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S Kobayashi

Publications and source records attributed to S Kobayashi.

At least 163 records · Page 9Linked to original sources
PubMed2001

A retrotransposon-derived gene, PEG10, is a novel imprinted gene located on human chromosome 7q21.

A novel paternally expressed imprinted gene, PEG10 (Paternally Expressed 10), was identified on human chromosome 7q21. PEG10 is located near the SGCE (Sarcoglycan epsilon) gene, whose mouse homologue was recently shown to be imprinted. Therefore, it is highly possible that a new imprinted gene cluster exists on human chromosome 7q21. Analysis of two predicted open reading frames (ORF1 and ORF2) revealed that ORF1 and ORF2 have homology to the gag and pol proteins of some vertebrate retrotransposons, respectively. These data suggest that PEG10 is derived from a retrotransposon that was previously integrated into the mammalian genome. PEG10 is likely to be essential for understanding how exogenous genes become imprinted.

Amino Acid Sequence↗
PubMed2001

Coexpression of microsomal-type prostaglandin E synthase with cyclooxygenase-2 in brain endothelial cells of rats during endotoxin-induced fever.

Fever is triggered by an elevation of prostaglandin E(2) (PGE(2)) in the brain. However, the mechanism of its elevation remains unanswered. We herein cloned the rat glutathione-dependent microsomal prostaglandin E synthase (mPGES), the terminal enzyme for PGE(2) biosynthesis, and examined its induction in the rat brain after intraperitoneal injection of pyrogen lipopolysaccharide (LPS). In Northern blot analysis, mPGES mRNA was weakly expressed in the brain under the normal conditions but was markedly induced between 2 and 4 hr after the LPS injection. In situ hybridization study revealed that LPS-induced mPGES mRNA signals were mainly associated with brain blood vessels, especially vein or venular-type ones, in the whole brain area. Immunohistochemical study demonstrated that mPGES-like immunoreactivity was expressed in the perinuclear region of brain endothelial cells, which were identified as von Willebrand factor-positive cells. Furthermore, in the perinuclear region of the endothelial cells, mPGES was colocalized with cyclooxygenase-2 (COX-2), which is the enzyme essential for the production of the mPGES substrate PGH(2). Inhibition of cyclooxygenase-2 activity resulted in suppression of both PGE(2) level in the CSF and fever (Cao et al., 1997), suggesting that the two enzymes were functionally linked and that this link is essential for fever. These results demonstrate that brain endothelial cells play an essential role in the PGE(2) production during fever by expressing COX-2 and mPGES.

Animals↗
PubMed2001

Molecular cloning of rat transmembrane domain protein of 40 kDa regulated in adipocytes and its expression in H9c2 cells exposed to ischemic hypoxia and reoxygenation.

We clone a 1230 bp complementary DNA encoding rat transmembrane domain protein of 40 kDa regulated in adipocytes (TPRA40), an orphan receptor, by reverse transcription-polymerase chain reaction using H9c2 cells derived from embryonic rat heart. The deduced amino acid sequence of rat TPRA40 consists of 369 amino acids and has a longer carboxyl terminus than that of the mouse protein. The level of TPRA40 mRNA decreases significantly throughout ischemic hypoxia and reoxygenation.

Adipocytes↗
PubMed2001

Up-regulation of inositol 1,4,5 trisphosphate receptor expression in atrial tissue in patients with chronic atrial fibrillation.

OBJECTIVES: We examined whether patients with atrial fibrillation (AF) have alterations in atrial inositol 1,4,5 trisphosphate receptors (IP3 receptors). BACKGROUND: Abnormal intracellular Ca2+ homeostasis occurs in chronic AF. The intracellular Ca2+ concentration is regulated by ryanodine and IP3 receptors. We recently reported alterations in ryanodine receptors in atrial tissue from patients in chronic AF. METHODS: We analyzed IP3 receptor expression in the right atrial myocardium from 13 patients with mitral valvular disease (MVD) with AF (MVD/AF), five patients with MVD who had normal sinus rhythm (MVD/NSR) and eight control patients with NSR (tissue obtained during coronary artery bypass surgery). Hemodynamic and echocardiographic data were obtained preoperatively, and an immunohistochemical study was performed on atrial tissue. RESULTS: The relative expression level of IP3 receptor protein was significantly greater in MVD/AF (0.75 +/- 0.26) than it was in MVD/NSR (0.42 +/- 0.13, p < 0.01), and both were significantly above control (0.14 +/- 0.08). The relative expression level of IP3 receptor messenger RNA was significantly greater in the MVD/AF group (0.028 +/- 0.008) than it was in the control group (0.015 +/- 0.004, p < 0.01), but patients with MVD/AF did not differ from patients with MVD/NSR (0.020 +/- 0.006). The relative expression levels of IP3 receptor protein and messenger RNA were higher in patients with left atrial dimension > or = 40 mm, pulmonary capillary wedge pressure > or = 10 mm Hg and right atrial pressure > or = 5 mm Hg. Inositol 1,4,5 trisphosphate receptors were over-expressed in the cytosol and at the nuclear envelope of atrial myocytes in MVD. CONCLUSIONS: Since chronic mechanical overload of the atrial myocardium increased IP3 receptor expression, especially in patients with chronic AF, up-regulation of IP3 receptors may be important in modulating intracellular Ca2+ homeostasis and initiating or perpetuating AF.

Aged↗
PubMed2001

Clinical value of the wild-type estrogen receptor beta expression in breast cancer.

To estimate the clinical value of estrogen receptor (ER) beta expression in breast cancer we used an immunohistochemical method to detect the wild-type ERbeta in 88 primary breast cancers. We used a highly specific polyclonal antibody to the carboxyl terminus of wild-type ERbeta. This antibody reacted with neither other variant forms of ERbeta nor any part of ERalpha. Slides were evaluated on a scale representing the estimated proportion and intensity of positive-staining tumor cells. Positive staining could be seen in 52 (59.1%) of 88 breast cancers; 36 (40.9%) were negative. Although there was no correlation between ERbeta staining and age, node status, tumor size, histological grade, or progesterone receptor (PgR)-enzyme immunoassay (EIA) status, we did observe a significant correlation with ERalpha-EIA (Fisher's exact probability test: P=0.0169). Moreover, ERbeta positive cases showed a better prognosis than negative cases in disease-free survival rate (Logrank test: P=0.0662, Breslow-Gehan-Wilcoxson test: P=0.0318). Our data demonstrated the possibility that wild-type ERbeta protein expression could be used as a good prognostic indicator for breast cancer.

Adenocarcinoma, Mucinous↗
PubMed2001

Total synthesis and structural elucidation of khafrefungin.

Total synthesis and structural elucidation of khafrefungin, a novel antifungal agent isolated from the fermentation culture MF6020, have been achieved. Unlike other inhibitors that inhibit the corresponding enzyme in fungi and mammals to the same extent, khafrefungin does not impair sphingolipid synthesis of mammals. The basic strategy for the structural elucidation is to prepare all stereoisomers of the structurally simplified khafrefungin mimics 1 and 2 that were designed for the elucidation of C10,11,12 and C2',3',4' relative stereochemistry, respectively. The comparison of their spectra with those of natural khafrefungin would result in the identification of eight possible stereoisomers, and the analytical details of these eight stereoisomers have led to the complete stereochemical assignment. On the basis of the structural elucidation, the total synthesis of khafrefungin has been accomplished by using tin(II)-catalyzed asymmetric aldol reactions as key steps.

Antifungal Agents↗
PubMed2001

Membrane recruitment of DOCK180 by binding to PtdIns(3,4,5)P3.

DOCK180 was originally identified as one of two major proteins bound to the Crk oncogene product and became an archetype of the CDM family of proteins, including Ced-5 of Caenorhabditis elegans and Mbc of Drosophila melanogaster. Further study has suggested that DOCK180 is involved in the activation of Rac by the CrkII-p130(Cas) complex. With the use of deletion mutants of DOCK180, we found that the C-terminal region containing a cluster of basic amino acids was required for binding to and activation of Rac. This region showed high amino-acid sequence similarity to the consensus sequence of the phosphoinositide-binding site; this led us to examine whether this basic region binds to phosphoinositides. For this purpose we used PtdIns(3,4,5)P(3)-APB beads, as reported previously [Shirai, Tanaka, Terada, Sawada, Shirai, Hashimoto, Nagata, Iwamatsu, Okawa, Li et al. (1998) Biochim. Biophys. Acta 1402, 292-302]. By using various competitors, we demonstrated the specific binding of DOCK180 to PtdIns(3,4,5)P(3). The expression of active phosphoinositide 3-kinase (PI-3K) did not enhance a DOCK180-induced increase in GTP-Rac; however, the expression of PI-3K translocated DOCK180 to the plasma membrane. Thus DOCK180 contained a phosphoinositide-binding domain, as did the other guanine nucleotide exchange factors with a Dbl homology domain, and was translocated to the plasma membrane on the activation of PI-3K.

Cell Line↗
PubMed2001

Efficient synthesis of piperidine derivatives. Development of metal triflate-catalyzed diastereoselective nucleophilic substitution reactions of 2-methoxy- and 2-acyloxypiperidines.

Nucleophilic substitution reactions of 2-methoxy- and 2-acyloxypiperidines were investigated. First, new and efficient methods for the preparation of the starting piperidine derivatives were developed. N-Benzyloxycarbonyl-2-methoxypiperidine (3) and 3-substituted-2-acyloxy-N-benzyloxycarbonylpiperidines (4a-d), which are recognized as the simplest imino-sugars, were prepared and were examined as substrates for nucleophilic substitution reactions with silyl enolates under the influence of catalytic amounts of metal triflates (Sc(OTf)3, Sn(OTf)2, Cu(OTf)2, Hf(OTf)4, etc). Among the triflates tested, Sc(OTf)3 gave the best results. It was found that 2-acetoxy-3-benzyloxy-N-benzyloxycarbonylpiperidine (4a) reacted with silyl enolates to afford the 2-alkylated adducts in high cis-selectivity, while 2,3-diacyloxy-N-benzyloxycarbonylpiperidines (4b-d) showed transselectivity. The stereochemical assignments were carefully performed using NMR analysis, X-ray crystallography, and synthetic transformations. Febrifugine (1), a potent antimalarial alkaloid, was successfully synthesized from 2,3-diacetoxy-N-benzyloxycarbonylpiperidine (4b) on the basis of these diastereoselective nucleophilic substitution reactions.

Journal Article↗
PubMed2001

Lewis acid-catalyzed ring-opening reactions of semicyclic N,O-acetals.

[figure: see text] Ring-opening reactions of semicyclic N,O-acetals with various nucleophiles such as silyl enol ethers are effectively catalyzed by a Lewis acid (TMSOTf). Reactions of 3-substituted N,O-acetals showed high diastereoselectivities. Synthetic utility of this method has been demonstrated in the stereoselective synthesis of an anti-malarial agent, isofebrifugine.

Journal Article↗
PubMed2001

Comparison of UDP-glucose:flavonoid 3-O-glucosyltransferase (UFGT) gene sequences between white grapes (Vitis vinifera) and their sports with red skin.

The expression of the UDP-glucose:flavonoid 3-O-glucosyltransferase (UFGT) gene has been shown to be critical for anthocyanin biosynthesis in the grape berry. Using white cultivars and bud sports with red skin, we examined the expression of seven anthocyanin biosynthetic genes including the UFGT gene and compared the coding/promoter sequences of the UFGT gene. Northern blot analysis showed that the seven anthocyanin biosynthetic genes were expressed coordinately at higher levels in the red-skin sports than in the white-skin progenitors of the sports. It was especially notable that UFGT gene expression was detected only in the red-skin sports and Kyoho. However, there were no differences in either coding or promoter sequences between Italia (Vitis vinifera) and its red-skin sport Ruby Okuyama, or between Muscat of Alexandria (V. vinifera) and the red-skin sport Flame Muscat. From these findings, the phenotypic change from white to red in the sports is thought to be the result of a mutation in a regulatory gene controlling the expression of UFGT.

Journal Article↗
PubMed2001

Enhancement of Rho/Rho-kinase system in regulation of vascular smooth muscle contraction in tachycardia-induced heart failure.

OBJECTIVE: The Rho/Rho-kinase system regulates Ca(2+) sensitivity in vascular smooth muscle. A new drug, Y-27632, specifically inhibits Rho-kinase and hence decreases the phosphorylation of myosin light chain, thus reducing contraction. Here, we compare the effects of Y-27632 and nifedipine on the vasoconstrictor response of the femoral artery in heart failure. METHODS: Heart failure (HF) was produced by chronic rapid RV pacing (250 bpm, 28 days, six dogs). Indo1-AM was loaded into endothelium-denuded femoral artery segments for measuring intracellular [Ca(2+)]. Tension and changes in intracellular [Ca(2+)] [the change in the ratio (418 nm/468 nm) of Indo1 fluorescence (F(ratio))] were simultaneously measured in Krebs-Ringer solution. RESULTS: In HF: (i) norepinephrine (10 microM) produced greater tension (784+/-52 g/cm(2)) than in control (502+/-64 g/cm(2)) despite a similar increase in F(ratio), indicating increased Ca(2+) sensitivity in vascular smooth muscle; (ii) nifedipine attenuated this enhanced response by only a maximum of 27% at 1 micromol/l with a 56% reduction in F(ratio); (iii) Y-27632 attenuated it by a maximum of 80% at 100 micromol/l without a significant change in F(ratio); (iv) RhoA protein and mRNA expression levels in the femoral artery were up-regulated by +110% and +56%, respectively, while those of Rho-kinase were unchanged. CONCLUSIONS: The Ca(2+)-sensitizing mechanism involving the Rho/Rho-kinase system may be deeply involved in the enhanced arterial vasoconstriction seen in HF. Since Y-27632 attenuated this response in small arteries, it shows potential as a novel, potent vasodilator for the treatment of HF.

Amides↗
PubMed2001

Nicotinamide-N-methyltransferase is higher in the lumbar cerebrospinal fluid of patients with Parkinson's disease.

Parkinson's disease (PD) may be initiated or precipitated by endogenous toxins with a structure similar to 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine in genetically-predisposed individuals. Nicotinamide N-methyltransferase (NNMT) catalyzes N-methylation of nicotinamide and other pyridines to form pyridinium ions. The protein amount of NNMT was measured in the lumbar cerebrospinal fluid of PD patients by immunoblot analysis using anti-human NNMT antibody. In younger (65 years old or younger) PD patients, the relative level of NNMT protein was significantly higher than that in younger controls. The NNMT protein was significantly affected by aging: the amount decreased along with aging in PD patients. These findings suggested that excess NNMT in the central nervous system might be implicated in the PD pathogenesis.

Aging↗
PubMed2001

2-Alkynyl-8-aryl-9-methyladenines as novel adenosine receptor antagonists: their synthesis and structure-activity relationships toward hepatic glucose production induced via agonism of the A(2B) receptor.

Novel adenosine antagonists, 2-alkynyl-8-aryl-9-methyladenine derivatives, were synthesized as candidate hypoglycemic agents. These analogues were evaluated for inhibitory activity on N-ethylcarboxamidoadenosine (NECA)-induced glucose production in primary cultured rat hepatocytes. In general, aromatic moieties at the 8-position and alkynyl groups at the 2-position had significantly increased activity compared to unsubstituted compounds. The preferred substituents at the 8-position of adenine were the 2-furyl and 3-fluorophenyl groups. In modifying the alkynyl side chain, change of the ring size, cleavage of the ring, and removal of the hydroxyl group were well tolerated. The order of the stimulatory effects of adenosine agonists on rat hepatocytes was NECA > CPA > CGS21680, which is consistent with involvement of the A(2B) receptor. In Chinese hamster ovary cells stably transfected with human A(2B) receptor cDNA, one of the compounds potent in hepatocytes, 15o (IC(50) = 0.42 microM), antagonized NECA-induced stimulation of cyclic AMP production (IC(50) = 0.063 microM). This inhibitory effect was much more potent than those of FK453, KF17837, and L249313 which have been reported to be respectively A(1), A(2A), and A(3) selective antagonists. These findings agree very well with the result that, compared to 15o, these selective antagonists for each receptor subtype showed only marginal effects in rat hepatocytes. These results suggest that adenosine agonist-induced glucose production in rat hepatocytes is mediated through the A(2B) receptor. Furthermore, 15o showed hypoglycemic activity in an animal model of noninsulin-dependent diabetes mellitus, the KK-A(y) mice. It is possible that inhibition of hepatic glucose production via the A(2B) receptor could be at least one of the mechanisms by which 15o exerts its in vivo effects. Further elaboration of this group of compounds may afford novel antidiabetic agents.

Adenine↗
PubMed2001

Eicosapentaenoic acid (EPA) induces Ca(2+)-independent activation and translocation of endothelial nitric oxide synthase and endothelium-dependent vasorelaxation.

Eicosapentaenoic acid (EPA), but not its metabolites (docosapentaenoic acid and docosahexaenoic acid), stimulated nitric oxide (NO) production in endothelial cells in situ and induced endothelium-dependent relaxation of bovine coronary arteries precontracted with U46619. EPA induced a greater production of NO, but a much smaller and more transient elevation of intracellular Ca(2+) concentration ([Ca(2+)]i), than did a Ca(2+) ionophore (ionomycin). EPA stimulated NO production even in endothelial cells in situ loaded with a cytosolic Ca(2+) chelator 1,2-bis-o-aminophenoxythamine-N',N',N'-tetraacetic acid, which abolished the [Ca(2+)]i elevations induced by ATP and EPA. The EPA-induced vasorelaxation was inhibited by N(omega)-nitro-L-arginine methyl ester. Immunostaining analysis of endothelial NO synthase (eNOS) and caveolin-1 in cultured endothelial cells revealed eNOS to be colocalized with caveolin in the cell membrane at a resting state, while EPA stimulated the translocation of eNOS to the cytosol and its dissociation from caveolin, to an extent comparable to that of the eNOS translocation induced by a [Ca(2+)]i-elevating agonist (10 microM bradykinin). Thus, EPA induces Ca(2+)-independent activation and translocation of eNOS and endothelium-dependent vasorelaxation.

Animals↗