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Biomedical subjects

H Hirai

Publications and source records attributed to H Hirai.

At least 127 records · Page 7Linked to original sources
PubMed2000

Targeted killing of carcinoembryonic antigen (CEA)-producing cholangiocarcinoma cells by polyamidoamine dendrimer-mediated transfer of an Epstein-Barr virus (EBV)-based plasmid vector carrying the CEA promoter.

The present study reports a novel nonviral method to efficiently and specifically target carcinoembryonic antigen (CEA)-producing cholangiocarcinoma (CC) cells in vitro. Epstein-Barr virus (EBV)-based and conventional plasmid vectors were constructed that possess the beta-galactosidase (beta-gal) or herpes simplex virus-1 (HSV-1) thymidine kinase (Tk) genes as well as tandem repeats of the human genomic sequence -82 to -42 bp from the transcriptional start site of the CEA gene. The plasmids were transfected by means of polyamidoamine dendrimer into CEA-positive (HuCC-T1) or -negative cell lines. Transfection of the conventional plasmid vector with the CEA promoter and beta-gal gene resulted in a very low or undetectable level of marker gene expression even in the CEA-positive cell line. Transferring the HSV-1 Tk gene by conventional plasmid did not affect the susceptibility of HuCC-T1 cells to ganciclovir. In marked contrast, strong beta-gal expression was specifically obtained in HuCC-T1 cells by transfecting the EBV-based plasmid in which the CEA promoter and a ubiquitous promoter (SRalpha) are employed to drive the EBV-encoded nuclear antigen 1 (EBNA1) and beta-gal genes, respectively (pTES.beta). Furthermore, CEA-positive but not -negative tumor cells were rendered highly susceptible to ganciclovir when transfected with the EBV-based vector that carries the CEA promoter-EBNA1 and SRalpha-HSV-1 Tk genes (pTES.Tk). These results strongly suggest that the EBV-based plasmid vector/cationic polymer system (EBV/polyplex) equipped with the CEA promoter provides an efficient nonviral method for the targeted gene therapy of CEA-producing malignancies.

Carcinoembryonic Antigen↗
PubMed2000

Generation of HLA-DRB1*1501-restricted p190 minor bcr-abl (e1a2)-specific CD4+ T lymphocytes.

A small population of cells in acute lymphoblastic leukaemia is characterized by a specific translocation of the c-abl oncogene on chromosome 9 to the break point cluster lesion (bcr) on chromosome 22, t(9; 22)(q34; q11) (e1a2). Theoretically, the junction-spanning sequences of oncogene fusion proteins might be ideal targets for immunotherapy because these are not present in normal cells. In this study, we show for the first time that in vitro immunization with a 17-mer e1a2 peptide representing the p190 minor bcr-abl fusion protein resulted in HLA-DRB1*1501-restricted peptide-specific proliferative CD4+ T lymphocytes, using peptide-pulsed monocyte-derived dendritic cells as the antigen-presenting cells.

CD4-Positive T-Lymphocytes↗
PubMed2000

Clustering of delta glutamate receptors is regulated by the actin cytoskeleton in the dendritic spines of cultured rat Purkinje cells.

The interaction of neurotransmitter receptors with the cytoskeleton is an important mechanism for the targeting of receptors to the postsynaptic membrane. Using cytoskeleton-perturbing agents, it was demonstrated that delta glutamate receptors, predominantly expressed on the dendritic spines of cerebellar Purkinje cells, are anchored to the actin cytoskeleton. The number of delta glutamate receptor-immunoreactive clusters was dramatically decreased following treatment of the Purkinje cells with the actin-disrupting agents, cytochalasin D or latrunculin A, without any significant effect on the number of presynaptic contacts of the granule cell axons. The clusters disrupted by latrunculin A were re-established 24 h after removal of the drug. These results suggest that morphological changes in the actin cytoskeleton regulate the delta glutamate receptor clustering on the dendritic spines, and may affect synaptic efficacy and plasticity.

Actin Cytoskeleton↗
PubMed2000

Anti-rat CD45RA monoclonal antibodies cross-react with glucagon.

Anti-rat CD45RA monoclonal antibody (MAb) has been used as a marker for rat B-lymphocytes (B-cells). When we applied the MAb to identify B-lymphocytes among immune cells infiltrating pancreatic islets of the BB rat, which is an animal model for human type 1 diabetes mellitus, a subpopulation of islet cells was immunostained as well. Our immunohistochemical investigations demonstrated that the stained islet cells are pancreatic alpha-cells and the MAb cross-reacts with glucagon.

Animals↗
PubMed2000

Association of insulin receptor substrate proteins with Bcl-2 and their effects on its phosphorylation and antiapoptotic function.

Insulin receptor substrate (IRS) proteins are docking proteins that couple growth factor receptors to various effector molecules, including phosphoinositide-3 kinase, Grb-2, Syp, and Nck. Here we show that IRS-1 associates with the loop domain of Bcl-2 and synergistically up-regulates antiapoptotic function of Bcl-2. IRS-2 but not IRS-3 binds to Bcl-2, and IRS-1 associates with Bcl-XL but not with Bax or Bik. Overexpression of IRS-1 suppresses phosphorylation of Bcl-2 induced by stimulation with insulin, and the hypophosphorylation may lead to its enhanced antiapoptotic activity. The binding site for Bcl-2 is located on the carboxyl half-domain of IRS-1. IRS-3, which lacks the corresponding region, dominant-negatively abrogates the survival effects of IRS-1 and Bcl-2. For the antiapoptotic activity of IRS-1, binding to Bcl-2 is more critical than activating phosphoinositide-3 kinase. Our results indicate that IRS proteins transmit signals from the insulin receptor to Bcl-2, thus regulating cell survival probably through regulating phosphorylation of Bcl-2.

Animals↗
PubMed2000

Real-time automated PCR for early diagnosis and monitoring of cytomegalovirus infection after bone marrow transplantation.

The purpose of this study was to assess the usefulness of real-time automated PCR as a quantitative, highly reproducible, and sensitive method to detect cytomegalovirus (CMV) DNA in blood specimens. Intra- and interassay precision rates were 0.89% (small number of copies [L]), 1.43% (middle number of copies [M]), and 1.12% (high number of copies [H]), and 4.46% (L), 1.51% (M), and 2.28% (H), respectively. The linearity of this assay was obtained between 10 and 10(7) copies/well, with a minimum detection limit of 20 copies/well. Specimens from 55 of 70 healthy subjects were found to be positive for CMV antibody, but CMV DNA was not detected in any of them. In the qualitative assessment of each specimen, the results of the CMV antigenemia assay and those of the real-time PCR assay agreed in 80% (plasma specimens), 79% (all nucleated cells), and 86% (blood) of the cases examined. For eight patients diagnosed as having CMV infection or disease, no sample was positive in the antigenemia assay earlier than in the real-time PCR assay. Furthermore, the results of this assay could be obtained within 8 h. We concluded that the real-time PCR assay is useful for rapid diagnosis of CMV infection and monitoring of clinical courses.

Adolescent↗
PubMed2000

Binding of Delta1, Jagged1, and Jagged2 to Notch2 rapidly induces cleavage, nuclear translocation, and hyperphosphorylation of Notch2.

Delta1, Jagged1, and Jagged2, commonly designated Delta/Serrate/LAG-2 (DSL) proteins, are known to be ligands for Notch1. However, it has been less understood whether they are ligands for Notch receptors other than Notch1. Meanwhile, ligand-induced cleavage and nuclear translocation of the Notch protein are considered to be fundamental for Notch signaling, yet direct observation of the behavior of the Notch molecule after ligand binding, including cleavage and nuclear translocation, has been lacking. In this report, we investigated these issues for Notch2. All of the three DSL proteins bound to endogenous Notch2 on the surface of BaF3 cells, although characteristics of Jagged2 for binding to Notch2 apparently differed from that of Delta1 and Jagged1. After binding, the three DSL proteins induced cleavage of the membrane-spanning subunit of Notch2 (Notch2(TM)), which occurred within 15 min. In a simultaneous time course, the cleaved fragment of Notch2(TM) was translocated into the nucleus. Interestingly, the cleaved Notch2 fragment was hyperphosphorylated also in a time-dependent manner. Finally, binding of DSL proteins to Notch2 also activated the transcription of reporter genes driven by the RBP-Jkappa-responsive promoter. Together, these data indicate that all of these DSL proteins function as ligands for Notch2. Moreover, the findings of rapid cleavage, nuclear translocation, and phosphorylation of Notch2 after ligand binding facilitate the understanding of the Notch signaling.

Animals↗
PubMed2000

CIZ, a zinc finger protein that interacts with p130(cas) and activates the expression of matrix metalloproteinases.

p130(cas) (Cas) is a docking protein that contains an SH3 domain and multiple tyrosine residues. p130(cas) is located at focal adhesions, is tyrosine phosphorylated in response to integrin stimulation, and is thought to transmit signals, via c-Crk and other proteins, for the remodeling of actin stress fibers and cell movement. In a search for the ligands of the SH3 domain of p130(cas) by far-Western screening, we cloned a novel protein named CIZ (for Cas-interacting zinc finger protein). CIZ consists of the following: a putative leucine zipper; a serine/threonine-rich region; a proline-rich sequence; five, six, or eight Krüppel-type C(2)H(2) zinc fingers; and the glutamine-alanine repeat. CIZ binds Cas in cells and is located in the nucleus and at focal adhesions. We showed that CIZ is a nucleocytoplasmic shuttling protein, by using the transient interspecies heterokaryon formation assay. In order to search for the targets of CIZ in nucleus, we determined the DNA binding consensus of CIZ as (G/C)AAAAA(A) by cyclic amplification and selection of targets analysis. The consensus-like sequences are found in several promoters of matrix metalloproteinases (MMPs), which are the enzymes used to degrade the extracellular matrix proteins. CIZ binds to a consensus-like sequence in the MMP-1 (collagenase) promoter. Overexpression of CIZ upregulates the transcriptions from MMP-1, MMP-3 (stromelysin), and MMP-7 (matrilysin) promoters, and this transactivation was enhanced in the presence of Cas. Furthermore, the stable overexpression of CIZ promoted the production of MMP-7 in culture medium. In summary, CIZ, a novel zinc finger protein, binds Cas, is a nucleocytoplasmic shuttling protein, and regulates the expression of MMPs.

Amino Acid Sequence↗
PubMed2000

Antisweet natural products. XV. Structures of Jegosaponins A-D from Styrax japonica Sieb. et Zucc.

From the fresh fruits of Strrax japonica Sieb. et Zucc., four new triterpenoid glycosides, named jegosaponins A-D (1-4), were isolated. Their structures were determined on the basis of spectroscopic data and chemical evidence. Compounds 1-4 are 3-O-tetraglycosides of barringtogenol C having an acetyl and a tigloyl or a (2Z)-hexenoyl groups at C-21, 22 and 28. The acylated saponins, 1-4, all showed antisweet activity.

Carbohydrate Sequence↗
PubMed2000

Chromosomal localization of 18S rDNA and telomere sequence in the aye-aye, Daubentonia madagascariensis.

Chromosomal localization of 18S rDNA and telomere sequence was attempted on the chromosomes of the aye-aye (2n = 30) using fluorescence in situ hybridization (FISH) and primed in situ labeling (PRINS), respectively. The rDNA was localized at the tip or whole of the short arm of acrocentric chromosomes 13 and 14 in all spreads observed. However, post-FISH silver-nitrate (Ag) staining showed that transcriptional activity of the rRNA genes was variable, particularly in chromosome 14, which was most frequently negative in one homologue carrying the smaller copy number of rDNA. This observation supports, at the molecular cytogenetic level, previous data concerning the relationship between the copy number of rDNA and its trancriptional activity. On the other hand, telomere sequence was localized only at the telomeric region of all chromosomes, the so-called telomere-only pattern, a characteristic similar to that of the greater bushbaby. These data may provide information on the chromosomal evolution of the lemur, because locations of rDNA and telomere sequences frequently offer important clues in reconstruction of karyotype differentiation.

Animals↗
PubMed2000

Osteometrical skull character in the four species of tree shrew.

The skull size and shape were osteometrically examined in the four species of the tree shrews (Tupaia tana, T. javanica, T. minor and T. dorsalis). We suggest that the skull characters were affected by the species specific behavior and terrestrial, arbo-terrestrial and arboreal life, among the genus Tupaia. The neurocranium was laterally narrower in the braincase area, and the splanchnocranium was longer only on dorsal side in T. tana, and these characters were opposite to T. minor. The principal component analysis confirmed the obviously separated clusters among T. tana, T. javanica and T. minor, affected by the adaptation for each behavior. T. dorsalis was considered as terrestrial species from the results of proportion analysis and the principal component analysis.

Animals↗
PubMed2000

Sympatric distribution of the two morphological types of the common tree shrew in Hat-Yai districts (South Thailand).

The two color types (grayish northern and reddish southern types) of the common tree shrew (Tupaia glis and Tupaia belangeri) were co-distributed in Hat-Yai region (South Thailand). Although the Isthmus of Kra in South Thailand has been considered as distribution barrier of the two types, the sympatric distribution of both types was confirmed in southern side of the Isthmus. In the principal component analysis, the skull measurement character from Hat-Yai region could also be separated into the northern and southern groups according to the skin color identification of corresponding individuals. We could generally distinguish the common tree shrew into two types by skull morphology as well as external skin color.

Animals↗
PubMed2000

In vitro and in vivo suppression of osteoclast function by adenovirus vector-induced csk gene.

The proto-oncogene c-src, which encodes a non-receptor-type tyrosine kinase c-Src, has been shown to be essential for osteoclastic bone resorption by the finding that the targeted disruption of the c-src gene induced osteopetrosis in mice. The csk (C-terminal Src family kinase) gene encodes a cytoplasmic protein-tyrosine kinase that specifically phosphorylates the negative regulatory site of c-Src (Tyr-527), thereby inhibiting its kinase activity. To regulate osteoclast function by modulating the kinase activity of c-Src, we constructed an adenovirus vector that carries this gene. The recombinant adenovirus vector carrying csk cDNA induced Csk expression in mouse osteoclast-like cells formed in vitro and clearly reduced c-Src kinase activity in a dose-dependent manner. The expression of Csk caused cytoskeletal disorganization of osteoclast-like cells and strongly suppressed pit-forming activity of the cells in vitro. In addition, the viral vector carrying csk gene dramatically suppressed interleukin-1 alpha-induced bone resorption in vivo. Conversely, kinase-inactive Csk caused an increase in c-Src kinase activity and bone resorbing activity of the cells both in vitro and in vivo, acting as a dominant negative molecule against intrinsic Csk. These findings indicate that the inhibition of c-Src activity by adenovirus vector-mediated csk expression offers an efficient means for inhibiting pathological bone resorption by suppressing osteoclast function.

Adenoviridae↗
PubMed2000

Analysis of cytokine mRNA expression in pancreatic islets of nonobese diabetic mice.

Nonobese diabetic mice develop type 1 diabetes in an age-related and gender-dependent manner. Th1 (IFN-gamma and TNF-beta) and Th2 (IL-4 and IL-10) cytokine mRNA expression was analyzed in pancreatic islets isolated from female NOD mice with a high incidence of diabetes and male NOD mice with a low incidence of diabetes. The levels were measured at 5 time points from the onset of insulitis until the development of overt diabetes, using a semiquantitative reverse transcriptase PCR (RT-PCR) assay. IFN-gamma mRNA levels were significantly higher in the islets obtained from females than those of males, from 10 weeks of age. TNF-beta mRNA was expressed in both females and males between 5 and 15 weeks of age. However, TNF-beta mRNA levels were decreased in males at 20 weeks of age. In contrast, IL-4 mRNA levels were lower in females than in males. These results suggest that islet beta-cell destruction and diabetes in female NOD mice correlates with IFN-gamma and TNF-beta production in the islets, and that male NOD mice may be protected from autoimmune beta-cell destruction by down-regulation of these cytokines. Furthermore, our findings also suggest that insulitis and beta-cell destruction are independently regulated: TNF-beta is more important in forming and maintaining the insulitis, while IFN-gamma has a more important role in beta-cell destruction.

Animals↗
PubMed2000

Serum levels of soluble interleukin-2 receptor after bone marrow transplantation: a true marker of acute graft-versus-host disease.

To examine whether serum levels of soluble interleukin-2 receptor (sIL-2R) may be a good marker of acute graft-versus-host disease (aGVHD), they were determined weekly in 56 patients receiving bone marrow transplantation (BMT). Because of wide variation in the pre-transplant sIL-2R levels (from 135 to 1918 IU/ml), we used a sIL-2R index in this study by comparing the peak levels with the pre-transplant levels. In agreement with previous reports, there was a significant correlation between the grade of aGVHD and the maximal sIL-2R index. The maximal sIL-2R index was 4.66 in patients with grade I to IV aGVHD, whereas it was 2.68 in patients without GVHD. This marker may be useful for monitoring the status of aGVHD. However, it was interesting that sIL-2R levels were elevated from the time of transplantation until the third week even in patients without GVHD or those who received autologous transplantation. Until the third week, no significant differences were observed in sIL-2R index between these patients and those who developed aGVHD during their clinical courses. After the fourth week, a higher sIL-2R index was observed in patients with aGVHD than in the other patients. Some factors other than GVHD contribute to the elevation of serum sIL-2R levels, and we should recognize the limitations of the measurement of this cytokine.

Adolescent↗
PubMed2000

[Essential thrombocythemia in transformation from myelodysplastic syndrome to acute myeloid leukemia with inv(3) after treatment for gastric cancer].

In March 1990, a 61-year-old man was given a diagnosis of essential thrombocythemia with a normal karyotype and subsequently treated with hydroxyurea. In November 1995, he underwent surgery for gastric cancer with thereafter received tegafur/uracil for 2 years. Refractory anemia with excess of blasts in transformation and chromosomal abnormalities including -5, -7, 20q-developed in August 1998. Combined chemotherapy with daunorubicin, cytarabine, mercaptopurine, and prednisolone, had only limited effectiveness. Acute myeloid leukemia was finally diagnosed in October 1998, and chromosomal analysis disclosed inv(3) in addition to -5 and -7. The appearance of inv(3) might be related to leukemic transformation of hematopoietic stem cell disease with an increase in the number of megakaryocytes and platelets.

Aged↗
PubMed2000

[Werner's syndrome associated with acute myelofibrosis].

A 47-year-old man was admitted to our hospital in June 1997 because of nasal bleeding. He presented with anemia in addition to physical characteristics of Werner's syndrome (WS). Peripheral blood examination disclosed pancytopenia with 4% blasts. Bone marrow aspiration was a dry tap; biopsy specimens revealed myelofibrosis. Chromosomal analysis of peripheral blood revealed hypodiploidy with complex abnormalities including -5 and del(7)(q21). Serum levels of PDGF, FGF, and TGF beta 1 were normal. A diagnosis of acute myelofibrosis was made. The patient's condition became quickly deteriorated and he died of pneumonia in October 1997. In the literature, we found 6 reported cases of myelofibrosis associated with WS. Considering that only approximately 1,100 cases of WS have been reported so far, the incidence of myelofibrosis in WS seems relatively high. This case suggested a link between WS and myelofibrosis, and the mechanism of myelofibrosis in WS was discussed.

Acute Disease↗