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Biomedical subjects

M Hamaguchi

Publications and source records attributed to M Hamaguchi.

At least 217 records · Page 12Linked to original sources
PubMed1985

Action of cobra venom cardiotoxin on chick embryonal fibroblasts transformed with a temperature-sensitive mutant of Rous sarcoma virus.

The cytolytic action of cardiotoxin analogue III from the venom of the Formosan cobra on chick embryonal fibroblasts transformed with a temperature-sensitive mutant of Rous sarcoma virus was investigated. The 50% effective dose of the toxin for the cells cultured at a non-permissive temperature (41 degrees C) or for noninfected normal cells was about 8 micrograms/ml whereas the value was 2 micrograms/ml for the cells cultured at a permissive temperature (36 degrees C). This indicates that the transformed cells became more susceptible to the cytolytic action of the toxin than the non-transformed cells.

Animals↗
PubMed1985

Expression of meta-vinculin associated with differentiation of chicken embryonal muscle cells.

We confirmed the existence of meta-vinculin, which cross-reacts immunologically with vinculin and has a slightly higher molecular weight than the latter. The immunological cross-reactivity between meta-vinculin and vinculin was confirmed with monoclonal antibodies against vinculin. Furthermore, we found that this protein is present only in either smooth or striated muscle, but is absent in non-muscular tissues and that the expression of this protein is associated with a differentiation of muscle cells either in vivo or in vitro. Microinjection experiments of fluorescently labelled vinculin and/or meta-vinculin into the cytoplasm of cultured myotubes suggested that meta-vinculin may be localized at sites similar to vinculin in muscle cells.

Animals↗
PubMed1985

Transcriptive complex of Newcastle disease virus. II. Structural and functional assembly associated with the cytoskeletal framework.

The transcriptive complex, the nucleocapsid with the viral RNA-synthesizing activity, of Newcastle disease virus (NDV) contains three protein components, the major structural subunit (NP) and two associated proteins (P and L) involved in the RNA synthesis. We studied the pathway of these proteins from synthesis to assembly into the complex by pulse-chase labeling of infected cells followed by detergent extraction of the cells to separate soluble and cytoskeletal fractions. Most molecules of NP and P (and probably L) became associated with the cytoskeletal framework immediately after their synthesis. Most of the remaining molecules were initially found in the soluble fraction, but joined the cytoskeletal framework within several minutes. Once attached, none of the proteins left the cytoskeleton, and it was here that they assembled with 50 S viral RNA into nucleocapsids. The nucleocapsids thus formed remained bound to the cytoskeletal framework and were found by in situ autoradiography to exhibit viral RNA synthesis on the framework. These results suggested that the cytoskeletal framework could actively participate in the structural and functional assembly of NDV transcriptive complex.

Animals↗
PubMed1984

Protein kinase associated with Newcastle disease virus.

Purified virions of Newcastle disease virus (NDV) were found to contain protein kinase activity which was, like other virion-associated kinases, stimulated by Mg2+, and totally independent of Ca2+ and cAMP. The kinase phosphorylated preferentially the P and NP polypeptides of NDV. Triton-KCl fractionation of the virions has shown that the protein kinase activity may be associated with glycoprotein-free subviral particles, but not with nucleocapsids containing either only NP or some L and P proteins together with NP as protein constituent.

Newcastle disease virus↗
PubMed1983

Monoclonal antibodies to the HN glycoprotein of Newcastle disease virus. Biological characterization and use for strain comparisons.

We have isolated nine hybridomas secreting monoclonal antibodies directed against the HN glycoprotein of D26 strain of Newcastle disease virus (NDV) and classified them into three groups by their biological reactivities. The group I antibodies from six clones had high hemagglutination inhibition (HI) and neutralization titers, and exhibited neuraminidase inhibition (NI) activity with substrates, both bovine fetuin and N-acetyl neuraminlactose, although the levels of maximum inhibition were greater with the former substrate. The group II antibodies from two clones also had high HI activity but showed a strongly substrate-dependent NI, with efficient inhibition against fetuin and a total lack of inhibition against neuraminlactose. These antibodies had neutralization capacity but the titers were much lower than those of group I antibodies. One clone (group III) secreted antibody which exhibited HI and NI with either substrate but no detectable neutralization activity. Frequency of isolation of antigenic variants and competitive binding assays have shown that group I and group II antibodies may recognize distinct regions not overlapping each other whereas antibodies placed within each of the two groups are to the respective same overlapping regions. The site of group III seemed to be close to or overlap the group I determinant but distinct from group II region. Using the isolated monoclonal antibodies we compared seven heterologous strains by HI and neutralization tests as well as by enzyme-linked immunosorbent assay. The results indicated that the regions recognized by group I antibodies would be conserved among the strains whereas those by the other two antibody groups might undergo considerable changes.

Antibodies, Monoclonal↗
PubMed1983

The uncoating of paramyxoviruses may not require a low pH mediated step.

The effect of lysosomotropic agents, chloroquine and NH4Cl, on the replication of paramyxoviruses was compared with that on the other enveloped RNA viruses whose uncoating is known to be blocked by the agents. Under the conditions where vesicular stomatitis or influenza virus infection was strongly inhibited at an early step, the agents exhibited little inhibitory effect on the progress of infection by Newcastle disease virus and Sendai virus, allowing viral primary transcription to occur and neither inhibiting the virus production significantly nor reducing the number of infected cells. These results are incompatible with the previous report showing that paramyxovirus and the other enveloped RNA viruses may have in common an intracellular step sensitive to lysosomotropic agents (Miller and Lenard, Proc. Nat. Acad. Sci. USA 78, 3605-3609, 1981) and suggest that paramyxovirus uncoating may not require a low pH mediated lysosomal step.

Ammonium Chloride↗
PubMed1983

Transcriptive complex of Newcastle disease virus. I. Both L and P proteins are required to constitute an active complex.

Virions of Newcastle disease virus (NDV) were disrupted with Triton X-100 in the presence of high salt and nucleocapsids were isolated by ultracentrifugation. The nucleocapsids had very low transcriptase activity and contained only NP as a prominent protein constituent, the bulk of L and P proteins not being retained. The L and P proteins were isolated by sequential treatment of the virions with low- and high-salt detergent followed twice by successive chromatography on phosphocellulose column and examined for their effect on RNA synthesis in a standard transcriptase system using the nucleocapsids as template. When both L and P proteins were added to the template, the RNA synthetic activity was greatly stimulated. P protein alone could not enhance but rather suppressed the activity. L protein exhibited stimulation to some extent but due to residual small amount of P protein in both L protein fraction and the template it has not been elucidated whether L protein could function as a polymerase by itself. These results indicate that both L and P proteins are required to reconstitute a fully active transcriptive complex with a functional template. Attempts have been made to isolate intracellular transcriptive complex from NDV-infected MDBK cells and to determine the protein species involved. The active complex has been recovered neither from cytoplasmic extract obtained by hypotonic disruption nor from Triton X-100 soluble fraction of the cells. However, we could isolate the complex from an extract by double detergents (Tween 40 and deoxycholate) solubilization. The complex contained L, P, and NP as virus specific proteins and several cellular proteins. These results support the concept that both L and P proteins are required for NDV-RNA synthesis and suggest further that the intracellular transcriptive complex may be associated with some cellular structure resistant to Triton X-100 but sensitive to the double detergents, presumably cytoskeletal frame work.

Animals↗
PubMed1983

Subcellular location of the major protein antigens of paramyxoviruses revealed by immunoperoxidase cytochemistry.

The major structural proteins of Newcastle disease virus and Sendai virus were localized in infected BHK-21 and MDBK cells by ultrastructural immunoperoxidase cytochemistry using antibodies against the individual viral protein antigens. The intracellular glycoproteins were strictly membrane bound, being localized in the rough endoplasmic reticulum (RER), perinuclear spaces, smooth membrane vesicles, and presumed Golgi apparatus. The nucleocapsid proteins were detected exclusively in membrane free cytosol and accumulated there, forming inclusions. The membrane (M) protein was found both in cytosol and on RER. The viral proteins on RER exhibited a distinct site specificity; the glycoproteins were facing the lumen of RER whereas M protein was present at the outer cytoplasmic surface. All the viral proteins were detectable at the plasma membrane where virus assembly takes place. However, their modes of distribution differed remarkably. The glycoproteins were spread widely over the entire cell surface including the areas of virus budding and those of normal morphology, whereas M protein was localized in restricted areas of the membrane, frequently forming a patch of virus specific membrane. The presence of nucleocapsids was confined to the virus particles budding from the plasma membrane. These results complement and extend the earlier morphological and biochemical data on the assembly or morphogenesis of paramyxoviruses.

Animals↗
PubMed1983

[Artificial liver support system: removal of bilirubin and bile acids by hemoperfusion with ionex-column].

Removal of toxic substances from the circulating blood of patients with severe hepatic failures may be life-saving, however, the substances should be clarified. IONEX is a new resin specially designed to adsorb bilirubin in the blood. The adsorption of bilirubin from the blood to IONEX was 2 to 3 times than that to the activated charcoal beads in vitro. Direct hemoperfusion with IONEX column was performed using 6 jaundiced dogs. Removal rate of bilirubin after 90 minutes direct hemoperfusion ranged from 49% (IONEX 70 gm) maximum to 9% (IONEX 24.1 gm) minimum. It has been found that the removal of bilirubin is disturbed by thrombus formation on the IONEX fibers. To prevent the platelet loss and aggregate formation during hemoperfusion with IOENX, Prostaglandin (PGE1 or PGI2) was administered continuously into the arterial line of the extracorporeal circulation using IONEX column during 90 min. direct hemoperfusion. Thrombus formation in the IONEX column was little observed and the platelet losses were also not observed. The average removal rate of bilirubin from the blood of the jaundiced dogs was as much as 40% (IONEX 70 gm) and that of bile acid was 61% in mean value. Using PGI2, slight hypotension could not be avoided in dog. In 7 patients with severe jaundice, direct hemoperfusion was performed. Mean removal rate of bilirubin and bile acids from the blood of these patients was 21% and 31.2% respectively after 60 to 90 minutes direct hemoperfusion with activated charcoal and IONEX.

Adsorption↗