cellular and tissue distribution.

We showed in an earlier study (Arita, H., Hanasaki, K., Nakano, T., Oka, S., Teraoka, H., and Matsumoto, K. (1991) J. Biol. Chem. 266, 19139-19141) that there is a high affinity and specific binding site for mammalian group I phospholipase A2 (PLA2-I) in Swiss 3T3 fibroblast cells. Analysis of the cellular distribution in rat using 125I-PLA2-I as a radioligand indicated the presence of this site in various cells, including vascular smooth muscle cells (VSMC), vascular endothelial cells, synovial cells, chondrocytes, and gastric mucosal cells. Scatchard analysis of rat VSMC revealed the existence of a single class of binding site with a Kd value of 1.60 nM and a Bmax value of 51.0 fmol/10(6) cells. The mammalian mature type of PLA2s-I derived from several animal species specifically recognized the same site in cells and stimulated DNA synthesis, whereas its inactive zymogen, mammalian group II PLA2s, and snake and bee venom PLA2s showed much lesser activities. 125I-PLA2-I bound to VSMC was rapidly internalized and subsequently released from the cells as trichloroacetic acid-soluble radioactivity. Down-regulation of the PLA2-I site was observed in the treatment of VSMC with cAMP-elevating agents, as well as glucocorticoids. Affinity labeling experiments indicated that PLA2-I binds to a single polypeptide with a mass of approximately 200,000 daltons. These results suggest a novel PLA2-I action on the cellular function via its specific binding site.

Animals↗
PubMed1992

[Treatment of liver metastases from gastric cancer].

Nine patients with liver metastases from gastric cancer were treated in our department since 1986. Hepatectomy was performed in 3 cases and hepatic arterial infusion chemotherapy was performed in 6 cases. In 3 patients in whom hepatectomy was performed, the extent of liver metastases showed 2 H1 and 1 H2. One has survived for 20 months, but the other 2 died after 5 and 7 months, respectively. In 6 patients in whom hepatic arterial infusion chemotherapy was performed, the extent of liver metastases was H3. These patients were treated with 5-FU.EPIR.MMC (3 cases), CDDP.MMC (1 case), MMC only (1 case) and 5-FU.ADM.MMC.CDDP (1 case). This treatment revealed a 50% response rate (CR 1, PR 2). The patient with CR has survived for 6 years and 2 patients with PR died after 8 and 12 months. The patient with CR showed high AFP level (55, 480 ng/ml), and 2 patients with PR showed high AFP level (24, 327 ng/ml) or high CEA level (3,903 ng/ml). The prognosis of hepatectomy for liver metastases from gastric cancer was not so good. Hepatic arterial infusion chemotherapy seemed to be a useful treatment for liver metastases from gastric cancer.

Aged↗
PubMed1991

Novel proliferative effect of phospholipase A2 in Swiss 3T3 cells via specific binding site.

Phospholipase A2 (PLA2), EC 3.1.1.4, which catalyzes the release of free fatty acids from the sn-2 position of glycerophospholipids, has been extensively studied from the viewpoint of eicosanoid production (Arita, H., Nakano, T., and Hanasaki, K. (1989) Prog. Lipid Res. 28, 273-301). Several lines of evidence suggest that extracellular PLA2 is pathophysiologically related to some disorders, including inflammation and hypersensitivity. Despite this, little is known of the precise mechanism of the pathological processes as well as their intrinsic correlation with dysfunction. Here, we report a novel PLA2 action on the proliferation of Swiss 3T3 fibroblasts via specific binding sites of approximately Mr 200,000. Pancreatic type PLA2 in the active form specifically recognized the sites and stimulated thymidine incorporation in DNA. Its inactive zymogen and other PLA2s from platelets, snake, and bee venoms showed much lesser activities. Although the physiological significance remains to be identified, our finding is the first to offer a new viewpoint on the effect of mammalian extracellular PLA2 on cellular function.

3T3 Cells↗
PubMed1991

Synergistic effect of thromboxane A2 and N-formylmethionylleucylphenylalanine on platelet-activating factor synthesis in human polymorphonuclear neutrophils.

The effects of thromboxane A2 (TXA2) on the synthesis of platelet-activating factor (PAF) and leukotriene B4 (LTB4) were studied using human polymorphonuclear neutrophils (PMN). Scatchard analysis for binding experiments using [3H]S-145, a specific TXA2/prostaglandin H2 (PGH2) receptor antagonist, revealed the existence of a single class of binding sites (Kd = 83.0 +/- 2.8 nM, Bmax = 113.0 +/- 3.1 fmol/2.10(6) cells) in human PMN. Upon stimulation with a combination of U46619, a TXA2 mimetic agonist, and N-formylmethionylleucylphenylalanine (FMLP, 1 microM), the synthesis of PAF was detected, although this was not significantly enhanced by U46619 or FMLP alone. The maximal production of PAF as well as the maximal activity of acetyl-CoA acetyltransferase was observed at approx. 20 min after addition of both stimuli. The effects of U46619 plus FMLP on PAF synthesis showed dose dependence to different concentrations of U46619 (0.1-10 microM), and were completely inhibited by S-145. Contrarily, no significant amounts of LTB4 were detected by radioimmunoassay during the stimulation with U46619 and FMLP. These results suggest that TXA2 and FMLP synergistically activate human PMN to induce PAF synthesis and this effect of TXA2 is mediated through its specific receptor.

15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5↗
PubMed1991

Endothelin-induced intracellular Ca2+ mobilization through its specific receptors in murine peritoneal macrophages.

We studied the presence of specific binding sites for endothelin (ET) and the effect of ET on cytosolic free Ca2+ concentration ([Ca2+]i) in murine thioglycolate-activated peritoneal macrophages. Scatchard analysis for binding experiments using [125I]ET-1 or [125I]ET-3 revealed the existence of a single class of binding sites. The binding parameters (Kd and Bmax) for [125I]ET-1 were almost identical to those for [125I]ET-3. In addition, unlabeled 3 ET isopeptides (ET-1, ET-2 and ET-3) inhibited the specific binding of both ET-1 and ET-3 with similar inhibitory potencies. All 3 ET isopeptides caused an increase in [Ca2+]i in the same dose-dependent manner (0.01-100 nM). These results demonstrate the existence of an ET receptor with the same affinity for all isoforms that mediates the ET-induced intracellular Ca2+ mobilization in murine peritoneal macrophages.

Animals↗
PubMed1991

Kinetic studies on stereospecific recognition by the thromboxane A2/prostaglandin H2 receptor of the antagonist, S-145.

1. The mechanism for the stereospecific recognition of the antagonist S-145 by the thromboxane A2 (TXA2)/prostaglandin H2 (PGH2) receptor was examined by ligand-binding techniques in rat vascular smooth muscle cells (VSMCs) and in human platelet membranes. 2. Scatchard analysis revealed the existence of a single class of binding sites with the same maximum number for both [3H]-(+)-S-145 and [3H]-(-)-S-145 in both cell types. The dissociation constants (Kd) for the binding of the (+)-isomer in rat VSMCs and human platelet membranes were, respectively, 0.40 +/- 0.03 and 0.20 +/- 0.02 nM, each value being lower than that for the (-)-isomer (3.57 +/- 0.74 and 2.87 +/- 0.08 nM, respectively). 3. The rank orders of potency (Ki) for a series of TXA2/PGH2 ligands at inhibiting [3H]-(+)-S-145 binding were highly correlated with those determined for [3H]-(-)-S-145 binding in both cell preparations. 4. Kinetic analysis of the binding of both radioligands revealed a much lower dissociation rate constant (k-1) and a slightly greater association rate constant (k1) for the (+)-isomer compared to those for the (-)-isomer. 5. These results suggest that it is at the stage of dissociation from the TXA2/PGH2 receptor that the stereochemistry of the optical isomers of S-145 confers their difference in affinity for these receptors in rat VSMCs and human platelet membranes.

Animals↗
PubMed1990

Receptor-mediated mitogenic effect of thromboxane A2 in vascular smooth muscle cells.

The effects of thromboxane A2 (TXA2) on the proliferation of vascular smooth muscles cells (VSMC) were examined using primary cultures of VSMC from rat aorta. U46619, a stable TXA2 mimetic, stimulated DNA synthesis of VSMC only in the presence of insulin. The effect was concentration-dependent with a half-maximal effect obtained at approximately 1 x 10(-8) M. The mitogenic effect of U46619 was larger than that of endothelin, another mitogen derived from endothelium. Among several TXA2/PGH2 analogs, the proliferative activity was detected only in the agonists, and not in the antagonists or in the metabolite of TXA2. A series of TXA2/PHG2 receptor antagonists completely suppressed the U46619-stimulated DNA synthesis as well as the [3H]SQ29,548 binding to the TXA2/PGH2 receptors in VSMC. The rank order of binding affinities to the receptors among the respective antagonists correlated well with the potencies for suppression of the proliferative effects of U46619. The mitogenic effects of U46619 were also attenuated by the presence of calcium antagonists. U46619 caused activation of phospholipase C with the production of inositol trisphosphate, leading to increases in the intracellular free Ca2+ concentration as measured with the fluorescent indicator fura-2. These results suggest that TXA2 induces mitogenic effects on VSMC through binding to its specific receptors. This effect of TXA2 on the proliferation of VSMC may be related to the development of atherosclerosis.

15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5↗
PubMed1990

Phorbol ester-induced expression of the common, low-affinity binding site for primary prostanoids in vascular smooth muscle cells.

We showed in an earlier study (Hanasaki, K., and Arita, H. (1989) Biochim. Biophys. Acta 1013, 28-35) that there is a common, low-affinity binding site for primary prostanoids in cultured vascular smooth muscle cells (VSMC). This site, called the "primary prostaglandin (PG) site," can be evaluated by radioreceptor assay using [3H]PGF2 alpha and [3H]PGE1. Comparison of the capacity of several PGF2 alpha analogs to displace both radioligand bindings indicated strict requirements of the 15-hydroxy group as well as the 13,14-double bond in the omega-side chain of prostaglandins for recognition of this site. Treatment of VSMC with phorbol 12-myristate 13-acetate (PMA), a known protein kinase C activator, led to concentration- and time-dependent increases in the binding activities of [3H] PGF2 alpha as well as [3H]PGE1, which could be completely suppressed by the addition of protein kinase C inhibitor, H-7. The PMA effects could be mimicked by phorbol 12,13-dibutylate, but not by inactive phorbol ester. Scatchard analyses revealed an approximately 8-fold increase in the binding density with unaltered binding affinity after PMA treatment. This expression of the primary PG site was blocked by the addition of cycloheximide and actinomycin D. In contrast, PMA did not affect the binding activity for the thromboxane A2/prostaglandin H2 receptor in VSMC. These results suggest that the expression of the primary PG site is regulated by a protein kinase C-dependent mechanism in VSMC.

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine↗