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Biomedical subjects

A Wada

Publications and source records attributed to A Wada.

At least 415 records · Page 23Linked to original sources
PubMed1986

Inhibition of Na+-pump enhances carbachol-induced influx of 45Ca2+ and secretion of catecholamines by elevation of cellular accumulation of 22Na+ in cultured bovine adrenal medullary cells.

In bovine adrenal medullary cells, we reported that 22Na+ influx via nicotinic receptor-associated Na+ channels is involved in 45Ca2+ influx, a requisite for initiating the secretion of catecholamines (Wada et al. 1984, 1985 b). In the present study, we investigated whether the inhibition of Na+-pump modulates carbachol-induced 22Na+ influx, 45Ca2+ influx and catecholamine secretion in cultured bovine adrenal medullary cells. We also measured 86Rb+ uptake by the cells to estimate the activity of Na+,K+-ATPase. Ouabain and extracellular K+ deprivation remarkably potentiated carbachol-induced 22Na+ influx, 45Ca2+ influx and catecholamine secretion; this potentiation of carbachol-induced 45Ca2+ influx and catecholamine secretion was not observed in Na+ free medium. Carbachol increased the uptake of 86Rb+; this increase was inhibited by hexamethonium and d-tubocurarine. In Na+ free medium, carbachol failed to increase 86Rb+ uptake. Ouabain inhibited carbachol-induced 86Rb+ uptake in a concentration-dependent manner, as it increased the accumulation of cellular 22Na+. These results suggest that Na+ influx via nicotinic receptor-associated Na+ channels increases the activity of Na+,K+-ATPase and the inhibition of Na+,K+-ATPase augmented carbachol-induced Ca2+ influx and catecholamine secretion by potentiating cellular accumulation of Na+. It seems that nicotinic receptor-associated Na+ channels and Na+,K+-ATPase, both modulate the influx of Ca2+ and secretion of catecholamines by accommodating cellular concentration of Na+.

Adrenal Medulla↗
PubMed1986

Elution-band relaxation method. A method to analyze isomerization kinetics by HPLC and application to protein denaturation-renaturation.

We present a novel method, the 'elution-band relaxation method', to analyze quantitatively reversible isomerization kinetics by elution chromatography, taking advantage of the high resolution and speed of high-performance liquid chromatography (HPLC). The kinetic information is obtained by measuring the first temporal moments of chromatograms of molecules undergoing isomerization and analyzing their dependence on the column length or flow rate. The major advantage of this method is that it is applicable to reactions as fast as the time of elution in HPLC, a speed which has not been attained previously in analysis of isomerization reactions based on the chromatographic property of molecules. We describe the method and report an experimental application to the denaturation-renaturation kinetics of bovine pancreatic ribonuclease A as an example.

Chromatography, High Pressure Liquid↗
PubMed1986

Effect of calmodulin on aldosterone synthesis by a cytochrome P-45011 beta-reconstituted system from bovine adrenocortical mitochondria.

Bovine adrenocortical calmodulin was purified and its general properties were examined. The latter were similar to those of bovine brain calmodulin. When added to a cytochrome P-450(11)beta-reconstituted system in the presence of dilauroylphosphatidylcholine, calmodulin decreased the rate of aldosterone production from corticosterone from 0.8 to 0.1 nmol/(min X nmol P-450), while it increased the rate of 18-hydroxycorticosterone production from 1.8 to 4.6 nmol/(min X nmol P-450). This effect of calmodulin on steroid production was maximum at a concentration of 1 microM, when 1 microM cytochrome P-450(11)beta was used. The effect was dependent on the presence of Ca2+, and maximal response was observed at less than 1 microM Ca2+. There was essentially no difference in the effect when bovine brain calmodulin was used. Calmodulin induced a change in the activity of cytochrome P-450(11)beta in the presence of a wide concentration range of corticosterone as a substrate. As for 18-hydroxycorticosterone production, calmodulin increased both the maximal activity and the apparent Km for corticosterone, but it decreased the apparent Km for adrenodoxin. Adrenodoxin at a concentration of less than 20 microM did not fully abolish the effect of calmodulin. A small type I difference spectrum appeared when calmodulin was added to cytochrome P-450(11)beta. The difference spectrum increased significantly in the presence of both Ca2+ and adrenodoxin. These results suggest that calmodulin interacts with cytochrome P-450(11)beta in the presence of adrenodoxin and then modulates the activity of aldosterone synthesis catalyzed by cytochrome P-450(11) beta.

Adrenal Cortex↗
PubMed1986

Analysis of Escherichia coli ribosomal proteins by an improved two dimensional gel electrophoresis. I. Detection of four new proteins.

Kaltschmidt and Wittmann's two dimensional gel electrophoresis was improved in the following points. Preruns using radical scavengers were carried out to eliminate free radicals remaining in gels. Gelation of sample solutions was not performed to avoid immobilization of proteins in the sample gels. Instead, for preparing sample gels, prior to the first dimension (1-D) electrophoresis, another electrophoresis was performed to charge proteins into gel pieces polymerized previously. Proteins migrated together with charged reductants to avoid formation of artificial disulfide bridges during migration. The second dimension (2-D) electrophoresis was carried out at a more acidic pH, 3.6, to get better separation of very small and basic proteins. With these modifications, quantitative yield and reproducibility became better, and many faint spots disappeared not only at the high molecular weight side but also in the region containing primary spots of ribosomal proteins. The proportionality of the migration distance to the logarithm of molecular weight was also increased. On the improved 2-D electrophoretogram of Escherichia coli ribosomal proteins, four new spots, called protein A, B, C, and D, were found in the basic region. Proteins A, B, and C belong to 50S subunits but D to 30S. Their molecular weights were determined electrophoretically as 6,400, 4,900, 8,200, and 5,900, respectively. Their copy numbers in crude ribosomes were estimated to be 0.6, 0.4, 0.3, and 0.1, respectively, by using 14C-labeling.

Bacterial Proteins↗
PubMed1986

Analysis of Escherichia coli ribosomal proteins by an improved two dimensional gel electrophoresis. II. Characterization of four new proteins.

Four new proteins, A, B, C, and D, found in Escherichia coli ribosomes by an improved two dimensional gel electrophoresis were characterized by oxidation, reduction, and carboxymethylation of cysteine residues, and CsCl fractionation. The cysteine contents of proteins A, B, C, and D were determined to be 1 +/- 0, 3 +/- 1, 5 +/- 1, and 0 +/- 0 by carboxymethylation with iodoacetic acid. The components of protein complexes, which formed numerously under non-reducing conditions, were analyzed. Including protein A, B, and C, every ribosomal protein (r-protein) having cysteine residue(s) except unconfirmed S1 was proved to form such complexes with various combinations. The cysteine residue in protein A, in particular, was highly reactive to make intermolecular S-S bridges so that spot A almost disappeared on the second dimension gel under the non-reducing conditions. Proteins B and C shifted their spots by reduction towards upper left side as do all known r-proteins having plural cysteine residues except S1. This suggests that proteins B and C change their conformation by intramolecular S-S bridges. The CsCl density gradient centrifugation of high salt washed 70S ribosomes showed that protein A belonged to the insoluble split proteins, proteins B and C to the core particles, and protein D and a small population of B to the soluble split proteins. The electrophoretic behaviors, CsCl fractionation and stoichiometry of the four new proteins suggested strongly that they were intrinsic ribosomal constituents different from known ribosomal proteins or factors.

Bacterial Proteins↗
PubMed1986

Halothane inhibits the cholinergic-receptor-mediated influx of calcium in primary culture of bovine adrenal medulla cells.

Adrenal medulla cells are cholinoceptive cells. Stimulation of the acetylcholine receptor causes the influx of Ca to the cells, and Ca acts as the coupler of the stimulus-secretion coupling. In this study, the authors investigated the effects of halothane on the receptor-mediated influx of 45Ca using cultured bovine adrenal medulla cells. Halothane at clinical concentrations (0.5-2%) inhibited the influx of 45Ca caused by carbachol, with simultaneous inhibition of catecholamine secretion. The influx of 45Ca and the secretion of catecholamines caused by K depolarization were inhibited by a large concentration of Mg, which competes with Ca at Ca channels, but not inhibited by halothane. Inhibition of the 45Ca influx by halothane was not overcome by increase in the carbachol concentration. Inhibition of the 45Ca influx by halothane was examined in comparison with that caused by a large concentration of Mg by the application of Scatchard analysis as the function of the external Ca concentration. Halothane decreased the maximal influx of 45Ca without altering the apparent kinetic constant of Ca to Ca channels. On the contrary, a large concentration of Mg increased the apparent kinetic constant without altering the maximal influx of 45Ca. Based on these findings, the authors suggest that inhibition of the 45Ca influx by halothane was not due to the direct competitive inhibition of Ca channels, nor to the competitive antagonism of agonist-receptor interaction. As a possibility, halothane seems to inhibit the receptor-mediated activation of Ca channels through the interference of coupling between the receptor and Ca channels.

Adrenal Medulla↗
PubMed1986

A new method for DNA surface-property display and its application to E. coli promoter sequences.

A personal computer program to visualize and compare the property of double-stranded DNA surface has been developed. Comparison of the surface property between Watson-Crick base-pairs in B-form DNA has elucidated that the base-pair replacement between a "degenerated base-pairs" conserves the pattern of potential hydrogen-bonding sites in both major and minor grooves. The idea of the "degenerated base-pairs" was applied for the problem of the base-sequence variation from the consensus sequence in the -35 region of E. coli promoter. The sequence variation is found to have tendency to occur among the degenerated base-pairs.

Computer Graphics↗
PubMed1985

Intra-arterial infusion chemotherapy as an induction therapy in multidisciplinary treatment for locally advanced breast cancer. A long-term follow-up study.

A multidisciplinary treatment including intra-arterial infusion chemotherapy as an induction therapy was administered to 55 patients with locally advanced breast cancer. Intra-arterial chemotherapy conducted preoperatively produced marked responses in primary and lymph node lesions with 78% complete + partial response (CR + PR), subsequently permitting extended radical mastectomy. Histologic examination of resected specimens also revealed that 33% of the patients had no viable cancer cells remaining in their lesions. Five-year and 10-year survival rates were 57% and 41%, respectively, compared with 24% and 18%, respectively for the 17 patients of historic control. Patients showing better local responses to intra-arterial chemotherapy had longer survival time with less frequent local recurrences. Intra-arterial chemotherapy is an effective modality for the treatment of locally advanced breast cancer.

Aclarubicin↗
PubMed1985

Neuronal control of brain microvessel function.

Cerebral capillary endothelium forms a barrier limiting and controlling the movement of ions and solutes between blood and brain. Recent anatomical, physiological and biochemical studies have suggested the possibility that capillary function may be directly controlled by neuronal structures. Alterations in neuronal systems involved in the regulation of microcirculation may account for microvascular dysfunctions which occur in different pathologic conditions.

Acetylcholine↗
PubMed1985

Gene expression in the polycistronic operons of Escherichia coli heat-labile toxin and cholera toxin: a new model of translational control.

A new model is proposed based on the suggestion that stable local secondary structures of mRNA interfere with ribosome movement on mRNA and consequently reduce the translation rate. This model accounts for a different level of translation for each cistron in the polycistronic mRNA of Escherichia coli heat-labile toxin (LT) and cholera toxin. We also conclude that the mRNA secondary structures have been conserved during the evolution of the toxin genes for its functional importance.

Bacterial Toxins↗
PubMed1985

Modulation by ouabain and diphenylhydantoin of veratridine-induced 22Na influx and its relation to 45Ca influx and the secretion of catecholamines in cultured bovine adrenal medullary cells.

The effects of ouabain and diphenylhydantoin on the secretion of catecholamines induced by veratridine were investigated in cultured bovine adrenal medullary cells with special reference to ion fluxes. Veratridine itself induced an influx of 22Na and 45Ca as well as secretion of catecholamines, which were antagonized by tetrodotoxin, a selective inhibitor of voltage dependent Na channels. The secretion of catecholamines caused by veratridine was not observed either in Na free or Ca free medium. Veratridine-induced influx of 45Ca did not occur in Na free medium, while veratridine-induced influx of 22Na occurred even in Ca free medium. Veratridine-induced influx of 22Na, 45Ca and secretion of catecholamines were all potentiated by ouabain, a potent inhibitor of Na,K-ATPase. Omission of K from the medium, a condition which suppresses the Na,K-ATPase activity, also augmented these cell responses caused by veratridine. On the contrary, diphenylhydantoin, which is known to decrease the intracellular concentration of Na, reduced the veratridine-induced influx of 22Na, 45Ca and secretion of catecholamines. The potentiating effects of ouabain on the veratridine-induced cell responses were all abolished by diphenylhydantoin. These findings imply that veratridine, ouabain and K removal as well as diphenylhydantoin modulate the intracellular accumulation of 22Na which is involved in the influx of 45Ca and the secretion of catecholamines.

Adrenal Medulla↗
PubMed1985

Heredofamilial factor as a cause of myocardial damage.

The existence in an inbred strain of Syrian hamster (BIO 14.6) of a genetically transmitted cardiomyopathy (CM) that results in congestive heart failure offers unique opportunities to study causes of myocardial damage and the sequence of events leading to cardiac insufficiency. It is well known that the clinical course and pathologic aspects of the cardiac condition of the CM hamster resemble a type of idiopathic CM in man. This paper presents the findings obtained from light- and electron-microscopic observations of the CM heart muscle of the BIO 14.6 hamster. The investigation included qualitative X-ray micro-analysis of intra- and extracellular calcific deposits in the damaged myocardium, biochemical data of calcium-activated neutral protease in cardiomyolysis, and morphometric analysis for cardiac hypertrophy. It is concluded that the myocardial damage is due to disturbed biosynthesis of contractile components and the cellular membrane system. This defect, being accompanied by persistent hypoxia together with a large amount of Ca2+ and Ca-activated protease, leads progressively to necroses and scar formation with calcification in the cardiomyopathic heart muscle of the left (and later also the right) ventricle. It also leads to hypertrophy as a compensatory mechanism of unaffected cardiomyocytes and ultimately to fatal heart failure.

Age Factors↗
PubMed1985

A hapten-induced conformational change accompanies the cryoprecipitation of an immunoglobulin.

By simultaneous spectroscopic measurements (absorbance, light scattering, fluorescence and circular dichroism) the temp-dependent behavior of a cryoglobulin, the mouse monoclonal immunoglobulin G to N-acetyllactosamine, has been observed. The results show that a hapten-induced conformational change in the structure of a cryoglobulin can accompany its intermolecular association. This association of the immunoglobulin increases after hapten saturation of its two antigen-combining sites.

Amino Sugars↗
PubMed1985

Influx of 22Na through acetylcholine receptor-associated Na channels: relationship between 22Na influx, 45Ca influx and secretion of catecholamines in cultured bovine adrenal medulla cells.

The effects of carbachol, veratridine and high K on the influx of 22Na were investigated in relation to the influx of 45Ca and the secretion of catecholamines in cultured bovine adrenal medulla cells, in which stimulation of nicotinic but not muscarinic acetylcholine receptor causes the secretory response. (1) Carbachol caused a rapid influx of 22Na, influx of 45Ca and secretion of catecholamines, all of which occurred within 1 min and leveled off thereafter. Influx of 45Ca and secretion of catecholamines caused by carbachol were not inhibited by tetrodotoxin, but were greatly reduced in Na-free medium. Nicotine evoked an influx of 22Na and it was antagonized by hexamethonium and d-tubocurarine but not by tetrodotoxin. Muscarine had no effect on 22Na influx. The concentration-response curve of carbachol for 22Na influx was quite similar to that for 45Ca influx. (2) Veratridine induced a sustained influx of 22Na, influx of 45Ca and secretion of catecholamines, all of which were antagonized by tetrodotoxin. Influx of 45Ca and secretion of catecholamines due to veratridine were not observed in Na-free medium. (3) High K caused an influx of 45Ca and secretion of catecholamines but did not cause an influx of 22Na. High K-induced influx of 45Ca and secretion of catecholamines were not inhibited by tetrodotoxin nor by Na removal. (4) Magnesium, an inhibitor of voltage-dependent Ca channels, inhibited the influx of 45Ca and secretion of catecholamines caused by carbachol, veratridine and high K. These results indicate that cultured bovine adrenal medulla cells have at least three distinct ion channels: (1) nicotinic acetylcholine receptor-associated Na channels which are not inhibited by tetrodotoxin, (2) voltage-dependent Na channels which are kept activated by veratridine and inhibited by tetrodotoxin and (3) voltage-dependent Ca channels. Influx of Ca through voltage-dependent Ca channels is the common ionic event for the secretion of catecholamines caused by either carbachol, veratridine or high K. It seems that the influx of Na through acetylcholine receptor-associated Na channels as well as voltage-dependent Na channels, activates voltage-dependent Ca channels which triggers the secretion of catecholamines.

Adrenal Medulla↗
PubMed1985

Probing stability and dynamics of proteins by protease digestion. I: Comparison of protease susceptibility and thermal stability of cytochromes c.

Protease susceptibility of homologous proteins in their native conformations was studied. This work aims to establish a broad and quantitative basis for the utilization of protease digestion to analyze the local stability of native proteins. Using high-performance liquid chromatography (HPLC) the time course of the proteolytic degradation of intact proteins was quantitatively traced. Rapid separation of peptide fragments with HPLC made possible the elucidation of sequential digestion originating from the cleavage at a very few sites which are locally unstable in the protein structure. Using four serine proteases, chymotrypsin, trypsin, elastase and subtilisin BPN', we found some common trends in proteolysis for a group of proteins of the cytochrome c family. By comparing of the proteolysis and thermal denaturation with ten homologous cytochromes c extracted from horse, beef, Candida krusei, Saccharomyces cerevisiae, chicken, tuna, pigeon, rabbit, dog and rat, protease susceptibility was related to locally unfolding states intrinsic to the native conformation.

Animals↗
PubMed1985

Histocytologic diagnosis of pancreatic cancer by percutaneous aspiration biopsy under ultrasonic guidance.

Percutaneous aspiration biopsy of the pancreas using a heparinized 22-gauge fine needle was performed under ultrasonic guidance in five patients with benign pancreatic diseases and in 18 patients with pancreatic cancer. Using a heparinized needle and syringe, it was possible to make good smears containing abundant tumor cells and to obtain small tissue specimens. Using egg albumin as binding material, a new cell-block technic was developed to conveniently obtain histologic specimens. In this way, a correct diagnosis was made cytologically in all 23 patients suspected of having a pancreatic malignancy. Histologic specimens were obtained in 22 (95.6%) our of 23 patients. A correct diagnosis was established histologically in all patients from whom histologic materials were obtained. This procedure thus has proved a very reliable method for diagnosing pancreatic cancer.

Adenocarcinoma↗