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Biomedical subjects

T Okada

Publications and source records attributed to T Okada.

At least 361 records · Page 20Linked to original sources
PubMed1998

Highly selective separation of rhodopsin from bovine rod outer segment membranes using combination of divalent cation and alkyl(thio)glucoside.

The micellization process of bovine rod outer segment (ROS) membranes is investigated utilizing a series of neutral detergents. It is found that when alkyl(thio)glucosides with an appropriate hydrophillic-lipophilic balance (e.g. octylthioglucoside) are used in combination with a divalent cation, rhodopsin is selectively extracted from ROS membranes at a specific detergent-to-membrane ratio. This allows remarkable purification of rhodopsin by a single-step solubilization, because the residual membranes are heavily aggregated in the presence of divalent cation and are therefore easily sedimented by low-speed centrifugation. The absorption spectrum of the supernatant reproducibly exhibits an A280/A500 value of 1.6, an excellent value that could rarely be obtained by chromatographic purification. The degree of purification also depends on the type of divalent cation included in the solubilization solution; specific binding of IIB-series cations (Zn2+ and Cd2+) to ROS membranes is suggested to play an important role in the solubilization process. The present result represents a unique example of selective solubilization of a specific membrane protein from highly aggregated membranes.

Animals↗
PubMed1998

Free-to-total prostate specific antigen ratio in clinical staging of prostate cancer.

The value of the free-to-total serum prostate-specific antigen (f/t PSA) ratio was compared with that of the total prostate specific antigen (tPSA) value for the prediction of clinical stage in patients with prostate cancer. The f/t PSA ratio was obtained from the frozen sera of 56 untreated patients with histologically proven BPH and 78 patients with prostate cancer. The clinical stage was organ-confined in 36, locally advanced in 20 and metastatic in 22 patients. Serum levels of free PSA (fPSA) and tPSA were determined using a chemiluminescent enzyme immunoassay. The f/t PSA ratio was calculated by dividing the fPSA value by the tPSA value and was compared with tPSA and fPSA in the correlation with clinical stage via the Spearman rank correlation test. Patients with prostate cancer had a significantly lower f/t PSA ratio than patients with BPH. The f/t PSA ratio did not differ between patients with clinically localized and metastatic cancer. tPSA and fPSA reflected the clinical stage and the extent of bone metastasis more accurately than the f/t PSA ratio. The extent of bone metastasis had no effect on the PSA ratio. The f/t PSA ratio had no additional value in clinical staging compared to tPSA. Our study suggests that the f/t PSA ratio does not reflect tumor load.

Aged↗
PubMed1998

[A case of Japanese B encephalitis with lesions of thalamus and substantia nigra revealed by MRI].

We report a 6-year-old girl with Japanese B encephalitis. The initial symptoms were high fever, headache and vomiting. On the second day of illness, she developed hemiconvulsion and was admitted to our hospital. Physical examination demonstrated a stiff neck. C-reactive protein elevated to 22.7 mg/dl. CSF examination showed a marked increase in the cell count (10,896/3 mm3). During the course of the treatment, she showed transient hemiparesis and dysphagia, followed by akinetic mutism lasting for about a month. The patient was left with severe cognitive and memory impairment and complex partial seizures but no motor dysfunction. Japanese B encephalitis was diagnosed by means of serological examination. Magnetic resonance imaging revealed cystic lesions in the medial and posterior thalamus and substantia nigra and severe atrophy of the hippocampus. Despite the involvement of substantia nigra, the patient had no parkinsonism. The cognitive impairment may in part be explained by the lesions in the medical and posterior thalamus.

Child↗
PubMed1998

[Acute mitral regurgitation caused by malignant lymphoma of the posterior mediastinum].

An 82-year-old man was admitted to our hospital in September 1996 due to dysphagia and cardiomegaly. Physical examination detected the fourth heart sound and a Levine III/VI systolic murmur in the cardiac apex. Surface lymph nodes were not palpable. LDH 662 IU/I was detected by laboratory examinations, and ultrasound cardiography showed grade 3 mitral regurgitation. Computed tomography revealed a huge mass in the posterior mediastinum, pressing the heart from the posterior direction. Thereafter, a left pleural effusion developed and aspiration was performed. Cytological examination of the fluid showed clusters of lymphoid cells with a positive immunophenotype for CD10, CD19 and HLA-DR. Chromosome analysis revealed complex abnormal karyotypes including t(8;14) (q24;32). A diagnosis of B cell lymphoma was made, and combination chemotherapy consisting of cyclophosphamide, THP-adriamycin, vincristine, and prednisolone was initiated. The patient's mass disappeared promptly, and his mitral reguration subsided. We reported this case because malignant lymphoma of the posterior mediastinum is rare, and because we are unaware of any previous reports of malignant lymphoma causing acute mitral regurgitation.

Acute Disease↗
PubMed1998

[Pulmonary aspergilloma clearly demonstrated by digital subtraction bronchography].

A 69-year-old woman was admitted because of persistent productive cough, low-grade fever and abnormal pulmonary infiltrates. Chest roentgenograms revealed right pulmonary cavitary lesions and infiltrates in both upper pulmonary fields. Digital subtraction bronchography (DSBG) was performed to detect for the presence of pulmonary cavitary lesions. Double-contrast DSBG images clearly revealed marked bronchiectasis in the right pulmonary cavity with aspergilloma in the cavitary lesions. Aspergilloma was not obvious in routine chest roentgenograms and computed tomograms. Asperillus fumigatus was cultured from sputum materials, and aspergilloma of the pulmonary cavity was clinically diagnosed. The infiltrative lesions were thought to be associated with invasive aspergillosis because the patient exhibited persistent low-grade fever and positive inflammatory reactions. Anti-fungal chemotherapy was highly effective in treating these lesions. DSBG is a new, less-invasive bronchographic technique for the evaluation of bronchial trees, and in this case proved useful for the detection of small aspergilloma in pulmonary cavities.

Aged↗
PubMed1997

Membrane recruitment of Raf-1 is not the only function of Ras in Raf-1 activation.

Ras interacts with Raf-1 and stimulates its kinase activity, which results in activation of the mitogen-activated protein (MAP) kinase cascade. It has been proposed that the main function of Ras in Raf-1 activation is to recruit Raf-1 to the plasma membrane, where a separate activation event such as phosphorylation takes place. Here, we examined the activities of various mutants of human Ha-Ras to induce membrane translocation of Raf-1 and to activate Raf-1 in vivo. Overexpression of an activator region mutant Ha-Ras(V45E) in COS7 cells induced membrane translocation of Raf-1 as effectively as wild-type Ha-Ras. However, the activity of this mutant to activate Raf-1 and extracellular signal-regulated kinase-2 (ERK2) was attenuated by approximately 70% compared to that of wild-type Ha-Ras. The decrease in the specific activity was further demonstrated by measuring the activity of the Ha-Ras(V45E)-associated Raf-1 purified from the membrane fraction. These results imply that the association of Ras with Raf-1 has another important consequence, presumably dependent on the interaction between its activator region and Raf-1, than the simple recruitment of Raf-1 to the plasma membrane.

Animals↗
PubMed1997

Endothelin enhances lipopolysaccharide-induced expression of inducible nitric oxide synthase in rat glial cells.

Lipopolysaccharide is known to stimulate production of nitrite via expression of inducible nitric oxide (NO) synthase in not only macrophages but also glial cells. We found that in glial cell cultures lipopolysaccharide-stimulated inducible NO synthase expression and nitrite accumulation were synergistically enhanced by pretreatment with endothelin, whereas endothelin itself did not induce these responses. Pretreatment with endothelin-1, endothelin-3, and the selective endothelin type B (ETB) receptor agonist IRL 1620 caused the same effect with similar potencies, suggesting that the synergism was mediated via the endothelin ETB receptor. A protein kinase C inhibitor, calphostin C, suppressed endothelin-3-enhanced inducible NO synthase expression. Pretreatment with either endothelin-3 or phorbol ester enhanced lipopolysaccharide-induced production of tumor necrosis factor-alpha (TNF-alpha). Simultaneous addition of TNF-alpha increased lipopolysaccharide-stimulated inducible NO synthase expression. These results suggest that the increase in inducible NO synthase expression by endothelin was due to the elevated TNF-alpha production via protein kinase C. Our findings present the possibility that endothelin is implicated in neurotoxicity via enhancement of inducible NO synthase expression.

Animals↗
PubMed1997

Presence of two rhodopsin intermediates responsible for transducin activation.

To identify how many rhodopsin intermediates interact with retinal G-protein transducin, the photobleaching process of chicken rhodopsin has been investigated in the presence or absence of transducin by means of time-resolved low-temperature spectroscopy. Singular value decomposition (SVD) analysis of the spectral data showed that a new intermediate called meta Ib is present between formally identified metarhodopsin I (now referred to as meta Ia) and metarhodopsin II (meta II). Since the absorption maximum of meta Ib (460 nm) is similar to that of meta Ia (480 nm), but considerably different from that of meta II (380 nm), meta Ib should have a protonated retinylidene Schiff base as its chromophore. Whereas transducin showed no effect on the conversion process between lumirhodopsin (lumi) and meta Ia, it affected the process between meta Ia and meta Ib and that between meta Ib and meta II. These results suggest that at least two intermediates (meta Ib and meta II) interact with transducin. The addition of GTPgammaS had no effect on the meta Ib-transducin interaction, while it abolished the ability of transducin to interact with meta II. Thus, meta Ib only binds to transducin, while meta II catalyzes a GDP-GTP exchange in transducin. These results suggest that deprotonation of the Schiff base chromophore is not necessary for the binding to transducin, while changes in protein structure including Schiff base deprotonation are needed to induce the GDP-GTP exchange in transducin.

Animals↗
PubMed1997

Inhibition of gene expression from the human c-erbB gene promoter by a retroviral vector expressing anti-gene RNA.

Anti-gene is a potent inhibitor of transcriptional promoter activity and subsequent gene expression. This property has been exploited to suppress the expression of a variety of oncogenes for regulating tumor proliferation or viral activities. In this paper, we describe a novel retroviral vector designed to express human c-erbB anti-gene RNA and to reduce the promoter activity in the cells. Mouse fibroblast NIH3T3 cells were stably transfected with an expression construct containing a truncated human c-erbB gene promoter fused to the firefly luciferase reporter gene. Infection into these cells of the c-erbB anti-gene retroviral vector targeted to the 26 bp pyrimidine-rich element in the human c-erbB gene promoter resulted in a dose-dependent decrease in the luciferase activity of the cells. Retroviral vector expressing anti-gene RNA may be useful as an alternative program of gene regulation in the cells.

3T3 Cells↗
PubMed1997

Distinct neuronal lineages of the ascidian embryo revealed by expression of a sodium channel gene.

The ascidian larva contains tubular neural tissue, one of the prominent anatomical features of the chordates. The cell-cleavage pattern and cell maps of the nervous system have been described in the ascidian larva in great detail. Cell types in the neural tube, however, have not yet been defined due to the lack of a suitable molecular marker. In the present work, we identified neuronal cells in the caudal neural tube of the Halocynthia embryo by utilizing a voltage-gated Na+ channel gene, TuNa I, as a molecular marker. Microinjection of a lineage tracer revealed that TuNa I-positive neurons in the brain and in the trunk epidermis are derived from the a-line of the eight-cell embryo, which includes cell fates to epidermal and neural tissue. On the other hand, TuNa I-positive cells in the more caudal part of the neural tissue were not stained by microinjection into the a-line. These neurons are derived from the A-line, which contains fates of notochord and muscle, but not of epidermis. Electron microscopic observation confirmed that A-line-derived neurons consist of motor neurons innervating the dorsal and ventral muscle cells. Isolated A-line blastomeres have active membrane excitability distinct from those of the a-line-derived neuronal cells after culture under cleavage arrest, suggesting that the A-line gives rise to a neuronal cell distinct from that of the a-lineage. TuNa I expression in the a-line requires signals from another cell lineage, whereas that in the A-line occurs without tight cell contact. Thus, there are at least two distinct neuronal lineages with distinct cellular behaviors in the ascidian larva: the a-line gives rise to numerous neuronal cells, including sensory cells, controlled by a mechanism similar to vertebrate neural induction, whereas A-line cells give rise to motor neurons and ependymal cells in the caudal neural tube that develop in close association with the notochord or muscle lineage, but not with the epidermal lineage.

Animals↗
PubMed1997

Heterodimeric phosphoinositide 3-kinase consisting of p85 and p110beta is synergistically activated by the betagamma subunits of G proteins and phosphotyrosyl peptide.

Phosphoinositide 3-kinase (PI 3-kinase) is a key signaling enzyme implicated in variety of receptor-stimulated cell responses. Receptors with intrinsic or associated tyrosine kinase activity recruit heterodimeric PI 3-kinases consisting of a 110-kDa catalytic subunit (p110) and an 85-kDa regulatory subunit (p85). We separated a PI 3-kinase that could be stimulated by the betagamma subunits of G protein (Gbetagamma) from rat liver. The Gbetagamma-sensitive PI 3-kinase appeared to be a heterodimer consisting of p110beta and p85 (or their related subunits). The stimulation by Gbetagamma was inhibited by the GDP-bound alpha subunit of the inhibitory GTP-binding protein. Moreover, the stimulatory action of Gbetagamma was markedly enhanced by the simultaneous addition of a phosphotyrosyl peptide synthesized according to the amino acid sequence of the insulin receptor substrate-1. Such enzymic properties could be observed with a recombinant p110beta/p85alpha expressed in COS-7 cells with their cDNAs. In contrast, another heterodimeric PI 3-kinase consisting of p110alpha and p85 in the same rat liver, together with a recombinant p110alpha/p85alpha, was not activated by Gbetagamma, although their activities were stimulated by the phosphotyrosyl peptide. These results indicate that p110beta/p85 PI 3-kinase may be regulated in a cooperative manner by two different types of membrane receptors, one possessing tyrosine kinase activity and the other activating GTP-binding proteins.

Amino Acid Sequence↗
PubMed1997

Suppression of autoimmune disease and of massive lymphadenopathy in MRL/Mp-lpr/lpr mice lacking tyrosine kinase Fyn (p59fyn).

MRL/Mp-lpr/lpr (MRL/lpr) mice suffer from a generalized autoimmune disease that includes autoantibody production and glomerulonephritis and develop massive lymphadenopathy characterized by an expanded population of CD4- CD8- B220+ T cells that is derived from autoreactive T cells in the periphery. Some of us previously reported that these atypical T cells overexpressed a gene for tyrosine kinase p59fyn (Fyn). To define the role of Fyn in the renal disease and lymphadenopathy in MRL/lpr mice, we have generated Fyn-deficient MRL/lpr mice whose fyn gene is replaced by the gene for beta-galactosidase. Fyn-deficient MRL/lpr mice developed markedly limited disease and lived more than twice as long as the conventional MRL/lpr mice. In the mutant mice, the production of IgG3 anti-DNA autoantibody was significantly (p < 0.005%) reduced, and glomerular deposits of IgG3 and C3 were remarkably diminished. Ag receptor-mediated proliferative responses of Fyn-deficient splenic T cells were markedly impaired. The mutant mice showed delayed accumulation of the atypical CD4- CD8- B220+ T cells that exhibited a significantly lower activity of ZAP-70 compared with those in the conventional MRL/lpr mice. These data demonstrated that Fyn is involved as a positive regulator in the disease of MRL/lpr mice. Fyn provides a signal for both the expansion of autoreactive T cells and the production of IgG3 anti-DNA autoantibody by B cells. Thus, manipulation of Fyn may improve systemic autoimmune disease in humans.

Animals↗
PubMed1997

Cytotoxicity of platinum (II) dinuclear complexes with 1-alkylthymine ligands against mouse sarcoma 180 cells.

We synthesized five platinum (II) dinuclear complexes containing 1-alkylated thymines. Two of the 1-alkylated thymine, 1-MeThy and 1-EtThy, complexes afforded good crystals. The X-ray structures of these complexes were determined. The 1-MeThy complexes has a head-to-head (H-H) arrangement, while the 1-EtThy complex has a head-to-tail (H-T) arrangement. The 1-MeThy complex (H-H) shows high electrophilicity against chloride anion (CI-) and high cytotoxicity against mouse sarcoma 180 (S-180) cells in vitro. The 1-EtThy complex (H-T) is inactive. The other 1-MeThy complex does not produce CDDP in the reaction with chloride ion and is inactive against the S-180 cell line. This complex is assumed to have an H-T arrangement. Similarly, two different 1-Pr(n)Thy complexes, one with high electrophilicity and cytotoxicity and the other without, must have an H-H and H-T arrangement, respectively. For comparison, we investigated six complexes, 1-methyluracil (1-MeUra) (H-H) dimer, alpha-pyridone (H-T) dimer, alpha-pyridone blue tetramer (PPB), 1-methylcytosine (1-MeCyt) (H-T) dimer, acetate dimer, and 1-MeUra monomer complexes. The alpha-pyridone (H-T), PPB, 1-MeCyt (H-T) dimer, and 1-MeUra monomer complexes are inert to chloride ion and inactive against mouse sarcoma S-180. The 1-MeUra (H-H) dimer and acetate dimer complexes show high electrophilicity and high cytotoxicity. Cellular accumulation of the platinum complexes phenomenally shows that all are incorporated to cancer cells to a lesser extent than CDDP. The relationships between the accumulation, the electrophilicity, and the interaction of these complexes with proteins are discussed.

Alkylation↗