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Biomedical subjects

I Saito

Publications and source records attributed to I Saito.

At least 55 records · Page 3Linked to original sources
PubMed2001

Glia-synapse interaction through Ca2+-permeable AMPA receptors in Bergmann glia.

Glial cells express a variety of neurotransmitter receptors. Notably, Bergmann glial cells in the cerebellum have Ca2+-permeable alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA)-type glutamate receptors (AMPARs) assembled without the GluR2 subunit. To elucidate the role of these Ca2+-permeable AMPARs, we converted them into Ca2+-impermeable receptors by adenoviral-mediated delivery of the GluR2 gene. This conversion retracted the glial processes ensheathing synapses on Purkinje cell dendritic spines and retarded the removal of synaptically released glutamate. Furthermore, it caused multiple innervation of Purkinje cells by the climbing fibers. Thus, the glial Ca2+-permeable AMPARs are indispensable for proper structural and functional relations between Bergmann glia and glutamatergic synapses.

Adenoviridae↗
PubMed2001

Possible involvement of EBV-mediated alpha-fodrin cleavage for organ-specific autoantigen in Sjogren's syndrome.

A cleavage product of alpha-fodrin may be an important organ-specific autoantigen in the pathogenesis of Sjogren's syndrome (SS), but the mechanisms of alpha-fodrin cleavage remain unclear. Since EBV has been implicated in the pathogenesis of SS, we determined whether EBV activation could induce the SS-specific 120-kDa autoantigen alpha-fodrin. ZEBRA mRNA expression, a marker for activation of the lytic cycle of EBV, was found in the salivary gland tissues from SS patients, but not in those from control individuals. ZEBRA-expressing lymphoid cells were also found in the SS glands in double-stained immunohistochemistry. Furthermore, a significant link between production of Abs against 120-kDa alpha-fodrin and reactivated EBV Ag was found in sera from patients with SS, but not in those from control individuals. EBV-activated lymphoid cells showed specific alpha-fodrin cleavage to the expected 120-kDa fragments in vitro. Pretreatment with caspase inhibitors inhibited cleavage of alpha-fodrin. Thus, an increase in apoptotic protease activities induced by EBV reactivation may be involved in the progression of alpha-fodrin proteolysis in the development of SS.

Amino Acid Chloromethyl Ketones↗
PubMed2001

Efficient gene activation in cultured mammalian cells mediated by FLP recombinase-expressing recombinant adenovirus.

A recombinant adenovirus (rAd) expressing Cre recombinase derived from bacteriophage P1 has already been extensively used for the conditional gene activation and inactivation strategies in mammalian systems. In this study, we generated AxCAFLP, a rAd expressing FLP recombinase derived from Saccharomyces cerevisiae and carried out quantitative comparisons with Cre-expressing rAd in both in vitro and in cultured cells to provide another efficient gene regulation system in mammalian cells. In the in vitro experiments, the relative recombination efficiency of FLP expressed in 293 cells infected with FLP-expressing rAd was approximately one-thirtieth that of Cre even at 30 degrees C, the optimum temperature for FLP activity, and was approximately one-ninetieth at 37 degrees C. Co-infection experiments in HeLa cells using a target rAd conditionally expressing LacZ under the control of FLP showed that an FLP-expressing rAd, infected at a multiplicity of infection (MOI) of 5, was able to activate the transgene in almost 100% of HeLa cells whereas the Cre-expressing rAd was sufficient at an MOI of 0.2. Since an MOI of 5 is ordinarily used in rAd experiments, these results showed that the FLP-expressing rAd is useful for gene activation strategies and is probably applicable to a sequential gene regulation system in combination with Cre-expressing rAd in mammalian cells.

Adenoviridae↗
PubMed2001

Synthesis and properties of peptide nucleic acids containing a psoralen unit.

We prepared the psoralen PNA unit from 8-methoxypsoralen and synthesized various PNAs containing psoralen by a typical (t)()Boc method. PNAs containing psoralen (P-PNA) at strand end formed a stable duplex with complementary DNA. The hybridization of P-PNA with complementary DNA resulted in a considerable decrease of the psoralen fluorescence.

Base Sequence↗
PubMed2001

Long-term acceptance of allografts by in vivo gene transfer of regulatable adenovirus vector containing CTLA4IgG and loxP.

CTLA4IgG was shown to inhibit the costimulatory signal for T cell activation by interfering with the ligation of CD28 and B7-1 or B7-2. To inhibit various immune responses including acute cellular rejection of allografts, a certain level of serum CTLA4IgG should be maintained for an appropriate period. We previously reported on an adenovirus vector containing CTLA4IgG, which we designated Adex1CACTLA4IgG. Adex1CACTLA4IgG was able to maintain a significant level of serum CTLA4IgG for a long period on intravenous injection, which in turn inhibited various immune responses including protective immunity against infectious agents. To overcome the inhibitory effect, we constructed a new adenovirus vector, Adex1CALoxCTLA4IgGLox, by cloning CTLA4IgG cDNA between two loxP sequences under the control of the CAG promoter. We demonstrated that the administration of adenovirus vector containing Cre recombinase gene (Adex1CACre) at the desired time induced Cre-mediated recombination within a gene derived from Adex1CALoxCTLA4IgGLox vector, and the cDNA of CTLA4IgG was excised from the transduced gene and terminated the expression of CTLA4IgG in vitro and in vivo. More importantly, we also demonstrated that the long-term acceptance of allografts was achieved after the termination of CTLA4IgG expression, while the immune response against adenovirus was restored.

Abatacept↗
PubMed2001

Specific binding of 2-amino-1,8-naphthyridine into a single guanine bulge as evidenced by photooxidation of GG doublet.

Photoirradiation of 2-amino-1,8-naphthyridines in the presence of duplex DNA containing the GG doublet opposite a single bulge was examined. After hot piperidine treatment, DNA cleavage was observed preferentially at the GG opposite a single bulge. The cleavage efficiency was highly dependent on the nature of bulged base. The G cleavage at the GG opposite a single G bulge was exceptionally weak, suggesting an intercalative binding of 2-amino-1,8-naphthyridine chromophore into the GG step.

Base Pair Mismatch↗
PubMed2001

Amplification of bleomycin-induced DNA cleavage at cytosine residues 3' to GGG sequences by pyrrole triamide.

We investigated the amplification of bleomycin-induced DNA cleavage by synthetic triamides containing N-methylpyrrole (Py) and/or N-methylimidazole (Im), PyPyPy, PyPyIm, PyImPy, and PyImIm, using 32P-labeled DNA fragments obtained from the human c-Ha-ras-1 and p53 genes. Peplomycin, a bleomycin analog, plus Fe(II) caused DNA cleavage at the 5'-GC-3' and 5'-GT-3' sequences (damaged bases are underlined). The addition of PyPyPy dramatically enhanced the cleavage, particularly at cytosine residues 3' to consecutive guanines. Alteration in the site specificity was not observed with other triamides (PyPyIm, PyImPy, and PyImIm). DNase I footprinting revealed that PyPyPy bound to the sites adjacent to the sites where DNA cleavage was enhanced by PyPyPy, and that PyPyPy enhanced DNase I-induced cleavage in GC-rich regions. These findings suggest that binding of PyPyPy to the DNA minor groove changes the DNA conformation to allow peplomycin to cleave DNA more efficiently at GC-rich sequences, resulting in intensive site-specific DNA cleavage particularly at cytosines at the 3'-side of polyguanines. The present study on amplifiers of antitumor drugs would appear to offer a novel approach to the establishment of more effective chemotherapy.

Antimetabolites, Antineoplastic↗
PubMed2001

Phenotypic changes associated with exogenous expression of p16INK4a in human glioma cells.

The tumor suppressor p16/CDKN2A/INK4a gene is frequently mutated, mostly by homozygous deletions in high-grade gliomas. Although the p16 protein suppresses cell proliferation primarily through inhibition of cell-cycle progression at the G1 phase, other phenotypic changes in glioma cells associated with p16INK4a alterations have not been fully described. To determine the roles of p16 alterations in glioma formation, we have established ecdysone-driven inducible p16 expression in the human glioblastoma cell line CL-4, which were derived from p16-null U87MG cells. Here we show that exogenous p16 expression in CL-4 cells results in morphological changes, with large and flattened cytoplasm, which are associated with increased formation of cytoplasmic actin-stress fibers and vinculin accumulation in the focal adhesion contacts. Adhesion of CL-4 cells to extracellular matrix proteins, such as laminin, fibronectin, and type IV collagen, significantly increased upon exogenous p16 expression, which correlated with increased expression of integrin alpha5 and alphav. Expression of a small GTP-binding protein, Rac, also decreased. Following epidermal growth factor stimulation, phosphorylation of MAP kinases ERK1 and 2 and induction of an early immediate gene product, c-Fos, were significantly reduced in CL-4 cells with p16 expression. These results suggest that the tumor suppressor p16 may exert its antitumor effects through modulation of multiple aspects of glioblastoma phenotypes, including proliferation, invasiveness, and responsiveness to extracellular growth stimuli.

Bradykinin↗
PubMed2001

Immunoglobulin D myeloma presenting as an extraosseous soft tissue tumor.

We report a 69-year-old man who suffered an extraosseous tumor of immunoglobulin D myeloma (lambda type) in the shoulder girdle, mimicking a primary soft tissue sarcoma. The tumor was isointense with adjacent muscle on T1-weighted MR images, and hyperintense on T2-weighted images. No continuity with the neighboring bone was noted. After administration of gadolinium, the central part of the tumor showed marked contrast enhancement. Although the tumor showed a complete response to the initial chemotherapy, the patient died of the disease 31 months after its initial manifestation. Several bone marrow aspirations and biopsies of the ilium and sternum had shown no increase in plasma cells (range 0.6-1.2%) until the disease became advanced 19 months after its initial manifestation.

Aged↗
PubMed2001

Calcification of the alar ligament of the cervical spine: imaging findings and clinical course.

Ligamentous calcification of the cervical spine has been reported in the yellow ligament, anterior and posterior longitudinal ligaments and interspinous ligament. Calcification in the upper cervical spine is rare, although some cases with calcification of the transverse ligament of the atlas have been reported. Two patients with calcification of the alar ligament with an unusual clinical presentation and course are described. Examination by tomography and computed tomography (CT) showed calcification of the alar ligament and the transverse ligament of the atlas. CT documented decreased calcification as symptoms resolved. There may be a role for CT in the search for calcifications in the upper cervical spine in patients presenting with neck pain and pharyngodynia if radiographs are normal.

Adult↗
PubMed2001

Gene therapy for hepatocellular carcinoma using two recombinant adenovirus vectors with alpha-fetoprotein promoter and Cre/lox P system.

Tissue-specific promoter has been used for cancer-specific suicide gene therapy, but its transcriptional activity is relatively low. For more efficient gene therapy of hepatocellular carcinoma, a simultaneous infection method of two recombinant adenoviruses was developed, in which one carried Cre gene under the control of alpha-fetoprotein promoter and the other a potent expression unit activated by Cre. When the vectors with lacZ reporter gene were introduced systematically into mouse models of disseminated tumors, specific and enhanced gene expression was observed exclusively in hepatocellular carcinomas both in the liver and in the lung. Next, using herpes simplex virus thymidine kinase, the anti-tumor effect was examined. Although in cultured cells, 60-300-fold expression of enzymatic activity and enhanced ganciclovir sensitivity was obtained compared with that of the single recombinant adenovirus directly driven by alpha-fetoprotein promoter, there was no significant anti-tumor effect for subcutaneous tumor on athymic mice. The lack of anti-tumor effect in mice could be explained by insufficient simultaneous transduction of the two vectors in the tumors, since it was found that a high multiplicity of infection was required to activate this system. Some strategies to overcome this dose limitation are needed, at least in the case of hepatocellular carcinoma.

Adenoviridae↗
PubMed2001

Improved selectivity for the binding of naphthyridine dimer to guanine-guanine mismatch.

Naphthyridine dimer composed of two 2-amino-1,8-naphthyridines and a connecting linker strongly binds to guanine-guanine (G-G) mismatch in duplex DNA. In order to improve G-G selectivity for the binding, we have examined structure modification of the linker. A new naphthyridine dimer possessing 3,6-diazaoctanedioic acid linker binds to G-G mismatch with an association constant of 1.18 x 10(7) M(-1), which is somewhat weaker than that of the original naphthyridine dimer having a shorter connecting linker. However, the binding of the modified naphthyridine dimer to G-A mismatch was almost negligible as compared to that of the original. This results in a net increase of the selectivity for the binding to G-G mismatch by 4-folds.

Base Pair Mismatch↗
PubMed2001

Scanning of guanine-guanine mismatches in DNA by synthetic ligands using surface plasmon resonance.

Here we have designed and synthesized ligands that specifically bind with high affinity (K(d) = 53 nM) to the guanine (G)-guanine mismatch, one of four types of single-nucleotide polymorphism (SNP). Detection of the G-G mismatch was performed by a surface plasmon resonance (SPR) assay using a sensor chip carrying the G-G specific ligand on its surface. The accuracy of the G-G mismatch detection by the SPR sensor was demonstrated by a marked SPR response obtained only for the DNA containing the G-G mismatch. DNAs containing G-A and G-T mismatches, as well as a fully matched duplex, produced only a weak response. Furthermore, this assay was found applicable for the detection of SNP existing in PCR amplification products of a 652-nucleotide sequence of the HSP70-2 gene.

Base Pair Mismatch↗
PubMed2001

Induction of photoreceptor-specific phenotypes in adult mammalian iris tissue.

We show that iris tissue in the adult rat eye, which is embryonically related to the neural retina, can generate cells expressing differentiated neuronal antigens. In addition, the Crx gene transfer induced the specific antigens for rod photoreceptors in the iris-derived cells, which was not seen in the adult hippocampus-derived neural stem cells. Our findings demonstrate a remarkable plasticity of adult iris tissue with potential clinical applications, as autologous iris tissue can be feasibly obtained with peripheral iridectomy.

Aging↗