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Biomedical subjects

M Komori

Publications and source records attributed to M Komori.

At least 73 records · Page 4Linked to original sources
PubMed1992

Stable expression of cytochrome P450IIIA7 cDNA in human breast cancer cell line MCF-7 and its application to cytotoxicity testing.

A mammalian cell expression plasmid containing cytochrome P450IIIA7 complementary DNA was constructed. Breast cancer cells (MCF-7) were transfected with the plasmid and neomycin-resistant selection marker plasmid. We established three cell lines, termed M13, M21, and M27, which expressed the cytochrome P450IIIA7 as examined by RNA blot and immunoblot analyses. These cell lines showed 8- to 10-fold higher sensitivity against aflatoxin B1 compared to parental MCF-7 cells, suggesting that cytochromes P450IIIA7 expressed in the cells were responsible for the production of the cytotoxic metabolite of aflatoxin B1.

Adult↗
PubMed1992

Toxicological significance of dog liver cytochrome P-450: examination with the enzyme expressed in Saccharomyces cerevisiae using recombinant expression plasmid.

A complementary DNA (cDNA) coding for a form of beagle dog cytochrome P-450 (Dah1), which is the orthologue to the CYP1A1 cDNA of rat, mouse and human, was inserted between the alcohol dehydrogenase (ADH) promoter and terminator regions of the yeast expression vector pAAH5. On introduction of the resulting recombinant plasmid pDC-1, Saccharomyces cerevisiae strain AH22 cells synthesized up to 1.5 x 10(5) molecules per cell of cytochrome P-450 protein (P-450(Dah1)). The carbon monoxide-bound reduced form of P-450(Dah1) showed an absorption peak at 447 nm and specific content of P-450(Dah1) was about 0.1 nmole P-450 per mg of microsomal protein. P-450(Dah1) cross-reacted with antibodies to rat P-448-H (CYP1A2) and dog P-450-D2 (CYP1A2). P-450(Dah1) activated 2-amino-3-methyl-imidazo[4,5-f]quinoline (IQ) and 2-amino-3,4-dimethylimidazo[4,5-f]quinoline (MeIQ) most efficiently in the umu test and exhibited a high activity of aryl hydrocarbon hydroxylase toward benzo[a]pyrene.

Animals↗
PubMed1992

Dynamic light scattering studies on hydrodynamic properties of fibrinogen-fibronectin complex.

A high molecular weight 'cryogel' was obtained as insoluble complexes by cold incubation at near-freezing temperatures from heparinized plasma of patients with rheumatoid arthritis. After the cryogel was solubilized at 37 degrees C, 1:1 complex of fibrinogen and fibronectin was purified at room temperature by affinity chromatography on a gelatin-Sepharose 4B. Hydrodynamic properties of the complex were investigated as a function of temperature and NaCl concentration using a dynamic light scattering. The diffusion coefficients of the complex at 20 degrees C decreased with increasing of NaCl concentration as free fibronectin. The complex appears to be a more compact form at low ionic concentration, which is associated with conformational changes of fibronectin. The diffusion coefficient of the complex at 20 degrees C in 0.05 M TrisHCl(pII7.4) containing 0.5 M NaCl was estimated as 8.5 x 10(-8) cm2s-1. The complex did not dissociate over the temperature range from 20 to 37 degrees C. The diffusion coefficients of the complex decreased significantly at 12 degrees C and 40 degrees C. The thermal denaturation of fibrinogen molecule in the complex was observed at 40 degrees C. The CONTIN analysis of the light scattering data showed that the complex associated to form higher aggregates at 15 degrees C, but not at near-freezing temperature. The equilibrium between the complex and higher aggregates appeared reversible.

Diffusion↗
PubMed1992

Quantitative assessment of digitized portal images: effect of sampling frequency on observer performance.

To investigate spatial resolution requirements for digitized portal images in radiation therapy, observer performance tests were performed. One hundred twenty portal images were digitized with sampling frequencies of 0.700, 0.350, and 0.175 mm for observation. Receiver operating characteristic analysis was used to determine the acceptable sampling frequency for clinical portal images. The detectability of setup errors was significantly better on the original images than on the digitized images with sampling frequencies of 0.700 mm (P = .005) and 0.350 (P = .046). Some clinical disadvantages might accrue with the use of a sampling frequency of 0.350 mm or larger.

Humans↗
PubMed1992

Human fetal liver cytochrome P-450: capacity to form genotoxic metabolites.

Unlike most experimental animals, human fetal liver possesses forms of cytochrome P-450. Thus, the purpose of this study was to clarify the toxicological significance of these forms of cytochrome P-450 to understand possible roles of these cytochromes in producing genotoxic metabolites from promutagens. In fact, human fetal livers showed considerable capacity to activate aflatoxin B1 and IQ (2-amino-3-methylimidazo [4,5-f] quinoline). Three of four forms of cytochrome P-450, P-450HFLa-d, which we could purify from human fetal livers were capable of activating promutagens to mutagens. One of these three forms, namely P-450HFLa, catalyzed the metabolic activation of aflatoxin B1 and IQ. An expression plasmid containing HFL33 cDNA encoding P-450HFLa was constructed and the protein expressed in insect (Sf9) cells and in human cancer cells, MCF-7. Aflatoxin B1 was efficiently activated to a mutagen upon addition of the lysate of Sf9 cells to the incubation mixture for the assay. Transformants of MCF-7 cells expressing P-450IIIA7 (HFLa) showed higher sensitivity to aflatoxin B1 than the parental MCF-7 cells as detected by cytotoxicity.

Animals↗
PubMed1992

Six-base deletion occurring in messages of human cytochrome P-450 in the CYP2C subfamily results in reduction of tolbutamide hydroxylase activity.

We isolated and expressed a clone, hPA6, possibly corresponding to the CYP2C9 cDNA. Compared with the other CYP2C9 cDNA clones, hPA6 showed a 6-nucleotide deletion near its middle. From the same cDNA library, we could also isolate another cDNA clone, named hPA22, which retained the 6 bases. For clarification of the effect of the 2-amino acid deletion resulting from the 6-base deletion on enzymatic activities, both clones were expressed in yeast. The expressed enzymes showed tolbutamide hydroxylase activities, and these activities were inhibited by antibodies against P-450-HM2, a probable CYP2C9. The activity of the enzyme encoded by hPA6 was lower than that encoded by hPA22; thus the 2-amino acid deletion in the CYP2C9 reduced the enzymatic activity.

Amino Acid Sequence↗
PubMed1992

[A case report of a burn produced by the probe of a pulse oximeter].

We experienced a case of burn produced by the pulse oximeter probe at the finger of a patient. Patient was a 75-year old man scheduled to transurethral prostatectomy. The pulse oximeter probe made by Physio-Control Co. was attached at his finger and was connected to a pulse oximeter manufactured by Ohmeda Co. Soon after the start of Spo2 measurement the anesthetist became aware of abnormal heating of the probe and removed it immediately. But unfortunately a small thermal injury was formed at patient's finger. Abnormally excessive current went through the detector diode and heated the finger probe. Attention to correct connection between the pulse oximeter probe and the finger is necessary before use.

Aged↗
PubMed1991

Effect of protonation on the isomerization properties of n-butylamine Schiff base of isomeric retinal as revealed by direct HPLC analyses: selection of isomerization pathways by retinal proteins.

Alumina adsorption chromatography and ion-pair reversed-phase chromatography were developed to analyze the isomers of unprotonated and protonated n-butylamine Schiff base of retinal (RSB and PRSB), respectively. Photoisomerization starting from the all-trans, 11-cis and 13-cis isomers was traced for RSB in n-hexane, acetonitrile, methanol and 1-butanol, and for PRSB in methanol, acetonitrile and 1-butanol. The quantum yields of photoisomerization for the all-trans, 9-cis, 11-cis and 13-cis isomers were determined for RSB and PRSB in the above solvents except 1-butanol. On the other hand, photoisomerization of isomeric retinal bound (through Schiff base linkage) to bovine serum albumin (RBSA) in aqueous solution (pH 3, 7 and 12) as well as thermal isomerization of RSB (in n-hexane), PRSB (in methanol) and RBSA (in aqueous solution, pH 7) were traced starting from the all-trans, 11-cis, and 13-cis isomers. Protonation of RSB drastically changes the pathway of photoisomerization and increases the quantum yields of isomeric RSB. The solvent polarity increases the quantum yields of RSB differently depending on the configuration. Protonation enhances thermal isomerization also. The results of the above model systems are compared with those of retinal proteins to rationalize their selection of the particular isomerization pathways.

Butylamines↗
PubMed1991

Sulphate conjugation enhances reversible binding of drug to human serum albumin.

Reversible binding of model compounds, their conjugated metabolites (sulphates and glucuronides), and also derivatives of the compounds, to human serum albumin (HSA) has been examined using an ultrafiltration method. p-Nitrophenol (p-NP), alpha-naphthol (alpha-NA) and beta-naphthol (beta-NA) were used as model compounds. Reversible binding of 500 microM p-NP sulphate to 4% HSA (96.6 +/- 0.35%, mean +/- s.d. n = 3) was significantly higher (P less than 0.001), whereas reversible binding of p-NP glucuronide to 4% HSA (33.3 +/- 9.82%) was much lower (P less than 0.001) than that of 500 microM p-NP (90.9 +/- 0.60%). Reversible binding of 500 microM p-NP glucopyranoside to 4% HSA (25.8 +/- 2.82%) was comparable with that of the glucuronide, with which it is structurally similar. In contrast, reversible binding of 500 microM p-NP phosphate, an anionic compound like p-NP sulphate, to 4% HSA (61.4 +/- 5.28%) was significantly lower than that of p-NP (P less than 0.001). Similar results were observed in reversible binding of sulphates of alpha-NA and beta-NA. Significant differences of dissociation constants for HSA binding were observed between the parent compound (alpha- or beta-NA) and its sulphate conjugate (P less than 0.005 for alpha-NA and alpha-NA sulphate, P less than 0.001 for beta-NA and beta-NA sulphate), but the number of binding sites was the same. These results indicated that sulphate conjugation enhances reversible binding of a parent compound to HSA by increasing the binding affinity of the parent compound to HSA. This enhancement appeared to be advantageous for preventing random distribution of this metabolite to organs in the body.

Acetaminophen↗
PubMed1991

Nicorandil reduces the basal level of cytosolic free calcium in single guinea pig ventricular myocytes.

Using fluorescent Ca2+ indicator fura-2 and whole-cell patch-clamp techniques, we examined the effect of 2-nicotinamidoethyl nitrate (nicorandil) on the intracellular free Ca2+ concentration ([Ca2+]i) and electrical properties in single guinea pig ventricular myocytes. Nicorandil (10 nM approximately 1 mM) reduced the resting level [Ca2+]i monitored as fura-2 fluorescence ratio in a concentration-dependent manner. Dibutyryl guanosine 3':5'-cyclic monophosphate (cyclic GMP), a membrane permeable cyclic GMP analogue, mimicked the nicorandil action. Neither application of caffeine (10 mM) nor deprivation of extracellular Na+ ions could prevent the nicorandil action on [Ca2+]i. In contrast, the nicorandil effect was virtually blocked by sodium orthovanadate (40 microM), a Ca2+ pumping ATPase inhibitor. During electrophysiological experiments, nicorandil shortened action potential durations (205 +/- 80 ms to 153 +/- 76 ms) by increasing a glibenclamide-sensitive outward K+ conductance. However, the drug produced little hyperpolarization (approximately 2 mV) because the resting potential of ventricular myocytes was close to the K+ equilibrium potential. The involvement of voltage-dependent Ca-channel current and Na-Ca exchanger was considered to be minimal under physiological conditions. It is thus concluded that nicorandil decreases basal [Ca2+]i via cyclic GMP-mediated activation of the plasma membrane Ca2+ pump in guinea pig ventricular myocytes.

Animals↗
PubMed1991

Molecular toxicology on cytochrome P-450 in human fetal livers.

Among mammalian species, humans and monkeys are sole species which possess cytochrome P-450 in the liver in fetal life. Cytochrome P-450 catalyzes oxidations of a wide variety of compounds including steroids, drugs and toxicants. Thus, the purpose of this study was to examine the toxicological significance of human fetal cytochrome P-450. Firstly, we found that human fetal livers were capable of activating numbers of carcinogenic compounds. Secondly, the capacity of human fetal cytochrome P-450 to activate mutagens and carcinogens was confirmed using purified forms of cytochrome P-450. For this, we purified four forms of cytochrome P-450 from human fetal livers. Lastly, we could isolate a cDNA clone coding for P-450HFLa which is one of the major forms of cytochrome P-450 in human fetal livers. Utilizing synthesized cD-NAs as probes, we found that this form of cytochrome was expressed specifically in fetuses.

Biotransformation↗
PubMed1990

Fetus-specific expression of a form of cytochrome P-450 in human livers.

The developmentally regulated expression of forms of cytochrome P-450, namely, those encoded by lambda HFL33 and NF25 or HLp cDNAs, which were isolated from respective fetal and adult human liver cDNA libraries, was investigated. When EcoRI fragments of cDNA clones of lambda HFL33 and NF25 were used as probes, these probes hybridized with RNA from both fetal and adult human livers. However, when oligonucleotides specific to the coding and 3'-noncoding region of lambda HFL33 (oli-HFL and oli-HFL3', respectively) were used as probes, these probes gave hybridizable bands with RNA from fetal but not adult livers. On the other hand, an oligonucleotide probe specific to the coding region of NF25 and HLp (oli-NF) gave positive bands with RNA only from adult livers. These results indicate that P-450(HFL33) is expressed specifically in fetal livers and that neither P-450NF nor HLp is expressed in fetal livers, but one or both are expressed in adult livers.

Adult↗
PubMed1990

Isolation of cDNAs coding for three different forms of liver microsomal cytochrome P-450 from polychlorinated biphenyl-treated beagle dogs.

Three different cDNA clones, namely DM1-1, Dah1, and Dah2, encoding hepatic cytochrome P-450, were isolated from a cDNA library in lambda gt11 constructed from liver RNA of polychlorinated biphenyl-treated beagle dogs. DM1-1 was 1857 base pairs (bp) long and encoded a polypeptide of 457 residues. Dah1 was 2394 bp long and contained an entire coding region for 524 amino acid residues. In addition, Dah2 was 1623 bp long and had an open reading frame consisting of 503 amino acid residues, although it lacked the translational initiation codon. Judging from the similarity of the nucleotide and amino acid sequences, forms of cytochrome P-450 encoded by DM1-1, Dah1, and Dah2 were judged to belong to the P450IIC, P450IA1, and P450IA2 subfamilies, respectively. Northern blot analysis of RNA from various tissues, using the specific 3' noncoding regions of Dah1 and Dah2 as probes, indicated that mRNAs for P-450(Dah1) and P-450(Dah2) were not detectable in tissues from untreated dogs, except for P-450(Dah2) in livers. Polychlorinated biphenyl induced both mRNAs in liver, kidney, and lung, especially in the kidney.

Amino Acid Sequence↗
PubMed1990

[Effect of ulinastatin on microcirculation under hemodilution and hypothermia].

Microcirculation during hemodilution and hypothermia under ether anesthesia treated with or without ulinastatin was studied by means of rabbit ear chamber. Sixteen rabbits were divided into 2 groups; 8 rabbits without ulinastatin treatment (C group) and 8 rabbits treated with ulinastatin (U group). Surface cooling to 20 degrees C and rewarming were performed. No significant differences between the groups were found in blood pressure and heart rate, but pulse pressure in U group was significantly larger (P less than 0.01) than that in C group. Pulmonary edema occurred in 3 rabbits in C group during rewarming. The results suggest that ulinastatin inhibited edema formation in U group. In C group, the blood flow velocity was very slow and the blood flow rate was very small at 20 degrees C in arterioles and venules. In U group the flow velocity and the flow rate were well maintained at 20 degrees C in arterioles and venules. It is concluded that ulinastatin is effective in maintaining microcirculation during hemodilution and hypothermia.

Animals↗
PubMed1989

Purification of cytochrome P-450 from polychlorinated biphenyl-treated crab-eating monkeys: high homology to a form of human cytochrome P-450.

Cytochrome P-450, designated as P-450-MK2, was purified to an electrophoretic homogeneity from polychlorinated biphenyl (PCB)-treated female crab-eating monkeys. P-450-MK2 catalyzed nifedipine and nilvadipine oxidations, at a rate comparable to human P-450-HM1. The N-terminal amino acid sequence of P-450-MK2 was highly homologous to those of P-450-HM1 and NF 25. The antibodies to P-450-HM1 recognized P-450-MK2 and effectively inhibited the activity of testosterone 6 beta-hydroxylase in monkey liver microsomes. These results suggest that a form of cytochrome P-450 corresponding to human P-450-HM1 or P-450NF which belongs to the P450 III gene family is also present in liver microsomes of crab-eating monkeys.

Amino Acid Sequence↗
PubMed1989

Effects of serum testosterone level with buserelin on the activities of drug and testosterone hydroxylase and on the content of a male-specific form of cytochrome P-450 in male rats.

The effects of administration of buserelin, a synthetic agonist of lutenizing hormone-releasing hormone (LH-RH), on the content of P-450-male, a male specific form of cytochrome P-450, and the activities of drug and testosterone hydroxylases were examined in liver microsomes of male rats. Administration of buserelin resulted in a decrease in the serum level of testosterone in a dose-dependent manner. Similar decreases were seen in the activities of aminopyrine N-demethylase, 7-propoxycoumarin O-depropylase and testosterone 2 alpha- and 16 alpha-hydroxylases, but not in the activity of aniline p-hydroxylase. Thus, correlations were observed between the serum level of testosterone and the activities of drug and testosterone hydroxylases except for aniline p-hydroxylase. These results indicate that the drug and testosterone hydroxylases are affected sensitively and sequentially by the changes in the testosterone levels in the body.

Aminopyrine N-Demethylase↗
PubMed1989

Interspecies homology of liver microsomal cytochrome P-450. A form of dog cytochrome P-450 (P-450-D1) crossreactive with antibodies to rat P-450-male.

P-450-male is a male specific form of cytochrome P-450 in rat liver microsomes. Cytochrome P-450 crossreactive with anti-P-450-male antibodies was purified to an electrophoretical homogeneity from liver microsomes of male beagle dogs. The specific content of the purified cytochrome P-450 (P-450-D1) was 16.9 nmol/mg protein. The apparent monomeric molecular weight of P-450-D1 was 48,000, which was smaller than P-450-male (51,000). P-450-D1 showed similarities in spectral properties, N-terminal amino acid sequence, and catalytic activities with some limited exceptions: P-450-D1 did not catalyze 2 alpha-hydroxylation of testosterone and progesterone and catalyzed 21-hydroxylation of progesterone. Based on these results, we propose that P-450-D1 is a form of cytochrome P-450 in the same gene subfamily as P-450-male.

Amino Acid Sequence↗