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K Machida

Publications and source records attributed to K Machida.

At least 73 records · Page 4Linked to original sources
PubMed1998

New evidence that the Tyr-157 and Tyr-159 residues of staphylococcal exfoliative toxin B are essential for its toxicity.

To determine the active site of exfoliative toxin B (sETB) of Staphylococcus aureus, the etb gene was cloned from an S. aureus SU strain obtained from a patient with impetigo. We prepared a frame shift mutant protein from a recombinant plasmid with a BglII linker inserted into the Tyr-155 codon of the ETB gene (pETB/BglIIL). The recombinant mutant protein (ETB/BglIIL) obtained from Escherichia coli containing pETB/BgIIIL showed no toxicity in neonatal mice and no agglutination activity. The 20-kDa ETB/BglIIL contained 185 amino acid residues. Site-directed mutagenesis was used to introduce mutations at either Tyr-155, Tyr-157, Tyr-159, or Tyr-162. Substitution of any of the Tyr residues decreased exfoliative activity compared with that of native sETB (4,000 EU/ml). Substitution of Tyr-155 with a Phe (ETBYN155) decreased activity 5-fold (800 EU/ml). Substitution of Tyr-157 with Leu (ETB/Y157) decreased activity 80-fold (50 EU/ml) and decreased agglutination titer 5-fold compared with that of native sETB (400,000). Substitution of Tyr-159 with Leu (ETB/Y159)decreased activity 4-fold (1,000 EU/ml). When both Tyr-157 and Tyr-159 were mutated (ETB/Y157-159), both toxicity and antigenicity were completely lost. On an immunodiffusion test, ETBNY157 showed a faint precipitation line, but ETB/BglIIL and ETB/Y157-159 had no activity, showing that the Tyr-157 and Tyr-159 residues are essential for the toxicity and antigenicity of ETB.

Amino Acid Sequence↗
PubMed1998

Farnesol-induced generation of reactive oxygen species via indirect inhibition of the mitochondrial electron transport chain in the yeast Saccharomyces cerevisiae.

The mechanism of farnesol (FOH)-induced growth inhibition of Saccharomyces cerevisiae was studied in terms of its promotive effect on generation of reactive oxygen species (ROS). The level of ROS generation in FOH-treated cells increased five- to eightfold upon the initial 30-min incubation, while cells treated with other isoprenoid compounds, like geraniol, geranylgeraniol, and squalene, showed no ROS-generating response. The dependence of FOH-induced growth inhibition on such an oxidative stress was confirmed by the protection against such growth inhibition in the presence of an antioxidant such as alpha-tocopherol, probucol, or N-acetylcysteine. FOH could accelerate ROS generation only in cells of the wild-type grande strain, not in those of the respiration-deficient petite mutant ([rho0]), which illustrates the role of the mitochondrial electron transport chain as its origin. Among the respiratory chain inhibitors, ROS generation could be effectively eliminated with myxothiazol, which inhibits oxidation of ubiquinol to the ubisemiquinone radical by the Rieske iron-sulfur center of complex III, but not with antimycin A, an inhibitor of electron transport that is functional in further oxidation of the ubisemiquinone radical to ubiquinone in the Q cycle of complex III. Cellular oxygen consumption was inhibited immediately upon extracellular addition of FOH, whereas FOH and its possible metabolites failed to directly inhibit any oxidase activities detected with the isolated mitochondrial preparation. A protein kinase C (PKC)-dependent mechanism was suggested to exist in the inhibition of mitochondrial electron transport since FOH-induced ROS generation could be effectively eliminated with a membrane-permeable diacylglycerol analog which can activate PKC. The present study supports the idea that FOH inhibits the ability of the electron transport chain to accelerate ROS production via interference with a phosphatidylinositol type of signal.

Electron Transport↗
PubMed1998

[Changes of physiological functions in rats induced by immobilization stress].

A study was conducted on the changes of physiological function in rats due to immobilization stress. Male Fischer rats (SPF) of 32 weeks of age were housed in individual cages for 4 weeks. Then all rats were immobilized by stainless wire mesh for 6 hours daily for 3 days. Blood was collected before the 1st stress, immediately after the 1st stress, immediately after the 3rd stress and the day after the 3rd stress. The results of this experiment were as follows: (1) The total leukocyte counts in the blood of the rats after the 1st trial was significantly higher than that before the 1st trial. (2) The percentage of lymphocytes in the blood after the 1st trial was significantly lower than that before the 1st trial, whereas that of neutrophils was significantly higher. (3) Correlations between phagocytic activity and superoxide production of neutrophils by histochemical NBT reduction assay showed significantly a positive correlation before the 1st trial. However, no significant correlations were observed in immediately after the 1st trial and the 3rd trial. The day after the 3rd trial, a positive correlation was observed again. These correlations showed that an unsuitable state of the neutrophil function was induced by the immobilization stress. (4) Serum biochemical profiles were affected by the immobilization stress. Also, GOT, GPT, LDH, CK and UA were increased after the 1st trial, whereas, TG, TP, ALB and ALP were decreased after the 1st trial. T-CHO was increased only immediately after the 3rd stress. These results suggest that immobilization stress affected blood cells and serum components, and then the host defense and physiological functions were damaged respectively.

Animals↗
PubMed1998

Suppression of cell growth by ectopic expression of N-cadherin.

We found that ectopic expression of N-cadherin in 3Y1 caused tight association of cells and, thereby, substantially suppressed cell growth. N-cadherin expression inhibited neither tyrosine phosphorylation of cellular proteins, GTP uptake onto Ras, nor activation of MAP kinase, suggesting that it does not directly interfere the Ras-MAP kinase pathway. However, co-expression of N-cadherin with dominant negative Ras, S17N Ras, showed synergestic growth inhibitory effect, suggesting that N-cadherin signaling antagonizes the Ras-MAP kinase signaling. In addition, we found that N-cadherin yielded cell-cycle arrest at G0/G1 phase. These results strongly suggest that N-cadherin cell adhesion machinery works as a negative controller of cell cycle in 3Y1 and this growth suppressive function of cadherin is distinct from the epithelial morphogenetic function.

Actins↗
PubMed1998

[Report of national survey on death due to tuberculosis in 1994 in national hospitals and the treatment and prognosis of tuberculous patients with mechanical ventilation].

1. National survey on died patients with active tuberculosis (tbc) or tbc sequelae had been held in national hospitals every five year from 1959 (3433 cases) to 1994 (688 cases). In 1994, 330 patients died due to pulmonary tbc. Recent study revealed the decreased rate of death due to operation, or far advanced cavitary cases, and the increased rate of nontuberculous death, aged people (> 60 yrs), and nontuberculous complications. Main causes of death in pulmonary tbc were lung insufficiency (about half) and general weakness (almost one fifth) in any survey. Rapid progression of pulmonary tbc had been increased cause of death (20.9% in 1994). Main attributable factors of death in 1994 in pulmonary tbc cases were severe condition on admission (38.4%), disturbed lung function (31.2%) and old age (33.2%). Delayed treatment (13.9%) and complications (12.1%) were increasing factors. Early death within 3 months from onset in 1994 was seen in patients < 60 yrs as well as in patients > 80 yrs. Severity due to delayed treatment and rapid progression were supposed to the causes of early death. 2. During 1994 to 1997, mechanical ventilation (MV; > 24 hours) was applied to 18 patients with active pulmonary tuberculosis; 10 acute respiratory failure (ARF), 5 chronic respiratory failure (CRF), 2 central nervous system tbc and 1 hemoptysis. Only one ARF case and three CRF ones survived. ARF cases had low PaO2/FIO2 (about 100), low albuminemia, short MV period (7 cases: < 7 days) and steroid therapy (9 cases). CRF cases had higher PaO2/FIO2 (294), longer MV period (4 cases: > 30 days) and all CO2 narcosis. 3. Noninvasive positive pressure ventilation (NIPPV) was applied to 23 patients with pulmonary tbc sequelae. In 13 patients with stable chronic respiratory failure (mean PaO2 91 mmHg, PaCO2 82 mmHg) 10 continued NIPPV and started home mechanical ventilation (HMV). In 10 patients with acute on chronic respiratory failure (mean PaO2 61 mmHg, PaCO2 92 mmHg) 2 patients fell into tracheal intermittent positive pressure ventilation (TIPPV). Eight patients recovered with NIPPV and 5 started HMV. NIPPV is supposed to be very effective to treat severe chronic hypercapmic respiratory failure.

Adult↗
PubMed1998

[MR cholangiography of congenital biliary malformations in infancy].

We presented MR cholangiography (MRC) of congenital biliary malformations in infancy. MRC was obtained during induced sleeping. In two cases of congenital dilation of bile duct, MRC revealed cystic or spindle dilatation of intra- and extra hepatic bile ducts. In one biliary atresia, MRC revealed the serpentine gall bladder and cystic dilatation of the extrahepatic bile duct without connection to the dilated hilar bile duct. MR cholangiography, which can be obtained noninvasively, is useful for the diagnosis and the preoperative assessment of congenital biliary malformations in infancy.

Bile Ducts↗
PubMed1998

[Clinical review of 74 cases with miliary tuberculosis].

Seventy-four cases of miliary tuberculosis were studied retrospectively. The mean age of the patients was 45.3 years. Twenty-two patients suffered from another underlying diseases. Six were infected with human immunodeficiency virus. Twelve had been treated with corticosteroids. Fever was present in 97.3 per cent of patients. Elevation of serum alkaline phosphatase was found in 67.6 per cent of cases. The skin reaction to tuberculin was positive in 61.2 per cent. Nodular shadows were found in the chest X-ray in 98.6 per cent of cases. The nodules were smaller than 2 mm in diameter in 52.7 per cent of cases. Other findings were enlargement of mediastinal lymph node (17.6%), cavities (23.0%), pleural effusion (27.0%), and consolidation (35.1%). Sputum cultures and urine cultures were positive for Mycobacterium tuberculosis in 76.8 per cent and 58.6 per cent of cases respectively. The diagnosis was confirmed by histopathological findings in some cases. The rate of positive biopsies was 61.5 per cent by bone marrow aspiration, 83.3 per cent by lymph node biopsy, 100 per cent by liver and lung biopsy. Antituberculosis therapy was successful in most of the patients. Seven patients died of miliary tuberculosis, 4 of them had adult respiratory distress syndrome.

Adolescent↗
PubMed1998

[Lung cancer in patients with sequelae of tuberculosis].

To clarify features of lung cancer in patients with tuberculosis sequelae, we analyzed data on 15 cases (5.1%) who were diagnosed with lung cancer before death among 294 deceased cases with tuberculosis sequelae at our hospital. There were 12 men and 3 women, with a mean age of 64 years. Most of the 15 patients had pulmonary dysfunction, and 4 had received home oxygen therapy. All 12 men had a history of smoking, and 10 of them had squamous cell carcinoma of the lung. There was no definite correlation between the locations of the tuberculosis lesion and those of lung cancer lesion on chest X-rays. Twelve patients had had thoracoplasty for tuberculosis, and in 6 of these patients the lung cancer occurred in the same lung. Lung cancer was apt to be diagnosed in an advanced stage. However, in patients who received home oxygen therapy, diagnosis had been made at an early stage because of the frequent chest X-ray follow-up. We conclude that lung cancer is an important complication in patients with tuberculosis sequelae, and early diagnosis of lung cancer by careful follow-up is essential in the care of cases with tuberculosis sequelae who have poor pulmonary function and/or systemic conditions.

Aged↗
PubMed1997

Isolation of RNA aptamers specific to the NS3 protein of hepatitis C virus from a pool of completely random RNA.

Hepatitis C virus (HCV) is a single-stranded RNA virus and its genome is translated into a single large polyprotein. The viral-encoded NS3 protein possesses protease, nucleoside triphosphatase, and helicase activities. Since these activities appear to be important for viral replication, efforts are being made to identify compounds that might inhibit the enzymatic activities of NS3 and serve as potential anti-HCV agents. We used a genetic selection strategy in vitro to isolate, from a pool of completely random RNA (120 random bases), those RNA aptamers that could bind to NS3. After six cycles of selection and amplification, 14% of the pooled RNAs could bind specifically to the NS3 protein. When the aptamers in the pool (cycle 6) were analyzed for binding and inhibition of the proteolytic activity of NS3 with the NS5A/NS5B peptide as substrate (S1), two aptamers, designated G6-16 and G6-19 RNA, were found to inhibit NS3 in vitro. Kinetic studies of the inhibition revealed that the aptamer G6-16 inhibited the NS3 protease with an inhibitory constant (Ki) of 3 microM. We also analyzed aptamers G6-16 and G6-19 for their action with a longer protein substrate (amino acid region 2203-2506) and found that these aptamers efficiently inhibited the proteolytic activity of NS3. In addition, both G6-16 and G6-19 aptamers were found to inhibit the helicase activity of NS3. Since these aptamers possesses dual inhibitory function for NS3, they could prove to be useful as anti-HCV drug leads.

Base Sequence↗
PubMed1997

Inhibitory effects of nucleoside 5'-alkylphosphates on sexual agglutination in Saccharomyces cerevisiae.

Among various nucleoside 5'-alkylphosphates, uridine 5'-hexadecylphosphate (UMPC16) and adenosine 5'-hexadecylphosphate (AMPC16) inhibited the sexual agglutination between a and alpha haploid cells of Saccharomyces cerevisiae. The effect of AMPC16 accompanied severe growth inhibition of the yeast cells but it was not observed with UMPC16. Sexual agglutination was not inhibited by the presence of UMPC16 or AMPC16 when the yeast cells had been pretreated with the mating pheromone. UMPC16 was characterized as a specific inhibitor of sexual agglutination without direct influence on the agglutinin function, being distinguishable from any of those ever known.

Adenine Nucleotides↗
PubMed1997

Rapid identification and typing of Staphylococcus aureus by nested PCR amplified ribosomal DNA spacer region.

We designed a polymerase chain reaction (PCR) assay for rapid detection of prokaryotic 16S-23S spacer regions. This PCR assay consisted of nested DNA amplifications. The first-step PCR was able to detect the general presence of eubacteriales with a unified set of universal primers. The universal primers were selected from highly conserved regions in 16S and 23S ribosomal RNA (rRNA) genes and amplified DNAs from all 62 different species of bacteria tested. In the second-step PCR, the identification primers could detect four important bacterial species through amplification of the rRNA spacer regions between the 16S-23S rRNA genes. For Staphylococcus aureus, intraspecies variation in spacer amplification products was observed with S. aureus specific primers. We suggest that the nested PCR assay could be used as a novel method for the identification and typing in epidemiological studies of S. aureus.

DNA Primers↗
PubMed1997

Inhibition of neutrophil apoptosis by antioxidants in culture medium.

Neutrophil apoptosis is an important mechanism that has implications for understanding the life span and toxic potentials of neutrophils at inflamed sights. In this study the authors examined the possibility that reactive oxygen species (ROS) released by neutrophils can regulate neutrophil survival. Cu,Zn-superoxide dismutase (Cu,Zn-SOD), Mn-SOD, and catalase in culture media were significantly effective in delaying the spontaneous apoptosis, suggesting that ROS play an important role in the resolution of inflammation by inducing neutrophil apoptosis. In this experiment, boiled Cu,Zn-SOD had no effect on inhibiting the apoptosis, but boiled Mn-SOD from Bacillus stearothermophilus was more effective in inhibiting the apoptosis than untreated Mn-SOD at the same dose. However, the boiled Mn-SOD showed only 80% of O2- inhibitable activity compared with the untreated Mn-SOD. This effect may be attributed to the partial liberation of manganese because MnCl2 inhibited the apoptosis effectively. Furthermore, Cu,Zn-SOD was effective in delaying apoptosis only when added to the culture within the first 3 h of incubation, suggesting that the isolation of neutrophils from peripheral blood enhances apoptosis of neutrophils.

Antioxidants↗
PubMed1997

Effect of angiotensin II on arteriovenous shunting assessed by hepatic arterial perfusion scintigraphy.

Angiotensin II (AT II) has been reported to improve drug delivery in intraarterial chemotherapy for hepatic tumors. We studied the effect of this agent on arteriovenous (AV) shunting in the liver. Eleven patients with hepatic tumors and an indwelling catheter in the hepatic artery underwent hepatic arterial perfusion scintigraphy with and without AT II infusion. At baseline, following the administration of technetium-99m macroaggregated albumin (Tc-99m MAA) through the catheter, static images of the upper abdomen and whole-body anterior and posterior images were obtained. Two to 14 days later, AT II was infused via the catheter one minute prior to Tc-99m MAA injection, and imaging was performed in the same way as the baseline study. Visual interpretation of the static images showed improvement of drug delivery to the hepatic tumors in eight of 11 patients and no changes in the remaining patients. The percent injected dose in the lungs assessed quantitatively using the whole-body images increased in all patients, suggesting enhancement of AV shunt flow. It was concluded that intraarterial infusion of AT II increases AV shunting in the liver with better targeting to hepatic tumors.

Adult↗
PubMed1997

Simultaneous detection of Streptococcus pneumoniae and Haemophilus influenzae by nested PCR amplification from cerebrospinal fluid samples.

Haemophilus influenzae and Streptococcus pneumoniae are often the cause of serious diseases such as meningitis. We designed a nested PCR assay to identify these pathogens from cerebrospinal fluid samples. The first-step PCR was able to detect eubacterial rRNA genes with a unified set of universal primers. In the second-step PCR, the identification primers, HI I and II and SP I and II, could detect H. influenzae and S. pneumoniae respectively through amplification of the rRNA spacer between the 16S and 23S rRNA genes. We suggest that the two-step PCR assay can be used as a novel method for the immediate and retrospective diagnosis of bacterial meningitis caused by H. influenzae and S. pneumoniae.

Cerebrospinal Fluid↗
PubMed1997

The use of bronchofiberscopy for diagnosis of allergic bronchopulmonary aspergillosis.

To assess the use of bronchofiberscopy for the diagnosis of allergic bronchopulmonary aspergillosis (ABPA), we retrospectively analyzed bronchofiberscopic and pathologic findings of specimens obtained by bronchofiberscopy in 8 patients who met the British criteria for ABPA. Two of the 8 patients completely met Rosenberg's primary criteria for ABPA, while the remaining 6 lacked only 1 or 2 immunologic and radiographic conditions of the criteria. Bronchofiberscopic inspection revealed the presence of mucus plugs in all patients. Pathologic examination of plug specimens obtained by suction from 6 of 7 of these patients showed that the plugs consisted of allergic mucin containing fungal hyphae, diagnostic of ABPA. Bronchofiberscopy appears to be a useful tool in the diagnosis of ABPA, if immunologic and radiographic findings are not typical of ABPA. This bronchofiberscopic method is easily applicable for diagnosing allergic bronchopulmonary fungal disease (ABPFD) in addition to ABPA.

Adolescent↗