Biomedical subjects

N Aoki

Publications and source records attributed to N Aoki.

At least 91 records · Page 5Linked to original sources

PubMed1999

Characterization of newly identified four isoforms for a putative cytosolic protein tyrosine phosphatase PTP36.

In the course of determining the expression profiles of protein tyrosine phosphatases in lactating mammary gland, we found the expression of an isoform for a putative cytosolic and cytoskeleton-associated protein tyrosine phosphatase PTP36. Further detailed RT-PCR and Northern blot analyses revealed the expression of several isoforms for PTP36 in a tissue-dependent manner. We have cloned the cDNAs encoding four truncated isoforms for PTP36 and designated PTP36-A, -B, -C, and -D, respectively. PTP36-A and -C had new sequences generated due to frameshift, whereas PTP36-B and -D were in-frame variants. Gly- and Glu-rich domains and a putative PTP domain were missing from PTP36-A, but the band 4.1 domain remained. PTP36-B retained the band 4.1 and PTP domains but lacked Pro-, Gly- and Glu-rich domains. Most domain structures were lacking in PTP36-C and -D. Interestingly, PTP36-C contained an incomplete band 4.1 domain, but the newly created sequence exhibited high homology to human nebulette, which was also suggested to associate with cytoskeletons. When transiently expressed in COS7 and HEK293 cells, not only the wild type but also all the isoforms were recovered in Triton X-100-insoluble cytoskeleton-associated fractions and this distribution was not affected by mechanical cell detachment and treatment with a kinase inhibitor staurosporine. Such cellular distribution of PTP36 was also observed in stable COS7 clones. Further studies using deletion mutants suggested that the first 30 amino acids as well as the band 4.1 domain of PTP36 were involved in association with Triton X-100 insoluble cytoskeletons. Tissue-dependent expression and deletion in domain structures might reflect the biological significance of the isoforms for PTP36 in certain physiological conditions.

Amino Acid Sequence↗

PubMed1999

Comparison of protective effects of preinfarction angina pectoris in acute myocardial infarction treated by thrombolysis versus by primary coronary angioplasty with stenting.

The protective effects of preinfarction angina were evaluated in acute myocardial infarction (AMI) treated by primary percutaneous transluminal coronary angioplasty (PTCA) and stenting. We studied 613 patients with AMI. Group 1 (n = 306) was treated by conventional medical therapies and coronary thrombolysis and group 2 (n = 307) was treated by primary PTCA supported by stenting. Each group was subdivided into those with and without preinfarction angina within 24 hours before the onset of AMI. There was no significant difference in clinical characteristics between the subgroups of groups 1 and 2. In group 1, there were differences between patients with preinfarction angina (n = 84) and those without (n = 222) in in-hospital mortality (11% vs 18%), pump failure (Killip classes 3 and 4) (11% vs 21%, p <0.05), left ventricular ejection fraction at discharge (52 +/- 13% vs 48 +/- 14%, p <0.05), and peak creatine kinase (2,106 +/- 1,637 vs 2,764 +/- 2,154 U/L, p <0.02). In group 2, however, there was no significant difference between those with preinfarction angina (n = 82) and those without (n = 225) in mortality (6% vs 6%), pump failure (12% vs 12%), left ventricular ejection fraction (50 +/- 13% vs 50 +/- 13%) and peak creatine kinase (3,285 +/- 2,306 vs 3,291 +/- 2,262 U/L). Multivariate analysis indicated that preinfarction angina was an independent determinant of in-hospital death and pump failure in group 1, but not in group 2. We conclude that the protective effects of preinfarction angina in AMI are not evident in those treated by primary PTCA and stenting, possibly because of the overwhelming protective effects of complete coronary revascularization provided by primary PTCA and stenting.

Aged↗

PubMed1999

Electron transfer from quinohemoprotein alcohol dehydrogenase to blue copper protein azurin in the alcohol oxidase respiratory chain of Pseudomonas putida HK5.

A blue copper protein was purified together with a type II quinohemoprotein alcohol dehydrogenase (ADH IIB) from the soluble fraction of Pseudomonas putida HK5 grown on n-butanol. The purified blue copper protein was shown to be azurin, on the basis of several properties such as its absorption maximum (623 nm), its low molecular mass (17 500 Da), its acidic nature (pI of 4.1), its relatively high redox potential (306 mV), the presence of an intramolecular disulfide bond, and N-terminal amino acid sequence homology with respect to azurins from other sources, especially from P. putida NCIB 9869 and Pseudomonas fluorescens. Direct electron transfer from ADH IIB to azurin was shown to occur at a rate of 48-70 s-1. The apparent Km value of ADH IIB for azurin, determined by steady-state kinetics, was decreased several-fold by increasing the ionic strength. Furthermore, the extent of fluorescence quenching of ADH IIB due to the interaction with azurin was increased by increasing the ionic strength, but the binding constant for binding between ADH IIB and azurin was unchanged. The redox potential of azurin was increased 12 mV by incubation with ADH but not vice versa. Furthermore, the redox potential gap between ADH and azurin was increased from 102 to 126 mV by increasing the ionic strength. It is conceivable that a hydrophobic interaction is involved in the electron transfer between both proteins, and it is also suggested that the electron transfer may occur by a freely reversible on and off binding process but may not be related to the global binding process of both proteins. Thus, the results presented here strongly suggest that azurin works as an electron-transfer mediator in a PQQ-dependent alcohol oxidase respiratory chain in P. putida HK5.

Alcohol Dehydrogenase↗

PubMed1999

Lactation-dependent down regulation of leptin production in mouse mammary gland.

Lactation-dependent regulation of leptin expression in mouse mammary gland and parametrial adipose tissue was estimated by RT-PCR analysis for virgin, pregnant, lactating and post-lactating mice, and the serum and milk leptin levels of these mice were also determined by ELISA. Leptin gene expression in mammary gland as well as in adipose tissue was obviously detected before pregnancy, markedly decreased to 30-50% after parturition and kept at the low level during lactation period, and restored to the original level after weaning. The leptin concentration of milk collected just before weaning was about two-fold higher than that of the milk collected at mid-lactating stages. The serum leptin levels of the mid- and late-lactating mice were not significantly higher than those of non-pregnant mice. These results suggested that the lactation-induced down regulation of leptin was associated with autocrine/paracrine action of leptin in mammary and adipose tissues, and that the milk leptin, especially at the latter stages of lactation, was not only ascribed to diffusive transport from maternal blood stream, but also regional production and secretion by mammary epithelial cells. This possible production of leptin by mammary epithelial cells was further supported by the fact that leptin was expressed by cultured cells of mammary epithelial cell line, COMMA-1D, in a manner negatively dependent on the lactogenic hormones.

Adipose Tissue↗

PubMed1999

Lactation-dependent expression of an mRNA splice variant with an exon for a multiply O-glycosylated domain of mouse milk fat globule glycoprotein MFG-E8.

Expression of mRNA encoding MFG-E8, a milk fat-associated glycoprotein was investigated in mouse mammary gland. Two forms of mRNA, long and short variants, were shown to be expressed in the mammary tissue by RT-PCR analysis. Sequence analyses of these two variants and an isolated MFG-E8 gene segment indicated that the long and short mRNA variants resulted from an alternative splicing of a single pre-mRNA through in-flame inclusion and skipping of one exon, which encodes a proline/threonine (Pro/Thr)-rich domain. The long variant was expressed predominantly in mammary gland and the expression level was remarkably increased at late gestation and kept high during lactation. On the contrary, the short variant was detected ubiquitously in various tissues and its expression in the mammary gland was rather decreased in a lactation dependent manner. Expression of the long variant was also detected in a mouse mammary epithelial cell line, COMMA-1D, and enhanced by incubation with lactogenic hormones. Glycosylation inhibition analyses using tunicamycin and alpha-benzyl-GalNAc were conducted with COS7 cells transfected with plasmids expressing each mRNA variant, demonstrating that a fully glycosylated product of the long mRNA variant was not only N-glycosylated but also multiply O-glycosylated, whereas a product of the short one had only N-glycan(s). These results suggest that the alternative splicing plays a critical role for the mammary-specific and lactation-dependent expression of the MFG-E8 isoform and that the multiply O-glycosylated Pro/Thr-rich domain of this isoform is functionally important for formation of milk fat globules in mammary epithelial cells.

Alternative Splicing↗

PubMed1999

Simple and rapid high-performance liquid chromatography analysis of propentofylline and its main metabolites in serum using a direct injection technique.

A rapid, simple and reliable high-performance liquid chromatography (HPLC) column-switching method with UV detection (270 nm) for the simultaneous determination of propentofylline and its metabolites in human and rat sera was developed. The method involves direct injection of serum onto an HPLC column, which contains a shielded hydrophobic stationary phase for the separation of analytes from proteins in serum, and then loading the analytes onto a short octadecylsilylated silica gel (ODS) column using a switching valve. Propentofylline and its three metabolites in serum were separated from the serum components within 30 min after the injection. The detection limits (S/N = 3) of analytes spiked in human and rat sera ranged from 0.08 to 0.57 nmol/mL, and the net volume of serum used was 20 microL. The relative standard deviations for within- and between-day variations using rat serum were less than 4.3 and 5.6%, respectively. The method was used to determine propentofylline and its main metabolites in rat serum after a single intravenous dose of propentofylline (5 mg/kg).

Animals↗

PubMed1999

Effects of repeated brief episodes of ischemia and reperfusion in isolated perfused rat hearts.

The effects of ischemia and reperfusion on the coronary endothelium and myocardium as well as tolerance to ischemia/reperfusion injury were assessed using isolated retrogradely perfused rat hearts. Repeated brief episodes of myocardial ischemia followed by reperfusion is known to have a protective effect against subsequent myocardial infarction. However, no studies have been performed with perfusion in the absence of blood cells to determine the effect of repeated ischemia and reperfusion on the coronary endothelium and myocardium. Using the Langendorff perfusion technique, rat hearts were subjected to a 30-, 10-, 5-, or 2-min period of low-flow perfusion by reducing the coronary flow to 3 ml/min followed by reperfusion at 20 ml/min for the same period of time. Control perfusion was then performed at a constant flow rate of 20 ml/ min for 60 min. Acetylcholine-induced coronary vasodilation was significantly (P < 0.05) lower in hearts subjected to 30 min of ischemia and 30 min of reperfusion when compared with the control hearts. Myocardial creatinine kinase (CK) activity was significantly reduced (P < 0.01) in hearts subjected to ischemia and reperfusion for either 30, 10, or 5 min. To assess the effect of repeated episodes of ischemia and reperfusion, the following protocols were used: a control study with constant perfusion for 60 min (group A), 30 min of ischemia and 30 min of reperfusion (group B), three 10-min episodes of ischemia and reperfusion (group C), six 5-min episodes of ischemia and reperfusion (group D), and 15 2-min episodes of ischemia and reperfusion (group E). Acetylcholine-induced coronary vasodilation was significantly inhibited in group B (80% +/- 12%, P < 0.05) and group C (70% +/- 13%, P < 0.01), but did not change significantly in either group D (123% +/-19%) or group E (142% +/- 15%), compared with the control group (group A; 127% +/- 15%, mean +/-SEM). Nitroglycerin-induced coronary vasodilation was not altered by ischemia/reperfusion in any group. In contrast, myocardial CK activity was significantly lower in group B (3.6 +/- 0.6IU/mg protein, P < 0.01), group C (3.2 +/- 0.1 IU/mg protein, P < 0.01), and group D (3.3 +/- 0.21U/mg protein, P < 0.01) than in group A (47 +/- 6.7 IU/mg protein). The myocardial CK activity of group E was not significantly different from that of group A, but was significantly higher than in groups B, C, and D (P < 0.01). In isolated perfused rat hearts, both the coronary endothelium and myocardium are damaged by repeated episodes of ischemia and reperfusion. However, the coronary endothelium is more resistant to such damage than is the myocardium.

Acetylcholine↗

PubMed1999

Dissection of the middle cerebral artery caused by invasion of malignant glioma presenting as acute onset of hemiplegia.

A 57-year-old, previously healthy man who developed acute onset of hemiplegia is presented. Neuro-imaging studies on admission suggested dissection of the middle cerebral artery producing infarction in the frontotemporal region. In contrast to his stable clinical course, serial neuro-imaging studies disclosed rapid growth of malignant glioma, which was confirmed at surgery. Microscopic examination of the surgical specimen demonstrated invasion of glioma cells into the arterial wall associated with intramural haematoma formation of the middle cerebral artery. This case is the first to document dissection of an intracranial artery caused by invasion of tumour cells.

Acute Disease↗

PubMed1999

Study of p53 in elderly patients with myelodysplastic syndromes by immunohistochemistry and DNA analysis.

We analyzed the tumor suppressor gene product, p53, in elderly patients with myelodysplastic syndromes (MDS) and in overt leukemia patients after transformation from MDS using immunohistochemical techniques. We examined 52 MDS patients (mean age 79 years, range 68 to 96) from the time of initial diagnosis to death or development of overt leukemia. p53 protein was detected by immunohistochemistry (IHC) in 8/52 patients (15%) at initial diagnosis: 1/26 with refractory anemia (RA), 0/4 with RA with ringed sideroblasts, 3/11 with RA with an excess of blasts (RAEB), 3/8 with RAEB in transformation, and 1/3 with chronic myelomonocytic leukemia. We also analyzed gene mutations in patients with positive IHC. p53 mutations were detected in 3/8 (38%) patients. IHC-positive patients had a significantly higher incidence of leukemic transformation and the presence of a complex karyotype with monosomy 17. IHC-positive cells included blasts as well as mature myeloid cells, erythroblasts, and megakaryocytes. Scrutiny of our data in combination with previous data revealed that patients with positive IHC in multilineage cells were older than those in whom positivity was noted mostly in myeloblasts. This suggests that p53 IHC positivity with a multilineage pattern may be a characteristic of MDS in older patients.

Aged↗

PubMed1999

Teratogenicity of thujaplicin in ICR mice.

Beta-thujaplicin (TP) was studied by in vitro and in vivo tests for teratogenicity using ICR mice. In the in vitro study, TP (0, 3.125, 6.25, 12.5 microg/ml medium) dissolved in dimethyl sulfoxide (DMSO) was administered to cultured embryos on 9 day of gestation. After 24 hr of exposure to TP, the embryos were examined for developmental parameters and external anomalies. Growth retardation and embryos with facial dysplasia or hydrocyst of the tail tip were observed among the embryos given 12.5 microg/ml. In the in vivo study, TP (0, 420, 560, 750 or 1000 mg/kg) dissolved in olive oil was administered orally to pregnant mice on day 9 of gestation. All foetuses were removed from the uterus on day 18 of gestation, and were examined for external and skeletal anomalies. Various types of malformations were observed in the mice given 560 mg/kg or more. The number of litters having foetuses with external or skeletal anomalies increased in proportion to the dose of TP. The regression lines of Y (probit response) on X (log dose) for external anomalies was Y = 4.87X-8.43 . The 1% effective dose (ED1) for the malformation was 190 mg/kg. The present study shows that TP has teratogenic effects on mice.

Abnormalities, Drug-Induced↗

PubMed1999

A 42-kDa glycoprotein from chicken egg-envelope, an avian homolog of the ZPC family glycoproteins in mammalian Zona pellucida. Its first identification, cDNA cloning and granulosa cell-specific expression.

A glycoprotein with molecular mass of 42 kDa was identified as the major component of the chicken egg-envelope, the filamentous, extracellular matrix known as the perivitelline layer. By using a DNA probe amplified with degenerative primers derived from the protein's partial amino acid sequences, a cDNA clone encoding the egg-envelope 42-kDa glycoprotein (gp42) was isolated from a hen's ovary cDNA library. The gp42 open reading frame encoded 435 amino acid residues, including a putative signal peptide of 20 amino acids. The deduced amino acid sequence of gp42 showed significant similarity to egg-envelope glycoproteins of the ZPC family of several other vertebrate species, including human ZP3, mouse ZP3, Xenopus laevis gp43 and medaka (Oryzias latipes) ZI3 (LS-F), which play important roles for sperm-egg interaction. A single N-glycosylation site present in chicken gp42 is conserved among all five of these proteins: carbohydrate analysis of gp42 revealed the presence of a complex type glycan chain at this site. N-terminal sequence analysis of the mature polypeptide suggests that C-terminal processing of the pro-protein occurs during synthesis and secretion. The 1.4-kb gp42 transcript was detected only in follicles, and was found to be accumulated in granulosa cells in a manner dependent on ovarian follicular development. Furthermore, a metabolically radio-labeled gp42 was immunopreciptated from both cell lysate and culture supernatant of the granulosa cells with specific anti-gp42 antibody, suggesting granulosa cell-specific synthesis and secretion of the glycoprotein.

Amino Acid Sequence↗

PubMed1999

Down-regulation of protein tyrosine phosphatase gene expression in lactating mouse mammary gland.

Detailed analysis of protein tyrosine phosphatase (PTP) expression in mouse mammary gland and mammary epithelial cells using a set of degenerate primers corresponding to the PTP core domain sequence revealed the presence of 16 different receptor-type and intracellular PTPs. Northern blot and RT-PCR analyses revealed that some PTPs were up-regulated during gestation, suggesting that these enzymes are involved in development of mammary gland. However, expression of most PTPs dramatically decreased during lactation, whereas the beta-casein gene expression was increased and remained at a high level. At the involution stage after weaning, most PTPs were up-regulated and their expression returned almost to the virgin level. Such up-regulation was also induced by forced weaning in lactating mother mice. These results suggest the possible contribution of PTPs to the development, involution, and remodeling of mammary gland and their possible inhibitory action on maintaining high expression of milk genes during lactation.

Animals↗

PubMed1999

Molecular cloning and expression analysis of a gene for a sucrose transporter in maize (Zea mays L.).

Here we report the cloning of a sucrose transporter cDNA from maize (Zea mays L.) and an analysis of the expression of the gene. A cDNA clone (ZmSUT1) was identified as a sucrose transporter cDNA from its sequence homology at the amino acid level to sucrose transporters that have been cloned from other higher plant species. Based on the results of genomic Southern hybridization, ZmSUT1 appears to be a single copy gene. A Northern blot analysis of seedlings and leaf blades suggests that the sucrose transporter is involved in the export of accumulated carbohydrates from source leaf blades. From the measurements of transcript levels and carbohydrate contents in mature leaf blades, we propose that the expression of the gene for the maize sucrose transporter is positively regulated by the amounts of soluble sugars such as sucrose and glucose in source leaves of maize. In addition, based on the tissue specificity of the expression of the gene in maize plants at the reproductive stage, it is possible that the sucrose transporter acts in sink tissues such as pedicles as well as in source tissues such as leaf blades.

Biological Transport↗

PubMed1999

Two germline missense mutations at codons 804 and 806 of the RET proto-oncogene in the same allele in a patient with multiple endocrine neoplasia type 2B without codon 918 mutation.

Multiple endocrine neoplasia (MEN) type 2B is a clinically distinct entity among the autosomal dominant MEN 2 syndromes. Most patients with MEN 2B carry a germline mutation (M918T) of the RET proto-oncogene, while a few carry A883F. We examined a patient with MEN 2B, but without M918T or A883F, and her relatives. Here, we report the presence in this patient of 2 germline mutations, V804M and Y806C in the same allele. While the novel Y806C was inherited from her father, its carriers (her father and brother) was not affected by MEN 2. In contrast, V804M was a de novo mutation, that has been reported in patients with familial medullary thyroid carcinoma. Combinations of mutations of the RET proto-oncogene may cause oncogenic activities different from those of single mutations.

Base Sequence↗

PubMed1999

Analysis and perception of spectral 1/f characteristics of amplitude and period fluctuations in normal sustained vowels.

Two kinds of fluctuations are always observed in the steady parts of normal sustained vowels. One is amplitude fluctuation, defined as the cyclic changes of maximum peak amplitudes. The other is period fluctuation, defined as the cyclic changes of pitch periods. The primary purpose of this paper is to present quantitative descriptions of amplitude and period sequences obtained from normal sustained vowels. These fluctuation sequences consisted of maximum peak amplitudes or pitch periods extracted successively from 512 consecutive pitch periods in the steady part. Results of the frequency analysis indicated that their frequency characteristics seemed to be subject to the spectral 1/f power law. In order to investigate the possibility that the frequency characteristics of the fluctuation sequences influence the voice quality of sustained vowels, psychoacoustic experiments were conducted. Amplitude and period sequences evaluated in the experiments were spectral 1/f0 (white noise), 1/f, 1/f2, and 1/f3 sequences, respectively. The experimental results indicated that the subjective voice quality of synthesized sustained vowels could reflect the differences in the frequency characteristics of the fluctuation sequences.

Adult↗

PubMed1999

Immunohistochemical detection of JC virus in nontumorous renal tissue of a patient with renal cancer but without progressive multifocal leukoencephalopathy.

We performed immunohistochemical staining on the nontumorous renal tissue of 45 patients with renal cancer but without progressive multifocal encephalopathy using JCV-specific antibody. For one patient we found positive staining of the nuclei of the renal collecting ducts. Immunoelectron microscopic examination of the positive cell nuclei revealed electron-dense polyomavirus-like particles.

Antibodies, Viral↗

PubMed1999

Prediction of coronary artery lesions in unstable angina by iodine 123 beta-methyl iodophenyl pentadecanoic acid (BMIPP), a fatty acid analogue, single photon emission computed tomography at rest.

Iodine 123 beta-methyl iodophenyl pentadecanoic acid (123I-BMIPP), a beta-methyl-branched fatty acid analogue, has been proven by experimental studies to reveal abnormalities in fatty-acid-related metabolism. This study was undertaken to validate the accuracy and limitations of 123I-BMIPP imaging at rest in detecting myocardial metabolic abnormalities and predicting coronary lesions in unstable angina (UA). One hundred UA patients without prior myocardial infarction were studied. 123I-BMIPP and thallium 201 chloride (201TlCl) imaging with single photon emission computed tomography (SPECT) and coronary and left ventricular cineangiography (LVC) were performed 1 week after the last episode of angina. There was reduced uptake of 123I-BMIPP imaging in 70 patients, reduced uptake of 201TlCl in 41, and abnormal LVC contraction in 49 patients. There were significant increases in severity scores of 123I-BMIPP imaging along with increases in the number of stenosed coronary arteries and the severity of stenosis in individual coronary arteries. There was a significant reduction in 123I-BMIPP severity scores 1 month after percutaneous transluminal coronary angioplasty (p < 0.01) and a significant correlation between the severity scores of 123I-BMIPP and LVC (r=0. 579, p<0.001). Overall rates of sensitivity and specificity in detecting significant coronary stenosis by 123I-BMIPP imaging were 74% and 86%, respectively, whereas rates of sensitivity and specificity in detecting significant coronary stenosis by 201TlCl were 31% and 91%, respectively. 123I-BMIPP sensitivity increases to 86% if only advanced coronary stenosis of >90% is included. In conclusion, 123I-BMIPP myocardial imaging is an effective method of predicting coronary artery lesions of UA patients without provocative test.

Adult↗

PubMed1999

High fat feeding of lactating mice causing a drastic reduction in fat and energy content in milk without affecting the apparent growth of their pups and the production of major milk fat globule membrane components MFG-E8 and butyrophilin.

Lactating mice were fed either a low fat or a high fat diet. Milk samples were collected and the composition was examined. Triglyceride and free fatty acid contents were greatly reduced in the milks of high fat diet group, while protein and lactose contents were almost the same between both diet groups. Although the energy content of each component was also lower in milk of high fat diet group, there was apparently no significant difference in the growth of the pups raised by either diet group. This discrepancy might be in part explained by a hypothesis that the pups might monitor calorie content in milk and keep suckling until the energy intake reaches their satisfaction. Moreover, nearly the same amounts of major milk fat globule membrane proteins MFG-E8 and butyrophilin were shown to be present in the milks from both diet groups and gene expression of both proteins in the mammary glands were also indistinguishable, suggesting that production of major MFGM components is not simply related to fat production and secretion.

Age Factors↗