Biomedical subjects

S Kurata

Publications and source records attributed to S Kurata.

At least 73 records · Page 4Linked to original sources

PubMed1996

Effects of hypertrichotic agents on follicular and nonfollicular cells in vitro.

Our previous studies revealed that topical minoxidil induced an increased rate of DNA synthesis in both dermal papilla and follicular germinal cells in early anagen and bulbar matrix as well as outer root sheath and perifollicular fibrocytic cells in mid and late anagen follicles in the bald scalp of the stump-tailed macaque. However, the epidermis and dermal fibrocytes showed no response. To determine the specific action of hypertrichotic agents on follicular cells, we examined the effects of two potent hypertrichotic agents, minoxidil and cyclosporin, on the DNA synthesis of cultured cells derived from either follicular cells (dermal papillar, perifollicular fibrocytic and outer root sheath cells) obtained from human and macaque scalps or nonfollicular cells (fibrocytes and epidermal keratinocytes) from human and macaque foreskin, palm and sole regions and the 3T3 cell line. Cultured subconfluent cells from the above follicular and nonfollicular specimens were incubated with either minoxidil (0.01-2 mM) or cyclosporin (0.01-100 mM) in medium (serum-free DMEM) for 48 h, then 3H-thymidine was added for the final 6 h. Minoxidil induced a significant increase in DNA synthesis in all follicular cells in a dose-specific manner (maximum rate at 0.5 mM for dermal papilla and perifollicular fibrocytic cells and 0.1 mM for outer root sheath cells). The perifollicular fibrocytic cells appeared to have a potentiality similar to that of the dermal papilla cells. Nonfollicular cells showed no response to minoxidil; 3T3 cells were rather suppressed. Cyclosporin appeared to have rather suppressive effects on both follicular and nonfollicular cells. These results suggest that minoxidil has a specific affinity to hair follicular cells and induced their cell proliferation. Although cyclosporin is known as a potent hypertrichotic agent, our studies on cultured follicular cells showed no direct proliferative effect. The hypertrichotic mechanism of cyclosporin appeared to be different from that of minoxidil.

3T3 Cells↗

PubMed1995

Regulation of the expression of cathepsin B in Sarcophaga peregrina (flesh fly) at the translational level during metamorphosis.

Previously, we reported that cathepsin B of pupal hemocytes participates in dissociation of the fat body of Sarcophaga larvae at metamorphosis. In this study, we demonstrated that the amount of cathepsin B in hemocytes was controlled at the translational level, i.e. larval hemocytes stored a significant amount of untranslated cathepsin B mRNA. When the larvae pupated, translation of the mRNA commenced, resulting in accumulation of cathepsin B. Our results indicate that the 3'-untranslated region (3'-UTR) of this mRNA is necessary for the repression of its translation in a rabbit reticulocyte lysate. We detected the 3'-UTR binding protein in the lysate of larval hemocytes, but not in the pupal hemocytes. The role of this protein in the translation of cathepsin B mRNA is discussed.

Animals↗

PubMed1995

Androgen induction of follicular epithelial cell growth is mediated via insulin-like growth factor-I from dermal papilla cells.

Androgen significantly stimulates the proliferation of outer root sheath cells that are cocultured with beard dermal papilla cells without cell contact. The expression of insulin-like growth factor I (IGF-I) mRNA in beard dermal papilla cells was stimulated by androgen and antagonized by cyproterone acetate. Outer root sheath cells did not express mRNA for IGF-I either in the presence or absence of androgen. Both of these two types of cells expressed mRNA for IGF-I receptor and the expression was not affected by androgen. Neutralizing antibody against IGF-I antagonized the stimulatory effect of androgen on the growth of outer root sheath cell cocultured with beard dermal papilla cells. These findings suggest that IGF-I is a candidate for androgen induced hair growth factors.

Antibodies↗

PubMed1995

Hyperbaric oxygenation reduces the cytostatic activity and transcription of nitric oxide synthetase gene of mouse peritoneal macrophages.

The effects of hyperbaric oxygenation (HBO) and hyperbaric air (HBA) on the cytostatic activity, peroxynitrite synthesis and transcription of the inducible nitric oxide synthetase (iNOS) gene of murine peritoneal macrophages were studied in vitro. Exposure of mice to HBO or HBA significantly reduced the cytostatic activity, peroxynitrite synthesis and transcription of iNOS mRNA of their macrophages stimulated with lipopolysaccharide (LPS) and interferon-gamma (IFN-gamma). These results indicate that HBO and HBA treatments of mice reduce the cytostatic activity of peritoneal macrophages by reducing iNOS mRNA synthesis.

Amino Acid Oxidoreductases↗

PubMed1995

Molecular cloning of cDNA for sapecin B, an antibacterial protein of Sarcophaga, and its detection in larval brain.

A cDNA clone for sapecin B, an antibacterial protein of Sarcophaga, was isolated. This cDNA encoded a precursor protein of sapecin B consisting of a signal sequence (24 residues), prosegment (30 residues) and mature sapecin B (34 residues). Sapecin B was synthesized almost exclusively in the fat body when the larval body wall was injured, but the brain of naive larvae was also demonstrated to contain a significant amount of sapecin B. These findings suggested that sapecin B is a bifunctional protein.

Amino Acid Sequence↗

PubMed1995

Molecular characterization of an insect transferrin and its selective incorporation into eggs during oogenesis.

A protein with a molecular mass of 65 kDa that was specifically taken up into eggs was purified from the hemolymph of adult female Sarcophaga peregrina flies. From cDNA analysis, this protein was shown to be a Sarcophaga transferrin. Unlike mammalian transferrin, the similarity between its N-terminal and C-terminal halves was only 19%, and it was suggested to conjugate one iron atom/molecule in its N-terminal half. Sarcophaga transferrin was found to transport iron ions into eggs during oogenesis and deliver them to another protein, thought to be ferritin. No significant activation of the transferrin gene was detected during embryogenesis, so probably maternal transferrin is used as an intercellular or intracellular iron-transporter during embryogenesis of this insect.

Amino Acid Sequence↗

PubMed1995

Protoplast fusion method for efficient transformation of mouse spermatogenic cells.

The efficiencies of methods for introducing DNAs into spermatogenic cells were investigated. The calcium phosphate method has been most commonly used to transfect DNA into cultured cells, but this method severely damages spermatogenic cells. Therefore, we investigated the efficiencies of transfection of spermatogenic cells by DEAE-dextran, modified liposome, and protoplast fusion methods, which seemed to cause less damage to spermatogenic cells than the calcium phosphate method. The protoplast fusion method was the most suitable for transfection of spermatogenic cells.

Animals↗

PubMed1995

Mode of action of an antibacterial peptide, KLKLLLLLKLK-NH2.

Previously, we reported that a synthetic undecapeptide, KLKLLLLLKLK-NH2, and its D-enantiomer have potent bactericidal activities against both Gram-positive and Gram-negative bacteria. Here we examined the mode of action of KLKLLLLLKLK-NH2 with special reference to its effect on bacterial membranes. We found that both the outer and inner membrane of Escherichia coli become permeable to low molecular mass substances when treated with this peptide. Under these conditions, the bacteria lost the ability to synthesize ATP and to transport proline, suggesting that their electrochemical membrane potential was disrupted. This peptide appears to form numerous channels in bacterial membranes that interfere with membrane functions, resulting in cell death.

Adenosine Triphosphate↗

PubMed1995

Purification and cDNA cloning of an antifungal protein from the hemolymph of Holotrichia diomphalia larvae.

An antifungal protein (AFP), holotricin 3, was purified from the hemolymph of the coleopteran insect Holotrichia diomphalia larvae. Analysis of its cDNA showed that holotricin 3 is a novel Gly- and His-rich protein consisting of 84 amino acid residues. This protein was similar to AFP, an antifungal protein of Sarcophaga peregrina that was reported previously, in terms of molecular size and high content of Gly and His residues. However, no appreciable sequence similarity was detected between the two proteins.

Amino Acid Sequence↗

PubMed1995

A sapecin homologue of Holotrichia diomphalia: purification, sequencing and determination of disulfide pairs.

We purified and characterized a sapecin homologue, named holotricin 1, from the hemolymph of immunized larvae of a coleopteran insect, Holotrichia diomphalia. We determined its complete amino acid sequence and three disulfide pairs. Holotricin 1 consisted of 43 amino acid residues and showed potent antibacterial activity against gram-positive bacteria, but antibacterial activity against gram-negative bacteria was not obvious.

Amino Acid Sequence↗

PubMed1995

Effects of mixed silane coupling agent on porcelain tooth material and various dental alloys.

Mixed salines, 3-methacryloxypropylsilytriisocyanate and 3-mercaptopropyltrimethoxysilane, were used as primers for adhesion of poly(methyl methacrylate) to porcelain teeth, precious alloys and nonprecious alloys. After thermocycle testing, the mean bond strength of the resin to porcelain, silver alloys and stainless steel were excellent. The fracture patterns of porcelain teeth after thermocycling followed by adherent testing were similar to those of specimens subjected to 48-hour water immersion at 37 degrees C. Two specimens failed cohesively in the porcelain and the other specimens failed in the resin. The mixed primer had no effect on cobalt-chromium alloy specimens.

Boron Compounds↗

PubMed1995

Interaction between dermal papilla cells and follicular epithelial cells in vitro: effect of androgen.

To investigate the mode of action of testosterone on human hair follicles, we studied the localization of androgen receptors in hair follicles and cultured dermal papilla cells from different body sites, using immunohistochemistry with a polyclonal antibody. Androgen receptors were detected in the dermal papilla cells of beard and axillary hair follicles, but not in those of occipital scalp hair follicles. Epithelial cells of the hair follicles were not stained by the antibody. Androgen receptors were also detected in the nuclei of cultured beard and axillary dermal papilla cells, but the dermal papilla cells from occipital scalp hair follicles showed little staining with this antibody. We also examined the effects of testosterone on the DNA synthesis and proliferation of cultured outer root sheath cells and dermal papilla cells. Testosterone did not have a proliferative effect on either type of cell when cultured alone. In contrast, testosterone significantly stimulated the proliferation of outer root sheath cells which were co-cultured with beard or axillary dermal papilla cells, without cell contact. No such effect was observed when dermal papilla cells from occipital scalp hair follicles were used in the cultures. This proliferative effect of testosterone was concentration-dependent, and was antagonized by cyproterone acetate. Our findings suggest that dermal papilla cells from the beard and axillary hair follicles produce androgen-dependent diffusible growth factors which stimulate follicular epithelial cells.

Adolescent↗

PubMed1994

Potential of azidothymidine to activate the HIV-1 promoter.

Azidothymidine (AZT) has been used for treatment of acquired immunodeficiency syndrome (AIDS), but a recent concord trial showed that it is ineffective for suppressing the development of AIDS. In this study, the effects of AZT on three cell lines transformed by pCD12 plasmid (human immunodeficiency virus-1 (HIV-1) promoter-chloramphenicol acetyltransferase) were examined. Results showed that AZT has the potential to activate the HIV-1 promoter.

Base Sequence↗

PubMed1994

Novel synthetic antimicrobial peptides effective against methicillin-resistant Staphylococcus aureus.

Previously, we identified a core undecapeptide of sapecin B having antimicrobial activity. Based on the structure of this peptide, we systematically synthesized peptides consisting of terminal basic motifs and internal oligo-leucine sequences and examined their antimicrobial activities. Of these peptides, RLKLLLLLRLK-NH2 and KLKLLLLLKLK-NH2 were found to have potent microbicidal activity against Staphylococcus aureus, Escherichia coli, methicillin-resistant S. aureus and Candida albicans in liquid medium. We also synthesized the D-enantiomer of KLKLLLLLKLK-NH2. This enantiomer was resistant to tryptic digestion and persisted longer in the culture medium, showing greater antimicrobial activity than the original peptide.

Amino Acid Sequence↗

PubMed1994

Purification, characterization, and cDNA cloning of procathepsin L from the culture medium of NIH-Sape-4, an embryonic cell line of Sarcophaga peregrina (flesh fly), and its involvement in the differentiation of imaginal discs.

A cysteine proteinase that preferentially hydrolyzes carbobenzyloxy-Phe-Arg-MCA (MCA, methylcoumaryl-7-amide) was purified from the culture medium of NIH-Sape-4 cells. The molecular mass of this enzyme was easily changed from 50 to 35 kDa, without appreciable loss of enzyme activity. From the analysis of its cDNA, this enzyme was concluded to be a procathepsin L of Sarcophaga. Only procathepsin L was found in the medium, while the mature 35-kDa cathepsin L was found exclusively in the cells, indicating that the cells have a mechanism to secrete procathepsin L selectively. An antibody against procathepsin L was found to inhibit the differentiation of imaginal discs cultured in vitro in the presence of 20-hydroxyecdysone. In fact imaginal discs were shown to secrete procathepsin L into the culture medium. These results suggest that procathepsin L is an essential proteinase for differentiation of imaginal discs. Northern blotting analysis revealed that unfertilized eggs contain a significant amount of mRNA for the procathepsin L as a maternal mRNA.

Amino Acid Sequence↗

PubMed1994

Synthesis and characterization of sapecin and sapecin B.

Two insect defencins, sapecin and sapecin B, were chemically synthesized to confirm their structure and antibacterial activity and also to examine the possibility that these peptides bind to the same site on the large conductance calcium-activated potassium channel as charybdotoxin. Both synthetic peptides showed the same antibacterial activity as native sapecins, indicating that the synthetic peptides folded correctly in the chemical synthesis. Synthetic sapecins did not show an inhibitory effect on [125I]charybdotoxin binding to rat brain synaptic membranes, suggesting that sapecin B recognizes a different binding site from that of charybdotoxin despite the similar structural motif.

Amino Acid Sequence↗

PubMed1994

Cloning and functional expression of poly(ADP-ribose) polymerase cDNA from Sarcophaga peregrina.

A cDNA spanning the entire coding region for poly(ADP-ribose) polymerase (PARP) of Sarcophaga peregrina was isolated and the nucleotide sequence was determined. The longest open reading frame encodes a polypeptide of 996 amino acid residues with a molecular mass of 113,033 Da. The similarities to the human PARP in amino acid sequence were relatively low in the DNA-binding and auto-modification domains, but very high in the C-terminal catalytic domain: identity of amino acids is 34% in the N-terminal DNA-binding domain (residues 1-369), 27% in the auto-modification domain (residues 370-507), and 56% in the C-terminal NAD-binding domain (residues 508-996). Two zinc-fingers (C-X2-C-X28-H-X2-C and C-X2-C-X31-H-X2-C)2 and a basic region in the N-terminal DNA-binding domain recognized in other PARP are conserved. Downstream of the basic region, another cysteine-rich motif (C-X2-C-X13-C-X9-C), a putative zinc-finger, was found to be well conserved in the PARP of Sarcophaga, Drosophila and human. A leucine-zipper motif (L-X6-L-X6-L-X6-L) which was found in the auto-modification domain of Drosophila PARP, is disrupted in the Sarcophaga enzyme: the second leucine is replaced by proline, and the third leucine by valine. Full-length cDNA for Sarcophaga PARP was cloned into an expression plasmid and expressed in Escherichia coli. A lysate of E. coli cells containing expressed protein reacted with antibody against Sarcophaga PARP, and PARP activity was detected. Thus, we conclude that isolated cDNA encodes a functional Sarcophaga PARP cDNA.

Amino Acid Sequence↗

PubMed1994

Inhibition of the Ca(2+)-activated K(+)-channel by sapecin B, an insect antibacterial protein.

Sapecin is an antibacterial protein of the flesh fly and sapecin B is its homologue structurally similar to charybdotoxin of scorpion venom, which is known to be a K+ channel inhibitor. We found that, like charybdotoxin, sapecin B inhibits part of the voltage pulse-induced K+ currents of rat cerebellar Purkinje cells. We suggest that this effect is due to inhibition of the Ca(+)-activated K+ channel. Probably, sapecin B is a naturally occurring K+ channel inhibitor as well as an antibacterial protein.

Animals↗