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Biomedical subjects

N Yamasaki

Publications and source records attributed to N Yamasaki.

At least 19 recordsLinked to original sources
PubMed1992

Isolation and characterization of isolectins from Erythrina variegata seeds.

Three isolectins were isolated from seeds of Erythrina variegata (Linn.) var. Orientalis by ion-exchange chromatography, followed by affinity chromatography on lactose-Sepharose 4B and acid-treated Sepharose 4B columns. The purified isolectins (EVLI, EVLII and EVLIII) are all specific for galactopyranosides and N-acetylgalactosamine, and their affinities for simple sugars are EVLIII greater than EVLII greater than EVLI. EVLI and EVLIII are homodimers made up of an A-subunit of molecular mass 36,000 and a B-subunit of molecular mass 33,000, whereas EVLII is a heterodimer composed of the A- and B-subunits. Upon treatment with trifluoromethansulphonic acid, the molecular masses of both subunits decreased to 31,000. Rechromatography of EVLII on the acid-treated Sepharose 4B column again produced the homodimeric lectins (EVLI and EVLIII). It is suggested that the constituent subunits of Erythrina variegata isolectins are eschangeable with each other in vitro.

Amino Acids↗
PubMed1992

Effect of amphipathic peptides with different alpha-helical contents on liposome-fusion.

The peptide-induced fusion of neutral and acidic liposomes was studied in relation to the amphiphilicities evaluated by alpha-helical contents of peptides by means of a carboxyfluorescein leakage assay, light scattering, a membrane intermixing assay and electron microscopy. An amphipathic mother peptide, Ac-(Leu-Ala-Arg-Leu)3-NHCH3 (4(3], and its derivatives, [Pro6]4(3) (1), [Pro2,6]4(3) (2), and [Pro2,6,10]4(3) (3), which have very similar hydrophobic moments, caused a leakage of contents from small unilamellar vesicles composed of egg yolk phosphatidylcholine and egg yolk phosphatidic acid (3:1). The abilities of the peptides to induce the fusion of the acidic liposomes increased with increasing alpha-helical content: in acidic liposomes the helical contents were in the order of 4(3) greater than 1 greater than 2 greater than 3 (Lee et al. (1989) Chem. Lett., 599-602). Electron microscopic data showed that 1 caused a transformation of the small unilamellar vesicles (20-50 nm in diameter) to large ones (100-300 nm). Based on the fact that these peptides have very similar hydrophobic moments despite of decreasing in the mean residue hydrophobicities to some extent, it was concluded that the abilities of the peptides to induce the fusion of liposomes depend on the extent of amphiphilic conformation evaluated by alpha-helical contents of the peptides in the presence of liposomes. For neutral liposomes of egg yolk phosphatidylcholine, all the proline-containing peptides showed no fusogenic ability but weak leakage abilities, suggesting that the charge interaction between the basic peptides and acidic phospholipid is an important factor to induce the perturbation and fusion of the bilayer.

Amino Acid Sequence↗
PubMed1992

A microassay for proteases using succinylcasein as a substrate.

A photometric assay for proteases has been developed. A chemically modified casein whose amino groups were succinylated was used as a substrate. After incubation with trypsin, chymotrypsin, thermolysin, and subtilisin, the extent of hydrolysis of the substrate was determined with trinitrobenzene sulfonate (TNBS). The whole procedure of the assay was performed in the microtiter plate wells and the increase in the absorbance resulting from the reaction between TNBS and newly formed amino groups in the substrate was able to be determined with a high sensitivity by a microtiter plate reader, enabling the simultaneous measurement of a number of samples. Application of this method to the measurement of proteolytic activity contained in the protein extract of Tapes philippinarum is demonstrated.

Animals↗
PubMed1992

Isolation and primary structure of proteinase inhibitors from Erythrina variegata (Linn.) var. Orientalis seeds.

The Kunitz-type trypsin inhibitors, ETIa and ETIb, and chymotrypsin inhibitor ECI were isolated from the seeds of Erythrina variegata. The proteins were extracted from a defatted meal of seeds with 10 mM phosphate buffer, pH 7.2, containing 0.15 M NaCl, and purified by DEAE-cellulose and Q-Sepharose column chromatographies. The stoichiometry of trypsin inhibitors with trypsin was estimated to be 1:1, while that of chymotrypsin inhibitor with chymotrypsin was 1:2, judging from the titration patterns of their inhibitory activities. The complete amino acids of the two trypsin inhibitors were sequenced by protein chemical methods. The proteins ETIa and ETIb consist of 172 and 176 amino acid residues and have M(r) 19,242 and M(r) 19,783, respectively, and share 112 identical amino acid residues, which is 65% identity. They show structural features characteristic of the Kunitz-type trypsin inhibitor (i.e., identical residues at about 45% with soybean trypsin inhibitor STI). Furthermore, the trypsin inhibitors show a significant homology to the storage proteins, sporamin, in sweet potato and the taste-modifying protein, miraculin, in miracle fruit, having about 30% identical residues.

Amino Acid Sequence↗
PubMed1992

[A role of interpectoral (Rotter's) lymph node dissection in modified radical mastectomy for breast cancer].

To study the influence of interpectoral lymph node (IPN) dissection on the prognosis of patients who underwent modified radical mastectomy, IPN was carefully dissected and studied pathologically on 168 cases of our breast cancer patients operated with modified radical mastectomy. There were 1.2 lymph nodes on an average in the interpectoral region, and they were almost 1-2mm in diameter. IPN metastases were found in 10 cases. (Tis: 0%, Stage I: 4.9%, Stage II: 5.7%, Stage III: 13%). Tumors located in outer quadrant in almost all these cases. Positive IPN were found in 6 (16%) of n1 alpha group, 1 (10%) of n1 beta group, and in 3 (50%) of n2 group. All these 3 cases of n2 died of distant metastasis and local recurrence. Two (1.7%) of axillary node (1a, 1b) negative patients had microinvolvement of cancer only in IPN, and are currently disease-free. These data suggest that IPN metastasis may occur even in the early breast cancer patients, and that may be controllable by lymph node excision. Therefore, routine and careful dissection of IPN through wide opening of sulcus interpectoralis is necessary for modified radical mastectomy and even for breast preserving operation.

Adult↗
PubMed1992

[Antitumor effect of carboplatin on in vitro three-dimensional tumor].

We studied the antitumor effect of Carboplatin on the three-dimensional tumor in vitro. First, we observed the three-dimensional growth of cultured tumor cell lines (Hep 2, KB and HT 29 cells), which were developed in the combined culture system of fibrin matrix and agar culture. The tumor cells developed a 3 x 3 mm tumor in diameter in the in vitro 10 day-culture system. This size was large enough to allow the histologic study. When we applied Carboplatin to the three-dimensional tumors, histologic change of the three-dimensional tumor developed from Hep 2 and KB cells were observed in a dose dependent manner. However, no remarkable histologic change was observed in the three-dimensional tumor developed from HT 29 cells. These results suggest the experimental system of three-dimensional tumor can predict histologically the antitumor effect of Carboplatin on various tumors at the solid tumor level.

Carboplatin↗
PubMed1991

In vitro anti-human immunodeficiency virus type 1 activity of biliverdin, a bile pigment.

Biliverdin (BV) is a bile pigment having anti-allergic properties. We examined the effect of BV on human immunodeficiency virus type 1 (HIV-1) in vitro. BV completely inhibited the cytopathic effect of HIV-1 in MT-4 cells at concentrations of 22.2 micrograms/ml or more. This inhibitory effect was also observed when BV was present during the adsorption period of HIV-1. However, BV was cytotoxic to MT-4 cells at concentrations above 800 micrograms/ml. At a concentration of 66.7 micrograms/ml, BV completely inhibited syncytia formation by HIV-infected and uninfected MOLT-4 cells. Moreover, after exposure of HIV-1 particles to BV for 2 h, the infectivity of the virus was reduced in a dose-dependent manner. It is speculated that the anti-HIV activity of BV is due to direct inactivation of virions and inhibition of virus binding to target cells.

Antiviral Agents↗
PubMed1991

[Analysis of flow cytometric nuclear DNA content of the thymoma and its relationship to malignant intensity].

Determination was made of the nuclear DNA content of paraffin-embedded specimens of resected thymoma using flow cytometry in 39 patients. Relationships among nuclear DNA content, clinicopathological findings and prognosis were studied. The frequency of DNA aneuploid tumors was 30.8%, 6.7% in stage I (15 patients), 28.6% in stage II (7 patients), 44% in stage III (9 patients) and 62.5% in stage IV (8 patients) according to Masaoka's classification, indicating more in increase with advancing stage and significant (p less than 0.05) more increase invasive thymoma than in noninvasive thymoma. The 5 year and 10 year survival rates of DNA diploid tumors were each 94%, while DNA aneuploid tumors, 75% and 45%. Patients with DNA aneuploid tumors showed less favorable prognosis than those with DNA diploid tumors. Similar results were found in patients with invasive thymoma and in those on whom subtotal or partial resection were performed. The present results indicate DNA aneuploid tumors to have greater malignant intensity than DNA diploid tumors in resected thymoma. Flow cytometric nuclear DNA content analysis provides useful biological data and new indices for evaluating the malignancy of resected thymoma.

Adult↗
PubMed1990

Design and synthesis of basic peptides having amphipathic beta-structure and their interaction with phospholipid membranes.

Basic amphipathic beta-structural peptides, Ac-(Ser-Val-Lys-Val)n-NHCH3 (1n, n = 1-3) and Ac-(Lys-Val)n-NHCH3 (2n, n = 2-4), were synthesized and their interaction with DPPC and DPPC-DPPG (3:1) bilayers was studied by CD, dye-leakage and fluorescence experiments. The CD data indicated that oligopeptides consisting of more than eight residues with alternating hydrophobic (Val) and hydrophilic amino acids (Ser and Lys) were able to form an amphipathic beta-structure in acidic phospholipid bilayers, but not or weakly in aqueous solution and in neutral phospholipid bilayers. The dye-leakage experiment showed that the basic amphipathic beta-structural peptides interact with acidic phospholipid bilayers to perturb them, but less effectively compared with basic amphipathic alpha-helical peptides. Fluorescent spectroscopic data suggest that hydrophobic side of the amphipathic peptides may immerse into membrane without deep penetration. Based on these results, we postulate that the formation of the basic amphipathic beta-structure on acidic lipid bilayers may be due to the combined effect of electrostatic and hydrophobic interactions between basic peptides and acidic lipid bilayers.

1,2-Dipalmitoylphosphatidylcholine↗
PubMed1990

Studies on medicinal resources from livestock. II. Anti-allergic effects of pig bile. (2).

Anti-allergic activities of lyophilized pig bile [( PB]) were examined in mice with picryl chloride-induced contact dermatitis (PC-CD), an experimental model of delayed-type hypersensitivity (DTH; type-IV allergy). PC-CD was markedly inhibited by an oral administration of [PB] within 4 h after but not during 8 to 16 h after challenge with picryl chloride. Anti-inflammatory activities of [PB] were also examined in acetic acid-induced mouse increased vascular permeability, hypotonic-hyperthermic lysis of rat erythrocytes and carrageenin-induced rat hind paw edema. [PB] had no effect on these models. The present study suggests that [PB] inhibits PC-CD through its immuno-modulation in the inductive phase of DTH rather than by an anti-inflammatory action.

Animals↗
PubMed1989

The spectroscopic analysis for binding of amphipathic and antimicrobial model peptides containing pyrenylalanine and tryptophan to lipid bilayer.

The binding of basic amphipathic fluorescent peptides to lipid bilayers was studied in relation to their antimicrobial activity. Four fluorescent peptides containing pyrenylalanine or tryptophan in an amphipathic basic peptide (4(4] consisting of four repeated units of tetrapeptide, -L-Leu-L-Ala-L-Arg-L-Leu-, were found to have antimicrobial activities against Gram-positive bacteria and to take conformations with fairly high alpha-helical content both in aqueous solutions and liposomes. The fluorescence spectroscopic data suggested that the pyrenylalanine-peptide existed as a monomer in methanol or liposomes but as an oligomer in aqueous solutions to form an excimer between pyrenylalanyl residues. Upon binding with liposomes, the fluorescence spectra of the tryptophan-containing peptide shifted to a shorter wavelength, indicating the change in the state of tryptophan from hydrophilic environment to hydrophobic one. The analytical data for the quenching of tryptophan fluorescence by I- anion suggest that the tryptophan residue in the peptide is not deeply buried in the hydrophobic core of the bilayers. Based on these findings, it is suggested that the peptides may interact with liposomes in such a manner that they lie parallel to the surface of the lipid bilayers with their hydrophobic regions shallowly in the amphipathic moiety of the bilayers.

Alanine↗
PubMed1989

Fluorescence spectroscopic studies on tryptophan at the saccharide-binding site of castor bean hemagglutinin.

The environment of tryptophan in castor bean hemagglutinin (CBH) was analyzed by fluorescence spectroscopy with regard to saccharide binding. Upon binding of specific saccharides, the fluorescence maximum of 333 nm of CBH shifted to a wavelength 2 nm shorter, owing to the change in the environment of tryptophan at the saccharide-binding site. By analyzing the change in the fluorescence intensity at 320 nm as a function of concentration of saccharides, the association constants for binding of saccharides to CBH were determined. The results suggest that the saccharide-binding site on each B-chain is actually composed of a subsite with which the saccharide residue linked to galactopyranoside at the non-reducing end can interact, and another site which recognizes the galactopyranoside moiety. Quenching data indicated that five out of 22 tryptophans in CBH are surface-localized and are available for quenching with both KI and acrylamide, and three other tryptophans are buried and are available only to acrylamide. Binding of raffinose to CBH decreased by 2 the number of tryptophan residues accessible to quenchers in the CBH molecule. We speculate that raffinose binds to CBH in such a manner as to shield the tryptophan located at the subsite from quenching by KI and acrylamide. The results also suggest that the tryptophan residue at the saccharide-binding site on each B-chain is localized near the surface, and present in the positively charged environment.

Acrylamides↗
PubMed1989

Binding of saccharides to ricin E isolated from small castor beans.

The binding of saccharides to ricin E isolated from small castor beans was studied by equilibrium dialysis and spectroscopy. Equilibrium dialysis data indicate that ricin E has two galactose-binding sites, a high affinity site (HA-site) and a low affinity site (LA-site). The binding of specific saccharides to ricin E induces a shift of the fluorescence spectrum to shorter wavelength by 3 nm and UV-difference spectra with a maximum at 290 nm and a negative intensity around 300 nm. The interaction of ricin E with its specific saccharides was analyzed in terms of the variation of the intensity at 320 nm in the fluorescence spectrum and the magnitude of the negative intensity at 300 nm in the UV-difference spectra as functions of saccharide concentration. The results indicate that these spectroscopic changes are representative of the binding of saccharides to the LA-site, which contains a tryptophan residue. By comparing the association constants of saccharides for ricin E with those for ricin D, isolated from the large castor beans, it was found that the HA of ricin E binds saccharides with an affinity of less than one-half that of ricin D, while the saccharide-binding abilities of the LA-site of the two ricins were about the same.

Carbohydrates↗
PubMed1989

The interaction of Abrus precatorius agglutinin with saccharides as analyzed by fluorescence spectroscopy.

The binding of saccharides to Abrus precatorius agglutinin (APA) was analyzed by fluorescence spectroscopy. Upon binding of specific saccharides, the fluorescence emission maximum of APA (338 nm) shifted to shorter wavelength by 5 nm, owing to the change in the environment of tryptophan. By analyzing the change in the fluorescence intensity at 338 nm as a function of concentration of saccharides, the association constants for binding of saccharides to APA were determined. The results suggest that in the saccharide binding site on each B-chain of APA, there may be a site which interacts with the saccharide residue linked to galactopyranoside at the non-reducing end, in addition to the site which recognizes the galactopyranosyl residue. Fluorescence quenching data indicate that 8 out of 24 tryptophans in APA are located at or near the surface of the protein molecule and are available for quenching with both KI and acrylamide, and 10 tryptophans are involved in the environment to which acrylamide has access but KI does not. Binding of lactose to APA reduced by 4 the number of tryptophan residues accessible to quenchers. Based on the results, it is suggested that the tryptophan residues at the saccharide binding site on each B-chain of APA are present on the surface of the APA molecule, and they are shielded from quenching by KI and acrylamide upon binding with specific saccharides.

Acrylamides↗
PubMed1989

Studies on medicinal resources from livestock. I. Anti-allergic effects of pig bile. (1).

Anti-allergic activities of animal biles and commercially available bile acids were evaluated in experimental allergic disease models. Pig bile exhibited marked preventive effects on the models of delayed-type hypersensitivity (type IV allergy), picryl chloride-induced contact dermatitis (PC-CD) and sheep red blood cells (SRBC)-induced footpad swelling in mice. Fel ursi (dried bear gallbladder) also had an inhibitory effect on PC-CD, whereas ox bile, chicken bile and bile acids had no effect on ether of the models.

Animals↗
PubMed1988

Production of heavy-chain class-switch variants of human monoclonal antibody by recombinant DNA technology.

We have previously established a human-mouse heterohybridoma (H6-3C4), which produced a human sperm-immobilizing antibody (mu, lambda of human type). The human rearranged immunoglobulin mu-chain and lambda-chain genes were cloned from the hybridoma H6-3C4. The cloned V region of the heavy chain (VH) gene was ligated to human immunoglobulin gamma 1-heavy chain constant region (C gamma 1) genes. This resulted in the heavy-chain class-switch from mu-chain to gamma 1-chain of H6-3C4 antibody. The class-switched heavy-chain gene as well as the cloned lambda-chain gene were introduced into mouse myeloma cell line X63Ag8.653 by protoplast fusion and electroporation. The stable transformants produced the human IgG monoclonal antibody, which fully retained specificity to human sperm cells and sperm-immobilizing activity.

Animals↗
PubMed1987

Sequence analysis of a cDNA clone of a gene encoding a component of a putative phosphorylcholine-specific T suppressor factor and functional property of its gene product.

We have isolated a putative phosphorylcholine (PC)-T cell suppressor factor (TsF) cDNA clone, p6-5, from a cDNA library of a T hybridoma which constitutively secretes a PC-TsF in vitro [8]. In the present study, we determined the nucleotide sequence of the p6-5 gene and found that the p6-5 sequence is 86% homologous to rat preproelastase 1 gene, one of the serine protease genes. An oligopeptide (14 mer, TsF14) deduced from the p6-5 sequence was synthesized and antisera against TsF14 were prepared in rabbits. Anti-TsF14-conjugated Sepharose 4B specifically absorbed the PC-TsF activity from the culture supernatant of PC-TsF-secreting T hybridomas. In contrast, the binding molecule eluted from the anti-TsF14-conjugated Sepharose suppressed the antibody response PC specifically. These results indicated that the p6-5 polypeptide is a component of the PC-TsF molecule.

Amino Acid Sequence↗