Biomedical subjects

Takeshi Watanabe

Publications and source records attributed to Takeshi Watanabe.

At least 73 records · Page 4Linked to original sources

PubMed2004

Crystallization and preliminary X-ray analysis of plant class I chitinase from rice.

We report here on crystallization and preliminary X-ray analysis of plant class I chitinase from rice (OsChia1b). Similar single crystals of full-length OsChia1b were obtained under two independent conditions. The crystals grown under these conditions diffracted up to 2.1 and 2.5 angstroms resolution, respectively, at a synchrotron beamline, and were found to belong to the tetragonal space group P4(3)2(1)2.

Chitinases↗

PubMed2004

Involvement of hypothalamic histamine H1 receptor in the regulation of feeding rhythm and obesity.

Histamine H(1) receptors (H(1)-Rs) are found in peripheral tissues and in regions of the hypothalamus that are concerned with regulating body composition. In the present study, we investigated the detailed mechanisms of histamine H(1)-Rs in the development of obesity. Histamine H(1)-R knockout (H1KO) mice gradually developed mature-onset obesity, which was accompanied by hyperphagia and decreased expression of uncoupling protein-1 (UCP-1) mRNA. Both younger nonobese (12-week-old) and older obese (48-week-old) H1KO mice exhibited impairment of the responsiveness to the leptin. In addition, disruption of the diurnal rhythm of feeding occurred before the onset of obesity in H1KO mice. Correction of these abnormal feeding rhythms by means of scheduled feeding caused a reduction in obesity and associated metabolic disorders in H1KO mice. Furthermore, central administration of a histamine H(1)-R agonist affected feeding behavior, body weight, and c-fos-like immunoreactivity in the hypothalamus. Taken together, these findings suggest that histamine H(1)-Rs are crucial for the regulation of feeding rhythm and in mediating the effects of leptin. Early disruption of H(1)-R-mediated functions in H1KO mice may lead to hyperphagia and decreased expression of UCP-1 mRNA, which may contribute to the development of obesity in these animals. In addition, centrally acting histamine H(1)-R may be a novel therapeutic target for the treatment of obesity and related metabolic disorders.

Adipose Tissue, Brown↗

PubMed2004

Structural analysis of human hair fibers under the ultra-high voltage electron microscope.

Three-dimensional structural analyses of human hair fibers and comparison of the different fibers were tried by using the Ultra-high Voltage Electron Microscope (UHVEM). The analysis condition, sample preparation, and a machine state were adjusted to the suitable condition for tilting observation of from -70 degree to +70 degree, at 2 degrees intervals. The tomography of hair fiber was successfully reconstructed from the different angle pictures with IMODE software in a computer. By using UHVEM, the various human hair fibers from Japanese and Caucasians were investigated and discussed about their structures.

Hair↗

PubMed2003

Complete sequencing and characterization of 21,243 full-length human cDNAs.

As a base for human transcriptome and functional genomics, we created the "full-length long Japan" (FLJ) collection of sequenced human cDNAs. We determined the entire sequence of 21,243 selected clones and found that 14,490 cDNAs (10,897 clusters) were unique to the FLJ collection. About half of them (5,416) seemed to be protein-coding. Of those, 1,999 clusters had not been predicted by computational methods. The distribution of GC content of nonpredicted cDNAs had a peak at approximately 58% compared with a peak at approximately 42%for predicted cDNAs. Thus, there seems to be a slight bias against GC-rich transcripts in current gene prediction procedures. The rest of the cDNAs unique to the FLJ collection (5,481) contained no obvious open reading frames (ORFs) and thus are candidate noncoding RNAs. About one-fourth of them (1,378) showed a clear pattern of splicing. The distribution of GC content of noncoding cDNAs was narrow and had a peak at approximately 42%, relatively low compared with that of protein-coding cDNAs.

Chromosomes, Human, 21-22 and Y↗

PubMed2003

Aromatic residues within the substrate-binding cleft of Bacillus circulans chitinase A1 are essential for hydrolysis of crystalline chitin.

Bacillus circulans chitinase A1 (ChiA1) has a deep substrate-binding cleft on top of its (beta/alpha)8-barrel catalytic domain and an interaction between the aromatic residues in this cleft and bound oligosaccharide has been suggested. To study the roles of these aromatic residues, especially in crystalline-chitin hydrolysis, site-directed mutagenesis of these residues was carried out. Y56A and W53A mutations at subsites -5 and -3, respectively, selectively decreased the hydrolysing activity against highly crystalline beta-chitin. W164A and W285A mutations at subsites +1 and +2, respectively, decreased the hydrolysing activity against crystalline beta-chitin and colloidal chitin, but enhanced the activities against soluble substrates. These mutations increased the K(m)-value when reduced (GlcNAc)5 (where GlcNAc is N -acetylglucosamine) was used as the substrate, but decreased substrate inhibition observed with wild-type ChiA1 at higher concentrations of this substrate. In contrast with the selective effect of the other mutations, mutations of W433 and Y279 at subsite -1 decreased the hydrolysing activity drastically against all substrates and reduced the kcat-value, measured with 4-methylumbelliferyl chitotrioside to 0.022% and 0.59% respectively. From these observations, it was concluded that residues Y56 and W53 are only essential for crystalline-chitin hydrolysis. W164 and W285 are very important for crystalline-chitin hydrolysis and also participate in hydrolysis of other substrates. W433 and Y279 are both essential for catalytic reaction as predicted from the structure.

Amino Acids, Aromatic↗

PubMed2003

The directionality of chitin biosynthesis: a revisit.

The molecular directionality of chitin biosynthesis was investigated by transmission electron microscopy (TEM) using electron crystallography methods applied to reducing-end-labelled beta-chitin microcrystals from vestimentiferan Lamellibrachia satsuma tubes and nascent beta-chitin microfibrils from the diatom Thalassiosira weissflogii. The data allowed confirmation that the microfibrils were extruded with their reducing end away from the biosynthetic loci, an orientation consistent only with elongation through polymerization at the non-reducing end of the growing chains. Such a chain-extension mechanism, which has also been demonstrated for cellulose and hyaluronan, appears to be general for glycosyltransferases that belong to the GT2 (glycosyl transferase 2) family. The data also allowed confirmation that in beta-chitin the chains are crystallized in a 'parallel-up' mode, in contrast with hypotheses proposed in previous reports.

Animals↗

PubMed2003

Improgan antinociception does not require neuronal histamine or histamine receptors.

Improgan, a chemical congener of the H(2) antagonist cimetidine, induces antinociception following intracerebroventricular (i.c.v.) administration in rodents, but the mechanism of action of this compound remains unknown. Because the chemical structure of improgan closely resembles those of histamine and certain histamine blockers, and because neuronal histamine is known to participate in pain-relieving responses, the antinociceptive actions of improgan were evaluated in mice containing null mutations in the genes for three histamine receptors (H(1), H(2), and H(3)) and also in the gene for histidine decarboxylase (the histamine biosynthetic enzyme). Similar to earlier findings in Swiss-Webster mice, improgan induced maximal, reversible, dose-related reductions in thermal nociceptive responses in ICR mice, but neither pre-improgan (baseline) nor post-improgan nociceptive latencies were changed in any of the mutant mice as compared with wild-type controls. Improgan also had weak inhibitory activity in vitro (pK(i)=4.7-4.9) on specific binding to three recently-discovered, recombinant isoforms of the rat H(3) receptor (H(3A), H(3B), and H(3C)). The present findings strongly support the hypothesis that neuronal histamine and its receptors fail to play a role in improgan-induced antinociception.

Analgesics↗

PubMed2003

Evaluation of the effects of anti-pruritic drugs on scratch responses using histamine H1 receptor-deficient mice.

The effects of anti-pruritic drugs on scratching behavior associated with passive cutaneous anaphylaxis in histamine H(1) receptor-deficient and wild-type mice were studied. Passive sensitization with mouse monoclonal anti-dinitrophenyl-immunoglobulin E (IgE) resulted in an increase in the incidence of scratching behavior induced by intravenous injection of dinitrophenyl-ovalbumin in both wild-type and histamine H(1) receptor-deficient mice. The histamine H(1) receptor antagonist diphenhydramine inhibited scratching behavior induced by antigen in passively sensitized wild-type mice, whereas no effect was observed in histamine H(1) receptor-deficient mice. On the other hand, oxatomide inhibited scratching behavior in both mice, although the effect in wild-type mice was more potent than that in histamine H(1) receptor-deficient mice. Tranilast inhibited scratching behavior with the same potency in both mice. We concluded that the scratching behavior associated with passive cutaneous anaphylaxis involves not only histamine H(1) receptors but also other chemical mediators. Furthermore, the results of the present study indicated that oxatomide has an antagonistic effect on histamine H(1) receptors as well as anti-pruritic effect in vivo.

Animals↗

PubMed2003

Involvement of noradrenergic system within the central nucleus of the amygdala in naloxone-precipitated morphine withdrawal-induced conditioned place aversion in rats.

RATIONALE: We previously reported that the central nucleus of the amygdala (CeA) plays a crucial role in the negative affective, rather than somatic, component of morphine withdrawal. However, numerous studies have reported that the central ascending noradrenergic system is implicated in morphine withdrawal syndrome, although the roles of the noradrenergic system within the CeA in the negative affective component remains less clear. OBJECTIVES: The possible role of the noradrenergic system within the CeA in the negative affective component of morphine withdrawal was investigated. METHODS: The extracellular noradrenaline level within the CeA during naloxone-precipitated morphine withdrawal was measured using an in vivo microdialysis experiment on unanesthetized and freely moving rats. The effects of microinjection of beta-adrenoceptor antagonists into the bilateral CeA on the naloxone-precipitated morphine withdrawal-induced conditioned place aversion (CPA) and somatic signs were examined. RESULTS: The extracellular noradrenaline level within the CeA was transiently elevated during morphine withdrawal. Intra-CeA injections of beta-adrenoceptor antagonists propranolol (30 nmol per side) and timolol (10 nmol per side) significantly attenuated the morphine withdrawal-induced CPA. Similarly, beta(1)-antagonist atenolol (30 nmol per side) or beta(2)-antagonist butoxamine (30 nmol per side) significantly attenuated the CPA. In contrast, they did not affect morphine withdrawal-induced somatic signs, except for propranolol. CONCLUSION: These results suggest that the activation of the noradrenergic system within the CeA contributes to naloxone-precipitated morphine withdrawal-induced CPA, rather than somatic signs, through beta(1)- and beta(2)-adrenoceptors.

Adrenergic beta-Antagonists↗

PubMed2003

Hematopoietic stimulation by a dipeptidyl peptidase inhibitor reveals a novel regulatory mechanism and therapeutic treatment for blood cell deficiencies.

In hematopoiesis, cytokine levels modulate blood cell replacement, self-renewal of stem cells, and responses to disease. Feedback pathways regulating cytokine levels and targets for therapeutic intervention remain to be determined. Amino boronic dipeptides are orally bioavailable inhibitors of dipeptidyl peptidases. Here we show that the high-affinity inhibitor Val-boro-Pro (PT-100) can stimulate the growth of hematopoietic progenitor cells in vivo and can accelerate neutrophil and erythrocyte regeneration in mouse models of neutropenia and acute anemia. Hematopoietic stimulation by PT-100 correlated with increased cytokine levels in vivo. In vitro, PT-100 promoted the growth of primitive hematopoietic progenitor cells by increasing granulocyte-colony-stimulating factor (G-CSF), interleukin-6 (IL-6), and IL-11 production by bone marrow stromal cells. Two molecular targets of PT-100 are expressed by stromal cells- CD26/DPP-IV and the closely related fibroblast activation protein (FAP). Because PT-100 was active in the absence of CD26, FAP appears to be the hematopoietic target for PT-100. Interaction of PT-100 with the catalytic site seems to be required because amino-terminal acetylation of PT-100 abrogated enzyme inhibition and hematopoietic stimulation. PT-100 is a therapeutic candidate for the treatment of neutropenia and anemia. The data support increasing evidence that dipeptidyl peptidases can regulate complex biologic systems by the proteolysis of signaling peptides.

Anemia↗

PubMed2003

Chemical kindling induced by pentylenetetrazol in histamine H(1) receptor gene knockout mice (H(1)KO), histidine decarboxylase-deficient mice (HDC(-/-)) and mast cell-deficient W/W(v) mice.

The role of brain histamine on seizure development of pentylenetetrazol (PTZ)-induced kindling was examined in H(1)-receptor gene knockout (H(1)KO), histidine decarboxylase-deficient (HDC(-/-)) and mast cell-deficient (W/W(v)) mice. All H(1)KO, HDC(-/-) and W/W(v) mice had accelerated seizure development of PTZ-induced kindling when compared to their respective wild-type mice. The daily PTZ-kindling increased histamine content in the cortex and diencephalon of H(1)KO mice, whereas the histamine content in the diencephalon of W/W(v) mice was decreased. The present study indicates that histamine plays a suppressive role in seizure development through H(1)-receptors.

Analysis of Variance↗

PubMed2003

Recent developments in marine indole alkaloid synthesis.

The manzamine alkaloids such as manzamine A, B, and C belongs to a unique family of cyctotoxic beta-carbline linked azacycles, which have been isolated from several marine sponges. Manzamine A has recently been shown to exhibit in vivo antimalarial activity against parasite Plasmodium berghei. Further progress was added recently by the isolation of the new and closely related members such as nakadomarin A, as well as an ingenious proposal for their biosynthetic pathway. Martefragin A, isolated from the Japan sea alga Martensia fragilis Harvey, has been shown to have a potent inhibitory activity against lipid peroxidation. Our recen, efforts to develop total synthesis entries to manzamine A, manzamine B, manzamine C, nakadomarin A, and martefragin A are presented.

Alkaloids↗

PubMed2003

Expression of apoptosis-associated protein RCAS1 in macrophages of histiocytic necrotizing lymphadenitis.

Receptor-binding cancer antigen expressed on SiSo cells (RCAS1), which is recognized by the 22-1-1 monoclonal antibody (MoAb) against human uterine adenocarcinoma cell line SiSo, has been identified on various kinds of cancer cells. RCAS1 appears to be an apoptosis-associated protein that induces apoptosis in activated T-cells and erythroid progenitor cells. We previously demonstrated that monocytes/macrophages express RCAS1. In the present study, we investigated RCAS1 expression by 22-1-1 MoAb in histiocytic necrotizing lymphadenitis (HNL), which is characterized by necrotic lesions consisting of T-cells undergoing apoptosis and macrophages in proliferation. Expression of RCAS1 was analyzed by immunohistochemical staining in 9 cases of HNL and in 9 cases of reactive lymphadenitis used as a control. The ratio of RCAS1+ cells to CD68+ cells (monocytes/macrophages) was significantly higher in the patients with HNL than in the patients with reactive lymphadenitis (P = .0002; paired t test). Our findings suggest that RCAS1 expressed on macrophages may play an important role in the induction of activated T-cell apoptosis in cases of HNL.

Adolescent↗

PubMed2003

Clinical significance of serum RCAS1 levels detected by monoclonal antibody 22-1-1 in patients with cholangiocellular carcinoma.

BACKGROUND/AIMS: The tumor-associated antigen, RCAS1, has been reported to be expressed in various types of cancer, including cholangiocarcinoma. We measured serum RCAS1 levels in patients with intrahepatic cholangiocellular carcinoma (CCC) and other hepatobiliary diseases, and examined the clinical significance of serum RCAS1 as a tumor marker. METHODS: Sera collected from the patients and healthy volunteers were used for ELISA for RCAS1. The values of RCAS1 for CCC patients were compared to those of other tumor marker proteins. RESULTS: Serum RCAS1 levels exceeded the normal limit in a high percentage (73.9%) of CCC patients. The positivity rate was higher than those of CA19-9 and CEA. No correlation was found between the RCAS1 and CA19-9 concentrations. Serum RCAS1 was positive in many cases that were negative for CA19-9. Surgical resection of CCC reduced the RCAS1 level to within the normal range. On the other hand, serum RCAS1 levels were elevated in very few cases of benign hepatobiliary disease. CONCLUSIONS: As a tumor marker in CCC, RCAS1 is, at least, of complementary value to CA19-9 and CEA. Measuring serum RCAS1 contributes to the diagnostic accuracy, and is useful for estimating tumor progression or therapeutic effect.

Adult↗

PubMed2003

The role of chemical mediators in eosinophil infiltration in allergic rhinitis in mice.

The involvement of chemical mediators other than histamine in eosinophil infiltration in the nasal mucosa was studied using histamine H(1) receptor-deficient mice. Histamine H(1) receptor-deficient mice and wild-type controls were immunized with ovalbumin and consecutive topical antigen instillation was performed. Histological alterations and eosinophil infiltration into the nasal mucosa of mice were examined. Diffuse infiltration of inflammatory cells and edema after sensitization with antigen were observed in the nasal mucosa in both wild-type and histamine H(1) receptor-deficient mice. The number of eosinophils in the nasal mucosa in mice sensitized with antigen was significantly increased as compared with controls. The number of eosinophils in the nasal mucosa was significantly decreased by cetirizine and epinastine, ramatroban and zafirlukast in wild-type mice. Not only histamine but also thromboxane A(2) and leukotrienes play important roles in allergic rhinitis, especially in the late phase participating in nasal eosinophilia.

Adjuvants, Immunologic↗

PubMed2003

Appearance of the LAT protein at an early stage of B-cell development and its possible role.

The linker protein LAT is expressed mainly in T and natural killer (NK) cells. LAT-deficient mice have an arrest of intrathymic T-cell development at the CD4+ CD8+ stage and lack mature T cells in the periphery. However, no gross abnormality in development and function of the B and NK cells has been described. Here we report that LAT is expressed in mouse progenitor B (pro-B) and precursor B (pre-B) cells, but not in immature or mature B cells. LAT in pre-B cells becomes tyrosine phosphorylated upon cross-linking of the pre-B-cell receptor (pre-BCR) by anti- micro antibody. Incubation of 1xN/2b (mouse pre-B-cell line) cells or bone marrow cells from microMT/ microMT mice, which lack B cells after the small pre-B-cell stage, with anti-Ig beta antibody resulted in the downregulation of LAT expression. Transgenic mice which expressed LAT protein in B-lineage cells showed an increased proportion of pro- and large pre-B cells in the bone marrow and a remarkable reduction in the numbers of mature B cells in peripheral lymphoid tissues. Collectively, the present results indicate that LAT is expressed in the cells at the early stages of B-lineage development, but is absent in immature and mature B cells. LAT may play a crucial role in the negative regulation of B-cell development at the transition from pre-B to mature B-cell stages, and signal(s) via the pre-BCR may extinguish LAT expression, thus allowing pre-B-cell differentiation towards the mature B-cell stage.

Adaptor Proteins, Signal Transducing↗

PubMed2003

In vivo cooperation between Bcl-xL and the phosphoinositide 3-kinase-Akt signaling pathway for the protection of epidermal keratinocytes from apoptosis.

To investigate the function of Bcl-xL in the skin, we established keratinocyte-specific Bcl-x gene-targeted mice under the keratin 5 promoter (K5). K5.Bcl-xL-/- mice were viable, devoid of alteration in the development of skin or appendages. However, they harbored spontaneous apoptotic keratinocytes in the epidermis. Bcl-xL-deficient keratinocytes cultured in vitro readily underwent apoptosis in the absence of growth factors, but the addition of HGF or EGF resulted in restoration of cell survival, which was reversed by treatment with wortmannin, an inhibitor of phosphoinositide-3 kinase (PI3K). Topical treatment of K5.Bcl-xL-/- mice with wortmannin sensitized the skin for apoptosis induced by UV (UV) B, although wild-type epidermis was only marginally affected by this treatment, suggesting that the resistance to UVB largely depended on PI3K-Akt signaling in Bcl-xL-deficient mice but not in wild-type mice. Furthermore, UVB irradiation resulted in redistribution of phosphorylated Akt from the basal layer to the suprabasal layer, indicating that Akt could spatially cooperate with Bcl-xL upon UVB exposure in the upper epidermis where Bcl-xL is normally localized. These results suggest that Bcl-xL and the PI3K-Akt pathway form a cooperative, intercompensatory axis for the protection of epidermal keratinocytes from apoptosis in vivo.

Androstadienes↗

PubMed2003

Expression and diagnostic evaluation of the human tumor-associated antigen RCAS1 in pancreatic cancer.

INTRODUCTION: Receptor-binding cancer antigen expressed on SiSo cells (RCAS1) is one of the membrane molecules expressed on human cancer cells and is presumed to play a protective role for tumor cells against immune surveillance by inhibition of clonal expansion and induction of cell death in immunocytes. AIMS: To address whether RCAS1 is expressed in pancreatic cancer and whether serologic diagnostic evaluation is useful compared with that of carbohydrate antigen 19-9 (CA19-9) and soluble Fas ligand. METHODOLOGY: Immunohistochemical expression of RCAS1 was examined by staining with a 22-1-1 monoclonal antibody, and serum RCAS1 concentrations were determined by an enzyme-linked immunosorbent assay in 20 cases of ductal adenocarcinoma of the pancreas and other pancreatic diseases. RESULTS: Immunohistochemically, RCAS1 detection occurred in 100% (20/20) of ductal adenocarcinoma of the pancreas cases, 100% (6/6) of intraductal papillary-mucinous adenoma of the pancreas cases, and 40% (2/5) of chronic pancreatitis cases. RCAS1 was found in the cytoplasm of cancer cells and ductal cells. Serum RCAS1 concentrations in patients with ductal adenocarcinoma of the pancreas were significantly higher than those in patients with chronic pancreatitis (p < 0.0001), acute pancreatitis (p < 0.005), and autoimmune pancreatitis (p < 0.001). RCAS1 concentrations in patients with intraductal papillary-mucinous adenoma of the pancreas were also significantly higher than those in patients with chronic pancreatitis (p < 0.05) and autoimmune pancreatitis (p < 0.05). Positive serum RCAS1 samples (concentration, > or = 10 U/mL) were found most often in cases of pancreatic neoplasm (80% [16/20], ductal adenocarcinoma of the pancreas; and 60% [3/5], intraductal papillary-mucinous adenoma of the pancreas). By contrast, in cases of pancreatic inflammatory diseases, raised concentrations occurred in 9.4% (3/32) of chronic pancreatitis cases, none (0/6) of acute pancreatitis cases, and none (0/8) of autoimmune pancreatitis cases. The sensitivity of CA19-9 for ductal adenocarcinoma of the pancreas was 75% and the specificity was 73.1% compared with chronic pancreatitis. On the other hand, the sensitivity of RCAS1 for ductal adenocarcinoma of the pancreas was 80% and the specificity was 96.2% compared with chronic pancreatitis. The specificity of RCAS1 for chronic pancreatitis was higher than that of CA19-9. Serum soluble Fas ligand concentrations were not considerably different among these patients. CONCLUSIONS: RCAS1 was highly expressed in ductal adenocarcinoma of the pancreas, and serum RCAS1 concentrations in patients with ductal adenocarcinoma of the pancreas were significantly higher than those in patients with other inflammatory pancreatic diseases. Our results indicate that serum RCAS1 concentrations could be a new marker in screening procedures for pancreatic cancer.

Aged↗