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Biomedical subjects

K Ono

Publications and source records attributed to K Ono.

At least 505 records · Page 28Linked to original sources
PubMed1995

Specific detection of the precursor of ras p21 with a mouse monoclonal anti-C-terminal peptide antibody, SARA-K1.

In an attempt to clarify the post-translational modifications of ras oncogene product p21, we have established a mouse monoclonal antibody specific for the precursor of p21. The C-terminal peptide (156-188) of K(4A)-ras oncogene product p21 (p21K(4A), termed K(4A)-peptide, was used as the immunogen. In Western blotting, monoclonal antibodies were examined for their differential reactivity between two types of p21K(4A) expressed in Escherichia coli (esh-p21K(4A)) and mammalian cell (mam-p21K(4A)). One monoclonal antibody, designated SARA-K1, reacted selectively with esh-p21K(4A). The epitope for SARA-K1 was defined on tryptic peptide (177-184), containing Cys180, of the K(4A)-peptide. Pulse-chase experiments of mam-p21K(4A) synthesis at 24 degrees C revealed that SARA-K1 precipitated a 21 kDa protein within a 7 min chase but not after a 10 min chase, indicating that SARA-K1 recognizes the precursor of mam-p21K(4A). Furthermore, in Triton X-114 partitioning experiments using mammalian cells pre-treated with Mevalotin, 3-hydroxy-3-methylglutaryl coenzyme A (HMG CoA) reductase inhibitor, SARA-K1 precipitated [35S]methionine-labeled, [3H]mevalonic acid-unlabeled mam-p21K(4A) in the aqueous phase, but did not precipitate [3H]mevalonic acid-labeled mam-p21K(4A) in either aqueous or detergent phase. The data presented clearly show that the SARA-K1 specifically recognizes the primary translational product pro-p21K(4A).

Amino Acid Sequence↗
PubMed1995

Distribution of the basic fibroblast growth factor and its receptor gene expression in normal and degenerated rat intervertebral discs.

STUDY DESIGN: Using a rat spondylosis model, the distributions of cells expressing the basic fibroblast growth factor and its receptor were investigated in normal and degenerated intervertebral discs. Cell-proliferating activity in degenerated discs was also assessed. OBJECTIVES: This study was conducted to determine whether basic fibroblast growth factor is related to intervertebral disc degeneration. SUMMARY OF BACKGROUND DATA: Basic fibroblast growth factor stimulates proliferation and matrix synthesis of cultured intervertebral disc cells. METHODS: Immunohistochemistry and in situ hybridization histochemistry were conducted to detect cells with basic fibroblast growth factor-like immunoreactivity and fibroblast growth factor receptor messenger RNA, respectively. Cell-proliferating activity was evaluated by AgNOR staining. RESULTS: In degenerated discs, round chondrocytes with basic fibroblast growth factor-like immunoreactivity and fibroblast growth factor receptor messenger RNA are scattered instead of spindle-shaped cells in the normal anulus (normal anular cells), which are devoid of basic fibroblast growth factor-like immunoreactivity and fibroblast growth factor receptor messenger RNA. The proliferating activity of these chondrocytes is suggested to exceed that of normal anular cells. CONCLUSION: Basic fibroblast growth factor is suggested to promote proliferation of chondrocytes in degenerated discs in an autocrine or paracrine manner. Basic fibroblast growth factor may be related to intervertebral disc degeneration as a proliferation-stimulating factor of chondrocytes that replace normal anular cells during disc degeneration.

Animals↗
PubMed1995

A sustained inward current activated at the diastolic potential range in rabbit sino-atrial node cells.

1. After blocking both the hyperpolarization-activated current and the membrane K+ conductance, depolarizations from -80 mV to between -70 and -50 mV induced a sustained current in sino-atrial node cells. We have tentatively designated this current Ist. 2. Ist was blocked by both organic and inorganic Ca2+ channel blockers, but was insensitive to tetrodotoxin (30 microM). Isoprenaline increased Ist. 3. The peak of Ist (at about -50 mV) was increased to 149 +/- 13% (n = 8, P < 0.01) by lowering the external Ca2+ concentration ([Ca2+]o) from 1.8 to 0.1 mM, in contrast to the marked depression of the L-type Ca2+ current. In 0.1 mM [Ca2+]o solution, the nicardipine-sensitive current-voltage relation showed the peak amplitude at about -50 mV and a reversal potential of +37.4 +/- 4.4 mV (n = 5). The isoprenaline-sensitive component also showed a reversal potential of about +30 mV. 4. Reducing [Na+]o from 140 to 70 mM in 0.1 mM [Ca2+]o decreased Ist to 53 +/- 5% (n = 7, P < 0.01). Increasing [Ca2+]o or [Mg2+]o decreased the amplitude of Ist. The half-maximum concentration of Mg2+ inhibition was 2.2 mM. 5. At 1.8 mM [Ca2+]o, Ist was inactivated by clamping for 5s at -10 mV, and gradually recovered after repolarization to -80 mV with a half-time of 1.36 +/- 0.4 s (n = 6). 6. The transitional sino-atrial node cell had minimal amplitude of Ist. 7. These characteristics of Ist are qualitatively comparable to those of the monovalent cation conductance of the L-type Ca2+ channel induced by depleting external divalent cations to the micromolar range. We conclude that Ist is generated by a novel subtype of L-type Ca2+ channel.

Action Potentials↗
PubMed1995

The immune reaction to heterologous serum causes osteonecrosis in rabbits.

Osteonecrosis (ON) was produced experimentally in rabbits by intravenous injection of horse serum. Eighty adult rabbits were used: 16 were injected twice with isotonic saline (Group A), 24 were injected once with saline and once with horse serum (Group B), and 40 were injected twice with horse serum (Group C). Both femurs of each rabbit were obtained from 2 h to 7 weeks after the final injection an were subjected to histological examination. No pathological changes were seen in Groups A and B. In Group C, 5 of 15 rabbits (33%) showed ON (necrosis of trabecula and bone marrow) in the femoral metaphysis. In Group C, the early major pathological findings in bone marrow are extravasation of erythrocytes in sinusoidal spaces and microthrombi in small arteries and arterioles near the lesion of extravasation. Immune complexes were demonstrated in the kidney within 24 h of the final injection of horse serum. The present study suggests that immunological reaction associated with serum sickness may play an important role in inducible ON and this model will contribute toward clarifying the pathogenesis of ON.

Animals↗
PubMed1995

A direct effect of forskolin on sodium channel bursting.

A long-lasting component of current through voltage-dependent Na channels is believed to contribute to the plateau phase of the cardiac action potential. Here we report that in cardiac ventricular myocytes forskolin increases the contribution of a very slow component of decay (tau = 36 +/- 16 ms, n = 13) in ensemble currents in response to step depolarizations to 0 mV. Long-lasting bursts of openings (mean duration of 27 +/- 14 ms, n = 10) accounted for this behavior. The slow time constant of decay was not altered by forskolin (5-50 microM). Rather, an increase in the probability of bursting behavior produced a forskolin concentration-dependent increase in the amplitude of this very slow component. This action of forskolin was not the result of stimulation of adenylyl cyclase because it was not affected when cAMP-dependent phosphorylation was inhibited by the protein kinase inhibitor H-89, and it could not be mimicked by addition of isoproterenol, membrane-permeant cAMP [8-(4-chlorophenylthio)-cAMP], or the phosphatase inhibitor okadaic acid. In addition, bursting was not augmented by guanosine 5'-O-(3-thiotriphosphate) (GTP [gamma S]) either applied to the bath or directly to the intracellular face of the channel in inside-out macropatches. Furthermore, 1,9-dideoxy-forskolin, which does not stimulate adenylyl cyclase and 6-(3-dimethylaminopropionyl)-forskolin, a hydrophilic derivative of forskolin, also augmented late channel activity.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1995

G-protein activation enhances Ca(2+)-dependent lipid secretion of the rat harderian gland.

We studied the secretory mechanism of the Harderian gland of rats. After perfusion with HEPES-buffered Ringer's solution containing NaF (10 mM) with AlCl3 (10 microM), a G-protein activator, the glandular cells of the Harderian gland showed massive exocytosis and apocrine-like protrusions on the luminal surface. Some of the secretory vacuoles aggregated within the cytoplasm, and large vacuoles were formed. Contraction of the myoepithelial cells covering the glandular endpieces caused a narrowing of the glandular lumina, which contained cytoplasmic fragments, and deformation of the basal contour of the glandular end-pieces. The basal regions of the glandular cells also bulged between the myoepithelial cells. Secretory vacuoles were also discharged to the lateral cell surface, and the intercellular spaces were dilated. The enhanced secretory activities of the glandular cells and the contraction of the myoepithelial cells were similar to those in rats stimulated with 10 microM carbachol (CCh). However, dilatation of the endoplasmic reticulum in glandular cells (type A cells), which leads to the formation of small vesicles, was observed in those glands stimulated by NaF+AlCl3, but not in those stimulated by CCh. Removal of Ca2+ from the perfusing HR or addition of EDTA (0.5 mM) diminished and inhibited NaF+AlCl3- or CCh-enhanced secretory activity of the glandular cells and also allayed the deformation of glandular cells caused by myoepithelial cell contraction. The present results demonstrate the involvement of G-proteins and Ca(2+)-influx in the lipid secretion of glandular cells and in the contraction of myoepithelial cells of the Harderian gland in rats.

Aluminum Chloride↗
PubMed1995

Natural killing activity is associated with progression of gastric cancer.

Previously, we proposed a new analysis of natural killing activity, in which an individual effector/target cell ratio was employed for comparison according to the peripheral number of effector cells. In 51 patients with gastric cancer, the activity was studied using that modified analysis. Natural killing activity was activated in patients with early cancer, where tumor-cell invasion was restricted to the mucosa or the submucosa, even though in well-differentiated adenocarcinoma with invasion of the mucosa alone, the activity remained at the level of controls. In contrast, the activity in advanced cancer, where tumor cells infiltrated beyond the submucosa, came to be inactivated as the cancer progressed. These facts suggest that natural killing activity in patients with gastric cancer is closely associated with tumor invasion and that reactive activation of the activity against tumor is induced, at least, in some patients with early stage.

Adenocarcinoma↗
PubMed1995

Usefulness of electron microscopy in the diagnosis of cardiac sarcoidosis.

A 49-year-old man with cardiac sarcoidosis is presented. He suffered from congestive heart failure, and left ventricular asynergy and reduced function was evident by echocardiogram and left ventriculogram. A light microscopic examination of the endomyocardial biopsy revealed nonspecific myocarditis without giant cells or noncaseating granulomas. Under an electron microscope, however, several epithelioid cells were found in the specimen. The serum level of lysozyme was elevated. The patient had a past history of sarcoidosis of the eyes and lungs 22 years previously. Cardiac diseases presenting epithelioid cells other than sarcoidosis were clinically ruled out. Thus, the diagnosis of cardiac sarcoidosis was made based on both clinical and ultrastructural findings, and corticosteroid therapy was initiated. In the second biopsy, performed 4 months later, a noncaseating granuloma was found. Generally, the incidence of histological diagnosis of cardiac sarcoidosis by light microscopy is relatively low in endomyocardial biopsy specimens. The present case suggests that the addition of an ultrastructural examination may improve the diagnostic usefulness of the endomyocardial biopsy in cardiac sarcoidosis, since electron microscopy can clearly identify the presence of even one epithelioid cell.

Biopsy, Needle↗
PubMed1995

Carbamylcholine- and catecholamine-induced intracellular calcium dynamics of epithelial cells in mouse ileal crypts.

BACKGROUND/AIMS: The intestinal epithelium is composed of various cells, and the enteric nervous system, which controls the epithelial functions, has different neurotransmitters and/or modulators. The aim of this study was to show whether the responses of intestinal epithelial cells to different neurotransmitters are elicited throughout the entire epithelium or are restricted to a certain cell. METHODS: The spatiotemporal dynamics of cytosolic calcium ion ([Ca2+]c) were measured by digital imaging analysis in isolated crypts of mouse ileum loaded with [1-[2-(5'-carboxyoxazol-2'-yl)-6-amino-benzofuran-5- oxy]-2-(2'-amino-5'-methylphenoxy)-ethane-N,N,N',N'-tetraacetic acid] pentakis (acetoxylmethyl) ester. Thereafter, the crypt cells were identified morphologically. RESULTS: Carbamylcholine elicited [Ca2+]c dynamics in Paneth cells, showing a biphasic increase, but neither cholecystokinin octapeptide nor nicotine had any effect on the [Ca2+]c of the crypt cells including the Paneth cells. Adrenaline and noradrenaline, but not isoproterenol, induced a transient elevation of [Ca2+]c of some enterochromaffin cells. Increases in the [Ca2+]c of most crypt cells were elicited by thapsigargin. Propagation of a [Ca2+]c wave in the crypts was not evident. CONCLUSIONS: Increases in [Ca2+]c can be induced by carbamylcholine in Paneth cells and catecholamines in some enterochromaffin cells. The digital imaging analysis showed the heterogeneity of the responses of intestinal crypt cells to different transmitters.

Animals↗
PubMed1995

Gammopathy with two M-components in a dog with IgA-type multiple myeloma.

A 12-year neutered male mixed-breed dog was referred to hospital for evaluation of chronic diarrhea. Cellulose acetate electrophoresis of its serum revealed two monoclonal peaks in the gamma-globulin fraction. On immunoelectrophoretic analysis, the two monoclonal peaks in the gamma-globulin region were strongly precipitated with anti-dog IgA serum. On sodium dodecyl sulfate-polyacrylamide gel electrophoretic analysis, the fractions corresponding to these two peaks were shown to be dimer and trimer or tetramer of immunoglobulin consisting of heavy and light chains. These results indicated that the studied dog had gammopathy with two M-components with dimer and trimer or tetramer of IgA. Accumulations of large amounts of these immunoglobulins with very high molecular weight in the serum were concluded to induce the hyperviscosity syndrome in this dog in the terminal stage.

Animals↗
PubMed1995

Immunochemical characterization of recombinant and native tropomyosins as a new allergen from the house dust mite, Dermatophagoides farinae.

BACKGROUND: Two predominant mite species, Dermatophagoides farinae and Dermatophagoides pteronyssinus, are responsible for the immediate hypersensitivity reaction. Although a large number of antigens and allergens was detected in a whole mite culture extract, only the major allergens of Der I and II groups have been extensively studied. Much information on other important allergens remains to be accumulated. METHODS: A new allergen complementary DNA (cDNA) clone was isolated from a D. farinae protein cDNA library. The expression product of the cDNA and native tropomyosin from D. farinae were purified, characterized, and evaluated. RESULTS: An amino acid sequence deduced from the cDNA clone revealed significant homology with tropomyosins conserved in a wide range of animals. The amino acid sequences of two fragments obtained by degradation of the purified protein with cyanogen bromide coincided completely with the corresponding portions of the deduced amino acid sequence. The native tropomyosin reacted with specific IgE in the 31 sera tested at a high frequency (80.6%), comparable to that of Der f I (90.3%) and Der f II (74.2%). CONCLUSIONS: The cloned cDNA encodes tropomyosin. The high binding response of specific IgE antibodies to recombinant and native tropomyosins demonstrates that mite tropomyosin is an additional important allergen in house dust mite.

Allergens↗
PubMed1995

Sphingolipid biosynthesis is necessary for dendrite growth and survival of cerebellar Purkinje cells in culture.

The requirement of complex sphingolipid biosynthesis for growth of neurons was examined in developing rat cerebellar Purkinje neurons using a dissociated culture system. Purkinje cells developed well-differentiated dendrites and axons after 2 weeks in a serum-free nutrient condition. Addition of 2 microM fumonisin B1, a fungal inhibitor of mammalian ceramide synthase, inhibited incorporation of [3H]galactose/glucosamine and [14C]-serine into complex sphingolipids of cultured cerebellar neurons. Under this condition, the expression of Purkinje cell-enriched sphingolipids, including GD1 alpha, 9-O-acetylated LD1 and GD3, and sphingomyelin, was significantly decreased. After 2 weeks' exposure to fumonisin B1, dose-dependent measurable decreases in the survival and visually discernible differences in the morphology were seen in fumonisin-treated Purkinje cells. The Purkinje cell dendrites exhibited two types of anomalies; one population of cells developed elongated but less-branched dendrites after a slight time lag, but their branches began to degenerate. In some cells, formation of elongated dendrite trees was severely impaired. However, treatment with fumonisin B1 also led to the formation of spinelike protrusions on the dendrites of Purkinje cells as in control cultures. In contrast to the alterations observed in Purkinje cells, morphology of other cell types including granule neurons appeared to be almost normal after treatment with fumonisin B1. These observations indicated strongly that membrane sphingolipids participate in growth and maintenance of dendrites and in the survival of cerebellar Purkinje cells. Indeed, these effects of fumonisin B1 were reversed, but not completely, by the addition of 6-[[N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)-amino dcaproyl]sphingosine (C6-NBD-ceramide), a synthetic derivative of ceramide. Thus, we conclude that deprivation of membrane sphingolipids in a culture environment is responsible for aberrant growth of Purkinje cells.

Animals↗
PubMed1995

A new algorithm for predicting splice site sequence based on an improvement of categorical discriminant analysis.

The splicing signals which govern the excision of introns are not yet well explained, since actual splice site sequences are to some extent different from the generally accepted consensus sequences. While a quantification method (categorical discriminant analysis: CDA) has been proposed to analyze splice site signals, sample sequences lying in the overlapping region of sample scores are not discriminated well by CDA, thus limiting the predictive ability of this analytical technique. In this paper, we propose a method to improve the performance of CDA, which is applicable to the analysis of 5'-splice site signals in various mammalian genes. The proposed algorithm has revealed a distinct enhancement in the predicting ability compared to that of traditional CDA and could explain point mutations in the 5'-splice site region of rabbit beta-globin gene fairly well.

Algorithms↗
PubMed1995

Mitochondrial alcohol dehydrogenase from ethanol-grown Euglena gracilis.

The inducing effects of ethanol on alcohol dehydrogenase and the key enzymes of the glyoxylate cycle, isocitrate lyase and malate synthase, in Euglena cells were investigated. Ethanol as the sole carbon source resulted in increases in alcohol dehydrogenase and the two glyoxylate cycle enzymes. The experimental results indicated that ethanol is assimilated by alcohol dehydrogenase and the glyoxylate cycle in Euglena. Mitochondria from aerobically grown Euglena contain a unique type of alcohol dehydrogenase that accounts for their ability to respire with ethanol as a substrate. This alcohol dehydrogenase was purified to homogeneity from ethanol-grown Euglena gracilis. The mitochondrial alcohol dehydrogenase was NAD(+)-specific but not NADP(+)-specific. Ethanol was the most active substrate, but the enzyme was also active towards 1-butanol, 1-heptanol, cinnamyl alcohol, and myristyl alcohol. These results indicated that mitochondrial alcohol dehydrogenase participated in alcohol metabolism in Euglena gracilis.

Alcohol Dehydrogenase↗
PubMed1995

Alteration in the reactivity of sphingomyelin in mitogen-stimulated lymphocytes.

The antigens for a monoclonal antibody, VJ-41, established by alloimmunization of B10.A(3R) mice with lymphocytes from B10.A(5R) mice and screening of its reactivity toward Con A-stimulated human T lymphocytes, were found to be phosphorylcholine-containing ceramides (sphingomyelin) and disaturated fatty acyl glycerol (phosphatidyl-choline, PC), but neither deacylated sphingomyelin nor unsaturated fatty acid-containing PC reacted with the antibody. Although the reactivity of disaturated fatty acyl PC increased with increasing chain length, that of sphingomyelin was stronger than that of di-20:0-PC. The binding of the antibody to Con A-stimulated lymphocytes was inhibited by sphingomyelin-containing liposomes, but not by di-18:0-PC-containing ones, and the concentration of sphingomyelin in Con A-stimulated human T-lymphocytes was the same as that in non-stimulated ones, indicating that the reactivity of sphingomyelin in lymphocytes is altered by Con A-stimulation.

Animals↗
PubMed1995

Isolation and characterization of sialodacryoadenitis virus (coronavirus) from rats by established cell line LBC.

The outbreak of sialoadenitis occurred in a laboratory rat colony and the causative agent was isolated from the affected salivary glands of diseased rats using the established cell line LBC. The isolate readily multiplied, producing clear cytopathic effects with syncytium formation, and it was identified virologically and serologically as rat sialodacryoadenitis virus. In attempts to isolate the virus by primary rat kidney (PRK) cells and suckling mice as well as LBC cells, the LBC cells showed higher susceptibility for the virus growth as compared with PRK cells or the brain of suckling mice. The isolation rate of virus was 100% (5/5) in LBC, 40% (2/5) in PRK cells and 60% (3/5) in suckling mice. After four passages in the LBC cells, the virus did not produce disease in adult rats, while the mouse brain-passaged virus did.

Animals↗