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M Nakata

Publications and source records attributed to M Nakata.

At least 73 records · Page 4Linked to original sources
PubMed2000

Solution structures of the N-terminal domain of yeast calmodulin: Ca2+-dependent conformational change and its functional implication.

We have determined solution structures of the N-terminal half domain (N-domain) of yeast calmodulin (YCM0-N, residues 1-77) in the apo and Ca(2+)-saturated forms by NMR spectroscopy. The Ca(2+)-binding sites of YCM0-N consist of a pair of helix-loop-helix motifs (EF-hands), in which the loops are linked by a short beta-sheet. The binding of two Ca(2+) causes large rearrangement of the four alpha-helices and exposes the hydrophobic surface as observed for vertebrate calmodulin (CaM). Within the observed overall conformational similarity in the peptide backbone, several significant conformational differences were observed between the two proteins, which originated from the 38% disagreement in amino acid sequences. The beta-sheet in apo YCM0-N is strongly twisted compared with that in the N-domain of CaM, while it turns to the normal more stable conformation on Ca(2+) binding. YCM0-N shows higher cooperativity in Ca(2+) binding than the N-domain of CaM, and the observed conformational change of the beta-sheet is a possible cause of the highly cooperative Ca(2+) binding. The hydrophobic surface on Ca(2+)-saturated YCM0-N appears less flexible due to the replacements of Met51, Met71, and Val55 in the hydrophobic surface of CaM with Leu51, Leu71, and Ile55, which is thought to be one of reasons for the poor activation of target enzymes by yeast CaM.

Amino Acid Sequence↗
PubMed2000

Induced antiferroelectric smectic- C(*)(A) phase by doping ferroelectric- C* phase with bent-shaped molecules

Doping of the ferroelectric Sm-C(*) phase with bent-shaped molecules induces the antiferroelectric Sm-C(*)(A) phase. The effect was observed by means of electro-optic and dielectric measurements in systems with weak interlayer interactions in which the relative strength of anticlinic-synclinic order between molecules in adjacent layers is easily controlled by external factors. Fourier-transform infrared spectroscopy studies suggest that the bent-shaped molecules are not flat. They reorient upon the electric field-induced antiferroelectric-ferroelectric transition to adopt a position in which the average direction of the carbonyl groups is in the smectic plane and a bending tip along the C2 symmetry axis.

Journal Article↗
PubMed2000

Cloning, characterization and tissue distribution of the rat ATP-binding cassette (ABC) transporter ABC2/ABCA2.

The ABC1 (ABCA) subfamily of the ATP-binding cassette (ABC) transporter superfamily has a structural feature that distinguishes it from other ABC transporters. Here we report the cloning, molecular characterization and tissue distribution of ABC2/ABCA2, which belongs to the ABC1 subfamily. Rat ABC2 is a protein of 2434 amino acids that has 44.5%, 40.0% and 40.8% identity with mouse ABC1/ABCA1, human ABC3/ABCA3 and human ABCR/ABCA4 respectively. Immunoblot analysis showed that proteins of 260 and 250 kDa were detected in COS-1 cells transfected with ABC2 having a haemagglutinin tag, while no band was detected in mock-transfected cells. After incubation with N-glycosidase F, the mobilities of the two proteins increased and a single band was detected, suggesting that ABC2 is a glycoprotein. Photoaffinity labelling with 8-azido-[alpha-(32)P]ATP confirmed that ATP binds to the ABC2 protein in the presence of Mg(2+). RNA blot analysis showed that ABC2 mRNA is most abundant in rat brain. Examination of brain by in situ hybridization determined that ABC2 is expressed at high levels in the white matter, indicating that it is expressed in the oligodendrocytes. ABC2, therefore, is a glycosylated ABC transporter protein, and may play an especially important role in the brain. In addition, the N-terminal 60-amino-acid sequence of the human ABC1, which was missing from previous reports, has been determined.

ATP-Binding Cassette Transporters↗
PubMed2000

Measurement of glucose uptake and intracellular calcium concentration in single, living pancreatic beta-cells.

There has been no method previously to measure both glucose transport and its effect on the various intracellular functions in single, living mammalian cells. A fluorescent derivative of d-glucose, 2-[N-(7-nitrobenz-2-oxa-1, 3-diazol-4-yl)amino]-2-deoxy-d-glucose (2-NBDG), that we have developed has made such measurements possible. COS-1 cells that overexpress the human glucose transporter GLUT2 show significantly greater 2-NBDG uptake than mock transfected cells. Using GLUT2-abundant mouse insulin-secreting clonal MIN6 cells, we found that 2-NBDG was incorporated into the cells in a time- and concentration-dependent manner. The 2-NBDG uptake was inhibited by high concentrations of d-glucose in a dose-dependent manner and also was almost completely inhibited by 10 micrometer cytochalasin B. We then measured both glucose uptake and the intracellular calcium concentration ([Ca(2+)](i)) in single, living pancreatic islet cells. 2-NBDG and fura-2 were used as the tracer of glucose and indicator of intracellular calcium, respectively. All of the cells that showed an increase in [Ca(2+)](i) in response to a high concentration of glucose (16.8 mm) rapidly incorporated significant 2-NBDG. Immunocytochemical examination confirmed these cells to be insulin-positive beta-cells. All of the cells that showed no significant, rapid 2-NBDG uptake lacked such glucose responsiveness of [Ca(2+)](i), indicating that these cells were non-beta-cells such as glucagon-positive alpha-cells. These results show the uptake of glucose causing a concomitant increase of [Ca(2+)](i) in beta-cells. Because 2-NBDG is incorporated into mammalian cells through glucose transporters, it should be useful for the measurement of glucose uptake together with concomitant intracellular activities in many types of single, living mammalian cells.

Animals↗
PubMed2000

Anandamide induces apoptosis of PC-12 cells: involvement of superoxide and caspase-3.

Anandamide (arachidonoylethanolamide), an endogenous cannabinoid receptor ligand has been suggested to have physiological role in mammalian nervous system. However, little is known about the role of anandamide on neuronal cells. Here, we demonstrate that anandamide causes death of PC-12 cells, showing marked DNA condensation and fragmentation, appearance of cells at sub-G(0)/G(1) and redistribution of phosphatidyl serine, the hallmark features of apoptosis. Anandamide raised intracellular superoxide level and CPP32-like protease activity in PC-12 cells markedly. Furthermore, antioxidant N-acetyl cysteine prevented anandamide-induced superoxide anion formation and cell death, implying that intracellular superoxide is a novel mediator of anandamide-induced apoptosis of PC-12 cells.

Acetylcysteine↗
PubMed2000

Polymyxin B binds to anandamide and inhibits its cytotoxic effect.

Anandamide (ANA), an endogenous cannabinoid, can be generated by activated macrophages during endotoxin shock and is thought to be a paracrine contributor to hypotension. We discovered that ANA in saline/ethanol solution and in serum was efficiently adsorbed in a polymyxin B (PMB)-immobilized beads column and eluted with ethanol. We confirmed the direct binding of PMB to ANA by using surface plasmon resonance. The adsorption of ANA by PMB may abolish the diverse effects of ANA such as hypotension, immunosuppression, and cytotoxicity, and may suggest a new therapeutic strategy for endotoxin shock.

Adsorption↗
PubMed2000

Pulmonary function after lobectomy: video-assisted thoracic surgery versus thoracotomy.

BACKGROUND: Whether video-assisted thoracic surgery (VATS) improves postoperative pulmonary function is still controversial. We compared postoperative pulmonary function after VATS lobectomy and standard lobectomy. METHODS: Eleven patients who had undergone standard lobectomy and 10 patients who had undergone VATS lobectomy were studied. Arterial blood gas analyses were performed on the 4th, 7th, and 14th postoperative days. Pulmonary function, including forced vital capacity (FVC), forced expiratory volume in 1 second (FEV1.0), and peak flow rate (PFR) were measured on the 7th and 14th postoperative days (early phase), and approximately 1 year after surgery (late phase). RESULTS: Pulmonary function, as assessed with arterial oxygen partial pressure (PaO2) (p = 0.054), arterial oxygen saturation (O2SAT) (p = 0.063), FVC (p = 0.10), and FEV1.0 (p = 0.08), was better after VATS lobectomy than after thoracotomy on the 7th postoperative day. PFR was significantly better after VATS on both the 7th and 14th postoperative days (p = 0.008 and p = 0.03, respectively). CONCLUSIONS: VATS lobectomy had advantages on early postoperative pulmonary function. We conclude that VATS lobectomy is a beneficial alternative to standard thoracotomy, especially for patients with poor pulmonary reserve.

Aged↗
PubMed2000

Induction and maintenance of immune effector cells in the gastric tissue of mice orally immunized to Helicobacter pylori requires salivary glands.

BACKGROUND & AIMS: Helicobactor pylori mostly colonizes the gastric mucus that contains salivary antibodies. We studied the role of saliva in the induction and maintenance of gastric immunity conferred by oral vaccination against H. pylori. METHODS: C57BL/6 mice underwent a sialoadenectomy before and after intragastric immunization using whole-cell sonicates of H. pylori and cholera toxin as an adjuvant. At 1 and 6 months after oral inoculation, we assessed the density of the H. pylori colonizing the stomach, specific antibodies in gastric secretion and sera, and the constituents of cellular infiltrates in the tissue. RESULTS: A sialoadenectomy before, but not after, immunization abrogated protection by the vaccination at 1 month after inoculation. Protected mice had more neutrophils, plasma cells, and lymphocytes, but fewer eosinophils, in the gastric tissue than nonprotected mice. Protected mice had a greater increase of immunoglobulin (Ig) G1 specific to H. pylori than IgG2a in sera. At 6 months after inoculation, oral immunization was less effective in mice who had a sialoadenectomy than in control immunized mice. The antibody titers in both gastric secretion and in sera did not correlate with the density of bacteria colonizing the stomach. CONCLUSIONS: It is suggested that, in intragastric immunization against H. pylori, saliva is necessary for both the induction and maintenance of optimal immunity in the stomach. Effective immunity was associated with an increased number of neutrophils and lymphocytes in gastric tissue.

Administration, Oral↗
PubMed2000

Embolotherapy for venous impotence: use of ethanol.

PURPOSE: To evaluate the usefulness of embolotherapy with ethanol for the treatment of venous impotence. MATERIALS AND METHODS: Twenty-three patients with venous impotence underwent embolotherapy. The diagnosis of venous impotence was made by means of pharmacocavernosometry and cavernosography. After exposure of the deep dorsal penile vein, a intravenous catheter was inserted directly into the deep dorsal penile vein and advanced into just front of the preprostatic plexus. Fifty percent ethanol was injected through the catheter and the endpoint of the procedure was determined based on results of venography immediately after injection. The procedure was finished when lack of venous leakage was confirmed. RESULTS: In all patients, the deep dorsal penile vein was successfully exposed surgically, the sclerosing agent successfully injected, and the endpoint successfully achieved. Immediate clinical therapeutic effect (restoration of erection) was obtained in 20 cases (87%). No severe complications were observed during or after the procedure. The follow-up period was 6-50 months. Long-term therapeutic effect was confirmed for 18 of 23 patients (78%). CONCLUSION: The authors' findings suggest that this treatment had satisfactory short-term and long-term clinical results and that longer follow-up is necessary to confirm its safety.

Adult↗
PubMed2000

Difference in tracks between habitual open and close mandibular movements at the condyle in children.

Although previous studies have paid much attention to the condylar movement in adults with permanent dentition, little attention has been paid to such movement in children. In this study, we therefore clarified the difference in habitual open and close movements at the condyle in children. Three groups of subjects were used; primary (10 children), early mixed (10 children), and permanent dentition (10 adults). The habitual open and close mandibular movement of each subject was measured using a TRIMET, which can three-dimensionally analyse the simultaneous movements of multiple points on the mandible of a subject. The measurements were then compared among the three groups. The three-dimensional analysis detected significant difference in all directions between children with primary dentition and adults with permanent dentition: primary dentition had the smallest anterior-posterior and superior-inferior directions, and the largest left-right direction. Coincidence of the open and close tracks occurred in the adults (adult group) but not in the children (primary and early-mixed dentition groups). The early-mixed dentition group showed tracks that were between those for the primary dentition group and the adult group. These results suggest that the regularity of the condylar track might be well established with dental development.

Adult↗
PubMed2000

Development of gastric emptying in the human fetus.

OBJECTIVE: To evaluate stomach size and the development of gastric emptying in human fetuses using ultrasound. DESIGN: Clinical observational study. METHODS: The motility and peristalsis of the fetal stomach were studied in 80 normal fetuses between 12 and 39 weeks of gestation. Fetal gastric motility was assessed by analysis of videotaped recordings of ultrasound images of the stomach taken in real time. RESULTS: Fetal maximum gastric area gradually increased and minimum gastric areas gradually decreased after 20 weeks of gestation. At term, the maximum and minimum gastric area ratios were approximately 13 and 5%, respectively. The change in fetal gastric area, defined as the difference of maximum and minimum gastric area ratios, was relatively constant at 3% from 12 to 15 weeks of gestation to 20-23 weeks of gestation. It increased significantly (to 8%) after 24-27 weeks of gestation until term. CONCLUSIONS: Fetal gastric emptying was quantified and its development assessed during pregnancy. A critical point of gastric development, associated with an increase in the change of gastric volume, was identified at 24-25 weeks of gestation.

Fetus↗
PubMed2000

Decreased superoxide dismutase expression and increased concentrations of lipid peroxide and prostaglandin F(2alpha) in the decidua of failed pregnancy.

To study the possible role of the superoxide radical and its scavenging system in the decidua of early pregnancy, superoxide dismutase (SOD) values and concentrations of lipid peroxide and prostaglandin F(2alpha) (PGF(2alpha)) were analysed in the decidua obtained from normal pregnancy and failed pregnancy. Failed pregnancy was divided into two groups; spontaneous abortion with or without vaginal bleeding. In the spontaneous abortion with vaginal bleeding, total SOD activities, Cu,Zn-SOD activities and Cu,Zn-SOD mRNA values in the decidua were significantly lower, and concentrations of lipid peroxide and PGF(2alpha) were significantly higher, than those in the normal pregnancy and the spontaneous abortion without vaginal bleeding. In contrast, activities and mRNA values of Mn-SOD were significantly higher in the spontaneous abortion with vaginal bleeding than the other two groups. There was no significant difference in all of these parameters between the normal pregnancy and the spontaneous abortion without vaginal bleeding. In conclusion, the decrease in Cu,Zn-SOD expression and the increase in lipid peroxide in the decidua could be involved in the termination of spontaneous abortion, mediated through the increase in PGF(2alpha) synthesis. In other words, Cu,Zn-SOD may contribute to the maintenance of pregnancy by preventing the accumulation of superoxide radicals that cause PGF(2alpha) synthesis.

Abortion, Spontaneous↗
PubMed2000

NMR studies of model peptides of PHGGGWGQ repeats within the N-terminus of prion proteins: a loop conformation with histidine and tryptophan in close proximity.

The N-terminal region of the prion protein from human and mouse contains five tandem repeats with the consensus sequence of PHGGGWGQ. NMR studies were performed in water for two cyclic peptides, cyclo-[C(1)R(2)Q(3)P(4)H(5)G(6)G(7)S(8)W(9)G(10)Q(11)R(12)D(13)C(14)] (C1) and cyclo-[C(1)R(2)D(3)P(4)H(5)G(6)G(7)G(8)W(9)G(10)Q(11)P(12)H(13)G(14)G (15)G(16)W(17)G(18)Q(19)R(20)D(21)C(22)] (C2), which are cyclized by a disulfide bridge between the Cys residues at the N- and C-termini, and for their corresponding linear peptides (L1 and L2) which are formed by reduction. The patterns of the C(alpha)H chemical shift difference of these four peptide mimetics were very similar to those observed for the tandem repeats of human prion protein reported by other researchers. The medium-range NOE connectivities were found between the C(beta)H of the H5 and the proton of the W9 side chain for L1. The corresponding NOEs were also observed in H5-W9 and H13-W17 of L2 with ambiguity. These observations indicate that histidine (i) is in close proximity to tryptophan (i+4). d(alphaN) (i,i+2) NOE connectivities were observed between W9 and Q11 of L1 and L2, and d(NN) (i,i+1) NOE connectivities were also observed for G10-Q11 of L1 and L2 and for G18-Q19 of L2. Significantly lower temperature coefficients of amide proton chemical shifts were obtained for Q11 and Q19 of L2 and C2. Structure calculations for L1 showed that HGG(G/S)W and (G/S)WGQ adopt a loop conformation and a beta-turn, respectively. These results strongly suggest that the tandem repeats within prion protein adopt a non-random structure.

Amino Acid Sequence↗
PubMed2000

Ferroelectric phases in a chiral bent-core smectic liquid crystal: dielectric and optical second-harmonic generation measurements

We report unambiguous evidence for ferroelectricity in a chiral bent-core liquid crystal. This evidence includes (i) a single current peak in the polarization reversal current, (ii) a strong dielectric response that is suppressed by applying a bias electric field, and (iii) strong optical second-harmonic generation in the absence of electric field even at normal incidence. We propose a phase structure that is anticlinic in tilt and ferroelectric in polar order, i.e., smectic-C(A)P(*)(F).

Journal Article↗
PubMed2000

Effect of leptin in platelet and endothelial cells. Obesity and arterial thrombosis.

We demonstrated that leptin showed effects on both platelets and endothelial cells through its functional receptor. These effects are the vector to inducing thrombotic tendency. Leptin concentrations used in our experiments correspond to that of leptin in the circulation of obese individuals. Thus we suggest that increased leptin may act as a risk factor for thrombosis in obese individuals.

Adenosine Diphosphate↗
PubMed2000

Insulinotropin PACAP potentiates insulin action. Stimulation of glucose uptake in 3T3-LI adipocytes.

PAC1 receptor was expressed in the rat fat tissue and 3T3-L1 adipocytes. PACAP-38 (10 nM) significantly enhanced insulin induced 2-deoxyglucose uptake by 3T3-L1 adipocytes. GLP-1 has a similar effect. PACAP-38 further increased insulin stimulated phosphatidylinositol (PI) 3-kinase activity, but has not effect on tyrosine phosphorylation of insulin receptor beta-subunit or IRS-1. These results reveal that PACAP-38 enhances insulin induced glucose uptake, an effect probably mediated by PI 3-kinase. In conclusion, PACAP potentiates not only insulin secretion but also insulin action in adipocytes, thereby exhibiting antidiabetic actions at two important steps of glucose metabolism (Fig. 2).

3T3 Cells↗
PubMed2000

[A case report of pneumococcal pneumonia diagnosed by urinary antigen].

Using a kit for the rapid detection of Streptococcus pneumoniae, we examined 3 clinical cases, clearly diagnosed as pneumococcal pneumonia. Case 1 was a 63-year-old man hospitalized for right middle lobular pneumonia. Streptococcus pneumoniae was detected by blood culture initiated on the day of admission. His urine sample was found to have Streptococcus pneumoniae antigen at hospital day 4, and positive test results were observed 3 times thereafter. The other 2 cases had negative sputum and blood cultures, but they were positive for urine antigen, with clinical courses consistent with those of pneumococcal pneumonia. The kit used was determined to provide a test result within 15 min for each urine sample, and it was easy to perform. Thus, this kit is expected to serve as a very useful clinical tool.

Antigens, Bacterial↗