PubMed Health⌕ Search

Biomedical subjects

S Matsuhashi

Publications and source records attributed to S Matsuhashi.

At least 37 records · Page 2Linked to original sources
PubMed1992

Primary and secondary culture of rat ameloblasts in serum-free medium.

Enamel is the hardest tissue in vertebrates. Ameloblasts are derived from epithelial cells and are responsible for enamel formation. They secrete enamel matrix components in which amelogenins are the major proteins, the biochemical properties of which are well known. However, little is known about the characteristics of ameloblasts themselves or about the functions of amelogenins. In this study, we developed a novel primary and secondary culture system for ameloblasts using a monoclonal antibody which recognized amelogenin (En3). The cell layer on dentine removed from rat mandibular incisors was isolated and cultured in low calcium, serum-free medium. Primary culture was performed on collagen-coated culture plates and typically, two types of cells appeared. One major type changed morphology after the addition of a high concentration of calcium to the medium. Expression of amelogenin was shown as cytoplasmic particles in these cells using En3. In the secondary culture, expression of amelogenins was also observed. In this system, the cells grew and maintained the expression of amelogenin for about 3 weeks.

Ameloblasts↗
PubMed1991

Suppression of murine leukemia virus-mediated 3Y1 cell fusion by expression of mouse MHC class I.

Rat 3Y1 fibroblasts transformed by adenovirus type 12 or its E1A gene formed syncytia by cocultivation with Friend murine leukemia virus (MuLV)-producing cells. On the other hand, parental 3Y1 cells and those derivatives induced by other tumor viruses or chemical carcinogen showed no MuLV-mediated syncytium formation [N. Momozaki et al. (1990) Arch. Virol. 115: 123-126]. The expression of major histocompatibility complex (MHC) class I mRNA and antigens was significantly reduced in these Ad12- and E1A-transformed 3Y1 cells. In contrast, other tumor virus-and chemical carcinogen-transformed 3Y1 cells expressed MHC class I almost in normal levels as did parental 3Y1 cells. Furthermore, Ad12-transformed 3Y1 cells which started to express the transfected exogenous MHC class I gene, H-2Ld, showed no more MuLV-mediated 3Y1 cell fusion. These results indicate that the expression of MHC class I on the cell membrane is closely related to the inhibition of 3Y1 cell fusion by MuLV.

Actins↗
PubMed1991

Monoclonal antibodies recognizing amino-terminal and carboxy-terminal regions of human aldolase A: probes to detect conformational changes of the enzyme.

Three monoclonal antibodies (MAbs1A2, 3C5, and 4C2) for human aldolase A [EC 4.1.2.13] were established. MAbs1A2, 3C5, and 4C2 were shown to belong to subclasses IgM, IgG1, and IgG2a, respectively. None of the MAbs inhibits aldolase A activity. Their epitopes were mapped in detail on the molecule by examining the reactivities of the MAbs to chimeric proteins between aldolases A and B [Kitajima et al. (1990) J. Biol. Chem. 265, 17493-17498] in ELISA and to the CNBr-cleaved fragments of aldolase A in immuno-blotting. MAbs1A2 and 3C5 reacted with sites located within amino acid residues 306-363 at the C-terminal region of the enzyme. MAb4C2 recognized an epitope of the enzyme present within amino acid residues 34-108 at the N-terminal region. In a competitive binding assay, MAbs1A2 and 3C5 competed with each other for binding to the antigen and also interfered with the binding of MAb4C2, whereas MAb4C2 failed to inhibit the binding of MAbs1A2 and 3C5 to the antigen. MAb3C5 showed a species-specificity in the reaction with the antigen; it reacted with human and rabbit aldolase A with similar reactivity but not at all with the rat and mouse enzymes, which differ from the human and rabbit enzymes in two amino acid residues at positions 328 and 348. Reactivities of MAbs to aldolase A were further examined with engineered enzymes containing an amino acid substitution.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗
PubMed1989

Preparation of monoclonal antibodies against nuclear antigens in a DNA affinity purified protein fraction from chick embryo extract.

Monoclonal antibodies (Mabs) were raised to components of a DNA binding protein fraction obtained by using native and denatured DNA-cellulose column chromatographies of chick embryo extract. Indirect immunofluorescence microscopic analysis revealed that 3 Mabs, Pr-28, Pr-123 (or Pr-122) and Pr-192 react to nuclear antigens of proliferating cells. Part of the cell nuclei in chick embryo fibroblast culture were stained in speckled patterns by all three of the Mabs. It seems however that they are different clones judging from the following criteria; 1. Pr-28 crossreacts to a P3U1 cell surface antigen but Pr-123 (or Pr-122) and Pr-192 do not. 2. The rate of nuclei stained by Pr-122 is different from that of Pr-192 in both growing and quiescent cultures.

Animals↗
PubMed1987

An antigen expressed in proliferating cells at late G1-S phase.

A monoclonal antibody Pr-28 was prepared, which recognized an antigen present only in proliferating cells. Immunofluorescence analysis of Pr-28 antigen showed that the antigen was localized mainly in perinuclear cytoplasm. Although Pr-28 antibody was produced against a chicken cell antigen, it reacts not only with chicken cells but also other cells of murine origin, such as L-cells and NIH 3T3 cells. The molecular weight (Mr) of the antigen recognized by Pr-28 antibody was 45,000 D as determined by SDS-PAGE run under reducing conditions. The antigen disappeared in NIH 3T3 quiescent cells, reappearing in quiescent cells stimulated by fetal calf serum (FCS). The synthesis of Mr 45,000 protein occurred at late G1 phase, just before DNA synthesis in serum-stimulated quiescent NIH 3T3 cells and ceased in S phase.

Animals↗
PubMed1987

Platelet proteins, including platelet-derived growth factor, specifically depress a subset of the multiple components of the response elicited by glutathione in Hydra.

Human serum more strongly depressed the feeding response of Hydra (ball formation) elicited by S-methylglutathione than plasma. On the basis of the effect of several proteins released by platelets, at least five apparent components of the response (R1-R5) were suggested. Each of the platelet proteins examined specifically depressed a subset of these components. Among the platelet proteins examined, platelet-derived growth factor (PDGF) specifically depressed the R2 response (the concentration at which the depressing effect was 50% of the maximum [ED50] was 0.17 pM), and basic fibroblast growth factor depressed the R3 and R5 responses (ED50 0.50 aM) and the R2 response (ED50 0.55 pM). With respect to the depression of the R2 response by PDGF, addition of an anti-PDGF IgG or chemical reduction of PDGF, both of which prevent PDGF from binding to its cell surface receptor on responsive cells, eliminated the depressing effect of PDGF on the hydra response. The implications of these observations are discussed.

Animals↗
PubMed1987

Human aldolase B gene: characterization of the genomic aldolase B gene and analysis of sequences required for multiple polyadenylations.

The chromosomal gene encoding human aldolase B was isolated. The gene is composed of nine exons interrupted by eight introns and spans 15 kb, and a single copy of it occurs per haploid human genome. The initiation of transcription occurs at three different sites. Two minor sites, m1 and m2, start at 49 and 21 nucleotides, respectively, upstream from the major site, M. The gene also carries poly(A) addition signals at two different sites, thereby another two distinct mRNA species are produced. We examined the sequences required for mRNA 3'-end formation in this gene carrying multiple poly(A) addition sites. By constructing deletion mutants as to the region distal to the poly(A) addition site and then assaying through transfection into COS-1 cells, we demonstrated that 8 nucleotides distal to the site of poly(A) addition is sufficient for proximal polyadenylation, but is not sufficient for distal polyadenylation.

Animals↗
PubMed1985

Determination of protein spots separated by two-dimensional polyacrylamide gel electrophoresis.

A simple method for quantitating proteins in the spots on two-dimensional polyacrylamide gel electropherograms is described. The system consists in three steps: (1) O'Farrell's two-dimensional gel electrophoresis of the proteins to be analysed; (2) staining of the gels with Coomassie brilliant blue; and (3) determination of the area and integrated density of the stained spots by the Joyce Loebl Magiscan-1 image analysis system. The method can be used for the determination of proteins in the range 0.5-100 micrograms/cm2; the amount of protein involved in most spots detected by the staining method actually falls within this range. As the minimum spot diameter that can easily be handled by the method is about 2 mm, as much as 30 ng of protein in such a spot can be determined. The method can also be applied to autoradiograms.

Autoradiography↗
PubMed1985

Stimulation of epidermal growth by the egg white and yolk.

It was found that white and yolk of the egg stimulated DNA synthesis of chick embryo epidermal culture. The epidermis cultured with yolk grew well in a manner similar to that with chick embryo extract, but the culture with egg white was poorer in growth than that with yolk. The mitogenic activity of egg white was inactivated by a heat treatment at 70 degrees C, while yolk was more heat-resistant. A heat-stable yolk fraction (hY fraction) which stimulated DNA synthesis of chick embryo epidermis was isolated by heat treatment at 98 degrees C. The mitogenic activity was enhanced by the addition of both egg white and hY fraction in the culture medium. The enhancement of epidermal growth by such an addition was confirmed by histological observations of the cultured epidermis. In embryonic skin, the growth and differentiation of epidermis is influenced by the nature of mesenchyme is proximity to the epidermal cells. The epidermis isolated from dermis does not grow or differentiate when cultivated in a chemically defined medium, although the tissue in whole skin does grow in the same medium. However, the isolated epidermis proliferates on non-living substrate such as membrane filter or collagen gel when cultured in a medium which contains chick embryo extract. This epidermal growth promoting property of chick embryo extract is inactivated by heat treatment or trypsin digestion.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1984

Development of chicken epidermis cultured with embryo extract.

The epidermis from 11-day-old chick embryo shank skin was cultured with 11-day-old chick embryo extract. The growth and the differentiation of the epidermis in culture were studied histologically, electron microscopically and with polyacrylamide gel electrophoresis of keratin proteins. The epidermis cultured with the chick embryo extract proliferated and stratum structures developed simultaneously with the increase in epidermal cell layers. Finally, a keratinized layer was observed after 10 days in culture. Electron microscopic observations revealed that tonofilaments were produced after 2 days in culture and increased thereafter with culture time, becoming condensed with desmosomes. Keratohyaline granules were observed in 7-day cultures. These keratinization characteristics occurring during culture showed the general characteristics of the alpha stratum observed in the skin of in ovo embryos during the early stages of development. However, the development of peridermal and subperidermal granules was poor and the stratum granulosum, which develops at the late stages between the stratum intermedium and the stratum corneum, was not observed. Polyacrylamide gel electrophoresis of S-carboxymethylated keratin proteins showed that the keratin protein band patterns of the culture differed from those of in ovo skin epidermis.

Animals↗
PubMed1974

Mechanism of action and development of resistance to a new amidino penicillin.

The mechanism of killing of Escherichia coli by a novel beta-lactam antibiotic, an amidino penicillin, has been investigated. This compound converts E. coli to relatively stable spherical forms at low concentration. However, the amidino penicillin caused no alteration in any of those parameters of peptidoglycan synthesis which can be studied. Above 10 mug of the antibiotic per ml the cells began to lyse, and a second mode of killing appeared. Mutants resistant to the amidino penicillin were isolated and several were studied in detail. Three mutant phenotypes were distinguished: (i) spherical shape and hypersensitive to lysis by either amidino penicillin or ampicillin; (ii) spherical shape and normally sensitive to lysis; (iii) rod shape, converted to viable spheres by amidino penicillin and normally sensitive to lysis.

Amidines↗