Biomedical subjects

T Asano

Publications and source records attributed to T Asano.

At least 433 records · Page 24Linked to original sources

PubMed1994

Proteolytic modification of membrane-associated phospholipase C-beta by mu-calpain enhances its activation by G-protein beta gamma subunits in human platelets.

Membrane-associated phosphoinositide-phospholipase C (PI-PLC)-beta (150 kDa) and its truncated forms (100 kDa and 45 kDa) were purified from human platelets. The 100 kDa PI-PLC-beta was found to be activated to a greater extent by brain G-protein beta gamma subunits compared to the intact 150 kDa enzyme. Furthermore, treatment with mu-calpain of the intact PI-PLC-beta (150 kDa) caused a marked augmentation of its activation by beta gamma subunits. This enhanced PLC activation by beta gamma subunits was due to truncation by mu-calpain, producing a 100 kDa PI-PLC, but not by another protease, thrombin.

Blood Platelets↗

PubMed1994

Overexpression of hexokinase I but not GLUT1 glucose transporter alters concentration dependence of glucose-stimulated insulin secretion in pancreatic beta-cell line MIN6.

The recently established pancreatic beta-cell line MIN6 retains the ability to secrete insulin in response to physiological glucose concentrations. To investigate the role of glucose transport and phosphorylation in glucose-stimulated insulin secretion by beta-cells, MIN6 cells were stably transfected with a rabbit GLUT1 glucose transporter cDNA or a rat hexokinase I cDNA cloned in an expression vector. Overexpression of GLUT1 increased 3-O-methylglucose uptake, but did not alter either glucose utilization or glucose-stimulated insulin secretion. In contrast, clones overexpressing hexokinase I exhibited enhanced glucose-stimulated insulin secretion at glucose concentrations below 10 mM with a concomitant increase in glucose utilization. Maximal insulin secretion as well as the maximal rate of glucose utilization were not altered in these clones. Insulin secretion stimulated by 2-ketoisocaproate, a non-glucose secretagogue, was not affected by hexokinase I expression. These results strongly suggest that the glucose phosphorylating step, but not glucose transport step, regulates glucose-stimulated insulin secretion by modulating the glycolytic rate in the beta-cell.

3-O-Methylglucose↗

PubMed1994

Characterization of vasoactive intestinal peptide receptors in canine liver membranes.

The binding characteristics of vasoactive intestinal peptide (VIP) in the liver membranes of the dog were examined using radioligand binding assay with 125I-VIP and unlabelled peptides and results were compared with those from the rat. The binding of VIP to canine liver membranes occurred in a reversible, saturable, specific and temperature-dependent manner. Guanine nucleotides dose-dependently inhibited VIP binding. The order of potency in competition experiments with unlabelled peptide was: VIP > pituitary adenylate cyclase activating peptide (PACAP)-27 > PACAP-38 >> peptide histidine isoleucine (PHI) = secretin in the dog, and PACAP-27 > PACAP-38 > VIP > PHI > secretin in the rat. PHI and secretin were about 5000 times less potent than VIP in the dog, but secretin was about 100 times less potent than VIP in the rat. The VIP binding sites in canine liver membranes have recognition sites for VIP which differ from those in rat liver membranes. As most of VIP in the portal vein was removed during its passage through the canine liver, the binding sites of canine liver may play a role in degradation of VIP.

Animals↗

PubMed1994

A brain-specific gamma subunit of G protein freed from the corresponding beta subunit under non-denaturing conditions.

It has generally been accepted that the beta and gamma subunits of G proteins are tightly associated and can only be dissociated under denaturing conditions. However, one form of the gamma subunit, free of the beta subunit, was isolated under non-denaturing conditions during the purification of beta gamma complexes from bovine brain. Amino acid sequence analysis revealed that the N-terminus of this gamma subunit was not blocked and its sequence was identical to that of a brain-specific gamma subunit, gamma 3. Among three forms of gamma subunits observed in bovine brain (gamma 2, gamma 3 and gamma 7), gamma 3 was the only form that was not modified at the N-terminus. Although the physiological significance of the free form of gamma 3 is not clear, these results suggest that N-terminal modification of gamma subunits may be important for interaction with beta subunits.

Adenosine Diphosphate Ribose↗

PubMed1994

Detection of minus-strand hepatitis C virus RNA in tumor tissues of hepatocellular carcinoma.

BACKGROUND: Recently, the detection of hepatitis C virus (HCV) antibody has been widely performed for clinical serum testing of HCV infection and can be identified in most hepatocellular carcinoma (HCC) cases in Japan. In the current study, the authors detected not only plus-strand but also minus-strand HCV RNA as a template for RNA replication in hepatocytes in the resected tumors of HCC, and they investigated those significant for HCC. METHODS: The plus-strand and minus-strand HCV RNA were detected using reverse transcription-polymerase chain reaction after the extraction of RNA from the tumor and nontumor tissues of hepatectomized liver, respectively. RESULTS: The detection of HCV RNA in liver tissues from hepatectomized specimens of 20 cases was as follows: in nontumor regions, both of the plus- and minus-strand HCV RNA were detected in 14 cases. Among those cases, plus- and minus-strand HCV RNA were detected in 11 cases, and in another 2 cases, only plus-strand HCV RNA was found in the tumor regions. CONCLUSIONS: These results suggest that HCV RNA is able to replicate in tumor tissues of HCC and may be involved in the development of HCC and also that HCV in remnant hepatocytes may cause the recurrence of HCC as secondary carcinogenesis after hepatectomy.

Adult↗

PubMed1994

Affinity binding of hamster oviductin to spermatozoa and its influence on in vitro fertilization.

The aim of this study was to determine the influence of hamster oviductal glycoprotein (oviductin) on in vitro gamete interaction. Oviductin was purified from the oviducts using lithium 3,5-diiodosalicylate, followed by phenol extraction. Immunocytochemistry using indirect fluorescence staining revealed that oviductin binds to the sperm anterior acrosomal region. The specific binding of oviductin resulted in inhibition of in vitro fertilization in studies using cumulus-free oocytes. The inhibitory effect was dependent on the concentration of oviductin and occurred in both ovarian and oviductal oocytes but not zona-free oocytes, indicating that sperm-zona interaction was interferred by oviduction. However, the inhibitory effect of oviductin in sperm-zona interaction was reduced when cumulus-enclosed oocytes from ovaries and oviducts were used, indicating that the egg investment including cumulus oophorus has some effect on oviductin-sperm complex and maintaining the fertilizing ability.

Animals↗

PubMed1994

Effects of protein kinase C activators on Na, K-ATPase activity in rat brain microvessels.

In order to study the possible role of C kinase (PKC) on sodium pump of cerebral vessels, we used diacylglycerol (diC8: sn-1,2-dioctanoylglycerol) and phorbol esters (PMA: phorbol 12-myristate 13-acetate; PDA: phorbol 12,13-diacetate; 4 alpha-P: 4-alpha phorbol) as PKC activators, and examined their effects on Na,K-ATPase activity in rat brain microvessels (MVs). Rats were divided into non-treated (control; n = 9), four-vessel occlusion (4VO; 30-30 minutes ischemia and recirculation, n = 5), and middle cerebral artery occlusion (MCAO, n = 3) groups. MVs were passed through nylon meshes and were obtained by ultracentrifuge at 58000 g. Na,K-ATPase activity in MVs was determined by the phosphomolybdate method. DiC8 enhanced Na,K-ATPase activity at 10(-4) M in the control group, the 4VO group and the contralateral hemispheres of the MCAO group (139% +/- 0.06, 135% +/- 0.2, 133% +/- 0.18, mean +/- SE, p < 0.05, p < 0.01, Wilcoxon rank sum) respectively, but had no effects on MVs in the ipsilateral hemispheres of MCAO group (74% +/- 0.04). This activation by diC8 was inhibited by PKC inhibitors, staurosporine (3 x 10(8) M) and H7 (10(-6) M) in the control MVs. By contrast, PMA suppressed Na, K-ATPase at 10(-5) M in the control group (-25% +/- 0.07), but it tended to activate Na,K-ATPase activity in the ipsilateral hemispheres of the MCAO groups (33% +/- 0.09). PDA and 4 alpha-P did not have any consistent effects at the concentration examined. The cause of difference between the effects of diC8 and PMA is unclear at present, but it may stem from the mode of lipid-membrane interaction in these agents and the difference in the condition of cells as well.

Animals↗

PubMed1994

Effect of endothelin-1 as growth factor on a human glioma cell line; its characteristic promotion of DNA synthesis.

In addition to its powerful vasoconstrictive activity, endothelin-1 (ET-1) has been recognized to stimulate DNA synthesis in some cell lines. In this study, we confirmed the existence of ET-1 receptor in YKG-1 human glioma cells, and investigated its effect on DNA synthesis in YKG-1 for 6 consecutive days, comparing it with that of epidermal growth factor (EGF), platelet-derived growth factor (PDGF), and insulin-like growth factor-I (IGF-I). Scatchard analysis of the binding data revealed the presence of a single class of high-affinity binding molecule. The apparent dissociation constant (Kd) was 5.2 x 10(-9) M and the maximal binding capacity (B max) was 4.7 x 10(4) sites/cell. The percentage of non-cycling cells was initially more than 85%, and decreased to 55.40%, 24.22%, 11.50%, and 7.51% on days 1, 2, 4, and 6, respectively, after ET-1 stimulation. Although ET-1 reduces the fraction of non-cycling cells more slowly than other growth factors such as EGF, PDGF and IGF-I, it reaches the same level as the others by day 6. These results indicate that YKG-1 human glioma cells have ET-1 receptors and that ET-1 initiates a peculiar slow induction of DNA synthesis in these cells, suggesting that secondary factors might exist to accelerate the DNA synthesis in response to ET-1.

Cell Cycle↗

PubMed1994

The effect of Nicorandil on chronic cerebral vasospasm.

The present study is aimed at examining the therapeutic effect of Nicorandil on chronic vasospasm using beagle dogs subjected to a "two-haemorrhage" insult, as well as its dilatory effect on the PDA [phorbol-12,13 diacetate]-induced contraction of the canine basilar artery. 1. A total of 12 animals of either sex, weighing 7 to 12 kg, were assigned into saline control and Nicorandil-treated groups. Immediately after the second induction of subarachnoid haemorrhage (SAH), animals started to receive the agent via the venous route at the constant rate of 10 micrograms/kg/minute for six hours (day 3). On days 4, 5, and 6, the drug was given twice at the same rate for three hours. After the final angiograms, animals were sacrificed by exsanguination. 2. Using ring specimen of the canine basilar artery at a resting tension of 3 g, isometric tension was monitored to examine the effect of Nicorandil on PDA induced contraction. Nicorandil significantly ameliorated chronic vasospasm and inhibited PDA-induced contraction in a dose-dependent fashion. The present data indicate that Nicorandil provides a useful way of treating chronic vasospasm after SAH.

Animals↗

PubMed1994

Liposome-encapsulated muramyl tripeptide up-regulates monocyte chemotactic and activating factor gene expression in human monocytes at the transcriptional and post-transcriptional levels.

Liposome-encapsulated muramyl tripeptide phosphatidylethanolamine (L-MTP-PE) is a novel immune modulator that is now under investigation against metastatic melanoma and osteosarcoma. We have already reported that L-MTP-PE induced monocyte-mediated tumoricidal activity and up-regulation of the tumor necrosis factor and interleukin-1 (IL-1) in vivo and in vitro. We now demonstrate that L-MTP-PE also induces monocyte chemotactic and activating factor (MCAF) mRNA expression at both the transcriptional and post-transcriptional levels. Monocyte chemotactic activity was also present in the supernatants of L-MTP-PE-stimulated cells. In monocytes, the increased expression of MCAF was induced rapidly (by 2 h) but was short-lived. By 4 h, MCAF mRNA had decreased to background level. We found no change in MCAF mRNA levels in lymphocytes exposed to L-MTP-PE. We therefore conclude that L-MTP-PE selectively up-regulates MCAF expression in monocytes and that MCAF may play a role in the tumoricidal and immune-stimulating activity of L-MTP-PE.

Acetylmuramyl-Alanyl-Isoglutamine↗

PubMed1994

CD3-negative lymphoproliferative disease of granular lymphocytes in a girl with an unusual pattern of anti-Epstein-Barr virus antibodies.

UNLABELLED: We describe an 11-year-old girl who initially had mild hepatosplenomegaly and then presented with abnormal expansion of CD3-negative granular lymphocytes in peripheral blood and Epstein-Barr virus (EBV) genome in the DNA obtained from the peripheral blood mononuclear cells (PBMNC). After approximately 3 years, she developed oedema, ascites, marked hepatosplenomegaly, and pancytopenia, and showed both a profile of anti-EBV antibodies of reactivated infection and a high titre of anti-cytomegalovirus antibody. Although she was treated with antibiotics, ganciclovir, and prednisolone, she died of hepatic failure. CONCLUSION: Careful clinical observation, periodic examination of anti-EBV antibodies, and the analysis of EBV genome from PBMNC are needed in young patients with CD3-negative lymphoproliferative disease of granular lymphocytes.

CD3 Complex↗

PubMed1994

Mitochondrial diabetes mellitus: prevalence and clinical characterization of diabetes due to mitochondrial tRNA(Leu(UUR)) gene mutation in Japanese patients.

Mutations in the mitochondrial gene were recently identified in a large pedigree of diabetes mellitus and deafness. As the mitochondrial gene is materially inherited, Japanese diabetic patients whose mothers were also diabetic were screened, using peripheral leucocytes, for an A to G transition at nucleotide pair 3243 of the mitochondrial gene, a tRNA(Leu(UUR)) mutation. This mutation was identified in four pedigrees from among 300 unrelated patients who were screened. Diabetes co-segregated with the mutation, except in one young subject, and was maternally inherited. The apparent onset of disease occurred between 11 and 68 years of age. Some of the affected members developed hearing impairment and congestive heart failure due to cardiomyopathy, though generally long after the onset of diabetes, and these patients had therefore not been diagnosed as having a specific form of diabetes. The duration of sulphonyl-urea treatment was not more than 8 years in these pedigrees and affected members were prone to progression to insulin-requiring diabetes. Thus, these patients were secondary sulphonylurea failures. Long-term follow-up revealed that the underlying disorder in affected members is a progressive impairment of insulin secretion. Some were initially diagnosed as having IDDM based on an apparent acute onset in youth and the clinical severity of their diabetes. Others were regarded as having MODY with an aggressive course. The mitochondrial gene mutation or diabetes is not transmitted to all offspring of the affected mothers.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗

PubMed1994

Vasoactive intestinal peptide released by acetylcholine in the dog ileum.

Release of vasoactive intestinal peptide (VIP) in response to acetylcholine (ACh) was characterized in the dog ileum using cholinergic antagonists. In blood-perfused ileum, ACh (2-200 nmol/min) produced a dose-dependent increase in venous VIP output, which was slightly reduced by hexamethonium (10 nmol/min) and blocked by hexamethonium and atropine (10 nmol/min) in combination. In isolated ileal tissues containing the submucous or myenteric plexus, excess KCl (75 mM), veratridine (0.1 mM) and ACh (0.1 mM) evoked the release of VIP. ACh-induced VIP output was decreased slightly by hexamethonium (0.1 mM), and blocked by atropine (0.1 mM) or pirenzepine (0.1 mM). Dimethylphenylpiperazinium (0.1 mM) also caused a small increase in VIP output sensitive to hexamethonium in the ileal tissues containing either the submucous or myenteric plexus. It is concluded that ACh evokes the release of VIP from VIP-containing neurons of the submucous and myenteric plexuses in the dog ileum mainly through the activation of M1 muscarinic receptors.

Acetylcholine↗

PubMed1994

G protein Gi2 alpha in the cochlea: cloning and selective occurrence in receptor cells.

A cDNA library was made from the mouse cochlea and screened with a G protein-cDNA like molecule obtained from cochlear tissue by polymerase chain reaction. The nucleotide sequence of a clone, named cochlear Gi2 alpha, had 99.2% identity to mouse macrophage Gi2 alpha. Using an antibody which is selective for Gi2 alpha, expression of the cochlear Gi2 alpha was localized in outer and inner hair cells of the organ of Corti. Possible functional roles of this G protein in hair cells are discussed.

Amino Acid Sequence↗

PubMed1994

Direct evidence of induction of interdigitated gel structure in large unilamellar vesicles of dipalmitoylphosphatidylcholine by ethanol: studies by excimer method and high-resolution electron cryomicroscopy.

Interaction of large unilamellar vesicle (LUV) of dipalmitoylphosphatidylcholine (DPPC) with ethanol was investigated by the excimer method developed by Yamazaki et al. (Yamazaki, M., M. Miyazu, and T. Asano. 1992. Biochim. Biophys. Acta. 1106:94-98) and the high-resolution electron cryomicroscope with a new cryostage (top-entry superfluid stage) (HiRECM) developed by Fujiyoshi, Y. et al. (Fujiyoshi, Y., T. Mizusaki, K. Morikawa, H. Aoki, H. Kihara, and Y. Harada. 1991. Ultramicroscopy. 38:241-251). The excimer method is based on the fact that the ratio of excimer to monomer fluorescence intensity (E/M) of pyrene PC is lowered in the membrane in the interdigitated gel structure (L beta I), because structural restriction of L beta I structure largely decreases collisions of pyrene rings of the pyrene PCs in the membrane. E/M of pyrene PC in DPPC LUV decreased largely at high concentrations of ethanol, which indicated the induction of L beta I structures in DPPC LUV. Frozen-hydrated DPPC LUVs in a vitreous ice were observed at 4K with HiRECM, and these images were characterized by a pair of concentric circles. The membrane thickness of DPPC LUV which was estimated from the distance between the two concentric lines decreased largely at high concentration of ethanol. The mean value of membrane thickness of the LUV in the absence of ethanol was 3.8 nm, while at 15% (w/v) ethanol was 3.0 nm. These values were almost same as those obtained from the electron density profile of DPPC MLV by the x-ray diffraction analysis in each structures, L beta' and L beta I structures, respectively. These results indicated directly the induction of L beta 1 structure in DPPC LUV at high concentration of ethanol.

1,2-Dipalmitoylphosphatidylcholine↗

PubMed1994

Isolation and characterization of Streptoverticillium anticoagulant (SAC), a novel protein inhibitor of blood coagulation produced by Streptoverticillium cinnamoneum subsp. cinnamoneum.

Three Streptoverticillium anticoagulants, SAC I, II, and III, which strongly inhibit human intrinsic blood coagulation, were each isolated in a homogeneous form from a culture fluid of Streptoverticillium cinnamoneum subsp. cinnamoneum IFO 12852. SAC I, II, and III are simple proteins with molecular weights of around 12,000, and with isoelectric points of 9.7, 9.7, and 9.9, respectively. Their amino acid compositions are similar and each SAC possesses two disulfide bonds. The COOH-terminal residue of each of these proteins is phenylalanine. Together with the similarity of their protein chemical properties, the results of NH2-terminal amino acid sequence analysis of these SAC proteins strongly suggested that the deletion of Ser-Leu and Ser-Leu-Tyr from the NH2-terminus of SAC I (Ser-Leu-Tyr-Ala-Pro-...) results in the generation of SAC II and III, respectively. The amount of each SAC necessary to double the partial thromboplastin time was around 5 micrograms/ml. SAC I inhibited activated human factor XII and human plasma kallikrein. It also inhibited, but to a lesser extent, activated factor X. The inhibition constants (Ki) of SAC I toward activated factor XII and plasma kallikrein were 5.3 x 10(-8) and 7.2 x 10(-9) M, respectively. The SACs also inhibited some microbial serine proteases such as subtilisin Carlsberg and, to a lesser extent, mammalian serine proteases including bovine trypsin and alpha-chymotrypsin. Of these three inhibitors, only SAC I inhibited metalloproteases such as thermolysin in addition to these serine proteases.

Amino Acid Sequence↗

PubMed1994

Primary structure and reactive site of Streptoverticillium anticoagulant (SAC), a novel protein inhibitor of blood coagulation produced by Streptoverticillium cinnamoneum subsp. cinnamoneum.

The complete amino acid sequence of SAC I, a novel protein inhibitor of blood coagulation produced by Streptoverticillium cinnamoneum subsp. cinnamoneum IFO 12852, was determined. Automated Edman degradation was employed for its fragment peptides, which were obtained by specific cleavage procedures including tryptic, chymotryptic, and peptic digestions, and cyanogen bromide treatment. SAC I is composed of 110 amino acid residues with a molecular weight of 11,642. It has two intramolecular disulfide bonds, Cys31-Cys46 and Cys68-Cys98, but no cysteine residues. The overall sequence homology of SAC I is 58% to plasminostreptin, a protein protease inhibitor produced by Streptomyces antifibrinolyticus, and 52% to S-SI, Streptomyces subtilisin inhibitor, produced by Streptomyces albogriseolus. Subtilisin [EC 3.4.21.14]-modified SAC I, which was prepared by incubating SAC I with subtilisin, had a newly-generated amino terminal sequence, Glu71-Trp72-Asn73-, in addition to the original amino terminal sequence, and a newly-generated carboxyl terminal arginine in addition to the original phenylalanine. These results clearly show that the Arg70-Glu71 bond was specifically cleaved on the limited proteolysis with subtilisin and that the bond is the relative site for subtilisin.

Amino Acid Sequence↗

PubMed1994

High frequency of cancer patients with abnormal assembly of the T cell receptor-CD3 complex in peripheral blood T lymphocytes.

Structural abnormality of T cell receptor (TCR)-CD3 complex on the cell surface was investigated in peripheral blood lymphocytes (PBL) from 55 cancer patients. In 24 of the 68 tests done on these patients, the CD3 zeta chain was not detected by immunoprecipitation with anti-CD3 epsilon monoclonal antibody (mAb), but was observed with anti-CD3 zeta mAb, suggesting that a high frequency of cancer patients possesses abnormal T cell receptor (TCR) complex in PBL. On the other hand, the total zeta chain was missing in several advanced cases. During follow-up of several patients, the zeta chain became undetectable after two or three months of cancer progression. It appears that immunosuppressive status can be monitored by analyzing the TCR-CD3 complex on the cell surface of PBL.

Disease Progression↗