Biomedical subjects

Takayuki Saitoh

Publications and source records attributed to Takayuki Saitoh.

9 recordsLinked to original sources

PubMed2005

Dimeric transthyretin variant assembles into spherical neurotoxins.

Familial amyloidotic polyneuropathy is a hereditary autosomal-dominant disease in which the deposited transthyretin fibrils are derived from amyloidogenic mutation. We investigated structure and stability of a human Ser112Ile transthyretin variant and showed that the Ser112Ile variant exists as a dimer having nonnative tertiary structure at physiological pH. In addition, the dimeric Ser112Ile assembles into a spherical aggregate and exerts cytotoxicity in a human neuroblastoma cell line. Our results suggest the importance of an unstable dimeric structure in forming spherical aggregates that will induce cell death.

Acrylamide↗

PubMed2005

Critical regions for assembly of vertebrate nonmuscle myosin II.

Myosin II molecules assemble and form filaments through their C-terminal rod region, and the dynamic filament assembly-disassembly process of nonmuscle myosin II molecules is important for cellular activities. To estimate the critical region for filament formation of vertebrate nonmuscle myosin II, we assessed the solubility of a series of truncated recombinant rod fragments of nonmuscle myosin IIB at various concentrations of NaCl. A C-terminal 248-residue rod fragment (Asp 1729-Glu 1976) was shown by its solubility behavior to retain native assembly features, and two regions within it were found to be necessary for assembly: 35 amino acid residues from Asp 1729 to Thr 1763 and 39 amino acid residues from Ala 1875 to Ala 1913, the latter containing a sequence similar to the assembly competence domain (ACD) of skeletal muscle myosin. Fragments lacking either of the two regions were soluble at any NaCl concentration. We referred to these two regions as nonmuscle myosin ACD1 (nACD1) and nACD2, respectively. In addition, we constructed an alpha-helical coiled-coil model of the rod fragment, and found that a remarkable negative charge cluster (termed N1) and a positive charge cluster (termed P2) were present within nACD1 and nACD2, respectively, besides another positive charge cluster (termed P1) in the amino-terminal vicinity of nACD2. From these results, we propose two major electrostatic interactions that are essential for filament formation of nonmuscle myosin II: the antiparallel interaction between P2 and N1 which is essential for the nucleation step and the parallel interaction between P1 and N1 which is important for the elongation step.

Amino Acid Sequence↗

PubMed2005

Histone deacetylase inhibitors profoundly decrease proliferation of human lymphoid cancer cell lines.

Methylation of tumor suppressor genes is frequently observed in human cancers. These genes are silenced by histone deacetylase (HDAC) recruited by methylated DNA in their promoter regions. HDAC removes acetyl groups from histones and prevents the basic transcriptional machinary access to the target gene, leading to transcriptional repression. HDAC inhibitors (HDACIs) can restore the expression of the tumor suppressor and/or cell cycle regulatory genes in cancer cells and block the cellular proliferation of these cells. In this study, we investigated the in vitro antiproliferative activities of the HDACIs, suberoylanilide hydroxamic acid (SAHA), and valproic acid against 14 human lymphoid cancer cell lines. All of these cell lines were sensitive to the antiproliferative effects of the HDACI. SAHA induced either G1 or G2-M arrest as well as apoptosis. SAHA downregulated cyclin D1 and D2, and upregulated p53, p21, and p27. Chromatin immunoprecipitation analysis revealed a remarkable increase in the level of acetylated histones associated with the p21 promoter after SAHA treatment. In nude mice, SAHA significantly inhibited growth of a mantle cell lymphoma without major toxic side effects. In summary, HDACIs are promising therapeutic agents for human lymphoid cancers.

Animals↗

PubMed2004

Contribution of cellular contractility to spatial and temporal variations in cellular stiffness.

Scanning probe microscopy and immunofluorescence observations indicated that cellular stiffness was attributed to a contractile network structure consisting of stress fibers. We measured temporal variations in cellular stiffness when cellular contractility was regulated by dosing with lysophosphatidic acid or Y-27632. This experiment revealed a clear relation between cellular stiffness and contractility: Increases in contractility caused cells to stiffen. On the other hand, decreases in contractility reduced cellular stiffness. In both cases, not only the stiffness of the stress fibers but also that of the whole of the cell varied. Immunofluorescence observations of myosin II and vinculin indicated that the stiffness variations induced by the regulation of cellular contractility were mainly due to rearrangements of the contractile actin network on the dorsal surface. Taken together, our findings provide evidence that the actin cytoskeletal network and its contractility features provide and modulate the mechanical stability of adherent cells.

Actins↗

PubMed2004

Expression of hepcidin is down-regulated in TfR2 mutant mice manifesting a phenotype of hereditary hemochromatosis.

Transferrin receptor 2 (TfR2) is a membrane glycoprotein that mediates cellular iron uptake from holotransferrin. Homozygous mutations of this gene cause one form of hereditary hemochromatosis in humans. We recently reported that homozygous TfR2(Y245X) mutant mice, which correspond to the TfR2(Y250X) mutation in humans, showed a phenotype similar to hereditary hemochromatosis. In this study, we further analyzed the phenotype as well as iron-related gene expression in these mice by comparing the TfR2-mutant and wild-type siblings. Northern blot analyses showed that the levels of expression of hepcidin mRNA in the liver were generally lower, whereas those of duodenal DMT1, the main transporter for uptake of dietary iron, were higher in the TfR2-mutant mice as compared to the wild-type siblings. Expression of hepcidin mRNA in the TfR2 mutant mice remained low even after intraperitoneal iron loading. In isolated hepatocytes from both wild-type and TfR2 mutant mice, interleukin-6 and lipopolysaccharide each induced expression of hepcidin mRNA. These results suggest that up-regulation of hepcidin expression by inflammatory stimuli is independent of TfR2 and that TfR2 is upstream of hepcidin in the regulatory pathway of body iron homeostasis.

Animals↗

PubMed2003

Suppressive activities of OGG1 and MYH proteins against G:C to T:A mutations caused by 8-hydroxyguanine but not by benzo[a]pyrene diol epoxide in human cells in vivo.

8-Hydroxyguanine (8OHG), an oxidatively damaged base, and benzo[a]pyrene-diol-epoxide (BPDE), a metabolite of benzo[a]pyrene found in cigarette smoke, are thought to be major causes for G:C to T:A transversions in DNA of human cells. In this study, we assessed the abilities of OGG1, MYH and APE1 proteins, which are components of a base excision repair pathway, to suppress G:C to T:A transversions caused by 8OHG or BPDE by a bacterial suppressor tRNA (supF) forward mutation assay using a shuttle plasmid, pMY189. The introduction of a single 8OHG residue at position 159 of the supF gene and treatment with BPDE led to a 65- and 34-fold increase in mutation frequencies of the pMY189 plasmid, respectively, after replication in the NCI-H1299 human lung cancer cell line. G:C to T:A transversions were predominantly induced in these plasmids. Both the mutation frequency of the 8OHG-containing plasmid in NCI-H1299 cells and the occurrence of G:C to T:A transversions at position 159 in the supF gene were significantly reduced by overexpression of OGG1 and MYH proteins, but not by that of APE1 protein. In contrast, neither mutation frequency nor the occurrence of G:C to T:A transversion of the BPDE-treated plasmid was reduced by overexpression of OGG1, MYH and APE1 proteins. These results indicate that OGG1 and MYH function as suppressors for G:C to T:A transversions by 8OHG but not by BPDE in human cells.

7,8-Dihydro-7,8-dihydroxybenzo(a)pyrene 9,10-oxide↗

PubMed2002

A single nucleotide polymorphism at the splice donor site of the human MYH base excision repair genes results in reduced translation efficiency of its transcripts.

BACKGROUND: Adenine paired with 8-hydroxyguanine, a major oxidatively damaged DNA lesion, is excised by mutY homologue (MYH) base excision repair protein in human cells. Since genetic polymorphisms of DNA repair genes associated with the activities and the expression levels of their products may modulate cancer susceptibility of individuals, we investigated the effect of a single nucleotide polymorphism (SNP) in the MYH gene on the difference in the expression levels of its products. RESULTS: An aberrant size of the beta type nuclear form transcript was detected in a lung cancer cell line, VMRC-LCD, by reverse transcriptase-polymerase chain reaction (RT-PCR) analysis. The transcript contained the intron 1 sequence, and it was due to alternative splicing resulting from IVS1+5G/C SNP. The presence of the upstream open reading frame (ORF) on the 5'-side of the native ORF in the beta type transcript from the IVS1+5C allele could reduce the translation efficiency of the transcript into the nuclear form protein. Thus, expression vectors bearing the 5'-untranslated region sequence of either the IVS1+5G or 5C allele were constructed. In vitro translation analysis, as well as Western blot and quantitative RT-PCR analyses of the H1299 lung cancer cell line transfected with these vectors, revealed that the translation efficiency of the IVS1+5C transcript into MYH protein was much lower (approximately 30) than that of the IVS1+5G transcript. CONCLUSIONS: The SNP at the splice donor site of the MYH gene resulted in reduced translation efficiency of its transcripts. This is the fourth case of single nucleotide variations that cause alterations in translation initiation sites and translation efficiencies in human cells.

Alleles↗

PubMed2002

Vitamin D3 analogs stimulate hair growth in nude mice.

The active form of vitamin D3 can regulate epidermal keratinization by inducing terminal differentiation; and mice lacking the vitamin D receptor display defects leading to postnatal alopecia. These observations implicate the vitamin D3 pathway in regulation of hair growth. We tested the ability of 1,25 dihydroxyvitamin D3 and its synthetic analogs to stimulate hair growth in biege/nude/xid (BNX) nu/nu (nude) mice exhibiting congenital alopecia. Nude mice were treated with different vitamin D3 analogs at doses that we had previously found to be the highest dose without inducing toxicity (hypercalcemia). The mice were monitored for hair growth and were scored according to a defined scale. Skin samples were taken for histological observation of hair follicles and for extraction of RNA and protein. Vitamin D3 analogs dramatically stimulated the hair growth of nude mice, although parental 1,25 dihydroxyvitamin D3 had no effect. Hair growth occurred in a cyclical pattern, accompanied by formation of normal hair follicles and increased expression of certain keratins (Ha7, Ha8, and Hb3). Vitamin D3 analogs seem to act on keratinocytes to initiate hair follicle cycling and stimulate hair growth in mice that otherwise do not grow hair.

Alopecia↗

PubMedDate not supplied

Ribosome-lamella complex of leukemic cells in a patient with aggressive NK cell leukemia.

A 36-year-old male was admitted to the hospital with anemia and leukopenia. A bone marrow specimen revealed the proliferation of leukemic cells with flower-like nuclei observed in adult T-cell leukemia. The leukemic cells were positive for HLA-DR, CD2, CD7, and CD56. A diagnosis of aggressive NK cell leukemia was made, the patient was treated with induction chemotherapy and cord blood stem cell transplantation, and he is well now. Seen with electron microscopy, some leukemic cells had ribosome-lamella complexes (RLC). This is the first reported case of leukemic cells with flower-like nuclei seen with light microscopy and RLC seen with electron microscopy.

Adult↗