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Biomedical subjects

K Fukuyama

Publications and source records attributed to K Fukuyama.

At least 217 records · Page 12Linked to original sources
PubMed1978

X-ray analysis of ferredoxin from Spirulina platensis. II. Chelate structure of active center.

A chloroplast-type ferredoxin from Spirulina platenis crystallized in an orthorhombic system, space group C2221, with cell dimensions a=62.32, b=28.51, and c=108.08 A. The electron density map at 2.8 A resolution was prepared by using the best phase angles determined by the single isomorphous replacement method coupled with the anomalous dispersion method. The chelating structure of the acitve center was revealed as follows. Of the six cysteinyl residues in the molecule, Cys 41, Cys 4k, Cys 49, and Cys 79 are involved in the active center. Cys 41 and Cys 46 are coordinated to one iron atom, and Cys 49 and Cys 79 to the other iron atom. Only one of these cysteinyl residues, Cys 79, is comparatively apart from the other three in the amino acids sequence of the molecule, as found in the case of bacterial ferredoxin. It appears that the NH....S hydrogen bonds are around the active center, as in other non-heme iron sulfur proteins.

Binding Sites↗
PubMed1978

Ultrastructural study of the nuclei in premitotic and repair DNA synthesis following UVB injury.

Ultrastructural changes in nuclei synthesizing DNA were studied by cytochemical technique. Guninea pigs ears were UVB irradiated and TdR-H3 was injected intradermally into the irradiated sites 1 hr before biopsy. Areas of the epidermis containing more than 80% of cells in DNA (repair or premitotic) synthesis identified by light microscopic autoradiography were selected and cut at 600 A. The glycolmethacrylate sections were stained with uranyl acetate and lead citrate, and consecutive sections were incubated with 0.01% pronase and 0.5% RNase before staining in order to observe DNA. In cells undergoing DNA repair, the zone of DNA became discontinuous and DNA was scattered throughout the entire karyoplasm as small aggregates and fine filaments. Nuclei in S-phase showed essentially the same change, but quantitatively the disappearance of DNA from the nuclear membrane and distribution in the karyoplasm became much greater. These changes were not seen in specimens treated without cytochemical technique.

Animals↗
PubMed1978

Effects of 4-tertiary butyl catechol on tissue cultured melanocytes.

Depigmenting effects of 4-tertiary butyl catechol (TBC) were investigated using tissue cultured melanocytes of adult guinea pig. Black guinea pig ear epidermis was trypsinized, suspended in BME Eagle media (5 X 10(6) cells) supplemented with 10% calf serum, and seeded in Petri dishes (2 ml/ea). On the 5th day of culture, TBC dissolved in DMSO was added to the media, and the central area of Petri dishes with large numbers of melanocytes (but not keratinocytes) present were examined by light and electron microscopy. After 6 hr of treatment with 1.5 X 10(-3) mg TBC per ml of media, about 15 to 30% of melanocytes detached and about half of the remaining cells showed reversible changes in the cell shapes. An increase of TBC concentration to more than 2 X 10(-3) mg per ml of media resulted in larger numbers of melanocyte deaths. The surviving cells contained numerous, but ultrastructurally unusual premelanosomes and melanosomes. In addition, microfilaments showed a wavy appearance.

Animals↗
PubMed1978

Two tris urea mercaptoethanol extractable polypeptides found uniquely in scales of patients with psoriasis.

This study was designed to chemically characterize the principal structural proteins of psoriatic scales. Cornified cells were obtained from 40 patients with psoriasis, 21 patients with other scaly diseases, and 13 normal individuals. Cells were washed with Tris-HCl buffer and incubated in 8 M urea containing 2-mercaptoethanol (pH 9.0) at 30 degrees C for 7 hr. Extracted proteins were subjected to SDS polyacrylamide gel electrophoresis and protein patterns from normal and diseased scales were compared. The 67,000 dalton constituent of normal cornified cells could not be identified in protein from psoriatic scale and instead, a pair of polypeptides of approximately 54,000 and 57,000 daltons appeared. These extra bands were not found in protein extractions from other skin diseases, uninvolved skin of psoriasis patients, or normal skin. In order to analyze further normal and psoriatic scale proteins, the immunoreaction of rabbit antisera to human 67,000 dalton polypeptide with extracted psoriasis protein and with frozen biopsy sections, was studied using immunoprecipitation tests and indirect immunofluorescence microscopy. Both techniques demonstrated the existence of the 67,000 dalton protein in psoriasis, but as a minor component. These results indicate that additional unique urea mercaptoethanol soluble proteins are formed in psoriatic lesions, and this unusual protein synthesis may reflect the morphological changes in this disease.

Humans↗
PubMed1978

Divalent cation stimulation of in vitro fibre assembly from epidermal keratin protein.

Keratin was extracted from purified cornified cells of newborn rats in Tris-HCl-buffered 8 M urea containing beta-mercaptoethanol. Microfilaments were assembled in vitro by reducing the ionic strength of buffer and the urea concentration. One millimolar concentration of KCl and NaCl did not affect filament formation, but the same concentration of divalent cations greatly altered this process. CaCl2 and MgCl2 induced gelation of keratin by formation of bundles of birefringent macrofilaments. ZnCl2, CuSO4 and HgCl2 formed greater numbers of macrofilaments and the protein aggregated.

Amino Acids↗
PubMed1977

Changes in nuclear DNA and RNA during epidermal keratinization.

The distribution of nucleic acids within nuclei of epidermal cells in skin from guinea-pig ear was investigated using an indirect enzyme digestion technique to observe both DNA and RNA, and a direct Schiff-Thallium reaction technique, to observe DNA alone. Similar results were obtained by both methods. The distribution of DNA and RNA change gradually in nuclei as epidermal cells differentiate. DNA in cells of the lower strata is localized in essentially the same areas in which electron-opaque components are seen by conventional electron microscopy. With the cytochemical treatments, however, we found that DNA is not present in all electron-opaque areas of nuclei in superficial granular cells. RNA is present in the nucleoli of cells in all layers, but its density is also lower in the upper granular cells. We postulate that nucleic acids in nuclei of granular cells gradually decrease and that the space is filled with newly synthesized electron-dense protein, as part of the differentiation process.

Animals↗
PubMed1977

Immunofluorescent studies of epidermal protein during UV induced carcinogenesis.

Previous studies with agar diffusion technique demonstrated that antibodies produced in rabbits by injection of urea extractable proteins of rat cornfied cells cross react with proteins extracted from normal epidermis of hairless mice using the same technique. In the present study we investigated by indirect immunofluorescence microscopy the immunoreactivity of epidermal proteins in normal and ultraviolet light (UVB) induced hyperplasia and malignant transformation. Reactivity to the antibody was seen over the entire epidermis of nontreated skin and hypertrophied epidermis which occurred at 6-8 weeks after initiation of UVB irradiation. However, the reactivity diminished when malignant changes took place in the epidermal cells. Almost complete disappearance of the immunoresponse was observed in squamous cell carcinoma produced by further UVB radiation. These results suggest that the reactivity of this urea extractable protein serves as an additional immunologic marker for normal epidermal cells. Alterations in the immunoreactivity parallels UVB induced carcinogenesis.

Animals↗
PubMed1976

Immunochemical studies of proteins in epidermal cornified cells of human and newborn rat.

1. Proteins were extracted from cornified cells of newborn rats and human palm with 8 M urea containing 0.1 M beta-mercaptoethanol. Two fractions, rat FIIIa and human F5.5 were obtained by acid precipitation for further study. 2. Antibodies raised in rabbits by injection of rat FIIIa gave two precipitin lines by agarose diffusion against rat FIIIa, but only one line against human F5.5. One of the antigenic determinants of rat FIIIa was found to be a protein of approximately 66 000 daltons. The other seems to be formed with two polypeptides in the range of 60 000 and 66 000 daltons. The antigenic determinant of human F5.5 was a protein of approximately 64 000 daltons which immunologically cross-reacted only with the antiserum to a protein of 66 000 daltons in rat FIIIa. 3. The antisera also cross-reacted with proteins extracted from epidermis of guinea pig, hamster, hairless mouse, dog ear, dog snout and dog foot pad, but did not react with the epidermis of either rabbit immunized with rat FIIIa or non-treated normal rabbit. 4. Indirect immunofluorescence demonstrated a reaction of rabbit anti-rat FIIIa serum over cornified cells as well as over granular, spinous and basal cells of the epidermis of newborn rat and human.

Animals↗
PubMed1976

Dibutyryl cyclic AMP-induced differentiation of epidermal cells in tissue culture.

Histochemical and biochemical techniques have been used to compare the effects of dibutyryl cyclic AMP on epidermal cells and dermal cells in primary tissue culture. Rhodamin B staining showed only scattered positive cells in nontreated epidermal cells and a few contaminating keratinizing cell foci in both nontreated and treated dermal cell cultures. In contrast, treated epidermal cells stained strongly and had many keratinizing cell foci. A significant increase in histidine, cystine, and arginine incorporation was noted in epidermal cells treated with dibutyryl cyclic AMP as compared to untreated epidermal cells and to dermal cell cultures both treated and untreated. Dibutyryl cyclic AMP had no significant effect on leucine and phenylalanine incorporation. These results seem to suggest that the intracellular level of cyclic AMP not only controls the synthesis of DNA by epidermal cells in culture but also induces the process of differentiation toward keratinization.

Arginine↗
PubMed1976

Inhibition of the in vivo effects of concanavalin-A on mammalian epidermis by alpha-methyl-D-glucopyranoside.

Concanavalin-A (Con-A) injected intradermally into newborn rats produces inhibition of granular-cell formation, accumulation of spinous cells, glycogen deposition, and a decrease followed by an increase in the number of basal cells in DNA synthesis. These changes were maximal with a dose of 0.1 mg Con-A, although 0.005, 0.01, and 0.05 mg caused some epidermal changes. The Con-A effects were partially blocked when 0.1 ml of 0.3 or 0.1 M alpha-methyl-D-glucopyranoside (alphaMG) solution was injected 2 hr after 0.1 mg Con-A and completely inhibited by injection of 0.1 ml of 3.0 M alphaMG solution. The inhibitory effects were not seen after injection of 0.1 ml of 3.0 M N-acetyl-galactosamine saline solution, or 0.1 ml normal saline. Injection of alphaMG alone did not cause any changes in epidermal cells. These results indicate that specific sugar inhibits Con-A effects on mammalian epidermis in vivo.

Acetylglucosamine↗
PubMed1976

The induction of human multinucleated monocytes in culture.

The formation of multinucleated giant cells in vitro has been studied using human monocyte cultures. Multinucleated monocytes appeared early in the course of the culture, indicating that they are not purely a phenomenon found only in aging cultures. In 4 of the 33 normal adults, more than 20% of multinucleated forms appeared in the cultures. Supernatants from early monocyte cultures of those subjects having a high incidence of multinucleated cells induced the formation of multinucleated cells in other cultures. This effect did not occur when supernatant from subjects with low counts of multinucleated cells were incubated with other monocyte cultures.

Cells, Cultured↗
PubMed1976

Keratinization.

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Animals↗
PubMed1975

Heterogenous proteins in keratohyaline granules studied by quantitative autoradiography.

Electron microscopic autoradiography with [3H]histidine, [3H]cystine, I13H]arginine, and [3H]proline was used to study protein synthesis in keratohyaline granules of newborn rats. All 3H-amino acids were incorporated into proteins in the granular cells, and the radioactive proteins appeared in the keratohyaline granules. However, the amount of radioactivity associated with the granules and the pattern of ultrastructural localization of the radioactive proteins differed considerably for each 3H-amino acid. "Histidine-labeled" protein was located mainly in the matrix portion of keratohyaline granules whereas "cystine-labeled" protein accumulated in the dense homogenous deposits. "Arginine-labeled" protein was distributed more diffusely in the organelles of granular cells, but that associated with keratohyaline granules seemed to localize mostly with "histidine-labeled" protein and partly with "cystine-labeled" protein. Large amounts of "proline-labeled" protein were also present in other areas of the cytoplasm than keratohyaline granules. This protein localized in the dense homogeneous deposits, but it seemed to turn over more rapidly than "cystine-labeled" protein, an indication that the dense homogenous deposits consist of at least two different polypeptide chains, one of which contains higher cystine and the other higher proline.

Amino Acids↗