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S Mita

Publications and source records attributed to S Mita.

At least 19 recordsLinked to original sources
PubMed2005

Constitutive IL-8 expression in cancer cells is associated with mutation of p53.

We previously reported that most cancer cell lines constitutively express various cytokines including IL-8. But how IL-8 gene expression is regulated in cancer cells is still unclear. p53 tumor suppressor gene plays an important role in the regulation of transcription and is mutated in cancer cell lines. We investigated whether p53 status affects the constitutive expression of IL-8 in human cancer cells. SUIT-2 and RERF-LCOK cancer cells constitutively produced high levels of IL-8 in culture medium. Both cell lines were shown to carry a p53 mutation, and constitutive NF-kappaB transcriptional activity. To analyze whether p53 status mediates IL-8 expression, the effect of wild-type p53 (wt-p53) gene transfer on activation of NF-kappaB was determined in both cell lines. ELISA showed that the IL-8 concentration in medium decreased dose dependently by transient expression of wt-p53. Western-blot analysis showed no marked change in NF-kappaB protein levels in cell nuclei. EMSA showed no repression of NF-kappaB binding activity after transient expression of wt-p53. In contrast, luciferase reporter studies indicated that transcriptional activity of NF-kappaB is suppressed by transfection of wt-p53. These results show that wt-p53 gene transfer inhibits IL-8 production and NF-kappaB transcription activity in cancer cells and suggest that constitutive IL-8 production in cancer cells is associated with mutation of p53.

Cell Line, Tumor↗
PubMed2001

Effects of cilnidipine on nitric oxide and endothelin-1 expression and extracellular signal-regulated kinase in hypertensive rats.

We evaluated the effects of cilnidipine, a long-acting Ca(2+) channel antagonist, on endothelial nitric oxide synthase (eNOS), preproendothelin-1 and endothelin ETA receptor expression in the left ventricle, and evaluated the relations between these effects and coronary microvascular remodeling and extracellular signal-regulated kinases belonging to one subfamily of mitogen-activated protein kinases in deoxycorticosterone acetate (DOCA)-salt hypertensive rats. Cilnidipine (DOCA-cilnidipine, 1 mg/kg/day, subdepressor dose) or vehicle (DOCA-vehicle) was given after induction of DOCA-salt hypertension for 5 weeks. The eNOS mRNA and protein expression in the left ventricle was significantly lower in DOCA-vehicle than in control rats and significantly higher in DOCA-cilnidipine than in DOCA-vehicle rats. Preproendothelin-1 and endothelin ETA receptor expression levels and phospho-p42/p44 extracellular signal-regulated kinase activities were significantly increased in DOCA-vehicle compared with control rats and significantly suppressed in DOCA-cilnidipine compared with DOCA-vehicle rats. DOCA-vehicle rats showed a significant increase in the wall-to-lumen ratio, perivascular fibrosis and myocardial fibrosis, with all these parameters being significantly improved by cilnidipine. These results led us to conclude that phospho-p42/p44 extracellular signal-regulated kinase activities may contribute to the coronary microvascular remodeling of DOCA rats and that protective effects of cilnidipine on cardiovascular remodeling may be at least in part mediated by an increased eNOS expression and a decreased endothelin-1 and endothelin ETA receptor expression in the left ventricle.

Animals↗
PubMed2001

Impaired production of naive T lymphocytes in human T-cell leukemia virus type I-infected individuals: its implications in the immunodeficient state.

Opportunistic infections frequently occur in patients with adult T-cell leukemia (ATL) and human T-cell leukemia virus type I (HTLV-I) carriers. However, the underlying mechanisms of such infections remain unknown. To clarify the mechanism of immunodeficiency in those infected with HTLV-I, this study analyzed the T-cell subsets in HTLV-I carriers and patients with HTLV-I-associated myelopathy/tropical spastic paraparesis and ATL using 3-color fluorescence with CD62L and CD45RA coexpression either with CD4(+) or CD8(+) T cells. The number of naive T lymphocytes was markedly suppressed in patients with ATL, particularly in those with acute form, compared with uninfected control individuals. The number of naive T cells was low in HTLV-I-infected individuals under 50 years old compared with uninfected individuals, whereas the number of memory T lymphocytes was greater in HTLV-I-infected individuals. Although the increase of memory T lymphocytes correlated with HTLV-I provirus loads, no relationship was found between naive T-cell counts and provirus loads. T-cell receptor rearrangement excision circles (TRECs), which are generated by DNA recombination during early T lymphopoiesis, were quantified to evaluate thymic function in HTLV-I-infected individuals. TREC levels were lower in HTLV-I-infected individuals than in uninfected individuals. In HTLV-I carriers less than 70 years old, an increase of Epstein-Barr virus DNA in peripheral blood mononuclear cells was observed in 6 of 16 (38%) examined, whereas it was detectable in only 1 of 11 uninfected controls. These results suggested that the low number of naive T lymphocytes was due to suppressed production of T lymphocytes in the thymus, which might account for immunodeficiency observed in HTLV-I-infected individuals.

CD4-Positive T-Lymphocytes↗
PubMed2001

Isolation of chymase complexed with physiological inhibitor similar to secretory leukocyte protease inhibitor (SLPI) from hamster cheek pouch tissues.

A low molecular weight protein complexed with chymase was isolated from hamster cheek pouch tissues. This protein had an apparent molecular mass of about 10 kDa on SDS-PAGE and the N-terminal sequence showed some homology to secretory leukocyte protease inhibitor (SLPI), which is known as the predominant inhibitor of neutrophil elastase and cathepsin G. Remarkably enhanced inhibition of chymase activity was achieved in the presence of heparin, indicating that the functional property was also similar to SLPI. These findings suggest that this SLPI-like protein is a candidate for a physiological inhibitor of chymase.

Amino Acid Sequence↗
PubMed2001

Bilateral pallidal stimulation for idiopathic segmental axial dystonia advanced from Meige syndrome refractory to bilateral thalamotomy.

Meige syndrome is an adult-onset dystonic movement disorder that predominantly involves facial muscles, while some patients with this syndrome develop spasmodic dysphonia and dystonia of the neck, trunk, arms, and legs. We report that all dystonic symptoms that had been refractory to both pharmacotherapy and bilateral thalamotomy were markedly alleviated by bilateral pallidal stimulation in a patient with segmental axial dystonia advanced from Meige syndrome.

Dominance, Cerebral↗
PubMed2001

Neurosteroids stimulate G protein-coupled sigma receptors in mouse brain synaptic membrane.

Dehydroepiandrosterone, its sulfate (DHEAS) and pregnenolone sulfate, representative neurosteroids as well as (+)-pentazocine concentration-dependently stimulated the [35S]GTPgammaS binding in synaptic membranes of mouse prefrontal cortex. These stimulations were blocked by NE-100, a sigma-receptor antagonist, and by progesterone, another type of neurosteroid. The DHEAS-induced stimulation was blocked by the pertussis toxin (PTX)-treatment, and completely recovered by reconstitution of PTX-treated membranes with recombinant G(i1), but not with G(oA). DHEAS also stimulated the [35S]GTPgammaS binding in the coronal sections of mouse brain in NE-100- or progesterone-reversible manner. These findings suggest that some neurosteroids may act on metabotropic sigma receptors, and this study may be the first to show the coupling of neurosteroid binding site and G(i).

Analgesics, Opioid↗
PubMed2001

Efficient repetitive gene delivery to skeletal muscle using recombinant adenovirus vector containing the Coxsackievirus and adenovirus receptor cDNA.

To improve adenovirus-mediated gene delivery to skeletal muscle, we have used a recombinant adenovirus vector encoding the human Coxsackievirus and adenovirus receptor (hCAR). Because CAR is expressed at a lower level in rodent myoblasts and muscle fibers than in other tissues, we expected that elevated expression of CAR in skeletal muscle would improve the efficacy of adenovirus-mediated gene transfer. Since the mouse myoblasts, C2C12 cells, showed low sensitivity to infection by recombinant adenovirus 5, we initially infected these cells at a high multiplicity of infection (MOI) of 250 with the recombinant adenovirus containing hCAR cDNA and LacZ gene. Subsequent infection by recombinant adenovirus containing the marker gene, green fluorescence protein, became efficient even at a low MOI of 25. Thus, elevated hCAR expression in mouse muscle fibers made a second virus inoculation at low doses possible. We also demonstrated that the elevated hCAR expression did not influence muscle membrane integrity. Our results suggest that co-expression of CAR and a therapeutic gene by adenovirus vector constitutes a novel strategy to advance gene therapy for hereditary muscle diseases.

Adenoviridae↗
PubMed2001

Bcl-2 expression by retrograde transport of adenoviral vectors with Cre-loxP recombination system in motor neurons of mutant SOD1 transgenic mice.

We investigated genes expression by retrograde axonal transport of replication-defective adenoviruses carrying genes for LacZ (AdLacZ) and Bcl-2 in motor neurons of transgenic mice expressing mutant human Cu/Zn superoxide dismutase (SOD1) gene containing a substitution of alanine for glycine at position 93. We found that intramuscular injection of AdLacZ into the tongue of mutant SOD1 transgenic mice and their wild-type littermates at various ages results in high expression of the transgene and similar time course of expression in hypoglossal cranial nerve nuclei, suggesting no difference in the behavior of the transgene expression between the two groups. Subsequently, we employed a molecular switching cassette for Bcl-2 designed to express Bcl-2 by Cre-loxP recombination using adenoviral vectors, and examined the COS7 and primary neuronal cells with the mutant SOD1 gene. The overexpression of Bcl-2 in both cells and the neuronal protection against staurosporine-induced apoptosis were observed, after dual infection of adenoviral vectors with cassette for Bcl-2 (AxCALNLBcl-2) and Cre recombinase (AxCANCre). After inoculation of AxCALNLBcl-2 followed by AxCANCre into the tongue of both mutant SOD1 transgenic mice and wild-type littermates, Bcl-2 was detected in both the injection site and the hypoglossal nuclei of brainstems, suggesting that this was the result of retrograde transport of AxCALNLBcl-2 and AxCANCre and expression of Bcl-2 by Cre recombinase in the hypoglossal nuclei. This strategy for delivery of exogenous genes such as Bcl-2 will be useful for studying neuronal death/survival and introducing foreign genes into postmitotic motor neurons, and in gene therapy for motor neuron diseases such as ALS.

Adenoviridae↗
PubMed2001

S1 proteins C2 and D2 are novel hnRNPs similar to the transcriptional repressor, CArG box motif-binding factor A.

S1 proteins A-D are liberated from thoroughly washed nuclei by mild digestion with DNase I or RNase A, and extracted selectively at pH 4.9 from the reaction supernatants. Here, we characterized the S1 proteins, focusing on protein D2, the most abundant S1 protein in the rat liver, and on protein C2 as well. Using a specific antibody, McAb 351, they were shown to occur in the extranucleolar nucleoplasm, and to be extracted partly in the nuclear soluble fraction. We demonstrate that the S1 proteins in this fraction exist constituting heterogeneous nuclear ribonucleoproteins (hnRNPs), through direct binding to hnRNAs, as revealed by centrifugation on density gradients, immunoprecipitation, and UV cross-linking. In hnRNPs, protein D2 occurred at nuclease-hypersensitive sites and C2 in the structures that gave rise to 40 S RNP particles. By microsequencing, protein D2 was identified with a known protein, CArG box motif-binding factor A (CBF-A), which has been characterized as a transcriptional repressor, and C2 as its isoform protein. In fact, CBF-A expressed from its cDNA was indistinguishable from protein D2 in molecular size and immunoreactivity to McAb 351. Thus, the present results demonstrate that S1 proteins C2 and D2 are novel hnRNP proteins, and suggest that the proteins C2 and D2 act in both transcriptional and post-transcriptional processes in gene expression.

Amino Acid Sequence↗
PubMed2001

Inhibition of thyroid hormone binding to the nuclear receptor by mobilization of free fatty acids.

Through binding to its nuclear receptor (TR), thyroid hormone (T3) activates the expression of the thyroid hormone-responsive genes that are essential for the regulation of energy consumption. Previously, we found that free fatty acids (FFAs) and their CoA esters strongly inhibited the binding of T3 to its nuclear receptor in vitro. In the present study, we have examined the physiological relevance of this inhibitory mechanism. TRs in isolated nuclei and in a solubilized free form were half-maximally inhibited with oleic acid at 120 and 2.8 microM, respectively. The lower sensitivity of the nuclear TR as compared with free TR was attributed to the nuclear envelope and the association of TR with chromatin. Among TRs in chromatin, those in the transcriptionally active chromatin exhibited the highest sensitivities to FFAs and were inhibited half-maximally by oleic acid at 10 microM. While the plasma concentration of FFAs in total was 0.4 to 1 mM, their nuclear concentration was about 5 microM. Thus, the sensitivities of TRs in active chromatin and in solubilized form were at physiological levels with respect to the nuclear FFA concentration. We further examined the effect of FFA mobilization on the T3-binding to TR in animals. Nuclear T3-binding was significantly inhibited when plasma and cellular FFAs were increased by norepinephrine in vivo. The increase in cellular FFAs and the TR-inhibition were well correlated, and much larger in the heart than in the liver and kidney. These results suggest that TR is negatively controlled by increased FFAs in a tissue-dependent manner.

Animals↗
PubMed2001

Clinical investigation of the lesions responsible for sensory disturbance in Minamata disease.

To clarify the lesions responsible for sensory disturbance in Minamata disease (MD), we clinically investigated the characteristics of sensory disturbance. In all patients with the classical type MD, two-point discrimination was severely disturbed, but the involvement of superficial sensation was relatively mild. On short-latency somatosensory evoked potential study, the component corresponding to N20 was completely absent with normal N9, N11, and N13 components. Although 14 of 38 chronic MD patients demonstrated intact superficial sensation, 10 of these 14 showed mild to moderate disturbance in two-point discrimination. The two-point discrimination in chronic MD patients was significantly high irrespective of the disturbance of superficial sensation. These findings suggest that the sensory disturbance of MD patients may mainly be caused by a lesion in the sensory cortex rather than in the peripheral nerves. However, other foci could be also responsible for the sensory impairment, since 9 of 38 chronic MD patients showed intact two-point discrimination.

Adult↗
PubMed2001

IL-6 and IFN-gamma regulation of IL-10 production by human colon carcinoma cells.

IL-10 has been shown to play a crucial role in immunosuppression in cancer patients. We explored the regulation of IL-10 production by TNF-alpha, IL-1 beta, IL-6, IL-8, and IFN-gamma in human colon carcinoma COLO205 cells. Northern analysis revealed a marked expression of IL-10 mRNA after stimulation by IL-6, and a marginal but significant expression by TNF-alpha, IL-1 beta or IFN-gamma. No IL-10 mRNA expression was observed when cells were untreated or incubated with IL-8. IL-10 in the culture supernatants showed good agreement with mRNA expression. In addition, IFN-gamma dose-dependently inhibited this IL-6-induced production of IL-10. MTT assay revealed that low dose IFN-gamma (1-10 ng/ml) had no effect on growth of COLO205 cells, but that high dose IFN-gamma (>100 ng/ml) significantly inhibited their proliferation. Northern analysis of COLO205 cells pretreated with IFN-gamma demonstrated that the IL-6R alpha chain was down-regulated. These results suggest that, in certain colon carcinoma cells, tumor-derived IL-10 production is directly regulated by systemic or local production of pro-inflammatory cytokines, such as IL-6 and IFN-gamma.

Blotting, Northern↗
PubMed2001

Metabotropic neurosteroid/sigma-receptor involved in stimulation of nociceptor endings of mice.

In peripheral nociceptive flexor test, SA4503, (+)-pentazocine, and (+)-3-(hydroxyphenyl)-N-(1-propyl)piperidine, representative sigma-receptor agonists, elicited dose-dependent flexor responses. These responses were blocked by sigma-receptor antagonists NE-100 or BD1063, but not by pretreatments with antisense oligodeoxynucleotide for sigma1 binding protein. The sigma-agonists' nociception is attributed to the substance P (SP) release from nociceptor endings through activations of Galpha(i1) and phospholipase C (PLC). On the other hand, attomolar doses of neurosteroids such as dehydroepiandrosterone sulfate (DHEAS) and pregnenolone sulfate caused similar nociception, and they were blocked by progesterone (PROG). However, DHEAS nociception was not affected by pertussis toxin, but was completely inhibited by a PLC inhibitor or thapsigargin. Although the nociception by lower doses of DHEAS was abolished by diphenhydramine (DPH), H1 antagonist, there were dose-dependent responses by high doses of DHEAS in the presence of DPH. The responses by DHEAS in the presence of DPH were blocked by NE-100, and those by (+)-pentazocine were blocked by PROG. All these findings suggest that two novel types of neurosteroid receptors exist, neuronal NS1/sigma-type, which mediates activation of Galpha(i1) by neurosteroids and sigma-agonists, followed by SP release from nociceptor endings; and NS2 type, which mediates histamine release from mast cells by very low doses of neurosteroids.

Animals↗
PubMed2000

Molecular cloning and characterization of GalNAc 4-sulfotransferase expressed in human pituitary gland.

We have previously cloned chondroitin-4-sulfotransferase (C4ST) cDNA from mouse brain. In this paper, we report cloning and characterization of GalNAc 4-sulfotransferase (GalNAc4ST), which transfers sulfate to position 4 of the nonreducing terminal GalNAc residue. The obtained cDNA contains a single open reading frame that predicts a type II transmembrane protein composed of 424 amino acid residues. Identity of the amino acid sequence between GalNAc4ST and human C4ST was 30%. When the cDNA was transfected in COS-7 cells, sulfotransferase activity toward carbonic anhydrase VI was overexpressed but no sulfotransferase activity toward chondroitin or desulfated dermatan sulfate was increased over the control. Sulfation of carbonic anhydrase VI by the recombinant GalNAc4ST occurred at position 4 of the GalNAc residue of N-linked oligosaccharides. The recombinant GalNAc4ST transferred sulfate to position 4 of GalNAc residue of p-nitrophenyl GalNAc, indicating that this sulfotransferase transfers sulfate to position 4 at the nonreducing terminal GalNAc residue. Dot blot analysis showed that the message of GalNAc4ST was expressed strongly in the human pituitary, suggesting that the cloned GalNAc4ST may be involved in the synthesis of the nonreducing terminal GalNAc 4-sulfate residues found in the N-linked oligosaccharides of pituitary glycoprotein hormones.

Amino Acid Sequence↗
PubMed2000

Binding of [35S]GTPgammaS stimulated by (+)-pentazocine sigma receptor agonist, is abundant in the guinea pig spleen.

Here we measured sigma receptor agonist, [3H](+)-pentazocine binding and (+)-pentazocine-stimulated [35S]GTPgammaS binding throughout brain regions and peripheral organs of mice and guinea pigs to investigate the distribution of G protein-coupled sigma receptors. There was no significant correlationship between both distributions, in which the [3H](+)-pentazocine binding is highest in the liver of each species, while the [35S]GTP-gammaS binding is highest in the guinea pig spleen. The agonist-stimulated [35S]GTPgammaS binding in the spleen was also confirmed by in situ autoradiography using sections. Thus it is suggested that there are at least two subtypes, metabotropic and nonmetabotropic sigma receptors, and the former ones are abundant in the guinea pig spleen.

Animals↗