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Biomedical subjects

Takeshi Ito

Publications and source records attributed to Takeshi Ito.

At least 19 recordsLinked to original sources

Activation process of the mosquitocidal delta-endotoxin Cry39A produced by Bacillus thuringiensis subsp. aizawai BUN1-14 and binding property to Anopheles stephensi BBMV.

Most delta-endotoxins produced by Bacillus thuringiensis require proteolytic processing in order to become active. The in vitro and in vivo activation processes of Cry39A, a delta-endotoxin that is highly toxic to Anopheles stephensi, were investigated. Cry39A with a molecular mass of 72 kDa was processed in vitro into a 60 kDa fragment by trypsin and gut extract from A. stephensi larvae. N-terminal amino acid sequencing of the 60 kDa fragment revealed that trypsin and the protease(s) in the gut extract cleaved Cry39A between Arg(61) and Gly(62). In contrast, 40 and 25 kDa polypeptides were generated in vivo by intramolecular cleavage of the 60 kDa fragment in A. stephensi larvae. Further, a co-precipitation assay was used to investigate the binding property of the activated Cry39A to A. stephensi BBMV. Cry39A bound to A. stephensi BBMV specifically and did not compete with the Cry4Aa toxin. This indicated that the binding molecule(s) for Cry39A might differ from those for Cry4A. In addition, Cry39A preferentially bound to the Triton X-100-insoluble membrane fraction.

Animals↗

Cloning and expression of two crystal protein genes, cry30Ba1 and cry44Aa1, obtained from a highly mosquitocidal strain, Bacillus thuringiensis subsp. entomocidus INA288.

Two novel crystal protein genes, cry30Ba and cry44Aa, were cloned from Bacillus thuringiensis subsp. entomocidus INA288 and expressed in an acrystalliferous strain. Cry44Aa crystals were highly toxic to second-instar Culex pipiens pallens (50% mortality concentration [LC50] = 6 ng/ml) and Aedes aegypti (LC50 = 12 ng/ml); however, Cry30Ba crystals were not toxic.

Aedes↗

[Incidence and risk factors of intraocular pressure elevation after triamcinolone acetonide administration for macular disorders].

PURPOSE: We investigated the incidence and risk factors of intraocular pressure (IOP) elevation following triamcinolone administration for macular disorders. METHODS: Intravitreal or posterior sub-Tenon's injections of triamcinolone acetonide were conducted on 119 eyes with macular disorders. The change in IOP was followed for all cases. Risk factors for IOP elevation were examined regarding causal disease, history of vitrectomy, methods of injection, frequency of injection, refractive power, and age. RESULT: The difference between the mean baseline IOP (13.0 +/- 2.9 mmHg) and the maximum mean IOP (21.5 +/- 9.8 mmHg) was statistically significant (p< 0.0001). The final mean IOP was 15.1 +/- 3.6 mmHg. Forty-one eyes (34.5%) had IOP elevation over 21 mmHg during the follow-up. Fifty-nine eyes (49.6%) showed pressure elevation of over 5 mmHg. One eye (1%) required trabeculotomy to control the IOP. Risk factors for the IOP elevation included younger age, high myopia, no history of vitrectomy, and multiple injections. CONCLUSION: Careful follow-up of the IOP is required after triamcinolone acetonide injections.

Age Factors↗

Cellular apoptosis susceptibility protein and proliferation in human hepatocellular carcinoma.

The cellular apoptosis susceptibility protein (CAS) is the human homologue of the product of the essential yeast chromosome segregation gene, CSE1, and has important roles in tumor necrosis factor (TNF)-induced apoptosis and cell proliferation. In this study, we used immunoblotting and immunohistochemistry to look at CAS expression in human hepatocellular carcinoma (HCC) cells. We also studied the correlation between CAS expression and cell proliferation. To do this, we studied the expression of proliferating cell nuclear antigen (PCNA) by immunostaining and at apoptosis by in situ nick end-labeling (TUNEL), followed by calculation of the PCNA labeling index (PCNA LI) and TUNEL labeling index (TUNEL LI). CAS was constitutively expressed in human HCC cell lines and was primarily confined to the cytoplasm of the cells. PCNA LI and TUNEL LI were significantly higher in HCC than in non-tumor tissue (p<0.01); however, the ratio of TUNEL LI/PCNA LI in HCC was significantly lower than that of non-tumor tissue. Immunohistochemistry revealed that the staining intensity score of CAS in HCC was significantly higher than that of non-tumor tissue (p<0.05). These results indicate that there is an augmentation of pro-liferative activity and apoptosis in HCC tissue, as compared to non-tumor tissue. There was a significant positive correlation between CAS and PCNA LI (p<0.05). In addition, we observed an inverse relationship between CAS expression and TUNEL LI, although the correlation did not reach statistical significance. These results suggest that CAS is expressed at higher levels in human HCC tissue than in non-tumor tissue. CAS may be associated with cell proliferation rather than apoptosis, and further, CAS might play an important role in the development of human HCCs.

Apoptosis↗

CD40 expression in HCV-associated chronic liver diseases.

CD40 is expressed primarily on B cells and plays an important role in antigen presentation, B cell proliferation, and T cell activation. It has been reported that the CD40 signal modulates apoptosis and has an anti-viral effect in certain cells. Therefore, we investigated the expression and the function of CD40 in HCV-associated chronic liver disease. The expression of CD40 on liver tissues was determined through immunohistochemistry on 50 liver specimens obtained from HCV-positive patients. The effect of CD40 signaling on apoptosis of HepG2 cells was assessed using the MTT assay. The effect of CD40 stimulation on NF-kappaB activation was determined in NF-kappaB reporter gene-transfected HepG2 cells with the Luciferase assay. CD40 positive hepatocytes were observed in both periportal and lobular areas, accompanied by inflammation. In both areas, CD40 staining intensity became significantly stronger, correlating with the histological grading. Similarly, it became stronger with the progression of the histological staging in F1, F2 and F3 cases; however, the expression level decreased in F4 cases. CD40 ligation induced apoptosis in HepG2 cells in the presence of 500 ng/ml of actinomycin D, while CD40 ligation alone could not. Anti-CD40 monoclonal antibody caused NF-kappaB activation in HepG2 cells in a dose-dependent manner. These results suggest that hepatocyte over-expression of CD40 might play an important role in regulating hepatocyte survival and death in HCV-associated chronic liver diseases.

Antibodies, Monoclonal↗

The right hepatic artery syndrome.

Various benign and malignant conditions could cause biliary obstruction. Compression of extrahepatic bile duct (EBD) by right hepatic artery was reported as a right hepatic artery syndrome but all cases were compressed EBD from stomach side. Our case compressed from dorsum was not yet reported, so it was thought to be a very rare case. We present here the first case of bile duct obstruction due to the compression of EBD from dorsum by right hepatic artery.

Cholestasis, Extrahepatic↗

Risk factors for cerebral palsy in preterm infants.

OBJECTIVE: To identify crucial factors that precipitate cerebral palsy by controlling confounding factors in logistic regression analyses. DESIGN AND PATIENTS: We retrospectively investigated a cohort of all 922 infants with gestational ages of less than 34 weeks (22-33 weeks), who were admitted to our neonatal intensive care unit between 1990 and 1998. Thirty (3.7%) were diagnosed to have cerebral palsy. We analyzed the prenatal and postnatal clinical variables of the cerebral palsy cases and compared them with 150 randomly selected controls. RESULTS: Risk factors for cerebral palsy identified in univariate analysis were: twin pregnancy, long-term ritodrine tocolysis, respiratory distress syndrome, air leak, surfactant administration, intermittent mandatory ventilation, high frequency oscillation, lowest PaCO2 levels, prolonged hypocarbia during the first 72 h of life, and postnatal steroid therapy. In a conditional multiple logistic model, long-term ritodrine tocolysis, prolonged hypocarbia and postnatal steroid therapy remained associated with an increased risk of cerebral palsy after adjustment for other antenatal and postnatal variables (OR [Odds Ratio] = 8.62, 95% CI [Confidence Interval], 2.18-33.97; OR = 7.81, 95% CI, 1.42-42.92; OR = 21.37, 95% CI, 2.01-227.29, respectively). CONCLUSIONS: Our results suggest that long-term ritodrine tocolysis underlines the development of cerebral palsy. Further assessments of the effect of ritodrine on fetal circulation and nervous system are required. Moreover, possible alternatives to systemic postnatal steroids are needed, and carbon dioxide levels should be more strictly controlled.

Cerebral Palsy↗

Hydrogen and ethanol production from glycerol-containing wastes discharged after biodiesel manufacturing process.

H2 and ethanol production from glycerol-containing wastes discharged after a manufacturing process for biodiesel fuel (biodiesel wastes) using Enterobacter aerogenes HU-101 was evaluated. The biodiesel wastes should be diluted with a synthetic medium to increase the rate of glycerol utilization and the addition of yeast extract and tryptone to the synthetic medium accelerated the production of H2 and ethanol. The yields of H2 and ethanol decreased with an increase in the concentrations of biodiesel wastes and commercially available glycerol (pure glycerol). Furthermore, the rates of H2 and ethanol production from biodiesel wastes were much lower than those at the same concentration of pure glycerol, partially due to a high salt content in the wastes. In continuous culture with a packed-bed reactor using self-immobilized cells, the maximum rate of H2 production from pure glycerol was 80 mmol/l/h yielding ethanol at 0.8 mol/mol-glycerol, while that from biodiesel wastes was only 30 mmol/l/h. However, using porous ceramics as a support material to fix cells in the reactor, the maximum H2 production rate from biodiesel wastes reached 63 mmol/l/h obtaining an ethanol yield of 0.85 mol/mol-glycerol.

Bioreactors↗

[Analysis of phenmedipham in agricultural products by HPLC].

An analytical method was developed for the determination of phenmedipham (PM) in agricultural products using reversed-phase high-performance liquid chromatography with UV detection. A sample was extracted with acetonitrile, and the acetonitrile layer was separated by salting-out. The acetonitrile phase was isolated and evaporated. The extract was dissolved in diethyl ether-hexane (1 : 1), and then cleaned up on a Florisil column. The column was washed with diethyl ether-hexane (1 : 1), and PM was eluted with acetone-hexane (3 : 7), and the eluate was evaporated. The residue was dissolved in acetone-hexane (2 : 8), and the sample solution was cleaned up on SAX/PSA cartridge. The SAX/PSA cartridge was washed with acetone-hexane (2 : 8), and PM was eluted with acetone-hexane (3 : 7). If required, the eluate of the Florisil column was cleaned up with SAX/PSA and ENVI-Carb/ NH2 cartridges. The SAX/PSA cartridge was washed with acetone-hexane (2 : 8), and connected to be ENVI-Carb/NH2 cartridge. The cartridges were washed with acetone-hexane (3 : 7), and then the SAX/PSA cartridge was removed. PM was eluted with acetonitrile-toluene (3 : 1) from the ENVI-Carb/NH2 cartridge. PM in the eluate was separated isocratically on an ODS column (4.6 mm i.d. x 150 mm, 5 microm) using acetonitrile-water (6 : 4) as a mobile phase (flow-rate 1.0 mL/min, temp. 40 degrees C), with monitoring at 235 nm. The calibration curve was linear from 0.005 microg/mL to 10 microg/mL of PM. The recoveries of PM from eight kinds of agricultural products spiked at levels of 0.1 and 0.02 microg/g were 80.8-98.7%. The determination limit was 0.01 microg/g.

Beta vulgaris↗

Different effects of bile acids, ursodeoxycholic acid and deoxycholic acid, on cell growth and cell death in human colonic adenocarcinoma cells.

Secondary bile acids have been implicated as an important etiological factor in colorectal cancer. We investigated the effects of ursodeoxycholic acid (UDCA) and deoxycholic acid (DCA) on the growth and cytotoxicity in HT29 human colonic adenocarcinoma cells. Proliferation assay, cell cycle analysis and cell death characterization by bile acids were performed. Both UDCA and DCA reduced their proliferation rate of HT29 over 48 h in a concentration- and time-dependent manner compared with control cultures. In terms of cell cycle effects, however, UDCA induced G2/M arrest, while DCA induced G1 arrest in a concentration- and time-dependent manner. As for the effects of each bile acid on cell toxicity, UDCA induced early apoptosis and DCA induced both early apoptosis and necrosis. Bile acids play an important role in regulating cell survival and cell death in colon adenocarcinoma cells.

Adenocarcinoma↗

Unliganded thyroid hormone receptor-beta1 represses liver X receptor alpha/oxysterol-dependent transactivation.

The thyroid hormone receptor (TR) and liver X receptor (LXR)-alpha are members of the nuclear hormone receptor family and are ligand-dependent transcription factors. Among the promoter target genes, TR and LXR recognize the T3 response element and LXR response element (LXRE), respectively. Because T3 response elements and LXREs have similar configurations, referred to as direct repeat 4, we investigated the possibility of cross-talk between the two ligand-dependent signal transduction pathways. We found that TRbeta1, a major isoform of TR in the liver, binds and transactivates LXREs derived from the mouse mammary tumor virus long-terminal repeat and the promoter of the sterol regulatory element binding protein 1c. Moreover, unliganded TRbeta1 suppresses promoter activity driven by LXRalpha and its ligand, whereas transactivation by T3-bound TRbeta1 is not affected by LXRalpha in the presence or absence of oxysterols. Gel shift, mammalian two-hybrid, and glutathione S-transferase pull-down assays demonstrated the direct binding of TRbeta1 to these LXREs and revealed that the interaction between TRbeta1 and corepressors is important to the unliganded TR-mediated suppression of LXRalpha-transactivation. Our findings suggest that T3 and TR influence lipid metabolism regulated by oxysterol/LXRalpha at the transcriptional level.

Animals↗

High-yield production of hydrogen by Enterobacter aerogenes mutants with decreased alpha-acetolactate synthase activity.

To enhance hydrogen (H2) production from glucose by Enterobacter aerogenes HU-101, two mutants, strains VP-1 and VP-2, with decreased alpha-acetolactate synthase activity, were isolated using the Voges-Proskauer (VP) test. In pH-uncontrolled batch culture, both mutants showed a lower 2,3-butanediol yield for the glucose consumed than that shown by the wild-type strain, although glucose remained in the medium after 12 h of culture. In the same cultures, compared to the H2 yield of 0.80 mol/mol-glucose of the wild-type strain, strain VP-1 showed a high H2 yield of 1.8 mol/mol-glucose with decreased lactate and increased succinate yields, while strain VP-2 showed an H2 yield of 1.0 mol/mol-glucose with an increased lactate yield. Increasing the phosphate buffer concentration, which contributes to maintaining the pH in the medium, increased the glucose consumption by both strains. However, in a pH-controlled batch culture at neutral pH, the H2 yield of strain VP-1 was decreased to 1.2 mol/mol-glucose due to the accumulation of formate, an intermediate of the H2-producing pathway, with the yield of H2 plus formate being 1.7 mol/mol-glucose.

Journal Article↗

Specific interaction between GroEL and denatured protein measured by compression-free force spectroscopy.

We investigated the interaction between GroEL and a denatured protein from a mechanical point of view using an atomic force microscope. Pepsin was bound to an atomic force microscope probe and used at a neutral pH as an example of denatured proteins. To measure a specific and delicate interaction force, we obtained force curves without pressing the probe onto GroEL molecules spread on a mica surface. Approximately 40 pN of tensile force was observed for approximately 10 nm while pepsin was pulled away from the chaperonin after a brief contact. This length of force duration corresponding to the circumference of GroEL's interior cavity was shortened by the addition of ATP. The relation between the observed mechanical parameters and the chaperonin's refolding function is discussed.

Adenosine Triphosphate↗

Human pituitary tumor transforming gene (hPTTG) inhibits human lung cancer A549 cell growth through activation of p21(WAF1/CIP1 ).

Pituitary tumor transforming gene (PTTG) is a proto-oncogene cloned from rat GH4 cells. This gene was able to induce cell transformation in vitro and is also associated with p53-dependent and -independent apoptosis. In this study, we cloned human PTTG (hPTTG) from a pituitary tumor and then stably transfected the hPTTG into HeLa and A549 cells. An overexpression of hPTTG significantly inhibited cell growth, which was determined by the adherent cell growth properties, colony formation in soft agar and [3H] thymidine incorporation, respectively, in HeLa and A549 cells. The inhibitory effect on cell growth was associated with the activation of p21WAF1/CIP1 in A549 cells, but not in HeLa cells. The hPTTG overexpression increased both the p21WAF1/CIP1 mRNA and protein expression levels as determined by both Northern and Western blot analysis, respectively, in A549 cells. The increased expression of p21WAF1/CIP1 mRNA was regulated at the transcription level and was independent on p53 expression because the luciferase activity increased after the co-transfection of hPTTG and p21WAF1/CIP1 promoter fragments with and without a p53 binding sequence. The subcellular distribution of hPTTG was dependent on cell type, and was predominantly in the nucleus in HeLa, Cos-7 and DU145 cells, but showed a diffuse distribution in both the nucleus and cytoplasm in A549, DLD-1 and NIH3T3 cells. These results indicate that an overexpression of hPTTG inhibits the cell growth due to different mechanisms, which are p21WAF1/CIP1 -dependent and -independent.

Animals↗

Expression of survivin during liver regeneration.

Survivin functions to suppress cell death and regulate cell division, and is observed uniquely in tumor cells and developmental cells. However, the expression and regulation of survivin in non-transformed cells are not well elucidated. Therefore, we investigated the expression of survivin in a murine liver regeneration model after partial hepatectomy and intraperitoneal carbon tetrachloride (CCl(4)) injection. We found that the expression of survivin transcript and protein were markedly elevated with the onset of DNA synthesis and remained elevated during G2 and M phases during liver regeneration. In a normal mouse liver cell line, over-expression of survivin resulted in a decrease in the G0/G1 phase and an increase in the S and G2/M phases, resulting in Rb phosphorylation. These findings suggest that survivin is dramatically expressed in a cell cycle-dependent manner during liver regeneration and provide a new insight into the regulation of cell proliferation and differentiation.

Animals↗

Effect of suplatast tosilate, a Th2 cytokine inhibitor, on cough variant asthma.

STUDY OBJECTIVE: Th2 cytokines play an important role in the pathogenesis of asthma. Our study objective was to determine the effect of suplatast tosilate, a Th2 cytokine inhibitor, on patients with cough-variant asthma. METHODS: Twenty patients with cough-variant asthma (CVA) were assigned to a suplatast tosilate (100 mg three times daily) group or a placebo group for 6 weeks in a double-blind randomized study. The cough scores, medication scores, pulmonary function, bronchial hyperresponsiveness to methacholine, cough threshold for capsaicin, percentage of eosinophils and concentrations of eosinophilic cationic protein (ECP) in hypertonic saline-induced (induced) sputum were evaluated. The main outcome measures were capsaicin cough threshold and concentrations of ECP in induced sputum. RESULTS: In the suplatast group, the cough scores and the medication scores decreased significantly over time. The percentage of eosinophils in induced sputum significantly decreased from 53.5+/-5.6% to 13.6+/-2.6%. The cough threshold for capsaicin improved significantly from 2.72+/-3.41 microM to 39.7+/-22.7 microM in the suplatast group. The concentrations of ECP in induced sputum decreased significantly from 435+/-123 microg/l to 56+/-34 microg/l in the suplatast group. The bronchial responsiveness to methacholine changed from 8.45+/-3.43 units to 11.4+/-3.76 units in the suplatast group. CONCLUSIONS: Suplatast improved the cough scores and the cough threshold for capsaicin in patients with CVA without significant side effects, suggesting the effectiveness of suplatast in the treatment of CVA. Suplatast also decreased the percentage of eosinophils and concentrations of ECP in induced sputum, suggesting improvement in eosinophilic inflammation in patients with CVA. Further pharmacodynamic research is needed to explain the precise mechanism.

Adult↗

Septic endophthalmitis and meningitis associated with Klebsiella pneumoniae liver abscess.

We report a female case of septic endophthalmitis and meningitis associated with Klebsiella pneumoniae liver abscess which was thought to be caused by duodeno-biliary reflux related to choledochoduodenostomy. We treated this patient by ultrasonography-guided percutaneous abscess drainage and intravenous administration of third generation antibiotics. However, the visual function of her left eye was eventually lost. Reports of liver abscess with metastatic lesions are rare, and our experience suggests that more physicians should be alert to septic metastatic lesions such as K. pneumoniae liver abscess or bacteremia with complaints of ocular or central nervous system symptoms.

Journal Article↗