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Y Shirakata

Publications and source records attributed to Y Shirakata.

51 records · Page 3Linked to original sources
PubMed1993

Multiple cis-acting elements are required for proper transcription of the mouse V delta 1 T cell receptor promoter.

To gain insight into the developmentally regulated expression of the mouse TCR V delta-gene segments, we have investigated the role of the 5' promoter region of the V delta 1-gene. Transient transfection assays showed that a construct encompassing 267 nucleotides upstream from the mapped transcriptional start site was capable of driving promoter activity when transfected into V delta 1+ T cells. The inclusion of an additional 459-bp 5' segment to this construct did not affect promoter activity. However, a deletion of 222 5' nucleotides from the same construct dramatically decreased promoter activity. In vivo genomic footprinting localized several protein-DNA interactions to the stretch of DNA shown to have transcriptional activity. A computer analysis revealed that the segments of DNA participating in these protein-DNA interactions were identical to the previously described cyclic AMP response element (CRE), E box, and leukemia virus E26 cis-acting elements. Transient transfection assays performed with -267 bp constructs containing mutations at each of the localized cis-acting elements revealed that the CRE, E box, and Ets elements work together in driving promoter activity and that the CRE and Ets elements are the most important for driving transcription. Gel mobility shift analyses showed that each of these cis-acting elements is capable of binding specific nuclear factors present in V delta 1-expressing cells. These data indicate that multiple transcription factors acting in concert are responsible for V delta 1 gene expression.

Animals↗
PubMed1993

High-dose tocopherol acetate therapy in epidermolysis bullosa siblings of the Cockayne-Touraine type.

We have employed a successful therapy for epidermolysis bullosa with high-dose oral tocopherol acetate. Two siblings with dominant dystrophic epidermolysis bullosa of the Cockayne-Touraine type were reported. Both siblings suffered from tense blisters and erosions healing with scars and transient milia on the extremities. Electron microscopic study of the blisters revealed a cleavage beneath the lamina densa in both siblings. High-dose oral tocopherol acetate therapy was administered to them with satisfactory clinical reduction of the blister formation.

Epidermolysis Bullosa Dystrophica↗
PubMed1992

Isolation and characteristics of scallop sarcoplasmic reticulum with calcium transport activity.

Sarcoplasmic reticulum with calcium transport activity has been isolated from the cross-striated adductor muscle of the scallop, which lives in cold (< or = 20 degrees C) sea water, by using pH 7.0 buffer solution both to homogenize the tissue and to sediment the membrane fraction. The yield of the preparation was 60-100 mg protein from 100 g of the scallop muscle. Ca(2+)-activated ATPase protein of about 100 kDa accounted for 40-50% of the protein preparation. The maximum activities of ATP-dependent, oxalate-facilitated calcium accumulation and Ca(2+)-ATPase were observed at a pH of about 7.0 and temperature of 20-30 degrees C, and their values were about 2 mumol Ca2+/mg of protein/min and about 3 mumol ATP hydrolysis/mg of protein/min, respectively. At 0 degree C, 10-20% of these activities was maintained, while at 37 degrees C, the activities were irreversibly lost. The Ca(2+)-ATPase activity was half-maximally activated at about 0.3 microM [Ca2+]. The ATPase activity exhibited non-Michaelian behavior with respect to ATP, with two different Km values of approximately 10 microM and 0.1-0.3 mM. GTP, CTP, and ITP were also hydrolyzed by the preparation at a rate of 10-30% of that of ATP. The preparation was stored at -80 degrees C with retention of function for about a year.

Adenosine Triphosphate↗
PubMed1991

Expression and characterization of membrane co-factor protein (MCP) in human skin.

Membrane co-factor protein (MCP; CD46) is an integral membrane protein with molecular weight (MW) of the two species of 63 kD and 55 kD, and regulates autologous complement activation, with the activity of factor I cofactor. The quantity of each species is genetically regulated, and two codominantly inherited allelic variants account for the three phenotypic patterns. By immunohistochemical study, MCP was found both in the intercellular spaces of the epidermis and on the endothelial cells in the dermis of normal human skin in vivo. The intensity of the staining pattern was higher in the basal layer than in the granular layer. By Western blot analysis with use of a monoclonal antibody, MCP in the epidermis appeared as several bands ranged from 60-50 kD, with a major band of 56 kD, which was different from those in either polymorphonuclear cells, platelets, and cultured keratinocytes. No other variants were found in the epidermis obtained from skin of 20 normal humans. Complement activation in human skin may be regulated at several steps, including DAF and HRF20, thereby protecting cells from autologous complement attack.

Antigens, CD↗
PubMed1991

Characterization of decay-accelerating factor (DAF) in human skin.

Decay-accelerating factor (DAF) is a 70-kD membrane glycoprotein that regulates autologous complement activation, by preventing assembly of alternative or classical C3/C5 convertases, and has been shown to have a wide tissue distribution. In this study, DAF antigen has been demonstrated at the intercellular spaces of normal human epidermis with monoclonal antibody against DAF using the peroxidase-anti-peroxidase method. The amount of DAF was greater at the granular layer than the basal cell layer as judged by intensity of the staining. Western blot analysis of DAF in the epidermis showed a 55-kD band, whereas that of buffy coat cells was approximately 67 kD. When DAF of the epidermis was treated with neuraminidase, the molecular weight was reduced to 53 kD, whereas that of buffy coat cells was 56 kD. These results indicated that the content of sialic acid of DAF in the epidermis was different from that of buffy coat cells. In phosphatidylinositol-specific phospholipase C (PIPLC)-treated normal human skin, DAF was not demonstrated in the epidermis, whereas DAF remained unchanged on the elastic fibers. After the treatment of the epidermis by PIPLC, DAF was released into the buffer shown by Western blot analysis. These results suggested that DAF on the epidermis was anchored to keratinocyte via phosphatidylinositol (PI), whereas the anchoring mechanism of DAF on the elastic fibers was not through PI.

Blotting, Western↗
PubMed1991

Diffuse neonatal hemangiomatosis with a giant cutaneous hemangioma on the axilla.

A case of diffuse neonatal hemangiomatosis is described. At birth, a giant tumor, 10 cm in diameter and 6 cm in height, was present on the left axilla and associated with thrombocytopenia. Numerous cherry-red papules were present on the skin and buccal mucosa. On the second day of life, the giant tumor was excised. Histological examination of the tissue revealed masses of endothelial cells embedded in fibrous tissues. Plain and enhanced computed tomography of the brain revealed a hemorrhage and two hemangiomas. The associations of intravascular papillary endothelial hyperplasia were discussed.

Axilla↗
PubMed1991

Complement allotypes in Japanese patients with pemphigus and bullous pemphigoid.

The allotypes of C6, C7, factor B (BF) and factor I (IF) of the human complement system were studied in 11 Japanese patients with pemphigus (5 with pemphigus vulgaris and 6 with pemphigus foliaceus) and 17 with bullous pemphigoid (BP) to investigate the genetic background of these diseases. The allotypes were detected by using isoelectric focusing and immunoblotting. The frequency for IF*A allele in the pemphigus patients was significantly higher (p = 0.009) than that in healthy controls (n = 60). A significant association of IF A allotype with pemphigus was also observed (p = 0.027), with a relative risk of 6.3. There was no association between the C6, C7, BF or IF allotypes and BP. These data suggest that IF A allotype may be an etiological genetic factor in the development of pemphigus.

Alleles↗
PubMed1990

Subclass characteristics of IgG autoantibodies in bullous pemphigoid and pemphigus.

Immunoglobulin (Ig) G subclasses in anti-basement membrane zone (BMZ) autoantibodies found in the sera of bullous pemphigoid (BP) and in anti-intercellular substance (ICS) autoantibodies of pemphigus were investigated using immunofluorescent (IF) staining. In BP, IgG4, IgG1, and IgG2 were detected in 13, 5 and 6 of 15 patients, respectively; IgG3 was not detected. In pemphigus, IgG4 was detected in all of 10 patients, IgG1 in 7, IgG2 in one, and IgG3 in one patient, respectively. In both BP and pemphigus, the most prominent subclass in intensity of IF staining was IgG4. Although one BP and one PV patient had only IgG4 autoantibodies, C3 deposition was detected. The quantification of IgG subclasses in the sera of the patients was performed by enzyme-linked immunosorbent assays (ELISA). Serum levels of IgG4 in both BP and pemphigus were elevated approximately 3-fold over those in normal controls; those of whole IgG and IgG1-3 were not significantly elevated. Using direct IF staining, the deposition of C3 at the BMZ and at the ICS was demonstrated in 9 of 10 BP and in 3 of 8 pemphigus patients, respectively. The prominent IgG subclasses of anti-BMZ and anti-ICS antibody were IgG4, a noncomplement-fixing antibody, suggesting that the deposition of C3 in the lesional skin occurred via the alternative pathway, or that small amounts of IgG1-3 subclass autoantibodies activated the classical pathway.

Autoantibodies↗
PubMed1990

Characterization of homologous restriction factor (HRF20) in human skin and leucocytes.

Homologous restriction factor with a molecular weight of 20 kD (HRF20) is a membrane protein that inhibits assembly of the membrane attack complex of homologous complement. Distribution of HRF20 in normal human skin was studied. The plasma membrane of keratinocytes was stained, and the intensity of the staining pattern was higher in the basal cell layer than in the granular layer. Endothelial cells of blood vessels in the dermis were also stained. The molecular weight of HRF20 on erythrocytes and epidermis is 16 kD, determined by Western blot analysis. Those of polymorphonuclear cells and lymphocytes appeared as two bands, a major band of 20 kD and a minor band of 16 kD. Susceptibility of HRF20 to phosphatidylinositol-specific phospholipase C (PIPLC) was examined. After PIPLC treatment of the sections, HRF20 was not detected on the epidermis and was very slightly expressed on the blood vessels. These results indicate that HRF20 attaches to keratinocytes and blood vessels via phosphatidylinositol, regulating the formation of membrane attack complexes of homologous complement on the cell membrane.

Blood Proteins↗
PubMed1989

The X gene of hepatitis B virus induced growth stimulation and tumorigenic transformation of mouse NIH3T3 cells.

To examine the transforming potential of the X gene product of hepatitis B virus (HBV), the X-gene-containing region (referred to as the HBx region) was introduced into mouse NIH3T3 cells. Each transformed cell line expressed X-coding mRNA at a different level. A positive correlation was found between the level of X-coding mRNA and the saturation density of the cells. The HBx-transformed cell lines exhibited X protein production and tumor formation in nude mice. The function of HBV in oncogenesis may involve the continuous expression of the X-gene-coded product in the HBV DNA-integrated cells.

Animals↗
PubMed1989

Oncogenic potential of hepatitis B virus.

The function of the X gene was clarified by examination of the transient production of hepatitis B virus (HBV) particles by transfected recombinant HBV DNA (pHBV-3 DNA) and its frameshift mutant (delta X) of the X open reading frame into hepatocellular carcinoma HuH-7 and HepG2 cells. No reduction of viral mRNAs was observed in the HuH-7 cells by the delta X mutant, whereas mRNAs underwent marked reduction in HepG2 cells. No reduction in core particle production was observed in HuH-7 cells, but in HepG2 cells reduction was considerable. To clarify the significance of the delta X mutation in the trans-acting function of the X gene in hepatoma cells, the chloramphenicol acetyl/transferase (CAT) assay was conducted. Transfection of plasmid pHBV-3 into HepG2 cells increased CAT activity of pSV2-CAT, while the delta X mutation clearly showed no stimulation of activity. On the other hand, in the HuH-7 cells, pHBV-3 exhibited no such stimulation. The trans-acting function of the X gene product in two different hepatoma cells was clearly shown to differ. Furthermore, transfection of X gene expression plasmid pKSV-HBx into mouse NIH3T3 cells increased the CAT activity of pSV2-CAT. Trans-activation was still detectable even following deletion of enhancer sequences in the pSV2CAT. The oncogenic potential of HBV is discussed with special attention to the X gene product, which may be able to activate a cellular transcription factor at the viral and cellular promotor sequences in the cells.

Animals↗
PubMed1987

Hepatitis B virus (HBV) particles are produced in a cell culture system by transient expression of transfected HBV DNA.

An in vitro system for the production of hepatitis B virus (HBV) particles was established by the transient expression of transfected HBV DNA using a human hepatocellular carcinoma cell, HuH-7, as a recipient. The 3.6- and 2.2-kilobase transcripts observed were similar to those in virus-infected liver cells. Both transcripts revealed the microheterogeneity of their 5' ends. The formation of virus-related particles subsequent to the RNA transcription was demonstrated. The core particles observed in the cytoplasm and the virus particles secreted in the culture medium contained the replicative intermediates of HBV DNA and banded at densities of 1.35-1.36 g/cm3 and 1.22-1.24 g/cm3, respectively. Furthermore, the in vitro mutagenesis of the template HBV DNA demonstrated that the P gene as well as the C gene products were essential for the production of HBV particles.

Amino Acid Sequence↗