Search PubMed⌕ Search

Biomedical subjects

M Takagi

Publications and source records attributed to M Takagi.

At least 451 records · Page 25Linked to original sources
PubMed1995

CYP52 (cytochrome P450alk) multigene family in Candida maltosa: identification and characterization of eight members.

Previously, we characterized three genes and presented evidence for an n-alkane-inducible cytochrome P450 (P450alk) multigene family in an n-alkane-assimilating and diploid-type yeast, Candida maltosa. In the present report, we isolated and characterized additional members of this gene family, including a total of thirteen P450alk-related sequences (eight genes and five of their alleles). Two sets, each consisting of two genes, were tandemly arranged in the genome. A gene replacement experiment showed that at least one gene had only a single allele in the genome. The determined nucleotide and the deduced amino acid sequences indicated that all had a characteristic constituent for P450s and exhibited amino acid identities from 94% to 37% to each other. Six genes showed relatively higher similarities to each other than to the other two genes and were thus classified into a subfamily. All the members of this subfamily were assigned to the same single chromosome, showing a good correlation between sequence similarity and chromosomal linkage. Although all the genes except for one were induced by n-alkane, their inducibilities by some other aliphatic carbon sources showed variabilities.

Alleles↗
PubMed1995

Effect of rice culturing practices on seasonal occurrence of Culex tritaeniorhynchus (Diptera: Culicidae) immatures in three different types of rice-growing areas in central Japan.

Seasonal occurrence of Culex tritaeniorhynchus Giles immatures was surveyed, together with detailed observations on the rice cultivation practices, weekly in three different types of rice field habitat from 1982 to 1985. Despite early water introduction and seedling transplantation, a late seasonal peak of density and abundance of first instars were observed at terraced rice fields (YE) with continuous water in- and outflow through sluices in a hilly area. In contrast, immature populations increased quickly in flat rice fields (AN, SK). High density and abundance of fourth instars were encountered before midseason drying in rice fields (AN) with less flow and little loss of water, because of construction of an independent irrigation and drainage system. The midseason drying and the alternating flooding and draining after the drying effectively suppressed the occurrence of immatures. Early final draining for harvesting eliminated mass production of mosquitoes, even if the density was high in some rice fields. Differences in timing of seasonal rice-culturing procedures and water management were assumed to be effective in controlling the seasonal occurrence of C. tritaeniorhynchus.

Agriculture↗
PubMed1995

Isolation of canCHS1A, a variant gene of Candida albicans chitin synthase.

A canCHS1A gene encoding the chitin synthase of Candida albicans was cloned. DNA sequencing and comparison with another canCHS1 gene described elsewhere indicated that the canCHS1A gene encoded a polypeptide with 775 amino acid residues, a protein with one less amino acid than that encoded by the canCHS1 gene. A six-base alteration was observed between the two genes, suggesting that the canCHS1A gene is a variant gene of the canCHS1. The pH profile/activity relationship of canChs1A in permeabilized cells was identical to that of canChs1. The canChs1A enzyme was competitively inhibited by polyoxin D (5.2 microM) and nikkomycin Z (12 microM). When the cloned gene was expressed in a Saccharomyces cerevisiae chs2 mutant that exhibited aberrant morphology, the normal structure was restored. We conclude that the function of the canCHS1A gene is similar to that of sacCHS2 in S. cerevisiae.

Aminoglycosides↗
PubMed1995

Detection of astroviruses from stool samples in Japan using reverse transcription and polymerase chain reaction amplification.

We developed a reverse transcription and polymerase chain reaction (RT-PCR) method for detecting astrovirus serotypes 1, 2, 3, 5, 6 and 7 (but not serotype 4). Furthermore, we developed the specific primers for detecting serotypes 1 and 2, the most predominant serotypes in the world. Sensitivity of the first PCR with serotype common primers was about 10 times higher than that of enzyme immunoassay with monoclonal antibody (EIA-MAb). Sensitivity of the second PCR with the serotype-specific primers was even higher. The RT-PCR method was useful for detecting astroviruses from clinical samples, especially serotypes 1 and 2.

Base Sequence↗
PubMed1995

Inducible expression of a gene encoding an L41 ribosomal protein responsible for the cycloheximide-resistant phenotype in the yeast Candida maltosa.

In a previous paper (S. Kawai, S. Murao, M. Mochizuki, I. Shibuya, K. Yano, and M. Takagi, J. Bacteriol. 174:254-262, 1992), we showed that in each genome of several yeast species, there is one of two types of L41 gene, one for an L41 (Q-type) protein which confers cycloheximide (CYH) resistance or one for an L41 (P-type) protein which does not. These genes have been suggested to be responsible for the CYH response used in taxonomy. For example, Saccharomyces cerevisiae, which is CYH sensitive, has a P-type L41 gene, while Kluyveromyces fragilis and Candida maltosa, which are CYH resistant, have Q-type L41 genes. However, in contrast to K. fragilis, which is constitutively resistant to CYH, C. maltosa is inducibly resistant to CYH. Here, we show that C. maltosa has both types of the L41 gene in its genome and that expression of the Q-type L41 gene is induced by CYH while the P-type L41 gene is constitutively expressed.

Base Sequence↗
PubMed1995

A transformation system for the yeast Candida utilis: use of a modified endogenous ribosomal protein gene as a drug-resistant marker and ribosomal DNA as an integration target for vector DNA.

We have developed a transformation system for the yeast Candida utilis. A novel strategy was applied to construct the transformation system, since auxotrophic mutants which could be used as hosts for transformation are not available. A gene encoding the ribosomal protein L41 was cloned from C. utilis, which is sensitive to cycloheximide, and used as a marker gene conferring cycloheximide resistance after modification of its amino acid sequence. The marker gene was constructed by substitution of the proline codon at position 56 with the glutamine codon by in vitro mutagenesis, as it had been reported previously that the 56th amino acid residue of L41 is responsible for the cycloheximide sensitivity of various organisms (S. Kawai, S. Murao, M. Mochizuki, I. Shibuya, K. Yano, and M. Takagi, J. Bacteriol. 174:254-262 1992). The ribosomal DNA (i.e., DNA coding for rRNA) of C. utilis was also cloned and used as a multiple-copy target for the integration of vector DNA into the genome, which resulted in a high transformation efficiency. Transformants were obtained by electroporation with a maximum efficiency of approximately 1,400 transformants per 1 microgram of linearized DNA carrying the gene for cycloheximide resistance and part of the ribosomal DNA. No transformants were obtained with intact plasmids. Multiple copies of the linearized plasmid were integrated into the host chromosome by homologous recombination. Southern analysis of the transformants in which vector DNA was integrated at the L41 gene locus indicated that there are two copies of gene for the L41 protein per cell, suggesting that C. utilis is diploid. Transformants were obtained from a variety of C. utilis strains, indicating that this method is applicable to the transformation of other C. utilis strains, even though there is significant heterogeneity in chromosomal karyotypes among these strains.

Amino Acid Sequence↗
PubMed1995

Myocardial infarct size by serum troponin T and myosin light chain 1 concentration.

The late troponin T (TnT) peak concentration, which is known to be independent of reperfusion of the infarcted zone in acute myocardial infarction (MI), has been suggested to be correlated with clinical estimates of cardiac function and myocardial infarct size. To refine the clinical application of the late TnT peak in infarct sizing, and to examine differences in this estimation in different infarct sites, we measured the serum concentrations of TnT and myosin light chain 1 (MLC1), and compared these values with left ventricular ejection fraction (LVEF) obtained from left ventriculography, and extent score (ES) and severity score (SS) obtained from 201Tl scintigraphy in patients with anterior and inferior myocardial infarction. The late TnT peak concentration was strongly correlated with the MLC1 peak value in patients with anterior MI (r = 0.67, p < 0.05) and in those with inferior MI (r = 0.92, p < 0.0005). Furthermore, there were strong linear correlations between the late TnT peak values and all of the clinical data (LVEF; r = -0.79, p < 0.01, ES; r = 0.75, p < 0.05, SS; r = 0.75, p < 0.05, respectively) in patients with anterior MI. However, these correlations were weak in patients with inferior MI. Similar correlations were observed between MLC1 and the clinical data. Thus, TnT and MLC1 have similar kinetics in the serum at the late phase and can be used to estimate the size of anterior infarct.

Aged↗
PubMed1995

Evidence that the expression of the gene for NADPH-cytochrome P-450 reductase is n-alkane-inducible in Candida maltosa.

A gene coding for NADPH-cytochrome P-450 reductase of an n-alkane-assimilating yeast, Candida maltosa, was isolated and sequenced. Northern analysis and assay of the expression of the reporter gene under the control of the promoter of this gene showed that the transcriptional level was induced 4 to 8-fold in cells grown on n-alkane relative to cells grown on glucose.

Alkanes↗
PubMed1995

Pseudomonas fluorescens KKL101, a benzoic acid degrader in a mixed culture that degrades biphenyl and polychlorinated biphenyls.

A mixed culture we had isolated, which degrades biphenyl/polychlorinated biphenyls, is composed of two strains, Pseudomonas fluorescens KKL101 and Pseudomonas sp. strain KKS102. KKS102 produces benzoic acid as a dead-end metabolite in the degradation of biphenyl. In this study we showed that KKL101 grew on benzoic acid as a sole source of carbon. This indicated a role of KKS102 in the growth of KKL101 and KKL101 for the growth of KKS102 in the mixed culture.

Bacteriological Techniques↗
PubMed1995

Human neutrophil collagenase (MMP-8), identified in bronchiectasis BAL fluid, correlates with severity of disease.

Collagenases in bronchoalveolar lavage fluid (BALF) of patients with bronchiectasis and healthy subjects were characterized using specific functional and immunologic assays. The BAL fluid contained interstitial collagenase and collagenolytic proteinases of bacterial origin. Collagenase activities, obtained after organomercurial activation, correlated with the severity of bronchiectasis. In severe cases, collagenase activities were 3.5 x 10(-7) IU/L/48 h or 4.8 x 10(-6) IU/g/48 h (p < 0.01), in moderate ones 1.74 x 10(-7) IU/L/48 h or 3.35 x 10(-6) IU/g/48 h (p < 0.05), and in mild cases 0.32 x 10(-7) IU/L/48 h or 0.7 x 10(-6) IU/g/48 h (p < 0.05). The corresponding activities in healthy control subjects were 0.08 x 10(-7) IU/L/48 h or 0.13 x 10(-6) IU/g/48 h. The cellular origin of interstitial collagenase was assessed with doxycycline inhibition test utilizing the differential sensitivity of fibroblast-type collagenase/MMP-1 (IC50 = 280 microM) and neutrophil-type collagenase/MMP-8 (IC50 = 26 microM) to the anticollagenolytic, nonantimicrobial doxycycline action. Interstitial collagenase, contained in BALF, was totally inhibited by 100 microM of doxycycline. It can therefore be concluded that most of mammalian collagenase presented in inflamed fluid of bronchiectasis originated from neutrophils. The molecular forms of neutrophil-type collagenase/MMP-8 were confirmed and analyzed by Western-blot, which showed evidence of the proteolytic conversion of the latent 85-kD MMP-8 proenzyme species into active 65-kD molecular weight species. These findings strongly suggest involvement of proteolytic activation pathway of proMMP-8, especially in severe and moderate forms of bronchiectasis. Furthermore, collagenolytic proteases of bacterial origins may also participate in tissue destruction of the lung.

Adolescent↗
PubMed1995

Cathepsin G and alpha 1-antichymotrypsin in the local host reaction to loosening of total hip prostheses.

The tissue localization and content of the proteolytic enzyme cathepsin G and its inhibitor alpha 1-antichymotrypsin were studied in the local host reaction to loosening of total hip-replacement prostheses in eleven patients and were compared with those in samples of non-inflammatory tissue from the synovial capsule obtained during arthroscopies of the knee. Immunostaining demonstrated cellular localization of cathepsin G in 71 per cent of monocyte or macrophage-like cells and in 46 per cent of fibroblast-like cells in the samples of interface tissue between the bone and the loose acetabular component obtained at the time of the total hip replacements, and in 59 and 42 per cent, respectively, in the samples of pseudocapsular tissue obtained at the same time, whereas the synovial lining cells in the samples of non-inflammatory tissue from the synovial capsule revealed only a slight immunoreactivity to cathepsin G. Cathepsin-G activity was also measured with synthetic succinyl-alanine-alanine-proline-phenylalanine-paranitroanilide as a substrate, the degradation of which was monitored spectrophotometrically. In accordance with results from immunohistochemical studies, cathepsin-G activity was found in the samples of interface tissue (31.6 international units per liter) and the samples of pseudocapsular tissue (15.5 international units per liter) obtained during the total hip replacements, whereas the level of cathepsin-G was low in the samples of non-inflammatory synovial capsular tissue (2.5 international units per liter). Cathepsin-G activity in the samples of pseudosynovial fluid obtained at the time of the total hip replacements was low (2.4 international units per liter), although immunoblot analysis showed marked immunoreactive cathepsin G in the samples of pseudosynovial fluid. This low activity of cathepsin G might be explained by the presence of alpha 1-antichymotrypsin, which was detected by laser nephlometric immunoassay and immunoblot analysis. These results demonstrate increased concentration of cathepsin G locally in the tissues around loose total hip-replacement prostheses. Because cathepsin G is not only able to act on extracellular matrix components (such as gelatin, proteoglycan, elastin, and laminin) at a physiological pH but also is able to activate collagenase, gelatinase, and stromelysin proenzymes, to inactivate tissue inhibitor of metalloproteinases, and to modulate tumor necrosis factor-alpha, it may play an important role in the degradation of periprosthetic connective tissue and in the lysis of bone around the implant, thus contributing to the loosening of prostheses.(ABSTRACT TRUNCATED AT 250 WORDS)

Aged↗
PubMed1995

Mark-release-recapture experiments to estimate the efficiency of the light trap in collecting Japanese encephalitis vector mosquitoes.

Efficiency of the light trap in collecting females of Culex tritaeniorhynchus, the principal vector mosquito of Japanese encephalitis, was evaluated by mark-release-recapture experiments. Females collected with dry ice in the field were marked and released in a pigsty. The recapture rate of marked females by the light trap was so low as around 1% or lower. Owing to this low efficiency, it is not likely that the light trap can be a useful tool in the control of Japanese encephalitis.

Animals↗
PubMed1995

Facilitation in Anopheles and spontaneous disappearance of filariasis: has the concept been verified with sufficient evidence?

The validity of recently much argued phenomenon of facilitation in the transmission of filariasis was considered by examining previously published papers. It was concluded that there was no clear evidence to support the existence of facilitation and facilitation-based unstable equilibrium in relation to microfilaria prevalence and density in human population below which filariasis would spontaneously disappear, even when the vector was Anopheles mosquitoes. Instead, the existence of a critical level of man/mosquito contacts for the disappearance of filariasis was suggested.

Animals↗
PubMed1995

Comparison of HIV-1 protease expression in different fusion forms.

Earlier observations showed that the expression of recombinant protease of human immunodeficiency virus type-1 (HIV-1 PR) was usually in a low level, and its proteolytic activity and hydrophobicity were believed to be toxic for the host cells. Various constructs were investigated that contained an N-terminal extended HIV-1 PR gene (PR107) in order to find a system which can express this protease in high level. The constructs of PR107 gene expressed as fusion proteins either with glutathione S-transferase (GST) by pGEX-PR107 or with maltose-binding protein (MBP) by pMAL-PR107 showed that the full length of fusion protein exhibited self-cleavage in E. coli. The results from expression experiments indicated that the size of the fusion portion does not affect the self-processing of fused HIV-1 PR to release its mature form, despite the attachment of only one subunit of the dimeric protease to GST or MBP. The construct, pET-PR107, under the control of strong bacteriophage T7 promoter system, did not show clear advantages for expression of this HIV-1 PR. Comparing these three constructs, the pGEX-PR107 system showed the highest expression level. Quantitative immuno-blotting indicated that the amount of HIV-1 PR expressed by pGEX-PR107 was twice that expressed by pMAL-PR107, and thrice that expressed by pET-PR107. More than 1 mg of pure HIV-1 PR from per liter culture of E. coli. DH5 alpha containing pGEX-PR107 can be obtained via the purification procedures [Biochem. Mol. Biol. International, (1995) 35:899-912].(ABSTRACT TRUNCATED AT 250 WORDS)

ATP-Binding Cassette Transporters↗
PubMed1995

[Bacterial intracavernous carotid aneurysm presented as massive epistaxis].

A 55-year-old man was admitted with complaints of headache, diplopia, fever and swelling of the right eyelid. Physical examination revealed low grade fever of 37.6 degrees C, but no evidence of infectious focus was detected. The neurological examination disclosed right abducens nerve palsy. An initial routine blood count showed an increased white blood cell count of 11,800/microliters. The erythrocyte sedimentation rate was 93mm and the C-reactive protein level was 3.6mg/dl. Massive epistaxis and hypovolemic shock developed on the 14th day while the patient was receiving antibiotics (CEZ, CMZ) for a possible intracranial infection. The magnetic resonance angiography of the brain and the cerebral angiography demonstrated an aneurysm within the cavernous sinus, which had not been present on the admission. Since epistaxis was caused by a rupture of the aneurysm, internal carotid artery ligation was done. The signs of inflammation improved as well as patients' symptoms. The improvement while using antibiotics strongly implies that the aneurysm was a result of bacterial infection of the cavernous sinus. There have been no reports of ruptured bacterial intracavernous aneurysms presented with massive epistaxis as this case.

Aneurysm, Infected↗
PubMed1995

[Bacteriological and clinical studies of SY5555 in pediatric field].

Clinical studies were carried out on SY5555, a new oral penem, in the field of pediatrics. The results obtained are summarized below. The clinical efficacies were examined in a total 31 patients consisting of 4 patients with pharyngitis, 10 with purulent tonsillitis, 4 with scarlet fever, 7 with impetigo, one with balanitis, one with cellulitis and 4 with UTI. The clinical efficacy rate was 96.8% (30/31). Bacteriological efficacies of SY5555 were examined on identified pathogens including 7 strains of Staphylococcus aureus, 6 of Streptococcus pyogenes, 3 of Enterococcus faecalis, 3 of Haemophilus influenzae, one of Escherichia coli and one of Citrobacter freundii. The bacteriological eradication rate was 81.0%. As for side effects, loose stool in one patient was noted. Abnormal laboratory findings test results included eosinophilia in 2 patients, eosinophilia and elevation of serum transaminase in one patient, and thrombocytosis in another.

Administration, Oral↗
PubMed1995

[Functional magnetic resonance imaging of the human primary visual cortex during visual stimulation].

Signal changes in the human primary visual cortex during visual stimulation were evaluated using non-invasive functional magnetic resonance imaging (fMRI). The experiments were performed on 10 normal human volunteers and 2 patients with homonymous hemianopsia, including one who was recovering from the exacerbation of multiple sclerosis. The visual stimuli were provided by a pattern generator using the checkerboard pattern for determining the visual evoked potential of full-field and hemifield stimulation. In normal volunteers, a signal increase was observed on the bilateral primary visual cortex during the full-field stimulation and on the contra-lateral cortex during hemifield stimulation. In the patient with homonymous hemianopsia after cerebral infarction, the signal change was clearly decreased on the affected side. In the other patient, the one recovering from multiple sclerosis with an almost normal visual field, the fMRI was within normal limits. These results suggest that it is possible to visualize the activation of the visual cortex during visual stimulation, and that there is a possibility of using this test as an objective method of visual field examination.

Adult↗