Biomedical subjects

T Aoki

Publications and source records attributed to T Aoki.

At least 523 records · Page 29Linked to original sources

PubMed1996

Primary culture of chicken hepatocytes in serum-free medium (pH 7.8) secreted albumin and transferrin for a long period in free gas exchange with atmosphere.

To study liver functions of chicken, we examined the primary culture of chicken hepatocytes, and found an easy method of long-term culture with free atmosphere exchange. Chicken hepatocytes were obtained by collagenase perfusion and cultured at 37 degrees C as a monolayer without substratum in serum-free L-15 medium (pH 7.8) with free atmosphere exchange. The amounts of albumin and transferrin in medium were assayed by ELISA. The culture of chicken hepatocytes was maintained in the serum-free L15-medium )pH 7.) and 37 degrees C with free atmosphere exchange for 20 days. The amount of albumin secreted in the medium decreased to low levels early in culture; however, this was followed by marked increase from day 9 to day 17 of culture. The amount of transferrin was constant until day 6, then it too increased with further culture. We reported an easy method for the simple monolayer culture of chicken hepatocytes in serum-free L12 medium (pH 7.8) with free atmosphere exchange over an extended period. Expression of liver-specific functions, viz. albumin and transferrin synthesis, was observed after 1 week of culture.

Air↗

PubMed1996

Purification of pepsinogens from human urine and electrophoretic analysis by caseogram print.

Pepsinogen (PG) A and C were purified from human urine, and analyzed by a highly sensitive detection method, "caseogram print". Purification was achieved by a series of conventional chromatographies and FPLC. A relatively large amount (13.2 mg) of PGA was purified from about 20 liters of urine. Purified PGA was separated by a Mono-Q column into each of its isozymogens. The elution order (PGA-5, 4+3, 2) corresponded to the order of electrophoretic migration. Although the concentration of urinary PGC was very low, a trace amount was purified and visualized by electrophoresis. The urinary and mucosal PGCs migrated at the same position, and urinary PGC was detected as two isozymogens similarly to mucosal PGC, suggesting that urinary and mucosal PGCs may be essentially identical.

Chromatography, Gel↗

PubMed1996

Modification of human neuron-specific enolase for application to radioimmunoassay.

A recombinant human neuron-specific enolase (R-NSE), isolated from Escherichia coli, could not be used in an RIA system because of instability upon labeling. To apply R-NSE to RIA and to simplify the purification procedure, the N- and C-terminals of R-NSE were modified by tyrosine- and histidine-tagging, respectively. SY-NSE, containing one additional tyrosine residue, was obtained from both soluble and insoluble fractions. More derivatives tagged by two or four tyrosine residues were expressed, but only in the insoluble fraction. SY-NSE and SY-NSE.H6 (containing six histidine residues at C-terminal of SY-NSE) purified from the soluble fraction were applicable to the RIA system, indicating that the addition of a tyrosine residue at the terminal is effective if the antigen is unstable during labeling.

Base Sequence↗

PubMed1996

Sulfonamide resistance gene in a transferable R plasmid of Pasteurella piscicida.

The sulfonamide resistance (SAr) determinant was cloned from a transferable R plasmid of Pasteurella piscicida, pSP9351, and the sequence was determined. The resistance gene (pp-sul) was localized to an approximately 1-kb region that includes the PstI-EcoRI site in the restriction map. An open reading frame coding a sul II-type gene composed of 810 nucleotides was identified. A direct repeat sequence was shown in the 5' flanking region of pp-sul, and a plasmid recombinational event may have occurred during the construction of pSP9351. In the 3' flanking region of the gene, a sequence homologous to the 5' noncoding sequence of the trimethoprim resistance gene, dhfr IX was found.

Amino Acid Sequence↗

PubMed1996

Sequence analysis of the florfenicol resistance gene encoded in the transferable R-plasmid of a fish pathogen, Pasteurella piscicida.

The florfenicol resistance gene (pp-flo) derived from a transferable R-plasmid of Pasteurella piscicida consisted of 1,122 nucleotides, and the predicted amino acid sequence showed 47.4% identity to that of a non-enzymatic chloramphenicol resistance gene (cmlA). The pp-flo gene was located in the downstream region of the sulfonamide resistance gene of the transferable R-plasmid.

Amino Acid Sequence↗

PubMed1996

Depression among cancer patients.

This study was done to investigate the frequency of co-morbidity and to demonstrate the best method for assessing depression among cancer patients. The subjects were 50 (25 male and 25 female) cancer patients and 50 (25 male and 25 female) medically ill patients. All subjects were interviewed by psychiatrists and were administered psychological tests such as SAS (self-rating anxiety scale), SDS (self-rating depression scale), POMS (Profile of Mood States), HADS (Hospital Anxiety and Depression Scale) and DRP (Depression-related personality traits). The psychiatric interview revealed that 44% of cancer patients and 38% of the medical patients had mental disorders according to DSM-IV. The most frequently observed disorder was depression, which was seen in 28% of the cancer patients and 30% of the medical patients. The cancer patients with depression scored significantly higher on the DRP and the Anger mood state of POMS than did the medically ill patients with depression. In addition, most psychological tests employed had no discrimination between depressed and normal subjects among the cancer and the medical patients. However, it was found that the Depression scale in HADS (HADS-D) split depressed patients from normal subjects since the HADS-D was composed of items that were not concerned with physically ill conditions.

Adult↗

PubMed1996

Hypoxanthine-guanine phosphoribosyltransferase gene analysis for Japanese patients with Lesch-Nyhan syndrome.

The Lesch-Nyhan syndrome results as a consequence of a severe deficiency of functional activity of purine salvage enzyme, hypoxanthine phosphoribosyltransferase (HPRT). We performed Southern blot analysis for five patients and their families using full length cDNA of the HPRT gene as a probe. Pst I digested Southern blot analysis revealed a large deletion that included exon 2 in patient 3. The size of this deletion was about 4.4 Kb. The mother of this patient had the same mutated allele and a normal one (heterozygote). This type of mutation from a Lesch-Nyhan syndrome patient has not been previously reported. The restriction fragment length polymorphism (RFLP) pattern was analyzed by Bam HI digested Southern blot analysis for one family who had no major gene abnormality. We determined from this analysis that the sister of the patient was a Lesch-Nyhan syndrome carrier and the fetus (brother) was normal for HPRT activity. This study shows RFLP analysis is still useful for carrier detection and prenatal diagnosis of Lesch-Nyhan syndrome.

Blotting, Southern↗

PubMed1996

Common acute lymphoblastic leukemia preceded by hypercalcemia in an infant.

A case of common acute lymphoblastic leukemia (ALL) preceded by hypercalcemia was observed in an infant. A high blood level of parathyroid hormone related protein (PTH-rP) was detected during the first examination at Ohashi Hospital. Treatment with antineoplastic agents led to complete remission. The blood PTH-rP level decreased as a result of treatment. PTH-rP produced by leukemic cells was though to be the cause of the hypercalcemia.

Antineoplastic Combined Chemotherapy Protocols↗

PubMed1996

Advantage of monitoring skin vibration as an index of arteriovenous fistular blood flow.

Monitoring of the skin vibration is disturbed little by environmental noise. Therefore, the skin vibration is more relevant than bruit on the skin by means of monitoring the fistular blood flow. Frequency analysis of the skin vibration generated by an internal arteriovenous fistula indicated two peaks of power spectral density (PSD), one in the frequency range of 4-10 Hz and the other from 100-300 Hz. The analysis of the skin vibration generated by an external fistula, however, showed only one peak in the range of 4-10 Hz and a moderate flat PSD level in the range of 100-300 Hz. The PSD level in the range of 100-300 Hz decreased dramatically or disappeared when the fistular blood flow diminshed or ceased. Therefore, the optimal frequency for monitoring skin vibration may be the range of 100-300 Hz.

Arteriovenous Fistula↗

PubMed1996

[Analysis of verotoxin-producing Escherichia coli (O157:H7) strains isolated in the Fukuoka area in 1994 by pulsed-field gel electrophoresis].

Nine verotoxin-producing Escherichia coli O157:H7 strains were isolated from 9 pediatric patients with sporadic enteritis between July and September 1994 at four clinics in the Fukuoka area. The patients included two families with two cases each. These strains were analyzed by pulsed-field gel electrophoresis for Xba I-digested DNA fragments. The restriction patterns were identical between each two strains within the two family outbreaks, but different among the seven strains of the distinct seven sporadic cases. It is strongly suggested that the seven sporadic cases were infected through distinct sources, and that the two family cases were due to a common source of infection or person to person infection.

Bacterial Toxins↗

PubMed1996

Inhibition of Trypanosoma cruzi growth in mammalian cells by purine and pyrimidine analogs.

Trypanosoma cruzi, the causative agent of Chagas' disease, exhibits two different developmental stages in mammals, the amastigote, an intracellular form that proliferates in the cytoplasm of host cells, and the trypomastigote, an extracellular form that circulates in the bloodstream. We have already established an in vitro culture system using mammalian host cells (HeLa) infected with T. cruzi in which the time course of parasite growth is determined quantitatively. We adopted this system for the screening of anti-T. cruzi agents that would ideally prove to be effective against trypanosomes with no toxicity to the host cell. Of the purine analogs tested, allopurinol markedly inhibited the growth of amastigotes in a dose-dependent manner, with no lethal effect on trypomastigotes. 3'-Deoxyinosine and 3'-deoxyadenosine also suppressed T. cruzi growth inside the host cell, with the concentrations causing 50% growth inhibition being 10 and 5 microM, respectively, in contrast to a concentration causing 50% growth inhibition of 3 microM for allopurinol. Among the pyrimidine analogs examined, 3'-azido-3'-deoxythymidine (zidovudine) significantly reduced the growth of the parasite at concentrations as low as 1 microM. The anti-human immunodeficiency virus agents 2',3'-dideoxyinosine and 2',3'-dideoxyadenosine caused a decrease in amastigote growth, while 2',3'-dideoxycytidine and 2',3'-dideoxyuridine had no inhibitory effect. When Swiss 3T3 fibroblasts were used as host cells, allopurinol, 3'-deoxyinosine, 3'-deoxyadenosine, and 3'-azid-3'-deoxythymidine also markedly inhibited T. cruzi proliferation. These results indicate that our culture system is useful as a primary screening method for candidate compounds against T. cruzi on the basis of two criteria, namely, intracellular replication by the parasite and host-cell infection rate.

3T3 Cells↗

PubMed1996

Endothelial modulation of pH-dependent pressor response in isolated perfused rabbit lungs.

With the use of isolated perfused rabbit lungs (n = 152), roles of endothelium-derived relaxing factor (EDRF) in pulmonary vascular responses to hypocapnia and hypercapnia were studied. Lungs were ventilated with a gas mixture containing 1, 5, or 10% CO2 and 21% O2, adjusting the perfusate pH to 7.8, 7.4, or 7.1, respectively. Methemoglobin (MetHb), hemoglobin (Hb), methylene blue (MB), and L-argininosuccinic acid (L-ASA) were used as modulators of EDRF. To eliminate augmented shear stress, we used papaverine during hypercapnia. As a measure of EDRF, we spectrophotometrically examined nitric oxide (NO) metabolites in the perfusate. Hypocapnia and hypercapnia evoked, respectively, unsustainable vasodilatation and vasoconstriction. Hb, MB, and L-ASA, but not MetHb, produced an increase in baseline pulmonary arterial pressure (Ppa). These agents also exacerbated vasoconstriction during hypercapnia. Hypercapnia and hypocapnia caused an increase and decrease, respectively, in EDRF production. L-ASA suppressed EDRF production in hypercapnic lungs. Papaverine did not suppress EDRF production under hypercapnia. In conclusion, 1) the effects of pH on pulmonary circulation are transient, 2) the increase in Ppa caused by hypercapnia is modulated by EDRF, and 3) the pulmonary EDRF genesis is activated by hypercapnic acidosis but suppressed by hypocapnic alkalosis.

Animals↗

PubMed1996

Lung tissue behavior in the mouse during constriction induced by methacholine and endothelin-1.

Recently, mice have been extensively used to investigate the pathogenesis of pulmonary disease because appropriate murine models, including transgenic mice, are being increasingly developed. However, little information about the lung mechanics of mice is currently available. We questioned whether lung tissue behavior and the coupling between dissipative and elastic processes, hysteresivity (eta), in mice would be different from those in the other species. To address this question, we investigated whether tissue resistance (Rti) and eta in mice would be affected by varying lung volume, constriction induced by methacholine (MCh) and endothelin-1 (ET-1), and high-lung-volume challenge during induced constriction. From measured tracheal flow and tracheal and alveolar pressures in open-chest ICR mice during mechanical ventilation [tidal volume = 8 ml/kg, frequency (f) = 2.5 Hz], we calculated lung resistance (RL), Rti, airway resistance (Raw), lung elastance (EL), and eta (= 2piRti/EL). Under baseline conditions, increasing levels of end-expiratory transpulmonary pressure decreased Raw and increased Rti. The administration of aerosolized MCh and intravenous ET-1 increased RL, Rti, Raw, and EL in a dose-dependent manner. Rti increased from 0.207 +/- 0.010 to 0.570 +/- 0.058 cmH2O.ml-1.s after 10(-7) mol/kg ET-1 (P < 0.01). After induced constriction, increasing end-expiratory transpulmonary pressure decreased Raw. However, eta was not affected by changing lung volume, constriction induced by MCh and ET-1, or high-lung-volume challenge during induced constriction. These observations suggest that 1) eta is stable in mice regardless of various conditions, 2) Rti is an important fraction of RL and increases after induced contriction, and 3) mechanical interdependence may affect airway smooth muscle shortening in this species. In mammalian species, including mice, analysis of eta may indicate that both Rti and EL essentially respond to a similar degree.

Airway Resistance↗

PubMed1996

Modulation of hypoxic pulmonary vasoconstriction by antioxidant enzymes in red blood cells.

To determine whether antioxidant mechanisms within red blood cells (RBCs) significantly contribute to preserving hypoxic pulmonary vasoconstriction (HPV) in both the absence and the presence of oxidative stress, we investigated HPV changes in isolated rabbit lungs perfused with solutions containing RBCs treated with various inhibitors of superoxide dismutase (SOD), anion channels, catalase (CAT), or glutathione peroxidase (GSH-Px). Perfusion was maintained at a constant flow rate of 70 ml/min, and lung temperature at 37 to 38 degrees C. Hematocrit was adjusted to 7%. In the absence of overt oxidative stress, HPV was significantly enhanced in the perfusate containing control RBCs (untreated RBCs) as compared with that in Krebs-Henseleit buffer. Inhibition of SOD, CAT, and GSH-Px within RBCs, as well as anion channels located on the RBC membrane, had little influence on HPV. Neither exogenous SOD nor CAT altered HPV. In the presence of high levels of reactive oxygen species (ROS), generated by addition of xanthine (100 microM) and xanthine oxidase (10 mU/ml) to the reservoir, HPV was considerably suppressed in the perfusate containing only buffer but was restored in the perfusate with control RBCs. Inhibition of CAT or GSH-Px in RBCs preserved the HPV, whereas inhibition of SOD or anion channels failed to preserve HPV obtained during exposure to high ROS levels. Addition of exogenous SOD, but not CAT, to the perfusate containing RBCs in which endogenous SOD had been inhibited restored HPV under high ROS conditions. In conclusion, (1) although RBCs augment HPV in the absence of ROS, this finding is not attributable to the antioxidants in RBCs. (2) RBCs restore HPV upon exposure to high ROS. This finding may well be explained by antioxidant mechanisms operating within RBCs, especially those of endogenous SOD.

Animals↗

PubMed1996

Roles of calcitonin gene-related peptide (CGRP) in hyperpnea-induced constriction in guinea pigs.

It has been reported that hyperpnea-induced bronchoconstriction in guinea pigs is a potential model for exercise-induced asthma in humans. We hypothesized that calcitonin gene-related peptide (CGRP) could modulate leukotriene D4 (LTD4)-induced responses and be involved in the pathophysiology in this asthma model. We measured tracheal (Ptr) and alveolar pressure (PA) using alveolar capsules in open-chested, mechanically ventilated (f = 1 Hz, VT = 9 ml/kg, PEEP = 4 cm H2O) guinea pigs. Animals were intravenously pretreated with saline (SAL), CGRP(8-37) (CGRP receptor antagonist), CGRP, MK-571 (LTD4 receptor antagonist), MK-886 (5-lipoxygenase inhibitor), or CGRP(8-37) + MK-571, and then underwent dry gas hyperpnea challenge (HC, 95% 02-5% CO2, 150 breaths/min, 7 min). We calculated resistance of lung (RL), tissue (Rti), and airway (Raw). HC increased RL, Rti, and Raw in SAL controls (322 +/- 27, 430 +/- 59, 299 +/- 23% baseline, respectively). MK-571, MK-886, and CGRP significantly reduced the responses to HC, while CGRP(8-37) enhanced HC-induced responses. Pretreatment with CGRP(8-37) and MK-571 in combination attenuated HC-induced constriction. In addition, pretreatment with CGRP reduced responses induced by intravenous administration of LTD4. These observations suggest that CGRP might be involved in the pathophysiology of hyperpnea-induced constriction in guinea pigs via modulation of LTD4-elicited responses.

Animals↗

PubMed1996

Modulation of ICAM-1 expression by extracellular glutathione in hyperoxia-exposed human pulmonary artery endothelial cells.

To investigate the mechanisms regulating hyperoxia-induced intercellular adhesion molecule-1 (ICAM-1) expression, we studied the effects of antioxidants on ICAM-1 expression, and the relationship between ICAM-1 expression and extracellular glutathione levels in human pulmonary artery endothelial cells (HPAEC) and human umbilical vein endothelial cells (HUVEC). Cells were cultured to confluence and exposed to hyperoxia (90% O2) for 48 h with or without various antioxidants, including superoxide dismutase (SOD), catalase, N-acetylcysteine (NAC), and glutathione. The levels of ICAM-1 expression in the endothelial cells and the concentrations of reduced (GSH) and oxidized glutathione (GSSG) in the media were examined by flow cytometry and spectrophotometry, respectively. After exposure to hyperoxia, ICAM-1 expression was increased, and the supernatant total glutathione was decreased as compared with those at normoxia. SOD did not change ICAM-1 expression. The hyperoxia-induced increase in ICAM-1 expression was even greater with the addition of catalase. The ICAM-1 expression was decreased and the GSH concentration was increased with the addition of NAC. There were negative relationships between the level of ICAM-1 expression and the supernatant total glutathione concentration in catalase-treated HPAEC (R = 0.822, P < 0.0005) and HUVEC (R = 0.567, P < 0.01). Negative relationships were also demonstrated between the level of ICAM-1 expression and the total extracellular glutathione concentrations in NAC-treated HPAEC (R = 0.877, P < 0.0005) and HUVEC (R = 0.727, P < 0.0005). Exogenous GSH decreased ICAM-1 expression in both hyperoxia-exposed HPAEC and HUVEC, while exogenous GSSG did not. These results suggest that extracellular GSH plays a role in regulating hyperoxia-induced ICAM-1 expression in HPAEC and HUVEC.

Acetylcysteine↗

PubMed1996

Endothelin-1 has a priming effect on production of superoxide anion by alveolar macrophages: its possible correlation with bronchopulmonary dysplasia.

The purpose of this study was to clarify the role of endothelin (ET)-1 in the development of bronchopulmonary dysplasia (BPD). Tracheal aspirates were obtained from 27 newborn babies with respiratory distress (13 with BPD and 14 without BPD) who were mechanically ventilated. Production of superoxide anion (O2-) by rabbit alveolar macrophages was determined by preincubation with the tracheal aspirate supernatant (TAS) and stimulation with phorbol myristate acetate (PMA). O2- production was demonstrated only when PMA was added to the experimental system and was enhanced with TAS of infants who later developed BPD compared with TAS from infants without BPD. The effects of ET-1 and ET-3 on O2- production and the blockade by anti-ET-1 antibody and BQ123 (ET A receptor antagonist) were also examined. The enhancing effect was blocked by either anti-ET-1 antibody or BQ123. PMA-stimulated production of O2- increased when cells were preincubated with several doses of ET-1 (5 x 10(-13) to 2 x 10(-12) M), whereas ET-3 was without effect. TAS contained significant amounts of immunoreactive ET-1, and there was a close positive correlation (r = 0.764) between the activity of O2- production and immunoreactive ET-1 levels in TAS samples. These results may be interpreted to indicate that ET-1 synthesized by and secreted from tracheal epithelial cells and/or alveolar macrophages has a priming effect on alveolar macrophages to produce O2-, thus possibly contributing to the development of BPD.

Albumins↗

PubMed1996

Relationship between the 13C-NMR chemical shift and the carcinogenic activity of benz[a]anthracenes.

A thorough NMR spectroscopic investigation was made on 7,12-dimethylbenz[a]anthracene (1) and the 1H- and 13C-NMR spectra were completely assigned. We compared the 13C-NMR chemical shifts (delta) of three benz[a]anthracenes, including compound 1, whose carcinogenicities vary largely, and investigated the relationship of the delta and carcinogenicity. It was found that the delta values at the C1, C2, and C6 positions differed significantly between the carcinogenic and noncarcinogenic compounds.

Benz(a)Anthracenes↗