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K Fukuyama

Publications and source records attributed to K Fukuyama.

At least 91 records · Page 5Linked to original sources
PubMed1992

Altered structure and expression of the p53 gene in human neuroepithelial tumors.

The p53 gene, located on chromosome 17p13.1, may be important in the pathogenesis of human neuroepithelial tumors, because it is a tumor suppressor gene and genetic alteration is essential for certain human cells to acquire the neoplastic phenotype. The structure and expression of the p53 gene were investigated in cultured human glioma cells and biopsied specimens of neuroepithelial tumors. Immunocytochemical examination of p53 gene expression revealed positive nuclear staining in six of seven glioma cell lines tested. Sodium dodecyl sulfate-polyacrylamide gel electrophoretic analysis demonstrated unequivocal heterogeneity of migration rate in p53 bands. Pulse-chase analysis clearly showed an increased half-life of p53 in cultured human glioma cells. These abnormalities are presumably due to genetic alterations in the p53 gene. Nucleotide substitutions in exon 5, 7, or 8 of the p53 gene could be detected by polymerase chain reaction-single strand conformational polymorphic analysis in four of seven (57%) human glioma cell lines, and nine of 29 (31%) biopsied specimens of neuroepithelial tumors examined. The present results indicate that genetic alterations in the p53 gene are responsible for the tumorigenesis of at least some human neuroepithelial tumors.

Astrocytoma↗
PubMed1992

Isolation and characterization of granuloma initiation factor.

A soluble component that transfers granulomatous tissue reaction was fractionated from Schistosoma mansoni egg-induced hepatic granulomas (SMHG) by Sephacryl S-300 column chromatography. The fractions separately bound to inert, Affi-Gel agarose beads were inoculated subcutaneously in naive mice. The low molecular weight fraction, consisting of proteins 23 kd, 20 kd, and 16 kd, produced organized granulomas 6 to 7 weeks after inoculation. This fraction was further purified by high-pressure liquid chromatography (HPLC) gel filtration and gave three fractions eluting at retention times of 44, 46, and 48 minutes. Each fraction contained all low-molecular-weight proteins in varying amounts and induced skin granulomas when inoculated subcutaneously. Amino acid sequence of the major 20-kd protein showed 11 N-terminal residues identical to those of cyclophilin. Antisera raised to the protein with retention time of 46 minutes, reacted with cells in the granulomas but not surrounding liver tissue as detected by immunofluorescence microscopy. The findings indicate a low molecular weight soluble fraction of SMHG can induce new granuloma formation when injected in an immobilized form into skin of naive mice. The results suggest granuloma initiation factor is a homolog of the cyclophilin gene family.

Animals↗
PubMed1991

Crystallization and preliminary X-ray crystallographic studies of bovine heart mitochondrial cytochrome bc1 complex.

Cytochrome bc1 complex (ubiquinol:ferricytochrome c oxidoreductase, EC. 1.10.2.2) from bovine heart mitochondria was crystallized by a batchwise method from protein solution containing sucrose monolaurate using polyethylene glycol-4000 as a precipitant. The red parallelepiped crystals grew to a size of approximately 1 mm x 1 mm x 1 mm. The crystalline protein showed enzymic activity catalyzing electron transfer from ubiquinol-2 to cytochrome c. The subunit composition and absorption spectrum of the crystalline enzyme were identical to those reported previously for the enzyme in solution. The crystal diffracted X-rays to 7.5 A resolution. The diffraction pattern indicated a monoclinic form, space group P2(1), and unit-cell constants of a = 196 A, b = 179 A, c = 253 A and beta = 97 degrees. Most probably four functional units are present in an asymmetric unit.

Animals↗
PubMed1991

Macrophage chemotactic factor partially purified from granulomatous inflammation.

Pathophysiological roles of macrophage chemotactic factor (MCF) in granulomatous inflammation were investigated. MCF was extracted in 10 mM phosphate-buffered saline, pH 7.4, from experimentally produced epithelioid cell granulomas in the liver and skin of mice. MCF activity reached a peak in the lesions prior to the time when granulomatous inflammation became maximal. MCF was then purified from 10-week-old hepatic granulomas and 2-week-old skin lesions by gel filtration, ion exchange column chromatography, and HPLC gel filtration. MCF from either liver or skin had a molecular weight about 650 kDa. MCF from hepatic granulomas was coupled to Affi-Gel beads and transplanted subcutaneously into naive mice. In vivo macrophage chemotaxis was observed around the beads and the cells formed a sheet, but organization of macrophages into granulomas did not occur with the MCF-active fractions. Macrophage chemotaxis alone is insufficient to elicit granulomatous inflammation.

Animals↗
PubMed1991

An improved noninfectious murine skin model of organized granulomatous inflammation.

An improved model of granulomatous inflammation in skin was developed by second passage skin grafting of isolated, lyophilized skin granulomas, originally elicited in naive mice by inoculations of lyophilized hepatic schistosome egg granulomas. The tissue reaction is caused by a single exposure to a noninfectious, acellular granulomagenic stimulus and occurs in healthy mice free of systemic disease. The model should prove useful for isolation of granuloma initiation factor(s). Furthermore, because there is a time lag before new granuloma formation begins, a window exists for analytical dissection of the initiation process. In this study we described the responses of host cells by autoradiography, and light and electron microscopy. The activity of angiotensin-converting enzyme and proline-specific endopeptidase showed a modulation during granuloma formation. In addition we found that severe immunosuppression with high dose cyclosporine therapy did not alter granuloma formation, supporting the idea that initiation of organized granulomas is T-cell independent.

Animals↗
PubMed1991

Immunogenetic influences on skin granuloma formation in mice.

Genetic influence on the development of granulomatous tissue reaction was investigated in C57BL/6 mice. Granulomas developed in the skin of euthymic C57BL/6 mice by transplantation of lyophilized hepatic granulomas were excised and lyophilized. The tissue mass free of parasite egg antigen and living cells was grafted into the skin of euthymic, athymic (nu/nu), and beige (bg/bg) C57BL/6 mice. Histological changes at the skin sites were studied weekly by light microscopy, and cells in newly developed granulomas at 6 weeks after grafting were examined by electron microscopy. Granulomatous inflammation occurred in all the variants but morphometric analysis showed that granulomatous inflammation was the most extensive in beige mice and least in athymic mice. The differences in the degree of tissue reaction were also quantified by measuring angiotensin converting enzyme and prolyl endopeptidase. Statistically significant differences among the animals with varying genetic background were confirmed by the marker enzyme activity. The findings confirm that initiation of a granulomatous response does not require T cells but T cell function is important for full expression of the reaction, while NK cell activity seems to suppress granuloma formation.

Animals↗
PubMed1991

Crystallization and preliminary X-ray diffraction study of cytochrome c552 from Hydrogenobacter thermophilus.

Cytochrome c552 from a thermophilic hydrogen-oxidizing bacterium, Hydrogenobacter thermophilus, exhibits remarkable thermostability. The oxidized cytochrome c552 has been crystallized in an ethanol/water mixture by means of the vapor diffusion method. The crystals belong to the orthorhombic system, space group P2(1)2(1)2, with unit cell dimensions of a = 93.4 A, b = 52.9 A, and c = 32.4 A. Most probably the asymmetric unit contains two molecules of cytochrome c552. The crystals diffract X-rays to better than 2.5 A resolution and are stable to X-ray irradiation.

Bacteria, Aerobic↗
PubMed1991

Dural arteriovenous malformation associated with occlusion of the superior sagittal sinus--case report.

The authors report a rare case of dural arteriovenous malformation (AVM) associated with occlusion of the superior sagittal sinus. A 78-year-old female developed transient aphasia, followed by generalized convulsion. Common carotid angiography showed a dural AVM fed by the bilateral middle meningeal arteries, draining to the superior sagittal sinus, and sinus occlusion. 123I-single photon emission computed tomography demonstrated decreased blood flow in the bifrontal-parasagittal regions. Transcatheter embolization via the feeding arteries improved the cerebral blood flow around the lesion, and the symptoms disappeared.

Aged↗
PubMed1990

Structure of the [2Fe-2S] ferredoxin I from the blue-green alga Aphanothece sacrum at 2.2 A resolution.

Crystals of a [2Fe-2S] ferredoxin (Fd) I with a relative molecular mass of 10,480 were obtained from the blue-green alga Aphanothece sacrum. Each asymmetric unit of the crystal contains four molecules. An electron density map calculated by the single isomorphous replacement method with the anomalous dispersion at 2.5 A resolution was refined by averaging the four molecules in the asymmetric unit. Positional and isotropic thermal parameters for the non-hydrogen atoms of the four molecules and 158 water molecules were refined to an R-factor (R = sigma[Fo-Fc[/sigma Fo) of 0.23 by the restrained least-squares method. The estimated root-mean-square (r.m.s.) error for the atomic positions is 0.3 A. The r.m.s. deviations of equivalent C alpha atoms of the asymmetric-unit molecules superposed by the least-squares method average 0.35 A. The Fd molecule has a structure like the beta-barrel in the molecule of the [2Fe-2S] Fd from Spirulina platensis. A [2Fe-2S] cluster is bonded covalently to the protein molecule by four Fe-S, in which three of the Fe-S bonds are in a loop segment from position 38 to 47. The hydrophobic core inside the beta-barrel is formed by seven conservative residues: Val15, Val18, Ile24, Leu51, Ile74, Ala79 and Ile87. The molecular surface around Tyr23, Tyr80 and the active center may interact with ferredoxin-NADP+ reductase. One of the two iron atoms of the [2Fe-2S] cluster should be more easily reduced than the other because of differences in the hydrogen-bonding scheme and the hydrophobicity around the atoms.

Amino Acid Sequence↗
PubMed1990

Tertiary structure of oxidized flavodoxin from an eukaryotic red alga Chondrus crispus at 2.35-A resolution. Localization of charged residues and implication for interaction with electron transfer partners.

The crystal structure of the oxidized form of a flavodoxin from an eukaryotic red alga, Chondrus crispus, has been determined by multiple isomorphous replacement and anomalous scattering methods. A model of the 173 residues and flavin mononucleotide (FMN) has been refined by a restrained least squares method to a crystallographic R-factor of 22.6% using 6236 reflections between 6.0 and 2.35 A with F greater than 3 sigma F. This molecule has a sheet consisting of five parallel beta-strands with two alpha-helices on one side of the sheet and three on the other side, and has a (beta alpha)5 structure. The molecule incorporates a substantial insertion in beta 5, as in Anacystis nidulans flavodoxin, which distinguishes these flavodoxins from the short-chain type. The isoalloxazine ring of FMN is sandwiched between the side chains of Trp-56 and Tyr-98, with its C-7 and C-8 methyl groups being exposed to solvent. The phosphate group of FMN is located at the N-terminal end of alpha 1, and forms extensive hydrogen bonds with the loop (T8-T13) between beta 1 and alpha 1 of the protein. Six of the total 11 lysine residues are clustered at the opposing face to the FMN-binding site, while about two-thirds of the total 35 acidic residues are located in the half of the molecule which includes the FMN-binding site. Such localization of charged residues produces a dipole within the molecule, which may be important in its recognition of the other proteins participating in electron transfer reactions.

Amino Acid Sequence↗
PubMed1990

Glycoconjugate expression of cells of human anagen hair follicles during keratinization.

Changes in the expression of glycoconjugates in cells of the inner root sheath (IRS) and outer root sheath (ORS) of human anagen hair follicles were investigated by lectin histochemistry. Concanavalin A (Con A) and Ricinus communis (RCA-I) stained hair follicle cells regardless of their differentiation stages. In IRS, Ulex europeaus-I (UEA-I) bound to the surface of the cells as soon as they were morphologically defined, and Glycine max (SBA) stained as their differentiation progressed. Innermost (IM) cells of ORS layers were reactive with UEA-I at the stage where Henle's cells were keratinized, while the reactivity of UEA-I was lost at the site of the completion of IRS keratinization where SBA reaction was detected. Staining of both UEA-I and SBA was prominent in other ORS cells at the levels where SBA binding in IM cells became strong. The staining intensity increased up to the position of the follicular isthmus. In addition, a sugar residue recognized by Dolichos biflorus (DBA) was detected in differentiated cells of ORS. In contrast, the DBA reaction was not found at all in cells of IRS, infundibulum, and epidermis. These findings identified a complexity of carbohydrate metabolism in the cells of different layers at various stages of keratinization. IM cells differentiate independently from other ORS cells but seem responsive to the degree of IRS keratinization. All ORS cells possess a unique sugar moiety not found in other keratinocytes either in the hair or epidermis.

Concanavalin A↗
PubMed1990

Angiotensin II-producing proteases from granulomatous tissue reaction in mice infected with Schistosoma mansoni.

1. Angiotensin I hydrolases, Mr 140,000 and Mr 70,000 were separated by gel filtration from Tris-HCl buffer extract of hepatic granulomas developed in mice with schistosomiasis. Two enzymes had different substrate specificity. 2. Mr 140,000 hydrolase activity was inhibited by captopril as reported for angiotensin converting enzyme (ACE), while that of Mr 70,000 hydrolase activity was inhibited by potato carboxypeptidase inhibitor. 3. An intermediary, des-Leu10-angiotensin I and then angiotensin II were formed from angiotensin I by Mr 70,000 hydrolase. 4. The findings suggest that Mr 70,000 enzyme is tissue carboxypeptidase A, and it generates angiotensin II in granulomatous inflammation as does ACE.

Angiotensin I↗
PubMed1990

Relationship between NK cells and granulomatous inflammation in mice.

The relationship between natural killer (NK) cells and granulomatous inflammation was investigated using two experimental granuloma models in C57BL/6 mice. The hepatic granuloma model was produced by infection with Schistosoma mansoni (S. mansoni), and the skin granuloma model was established by subcutaneous inoculation of the isolated hepatic granulomas. NK cell activity in lymph nodes and spleen, measured by 51Cr-release assay against YAC-1 cells, was compared to that in age-matched control mice. The activity decreased progressively as granulomas developed without changing the number of NK-1.1+ cells. The reduced NK cell activity was not reversed by administration of indomethacin. In order to further substantiate whether NK cells contribute to granulomatous inflammation, NK cells were depleted in the mice by injection of anti-NK-1.1 mAb. Reduction of NK cell activity (70-90%) was achieved during granuloma formation. An increase of about 20% in the mean granuloma diameter was detected in the mAb-treated mice in both models. Moreover, the percentage of granuloma takes in the skin model was enhanced (65% increase). The mAb treatment did not alter T cell counts in granulomas, T cell subset numbers or proliferative response in spleen. These findings indicate that NK cells directly play a regulatory role in granulomatous inflammation.

Animals↗
PubMed1990

Local effects of granulomatous inflammation on functional activation of T cells in athymic mice.

We investigated the effect of granulomatous inflammation in skin on lymphocyte maturation in athymic (nu/nu) mice. Hepatic egg granulomas developed in euthymic (nu/+) mice with schistosomiasis were transplanted into skin of nu/nu mice. During skin granuloma development the rate of DNA synthesis and interleukin 2 activity of lymphocytes from lymph nodes, with and without concanavalin A stimulation, showed that the nu/nu cells were activated to levels of untreated nu/+ lymph node cells. Activation of splenic lymphocytes was not detected in the grafted nu/nu mice. Also, immunohistochemical staining demonstrated an increase in cells expressing Thy 1.2, Lyt-1 or L3T4 surface markers in the skin and lymph nodes, but not in spleen. The findings indicate that a granulomatous reaction in nu/nu mouse skin induces local, but not systemic, proliferation and differentiation of lymphocytes, to a low degree compatible with resting nu/+ mice.

Animals↗
PubMed1989

Structure of [4Fe-4S] ferredoxin from Bacillus thermoproteolyticus refined at 2.3 A resolution. Structural comparisons of bacterial ferredoxins.

The structure of a low-potential ferredoxin isolated from Bacillus thermoproteolyticus has been refined by a restrained least-squares method. The final crystallographic R factor is 0.204 for 2906 reflections with F greater than 3 sigma F in the 6.0 to 2.3 A resolution range. The model contains 81 amino acid residues, one [4Fe-4S] cluster, and 59 water molecules. The root-mean-square deviation from ideal values for bond lengths is 0.018 A, and the mean coordinate error is estimated to be 0.25 A. The present ferredoxin is similar in the topology of the polypeptide backbone to the dicluster-type ferredoxins from Peptococcus aerogenes and Azotobacter vinelandii, but has considerable insertions and deletions of the peptide segments as well as different secondary structures. Although all but the C-terminal C zeta atoms of P. aerogenes ferredoxin superpose on the C alpha atoms of A. vinelandii ferredoxin, only 60% superpose on the C alpha atoms of B. thermoproteolyticus ferredoxin, with a root-mean-square distance of 0.82 A for each pair. The conformations of the peptide segments surrounding the [4Fe-4S] clusters in these three ferredoxins are all conserved. Moreover, the schemes for the NH...S hydrogen bonds in these ferredoxins are nearly identical. The site of the aromatic ring of Tyr27 in B. thermoproteolyticus ferredoxin is close spatially to that of Tyr28 in P. aerogenes ferredoxin with reference to the cluster, but these residues do not correspond in the spatial alignment of their polypeptide backbones. We infer that in monocluster-type ferredoxins, the side-chain at the 27th residue has a crucial effect on the stability of the cluster. Of the four cysteine residues that bind to the second Fe-S cluster in the dicluster-type ferredoxins, two are conserved in the monocluster-type ferredoxins from Desulfovibrio gigas. D. desulfuricans Norway, and Clostridium thermoaceticum. The tertiary structure of B. thermoproteolyticus ferredoxin suggests that in such monocluster-type ferredoxins these two cysteine residues, which in it correspond to Ala21 and Asp53, form a disulfide bridge.

Amino Acid Sequence↗