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Biomedical subjects

N Fujii

Publications and source records attributed to N Fujii.

At least 19 recordsLinked to original sources
PubMed1999

T134, a small-molecule CXCR4 inhibitor, has no cross-drug resistance with AMD3100, a CXCR4 antagonist with a different structure.

T22, an analog of polyphemusin II (18 amino acid residues), was found to block T-tropic human immunodeficiency virus type 1 (HIV-1) entry into target cells as a CXCR4 inhibitor. We synthesized T134, a small analog (14 amino acid residues) of T22 with reduced positive charges. T134 exhibited highly potent activity and significantly less cytotoxicity in comparison to that of T22. T134 prevents the anti-CXCR4 monoclonal antibody from binding to peripheral blood mononuclear cells but has no effect on the binding of anti-CCR5 monoclonal antibodies. Since T134 inhibits the binding of stromal cell-derived factor-1 (SDF-1) to MT-4 cells, it seems that T134 prevents HIV-1 entry by binding to CXCR4. The bicyclam AMD3100 has also been shown to block HIV-1 entry via CXCR4 but not via CCR5. Both T134 and AMD3100 are CXCR4 antagonists and low-molecular-weight compounds but have different structures. Our results indicate that T134 is active against wild-type T-tropic HIV-1 strains and against AMD3100-resistant strains.

Anti-HIV Agents↗
PubMed1999

Biological roles of angiotensin II via its type 2 receptor during rat follicle atresia.

Type 1 angiotensin II (ANG II) receptors play crucial roles in the regulation of blood pressure and fluid osmolarity, whereas the physiological roles of type 2 ANG II receptors (AT2) remain unclear. Because AT2 is expressed in atretic follicles where granulosa cells undergo apoptosis, we examined the space and time relationship between AT2 expression and follicle atresia in vivo and the effect of AT2 on follicle-stimulating hormone (FSH) actions in vitro. Binding studies, autoradiography, and RT-PCR of AT2 revealed that the AT2 content in granulosa cells was time dependently increased at both protein and mRNA levels in equine chorionic gonadotropin-treated immature female rats. This increase paralleled the progression of atresia. ANG II suppressed FSH-caused prevention of DNA fragmentation, increases in luteinizing hormone receptor content, and estrogen production through AT2 in cultured granulosa cells. Moreover, FSH-induced stimulation of extracellular signal-regulated kinase activity, critical for cell survival, was inhibited by AT2 stimulation. These results suggest that AT2 mediates the progression of follicle atresia through granulosa cell apoptosis by inhibiting FSH actions.

Angiotensin II↗
PubMed1999

Microstimulation of the lateral wall of the intraparietal sulcus compared with the frontal eye field during oculomotor tasks.

We compared the effects of intracortical microstimulation (ICMS) of the lateral wall of the intraparietal sulcus (LIP) with those of ICMS of the frontal eye field (FEF) on monkeys performing oculomotor tasks. When ICMS was applied during a task that involved fixation, contraversive saccades evoked in the LIP and FEF appeared similar. When ICMS was applied to the FEF at the onset of voluntary saccades, the evoked saccades collided with the ongoing voluntary saccade so that the trajectory of voluntary saccade was compensated by the stimulus. Thus the resultant saccade was redirected and came close to the endpoint of saccades evoked from the fixation point before the start of voluntary saccade. In contrast, when ICMS was applied to the LIP at the onset of voluntary saccades, the resultant saccade followed a trajectory that was different from that evoked from the FEF. In that case, the colliding saccades were redirected toward an endpoint that was close to the endpoint of saccades evoked when animals were already fixating at the target of the voluntary saccade. This finding suggests that the colliding saccade was directed toward an endpoint calculated with reference to the target of the voluntary saccade. We hypothesize that, shortly before initiation of voluntary saccades, a dynamic process occurs in the LIP so that the reference point for calculating the saccade target shifts from the fixation point to the target of a voluntary saccade. Such predictive updating of reference points seems useful for immediate reprogramming of upcoming saccades that can occur in rapid succession.

Animals↗
PubMed1999

Simultaneous amplification of Bordetella repeated insertion sequences and toxin promoter region gene by polymerase chain reaction.

A polymerase chain reaction was devised to simultaneously detect repeated insertion sequences and the pertussis toxin promoter gene for the diagnostic identification of Bordetella pertussis, B. parapertussis, and B. bronchiseptica. The sensitivity of this method was sufficient to detect one B. pertussis organism using the following cycles and temperatures: 95 degrees C for 15 min, followed by 32 amplification cycles (1 min at 95 degrees C, 1 min at 66 degrees C, 1 min at 72 degrees C), and finally 5 min at 72 degrees C. Using the primers as a combined set did not affect sensitivity, but required an increased temperature for optimal annealing compared with a single-sequence assay. As nasopharyngeal aspirate and swab materials sometimes contain hemoglobin, we also tested the inhibitory effect of hemoglobin on this assay, which was inhibited completely when using DNA extracts from samples containing hemoglobin at a final concentration >0.015 g/L: this inhibition was reversed by addition of bovine serum albumin to the buffer. Our assay shows promising sensitivity and specificity for clinical use.

Bordetella Infections↗
PubMed1999

Molecular characterization of total kininogen deficiency in Japanese patients.

Kininogens are multifunctional plasma glycoproteins. There are two forms of human kininogen: low molecular weight kininogen (LK) and high molecular weight kininogen (HK). Both are derived from the same gene by alternative splicing. Some patients with kininogen deficiency have been reported to be deficient only in HK while others are deficient in both HK and LK (total kininogen deficiency). We analyzed three Japanese patients with total kininogen deficiency by the Csp45I digestion study of exon 5 as previously reported in Williams trait and found that two had the same point mutation of C to T at base 22 of exon 5, resulting in a transition of CGA (Arg) codon to TGA (Stop) codon. This is the first report of molecular characterization of total kininogen deficiency in the Japanese population.

Codon↗
PubMed1998

A low-molecular-weight inhibitor against the chemokine receptor CXCR4: a strong anti-HIV peptide T140.

T22 ([Tyr5,12, Lys7]-polyphemusin II) is an 18-residue peptide amide, which has strong anti-HIV activity. T22 inhibits the T cell line-tropic (T-tropic) HIV-1 infection through its specific binding to a chemokine receptor CXCR4, which serves as a coreceptor for the entry of T-tropic HIV-1 strains. Herein, we report our finding of novel 14-residue CXCR4 inhibitors, T134 and T140, on the basis of the T22 structure. In the assays we examined, T140 showed the highest inhibitory activity against HIV-1 entry and the strongest inhibitory effect on the binding of an anti-CXCR4 monoclonal antibody (12G5) to CXCR4 among all the CXCR4 inhibitors that have been reported up to now.

Amino Acid Sequence↗
PubMed1998

Acein-1, a novel angiotensin-I-converting enzyme inhibitory peptide isolated from tryptic hydrolysate of human plasma.

A novel angiotensin-I-converting enzyme (ACE) inhibitory peptide, designated acein-1, was isolated from the tryptic hydrolysate of human plasma. Gel filtration and cation exchange chromatography were performed to purify this peptide, followed by reversed-phase gradient and isocratic high-performance liquid chromatography. Acein-1 was found to be a heptapeptide, Tyr-Leu-Tyr-Glu-Ile-Ala-Arg, corresponding to f(138-144) of human serum albumin. The synthetic heptapeptide, hexapeptide (Tyr-Leu-Tyr-Glu-Ile-Ala, des-7R acein-1) and octapeptide (Tyr-Leu-Tyr-Glu-Ile-Ala-Arg-Arg, acein-1R) showed dose-dependent inhibitions of ACE, and their IC50 values were 16 micromol/l, 500 micromol/l and 86 micromol/l, respectively. Acein-1 might be a non-competitive inhibitor, while acein-1R may be an uncompetitive inhibitor, as shown by Lineweaver-Burk plots.

Amino Acid Sequence↗
PubMed1998

Carbon dioxide fixation by reversible pyrrole-2-carboxylate decarboxylase from Bacillus megaterium PYR2910.

Pyrrole-2-carboxylate decarboxylase from Bacillus megaterium PYR2910 attains a balanced reaction equilibrium with an equilibrium constant of 0.3-0.4 M. Therefore, the enzyme catalyzes the reverse carboxylation of pyrrole after addition of bicarbonate. For the synthesis of pyrrole-2-carboxylate, the reverse reaction was optimized and the equilibrium was shifted towards the carboxylate. The product yield was 230 mM (25.5 g/l) pyrrole-2-carboxylate from 300 mM pyrrole in a batch reaction and 325 mM (36.1 g/l) from 400 mM pyrrole in a fed-batch reaction, using both whole cells and the purified enzyme in a pH 8.0 reaction mixture with bicarbonate saturation of 1.9 M. Kinetic studies indicated, that bicarbonate is the reactive species used by this carbon dioxide-fixation enzyme.

Bacillus megaterium↗
PubMed1998

An oculomotor representation area within the ventral premotor cortex.

We explored the ventral part of the premotor cortex (PMV) with intracortical microstimulation (ICMS) while monkeys performed a visual fixation task, to see whether the PMV is involved in oculomotor control. ICMS evoked saccades from a small-restricted region in the PMV, without evoking movements in the limbs, neck, or body. We found the saccade-evoking site in the PMV in a total of three hemispheres in two monkeys. Quantitative analysis of the effects of eye position on saccades evoked by microstimulation of the PMV characterized the evoked saccades as goal directed. The nature of the saccades evoked in the PMV contrasted with the fixed vector nature of saccades evoked by ICMS of the frontal eye field. We also found that neurons in this restricted area of the PMV were active while the animals were performing a saccade task that required them to make saccades toward targets without arm movements. These data provide evidence for the presence of an oculomotor-specific subregion within the PMV. This subregion and the surrounding skeletomotor-representing regions of the PMV seem to coordinate oculomotor and skeletomotor control in performing goal-directed motor tasks.

Animals↗
PubMed1998

The gene for component-II of botulinum C2 toxin.

The gene encoding component-II of the Clostridium botulinum C2 toxin was cloned from the chromosomal DNA of C. botulinum type C strain (C)-203U28, and the nucleotide sequence was determined. The gene (bc2h) encodes a protein with 721 amino acid residues and is located at 247 bp downstream of the gene for component-I. The N-terminal 16 amino acids were identical to those obtained by analysis of the purified component-II toxin. The ORF for bc2h had only 39% homology at the amino acid level with the C.perfringens iota-Ib protein and an ATP/GTP binding site which is present in the iota-Ib protein is missing from the protein encoded by bc2h. Both genes had a homologous region that predicts a transmembrane segment.

Amino Acid Sequence↗
PubMed1998

Evidence for presenilin-1 involvement in amyloid angiopathy in the Alzheimer's disease-affected brain.

Presenilin-1 (PS-1) has been identified as the protein encoded by the chromosome 14 locus that, when mutated, leads to familial Alzheimer's disease (FAD). The role PS-1 plays in the pathogenesis of Alzheimer's disease (AD) remains unclear. Using a set of antibodies raised against PS-1 synthetic peptides, polyclonal antibody to amyloid beta protein (Abeta) and end-specific antibodies against Abeta40, and Abeta42, immunohistochemical studies were performed on brain sections obtained from AD cases and controls. The PS-1 antibodies clearly stained amyloid angiopathies in AD-affected brains, but no recognizable immunoreactions were observed in any other vessels free from amyloid involvement in either AD-affected brains or controls. Abeta antibodies and the end-specific antibody against Abeta40 also decorated amyloid angiopathies, showing localization similar to that of PS-1. Western blot analyses predominantly detected protein band polypeptide species of a 50 kDa, band, presumably full-length PS-1 protein with N-terminus antisera, since these antibodies turned out to recognize a 50-kDa full-length band in cell lysate of transfected HeLa cell overexpressing PS-1. In addition, we recognized 30, 27 and 25 kDa proteins in both AD and control brain homogenate with these antibodies. In microvessel fractions extracted from brain homogenates, the 50, and 27 kDa fragments were observed in AD-affected brains but not in those of controls. C-terminus rabbit antisera reacted strongly with the 33 and 27 kDa bands, and additionally detected a small amount of full-length PS-1 protein in extracts from AD and control brains. Our present data indicate that PS-1 might be involved in the pathogenesis of amyloid angiopathy in the AD brain.

Alzheimer Disease↗
PubMed1998

Negative chronotropic effect of botulinum toxin on neonatal rat cardiac myocytes.

We demonstrated that botulinum neurotoxin attenuated the spontaneous beating rate of cultured cardiac myocytes. Primary cultured cardiac myocytes were prepared from the ventricles of neonatal Wistar rats (1-3 days old). On 7 days after cell seeding, botulinum toxin type A incorporated into liposomes was added to the culture medium. At a final concentration of 5.0 micrograms/ml, botulinum toxin markedly attenuated the beating rate of cardiac myocytes within 2-4 hours. These results demonstrated the effect of SNARE-complex proteins on the spontaneous beating of cardiac myocytes.

Animals↗
PubMed1998

Gene arrangement in the upstream region of Clostridium botulinum type E and Clostridium butyricum BL6340 progenitor toxin genes is different from that of other types.

The cluster of genes encoding the botulinum progenitor toxin and the upstream region including p21 and p47 were divided into three different gene arrangements (class I-III). To determine the gene similarity of the type E neurotoxin (BoNT/E) complex to other types, the gene organization in the upstream region of the nontoxic-nonhemagglutinin gene (ntnh) was investigated in chromosomal DNA from Clostridium botulinum type E strain Iwanai and C. butyricum strain BL6340. The gene cluster of type E progenitor toxin (Iwanai and BL6340) was similar to those of type F and type A (from infant botulism in Japan), but not to those of types A, B, and C. Though genes for the hemagglutinin component and P21 were not discovered, genes encoding P47, NTNH, and BoNT were found in type E strain Iwanai and C. butyricum strain BL6340. However, the genes of ORF-X1 (435 bp) and ORF-X2 (partially sequenced) were present just upstream of that of P47. The orientation of these genes was in inverted direction to that of p47. The gene cluster of type E progenitor toxin (Iwanai and BL6340) is, therefore, a specific arrangement (class IV) among the genes encoding components of the BoNT complex.

Botulinum Toxins↗
PubMed1998

Poor induction of interferon-induced 2',5'-oligoadenylate synthetase (2-5 AS) in cells persistently infected with mumps virus is caused by decrease of STAT-1 alpha.

Poor induction of interferon-induced 2',5'-oligoadenylate synthetase (2-5AS) has been demonstrated in cells persistently infected with mumps virus as compared with uninfected cells. As for the number of interferon (IFN) receptors and the level of IFN regulatory factors (IRF-1 and IRF-2) mRNAs, there was little difference between them. Therefore, it is suggested that the IFN-alpha signaling system is ineffective in the persistently infected cells. Components of IFN-stimulating gene factor 3 alpha (ISGF-3 alpha), STAT-1 alpha (p91) and STAT-2 (p113), were investigated in human amnion (FL), human nasopharyngeal cancer (KB), human T-lymphoid (HUT 78), and human B-lymphoid (Akata) cells persistently infected with mumps virus. STAT-1 alpha, but not STAT-2, disappeared in these persistently infected cells, and this factor was not restored by treatment of these cells with IFN. However, no difference was observed between the levels of STAT-1 alpha mRNA transcript in persistently infected and uninfected control cells. It is reasonable to infer that the poor induction of 2-5AS activity is due to the decrease of STAT-1 alpha in correlation with the IFN-signal transduction pathway. Furthermore, induction of other IFN-stimulated genes (ds-RNA activated protein kinase, PKR, and MxA protein) was also reduced in the cells persistently infected with mumps virus.

2',5'-Oligoadenylate Synthetase↗
PubMed1998

Advanced glycation end products in Alzheimer's disease and other neurodegenerative diseases.

Advanced glycation end products (AGEs) have been implicated in the chronic complications of diabetes mellitus and have been reported to play an important role in the pathogenesis of Alzheimer's disease. In this study, we examined the immunohistochemical localization of AGEs, amyloid beta protein (A beta), apolipoprotein E (ApoE), and tau protein in senile plaques, neurofibrillary tangles (NFTs), and cerebral amyloid angiopathy (CAA) in Alzheimer's disease and other neurodegenerative diseases (progressive supranuclear palsy, Pick's disease, and Guamanian amyotrophic lateral sclerosis/Parkinsonism-dementia complex). In most senile plaques (including diffuse plaques) and CAA from Alzheimer's brains, AGE and ApoE were observed together. However, approximately 5% of plaques were AGE positive but A beta negative, and the vessels without CAA often showed AGE immunoreactivity. In Alzheimer's disease, AGEs were mainly present in intracellular NFTs, whereas ApoE was mainly present in extracellular NFTs. Pick's bodies in Pick's disease and granulovacuolar degeneration in various neurodegenerative diseases were also AGE positive. In non-Alzheimer neurodegenerative diseases, senile plaques and NFTs showed similar findings to those in Alzheimer's disease. These results suggest that AGE may contribute to eventual neuronal dysfunction and death as an important factor in the progression of various neurodegenerative diseases, including Alzheimer's disease.

Adult↗