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Biomedical subjects

T Okubo

Publications and source records attributed to T Okubo.

At least 37 records · Page 2Linked to original sources
PubMed2001

Cigarette smoking depletes cells spontaneously secreting Th(1) cytokines in the human airway.

Cigarette smoking may modify the immune balance in the airway since it alters the course of diseases in which immune system has an important role. This study examined whether cigarette smoking could affect the distribution of cells secreting Th(1) or Th(2) cytokines in the human airway. We utilized cytokine ELISPOT assay to detect and quantitate the frequencies of cells spontaneously secreting cytokines in bronchoalveolar lavage fluid (BALF). BALF was collected from six non-smokers and four heavy cigarette smokers without clinical airway symptoms. Cytokine ELISPOT assay was performed to quantitate cells secreting interleukin (IL-)2, IL-4 and interferon (IFN-)gamma with or without phorbor 12-myristate 13-acetate (PMA) stimulation. There were no cells spontaneously secreting IL-2 detected in all samples from smokers whereas most of non-smokers had detectable IL-2-secreting cells. The number of IFN-gamma-secreting cells was also extremely decreased in smokers. Mitogen-stimulated Th(1) cytokine-secreting cells were again significantly decreased in smokers' airways. The frequency of IL-2-secreting cells and CD4/CD8 ratio in BALF had a weak positive correlation. IL-4-secreting cells were not detected in any samples from both groups. These results show that cigarette smoking depletes Th(1) cytokine-secreting cells in the human airway. It may explain the susceptibility of smokers to certain airway disease conditions such as viral or mycobacterial infections and allergic diseases.

Bronchoalveolar Lavage Fluid↗
PubMed2001

Down-regulation of promoter 1.3 activity of the human aromatase gene in breast tissue by zinc-finger protein, snail (SnaH).

Aromatase (estrogen synthetase) is expressed in breast cancer tissue, and in situ expression of the enzyme stimulates breast cancer growth. Promoter I.3 is one of the major promoters that control the expression of aromatase in breast cancer tissue. Using the yeast one-hybrid approach to screen a human breast tissue hybrid cDNA expression library, we found that the zinc-finger transcriptional factor Snail (SnaH) interacted with a regulatory region near promoter I.3 of the human aromatase gene. DNA mobility shift assays and mutation analyses using recombinant SnaH protein expressed in Escherichia coli have revealed that this protein interacts with a segment, 5'-CTGATGAAGT-3', which is between 66 and 76 bp upstream from the transcriptional start site of promoter I.3. Using mammalian cell transfection experiments, SnaH was found to act as a repressor of promoter I.3 activity. Site-directed mutagenesis experiments have revealed that the NH2-terminal SNAG domain is important for the repressor activity of SnaH. To demonstrate the inhibitory activity against aromatase expression, a stable SnaH-expressing MDA-MB-231 breast cancer cell line was generated, and the aromatase RNA messages in the SnaH-transfected cell line were found to be 30% of those in the vector-transfected cell line. Reverse transcription-PCR analysis on RNAs isolated from 12 cell lines has confirmed that SnaH is expressed at a higher level in normal breast epithelial cell and stromal fibroblast cell lines than in breast cancer cell lines. In addition, SnaH mRNA was detected in only 16 of 55 breast cancer specimens. On the other hand, aromatase mRNA was detected in 54 of the 55 specimens. Our results indicate that SnaH acts as a repressor that down-regulates the expression of aromatase in normal breast tissue by suppressing the function of promoter I.3. A reduction of the expression of SnaH in breast cancer tissue further suggests a cancer-protective role for this protein in normal breast tissue.

Amino Acid Sequence↗
PubMed2001

Association analysis of polymorphisms in the prepronociceptin gene and schizophrenia.

The human prepronociceptin (ppNoc) gene is located on chromosome 8p21, and several linkage studies have suggested that the susceptibility locus for schizophrenia is present in this chromosomal region. We investigated genetic variations in the promoter and coding regions of the ppNoc gene to determine if there may be an association between naturally occurring mutations and the manifestation of schizophrenia. Association analyses for the variations found were conducted between 87 unrelated schizophrenic patients and 100 healthy controls. Polymorphisms were found at both the -503 locus (G/A) in the promoter region and the 353 locus (Ala118Gly) in exon 3. Additionally, rare variants were identified at the -261 locus (A/G) in the promoter region, the 515 locus (Gln172Arg) in exon 3, and the 561 locus (G/A) in the untranslated region of exon 3. A significant difference was observed in allele frequency at the 353 locus between controls and patients with continuous-course schizophrenia (P = 0.0237), and between patients with and without prominent negative symptoms (P = 0.0201). However, neither difference was significant after Bonferroni correction. In addition, there were no significant differences in genotype and allele frequencies at either the -503 or 353 locus between the control group and patients with schizophrenia. These results suggest that ppNoc gene polymorphisms have no association with schizophrenia.

Adult↗
PubMed2001

ER-dependent estrogenic activity of parabens assessed by proliferation of human breast cancer MCF-7 cells and expression of ERalpha and PR.

Estrogenic activities of the phenolic preservatives methylparaben, ethylparaben, propylparaben, butylparaben, isopropylparaben and isobutylparaben were examined by assaying estrogen-receptor (ER)-dependent proliferation of MCF-7 cells. All the compounds stimulated the proliferation to about the same level as the maximal cell yield attained with 3x10(-11) M 17beta-estradiol, but at a concentration in the order of 10(5) to 10(7) higher than 17beta-estradiol. The cell-proliferative effects of parabens were completely suppressed by anti-estrogen ICI 182,780. MCF-7 cells treated with butylparaben and isobutylparaben exhibited a decrease in gene expression of ERalpha and an increase in that of progesterone-receptor (PR), but the effects of these parabens were not as prominent as those of 17beta-estradiol. Western blot analysis indicated that these parabens caused a slight decrease in expression of ERalpha protein. Competitive binding to human ERalpha and ERbeta in vitro revealed that the parabens with longer side-chains showed greater affinity for estrogen receptors, and that they had similar relative binding affinity (RBA) values to both ERalpha and ERbeta. RBA values were much smaller than that of diethylstilbestrol. In conclusion, parabens have ER-dependent estrogenic activities, and their effects on the intracellular signaling pathway might be different from that of 17beta-estradiol.

Binding, Competitive↗
PubMed2001

Modulation of aromatase expression in human breast tissue.

Aromatase plays an important role in breast cancer development through its role in the synthesis of estrogen. Aromatase expression in breast tissue can be regulated by several mechanisms. The major promoter usage for aromatase expression in breast tumors (i.e. cAMP-stimulated promoters I.3 and II) is different from that in normal breast tissue (i.e. glucocorticoid-stimulated promoter I.4). Recent characterization of transcription factors that interact with the two important regulatory elements near promoters I.3 and II, i.e. S1 and CREaro, helps us better understand the mechanism of the switch of promoter usage between normal breast tissue and cancer tissue. It is thought that in normal breast tissue, the function of promoters I.3 and II is suppressed through the binding of EAR-2, COUP-TFI, and EARgamma to S1, and through the binding of Snail/Slug proteins to their binding site that quenchs the CREaro activity. In cancer tissue, the expression levels of EAR-2, COUP-TFI, EARgamma, Snail, and Slug decrease, and aromatase expression is then up regulated through the binding of ERRalpha-1 to S1 and the binding of CREB or related factors to CREaro. Results from this and other laboratories reveal that aromatase activity in aromatase expressing cells can also be modified by treatment with aromatase inhibitors and the antiestrogen ICI 182, 780. While aromatase inhibitors are used to treat breast cancer, the treatment has been found to increase the level of aromatase in the breast tissue of some patients. The enhancement of aromatase activity by aromatase inhibitors is thought to be due to a decrease of aromatase protein degradation by enzyme-inhibitor complex formation, up-regulation of the aromatase gene transcription through a cAMP-mediated mechanism, and an induction of aromatase expression by gonadtropins that are released from the pituitary in response to a reduction of estrogen levels in circulation in premenopausal women. Antiestrogen ICI 182, 780 has been found to suppress aromatase expression, but the mechanism has not yet been determined. In addition, aromatase activity and expression can be affected by environmental chemicals. A detailed structure-function study has revealed that flavones, but not isoflavones, are inhibitors of aromatase. It was found that flavones bind to the active site of aromatase in an orientation in which their rings-A and -C mimic rings-D and -C of the androgen substrate. The modulation of aromatase expression by endocrine disrupting chemicals is exemplified by two organochlorine pesticides (i.e. toxaphene and chlordane) that have been found to be antagonists of ERRalpha-1 orphan receptor. These compounds reduce ERRalpha-1 activity, resulting in a suppression of aromatase expression.

Aromatase↗
PubMed2001

Synthesis and structure-affinity relationships of 4-(5-Aryl-1,2,3,6-tetrahydropyridino)pyrimidine derivatives as corticotropin-releasing factor(1) receptor antagonists.

Recently, various non-peptide corticotropin-releasing factor(1) (CRF(1)) receptor antagonists have been reported. Structure-affinity relationships (SARs) of non-peptide CRF(1) antagonists suggest that such antagonists can be constructed of three units: a hydrophobic unit (Up-Area), a proton accepting unit (Central-Area), and an aromatic unit (Down-Area). We previously presented 4-aryl-1,2,3,6-tetrahydropyridinopyrimidine derivatives including potent CRF receptor ligands 1a and 1b and proposed that the 4-aryl-1,2,3,6-tetrahydropyridino moiety might be useful as a substituent in the Up-Area. Our interest shifted to 5-aryl-1,2,3,6-tetrahydropyridinopyrimidine derivatives 2, among which compound 2m (CRA0165) had highest affinity for CRF(1) receptors (IC(50)=11nM). We report here the design, synthesis and SARs of derivatives 2.

Animals↗
PubMed2001

Chemical modification of aryl-1,2,3,6-tetrahydropyridinopyrimidine derivative to discover corticotropin-releasing factor(1) receptor antagonists: aryl-1,2,3,6-tetrahydropyridino-purine, -3H-1,2,3-triazolo[4,5-d)pyrimidine, -purin-8-one, and -7H-pyrrolo[2,3-d]pyrimidine derivatives.

Structure-affinity relationships (SARs) of non-peptide CRF(1) antagonists suggest that such antagonists can be constructed of three units: a hydrophobic unit (Up-Area), a proton accepting unit (Central-Area), and an aromatic unit (Down-Area). Recently, various non-peptide corticotropin-releasing factor(1) (CRF(1)) receptor antagonists obtained by modification of the Central-Area have been reported. In contrast, we modified the Up-Area and presented 4- or 5-aryl-1,2,3,6-tetrahydropyridinopyrimidine derivatives including potent CRF receptor ligands 1a-c, and proposed that the 4- or 5-aryl-1,2,3,6-tetrahydropyridino moiety might be useful as a substituent in the Up-Area. Our interest shifted to the chemical modification in which the pyrimidine ring of 1a-c was replaced by other heterocycles, purine ring of 2, 3H-1,2,3-triazolo[4,5-d]pyrimidine ring of 3, purin-8-one ring of 4 and 7H-pyrrolo[2,3-d]pyrimidine ring of 5. Among them, 5-aryl-1,2,3,6-tetrahydropyridinopurine compound 6j (CRA0186) had the highest affinity for CRF(1) receptors (IC(50)=20nM). We report here the synthesis and SARs of derivatives 6-9.

Affinity Labels↗
PubMed2001

Nitric oxide donor FK409 and 8-bromoguanosine-cyclic monophosphate attenuate cardiac contractility assessed by Emax.

FK409 decomposes and releases nitric oxide (NO) spontaneously when it is dissolved in phosphate buffer solution at 37 degrees C. With the use of this NO donor, the effect of exogenous NO on cardiac contractility was examined by assessing Emax. alpha-chloralose-anaesthetized dogs were instrumented for measurements of left ventricular (LV) pressure and volume and coronary blood flow (CBF) in the left anterior descending artery (LAD). FK409, 8-bromoguanosine-cyclic-monophosphate (8-Br-cGMP) and papaverine were infused into the LAD, and Emax was determined by transient inferior vena cava occlusion when CBF was increased and reached its peak. Neither drug affected heart rate nor LV pressure just before the measurement of Emax. FK409 increased CBF and decreased Emax in a dose-dependent manner. 8-Br-cGMP also increased CBF and decreased Emax in a dose-dependent manner. Pretreating with propranolol did not affect the effects of FK4098-Br-cGMP on CBF and Emax. Papaverine increased mean CBF but did not affect Emax. In conclusion NO attenuates cardiac contractility in vivo, while increasing CBF. This effect seems to be mediated by cyclic-guanosine monophosphate, a second messenger of NO.

Adrenergic beta-Antagonists↗
PubMed2001

Polymorphism of the neuropeptide Y gene: an association study with alcohol withdrawal.

BACKGROUND: Neuropeptide Y (NPY) exists in both the central and peripheral nervous system and is thought to modulate many functions such as feeding behavior, anxiety-associated behavior, circadian rhythm, seizure modulation, and hormone secretion. Recent studies have revealed that NPY influences alcohol consumption in mice and that alcohol-preferring rats showed lower concentrations of NPY-like immunoreactivity compared with alcohol-nonpreferring rats in several brain regions. METHODS: In the present study, we analyzed the whole coding region and 5'-untranslating region of the NPY gene for 163 Japanese male alcoholics with different withdrawal symptoms (93 with delirium tremens, 71 with seizures, 49 with hallucinations) and 98 Japanese male controls. ALDH2 genotypes of all subjects tested were ALDH2*1 homozygote. RESULTS: Three polymorphic nucleotide substitutions, namely -121C/A (promoter), 1258G/A (exon 2), and 5671C/T (exon 3), were detected in both groups. Substituting C to A in the -121 locus produced a putative binding site of GATA-1 and GATA-2. Also, -90G/A (promoter) as a rare variant and 5642-5651 single nucleotide repeats T10/T11 (intron 2) were found. Polymorphism (C/T) at the 1128 locus has been reported to be associated with a higher serum cholesterol level in obese white subjects, but such a polymorphism was not found in our samples. The genotypical distributions for these polymorphic loci (-121C/A, 1258G/A, 5642-5651 T10/T11, and 5671C/T) were not significantly different between the alcoholics and controls. However, frequency of the T allele and frequency of the genotype that possessed T alleles (CT, TT) at the 5671 locus were significantly higher in patients with seizure than in those without seizure (p < 0.05, p < 0.02). CONCLUSION: Our data suggested that a C to T substitution at the 5671 locus of the NPY gene may be associated with seizure during alcohol withdrawal.

5' Untranslated Regions↗
PubMed2001

Identification of aminoglycoside-modifying enzymes by susceptibility testing: epidemiology of methicillin-resistant Staphylococcus aureus in Japan.

A multiple-primer PCR was used to identify genes encoding aminoglycoside-modifying enzymes in 381 clinical isolates of methicillin-resistant Staphylococcus aureus (MRSA). The technique used three sets of primers delineating specific DNA fragments of the aph(3')-III, ant(4')-I, and aac(6')-aph(2") genes, which influence the MICs of gentamicin, tobramycin, and lividomycin. Isolates with none of the three genes detected were susceptible to all three agents. Isolates with the aph(3')-III gene showed resistance to lividomycin (MIC > 1,024 microg/ml), and those with the ant(4')-I gene were resistant to tobramycin (MIC > or = 8 microg/ml). Isolates with only the aac(6')-aph(2") gene were resistant to gentamicin (MIC > or = 8 microg/ml) and tobramycin in decreasing order; those with both the ant(4')-I and aac(6')-aph(2") genes also were resistant to gentamicin and tobramycin, but in increasing order. Susceptibility testing, then, could detect specific genes. In 381 Japanese MRSA isolates, the ant(4')-I, aac(6')-aph(2"), and aph(3')-III genes were prevalent in 84.5, 61.7, and 8.9%, respectively. Isolates with only the ant(4')-I gene had coagulase type II or III, but isolates with both the ant(4')-I and aac(6')-aph(2") genes included all coagulase types. Most isolates with coagulase type IV or VII carried the aac(6')-aph(2") gene. Of the MRSA isolates with ant(4')-I and/or aac(6')-aph(2") genes, 97% were resistant to aminoglycosides in clinical use, but a new aminoglycoside, arbekacin, had excellent activity against these isolates.

Acetyltransferases↗
PubMed2001

Polymorphism of the 5'-upstream region of the human SNAP-25 gene: an association analysis with schizophrenia.

Recent studies have suggested that synaptic abnormalities may be part of the pathophysiology of schizophrenia. SNAP-25 (synaptosomal-associated protein of 25 kD) is one of the synaptic proteins responsible for presynaptic neurotransmission, axonal elongation and synaptogenesis. Genetic variation in the 5'-upstream region of the SNAP-25 gene was analyzed in 87 unrelated schizophrenic patients and 100 healthy controls. A novel polymorphic (TAAA)(n) tandem repeat was identified in the 5'-upstream region. There were no significant differences between the patient and the control groups in the distribution of repeat numbers of alleles or genotypes. In addition, no associations were found between the polymorphism for subtypes, longitudinal courses or positive family history of the patients. Our results suggest that polymorphisms in the 5'-upstream region of the SNAP-25 gene have no association with schizophrenia.

Adult↗
PubMed2001

Polymorphisms in the promoter and coding regions of the synapsin III gene. A lack of association with schizophrenia.

The human synapsin III gene, located on chromosome 22q12-13, has previously been reported to indicate a susceptibility for schizophrenia. Noval rare variants (Thr136Thr in exon 3, Pro468Ser, Glu525Gln and Pro534Leu in exon 12, and 1769 G/C in the untranslated region of exon 13) were found in addition to the polymorphic variant (-196 G/A in the promoter region). No significant differences in genotypic or allelic frequencies of the -196 G/A polymorphism were found between 87 unrelated schizophrenic patients and 100 healthy controls, even when the patients were diagnostically subdivided into subtypes and course specifiers. Furthermore, allelic frequencies of the GATG repeat in intron 1 were not significantly different between the patients and the controls. These results suggest that synapsin III gene polymorphisms are not associated with schizophrenia.

Adult↗
PubMed2001

Aromatase p450 expression in a feminizing adrenal adenoma presenting as isosexual precocious puberty.

A 7-yr-old girl presented with isosexual precocious puberty secondary to a feminizing adrenal adenoma. The adrenal tumor was found to express aromatase messenger ribonucleic acid. Enzyme kinetic studies revealed a high level of aromatase activity in the adrenal tumor, with a K(m) of 45 nmol/L and a maximum velocity of 25.6 pmol/mg.h. Aromatase activity was approximately 500-fold higher in the tumor than in adjacent normal adrenal tissue. Although histopathological examination of the tumor was most consistent with a benign adenoma, the aromatase transcripts present in the tumor corresponded to those previously associated with malignant as well as benign tumors. We consider the pattern of aromatase expression sufficient to warrant continued follow-up for tumor recurrence. Our case demonstrates that isosexual precocious puberty secondary to a feminizing adrenal tumor can be due to estrogen synthesis from the tumor itself rather than peripheral aromatization as had been previously theorized.

Adenoma↗
PubMed2001

Stromal cells provide signals different from cytokines for STAT5 activation in hematopoietic cells.

After detachment from the stromal cells, hematopoietic stem cells are thought to differentiate to the cytokine-dependent stages where their growth and differentiation are promoted by these cytokines. To examine the stromal regulation of hematopoietic stem cells, we previously established a primitive hematopoietic stem-like cell line, THS119, whose growth was dependent on the bone marrow stromal cell line, TBR59, and from which IL-3- (THS119/IL-3) or IL-7- (THS119/IL-7) dependent cell lines were then generated. Using these cell lines, we examined the difference in signals mediated by the stromal cells and cytokines. The cytokine-dependent cell lines (THS119/IL-3 and THS119/IL-7) showed induction of STAT5 phosphorylation and target genes for STAT5 such as CIS, pim-1, p21 and bcl-xL upon addition of IL-3 or IL-7. IL-3 or IL-7 also induced STAT5 phosphorylation and STAT5 target genes of the stromal cell-dependent cell line, THS119, in the absence of stromal cells at levels similar to the cytokine-dependent cell lines. However, quite interestingly, TBR59 stromal cells could not induce STAT5 phosphorylation of THS119 cells, although they did induce STAT5 target genes in THS119 cells. In addition, the mRNAs for STAT5 target genes in THS119 cells on the stromal cells seemed to be more stable than those in the cytokine-dependent cell lines. Expression of the antiapoptotic genes bcl-2 and bcl-xL was higher in the stromal cell-dependent cell line than in the cytokine-dependent cell lines. These results suggested that stromal cells and cytokines may provide different signals for growth and differentiation of the hematopoietic cells.

Cell Division↗
PubMed2001

Repeated postpericardiotomy syndrome following a temporary transvenous pacemaker insertion, a permanent transvenous pacemaker insertion and surgical pericardiotomy.

The postpericardiotomy syndrome is a well-known complication of opening and manipulating the pericardium. The occurrence of this syndrome following transvenous pacemaker insertion is very rare, and only 5 cases have been reported to date. The present patient repeated this syndrome 3 times in a short period following 3 different interventional techniques: a temporary transvenous pacemaker, a permanent transvenous pacemaker and surgical pericardiotomy.

Aged↗
PubMed2001

Intramyocardial delivery of basic fibroblast growth factor-impregnated gelatin hydrogel microspheres enhances collateral circulation to infarcted canine myocardium.

The present study examined whether basic fibroblast growth factor (bFGF)-impregnated acidic gelatin hydrogel microspheres (AGHM) would enhance collateral development to the infarct area in dogs with coronary occlusion. Studies were conducted in 28 dogs with a 2-week occlusion of the proximal left anterior descending coronary artery (LAD). The dogs were divided into 3 groups according to treatment: Group A treated with bFGF-impregnated AGHM in the infarct area; Group B with free-form bFGF; Group C with AGHM alone. Coronary angiography (n=15; Group A, 7 dogs; Group B, 5 dogs; Group C, 3 dogs) and a regional myocardial blood flow study (n=13; Group A, 6 dogs; Group B, 4 dogs; Group C, 3 dogs) were repeated at a 2-week interval. Coronary angiography revealed that in Group A, antegrade flow in the LAD distal to the occlusion, which was assessed by Thrombolysis in Myocardial Infarction (TIMI) grade, was significantly increased after treatment. In contrast, in Groups B and C, the treatment did not change the flow grade in the LAD. In Group A, the regional myocardial blood flow in the collateral dependent area was significantly increased after treatment, and the regional myocardial blood flow reserve after adenosine injection was also significantly increased. These measurements remained after treatment in Groups B and C. The immunohistochemical study with factor VIII-related antigen revealed an increase of vascular density in the ischemic region in Group A. Intramyocardial delivery of bFGF-impregnated AGHM, but not free-form bFGF, improves the collateral circulation to the infarct area of a coronary occlusion in dogs.

Animals↗
PubMed2001

Polymorphisms of the CCK, CCKAR and CCKBR genes: an association with alcoholism study.

OBJECTIVE: Cholecystokinin (CCK) plays an important role in the functioning of the central nervous system via an interaction with dopamine and other neurotransmitters. The dopaminergic system has been implicated in the maladaptive behaviors associated with drugs of abuse (e.g., alcohol and cocaine). We analyzed genetic variations in the promoter and coding regions of the CCK, CCKA-receptor (CCKAR) and CCKB-receptor (CCKBR) genes, and performed association analyses with alcoholism. METHOD: A total of 214 Japanese male patients with alcoholism (93 with delirium tremens, 49 with hallucinations, 38 with seizures and 83 without any of these symptoms) and 98 age-matched Japanese male controls were examined using PCR-based Single Strand Conformational Polymorphism (SSCP) analysis. RESULTS: A total of 8 variants in the CCK gene, 11 variants in the CCKAR gene and 9 variants in the CCKBR gene were detected in the present study. Nominally significant differences between alcoholics and controls were found at the -85 locus of the CCKAR gene (p = .035). In addition, patients displaying hallucinations showed a higher frequency of the homozygous genotype of the allele (GT)8 at the -388 locus of the CCKAR gene (p = .042) and homo- and heterozygous genotypes of the T allele at the -333 locus (p = .025), relative to other patients. However, these differences were not significant after Bonferroni correction. CONCLUSIONS: Our data suggest that polymorphisms of the CCK, CCKAR and CCKBR genes do not play a major role in alcohol withdrawal symptoms (even though significant associations were found among polymorphisms at the -388 and -333 loci of the CCKAR gene and hallucinations, the rate was nonsignificant after Bonferroni correction).

Adult↗