Biomedical subjects

R Hirata

Publications and source records attributed to R Hirata.

At least 19 recordsLinked to original sources

PubMed2005

Prevalence of IgG diphtheria antitoxin in blood donors in Rio de Janeiro.

The lack of information on the immunity of adults in Brazil against diphtheria prompted us to analyse sera from 234 blood donors aged 18-61 years (30.3% females and 69.7% males). IgG diphtheria antitoxin levels determined by means of an ELISA, validated by toxin neutralization test in Vero cells, showed that 30.7% (95% CI 25.0-37.1) of the population was fully protected (>or=1 IU/ml). The highest percentage of subjects fully protected was in the 31-40 years age group. Most of the subjects with uncertain or no protection (<1 IU/ml) were found in the 18-30 years age group (43.8%, OR 2.18, P=0.01). Antitoxin levels were not influenced by the increase in age. Males were more protected than females (80.5%, OR 0.44, P=0.01). The prevalence of 30% of individuals fully protected against diphtheria in blood donors in Rio de Janeiro supports the fact that immunity to diphtheria among healthy Brazilian adults is inadequate. To avoid diphtheria epidemics in the future the immunity among adults should be raised in the coming years.

Adolescent↗

PubMed2003

The influence of organic load on the antimicrobial activity of different concentrations of NaOCl and chlorhexidine in vitro.

AIM: To evaluate bacterial growth after contact with sodium hypochlorite (NaOCl; 1 and 5%) and chlorhexidine (CHX; 0.12, 0.5 and 1%) in vitro with or without the addition of organic material (bovine serum albumin (BSA) 0.5%). METHODOLOGY: Bacterial samples (American Type Culture Collection (ATCC)) of Staphylococcus aureus, Enterococcus faecalis, Escherichia coli, Porphyromonas gingivalis and Fusobacterium nucleatum were kept in contact with each irrigating solution for varying intervals of time: immediately (t0), 5 min (t5), 15 min (t15) and 30 min (t30). Each test was repeated 10 times. In half of the specimens, 0.5% BSA was added as organic material in an attempt to simulate the organic tissue present in the root canal system. Bacterial growth under appropriate condition of incubation was evaluated and compared for each microorganism at all time intervals. RESULTS: A 0.12% CHX solution did not eliminate E. faecalis at any time interval. One percent CHX eliminated all strains, as did both NaOCl concentrations. BSA did not interfere substantially with the antimicrobial activity of any of the irrigating solutions. CONCLUSIONS: Under the condition of this study, a 0.12% CHX solution was ineffective at killing E. faecalis.

Anti-Infective Agents, Local↗

PubMed2001

Corynebacterium diphtheriae surface proteins as adhesins to human erythrocytes.

Corynebacterium diphtheriae strains express non-fimbrial surface proteins able to recognize and bind to specific host cells receptors. Protein extracts were obtained from bacterial cells by mechanical process and ammonium sulfate precipitation at 25 and 45% (w/v) saturation. SDS-PAGE analysis of the extracts detected two polypeptide bands of 67 and 72 kDa, named 67-72 p. The 67-72 p, rabbit anti-67-72 p IgG antibodies as well as human gastric mucin, N-acetylneuraminic acid and N-acetyl D-glucosamine molecules were able to inhibit bacterial hemagglutination. Hemagglutination assays using 67-72 p-coated latex beads and Western blot analysis of biotin-labeled 67-72 p and erythrocyte receptors demonstrated the binding of 67-72 p to human erythrocyte membranes. Immunolabeled colloidal gold-A protein transmission electron microscopy using anti-67-72 p revealed a diffuse distribution of non-fimbrial 67-72 p on the surface of C. diphtheriae strains of both sucrose-fermenting and non-fermenting biotypes. Non-fimbrial lectin-like surface 67-72 p may play a role as adhesins in bacterial attachment thereby facilitating the early steps in pathogenesis of both toxigenic and non-toxigenic C. diphtheriae.

Acetylglucosamine↗

PubMed2001

Effects of quercetin on the heat-induced cytotoxicity of prostate cancer cells.

BACKGROUND: It has been demonstrated that prostate cancer cells are relatively sensitive to heat stress. We have reported that heat treatment at 43 degrees C increases the expression of heat shock protein 70 (hsp70) in prostate cancer cells, leading to apoptosis. Hsp70 is a protein that protects cells against heat damage. Cells with lower levels of hsp70 have been shown to have a higher sensitivity to heat stress. Therefore, downregulation of hsp70 is expected to enhance heat-induced inhibitory effects on cell growth. Quercetin has been reported to be an agent that inhibits hsp70 expression. The present study was undertaken to investigate the effects of quercetin and/or heat on the growth of prostate cancer cells in vitro. METHODS: Three human prostate cancer cell lines were used: Lncap; PC-3; and JCA-1. The cells were treated with quercetin and/or heat. Alterations in the cell cycle and hsp70 expression were examined by means of flow cytometry (FCM). The apoptotic cells were detected by FCM using fluorescein isothiocyanate (FITC) labeled annexin V. RESULTS: Treatment with quercetin alone resulted in an apparent decrease of hsp70-positive cells and an increase of subG1 cells in JCA-1 and LNcap cells. Quercetin inhibited an increase of hsp70 expression after heat treatment and increased the number of subG1 cells with lower levels of hsp70 in JCA-1 and LNcap cells. Quercetin was found to enhance heat-induced inhibitory effects on cell growth and heat-induced apoptosis in both JCA-1 and LNcap cells. CONCLUSION: These results suggest that quercetin may enhance heat-induced cytotoxicity in prostate cancer cell lines through the inhibition of hsp70 production.

Apoptosis↗

PubMed2001

Identification of a 43-kDa outer-membrane protein as an adhesin in Aeromonas caviae.

Aeromonas spp. are associated with intestinal and extra-intestinal infections. However, the virulence factors of A. caviae remain, for the most part, poorly known. This study examined the interactions involved in the adherence of A. caviae isolates Ae56, Ae391 and Ae398 to HEp-2 cells. All strains expressed high levels of aggregative adherence. Maximum adhesion occurred with bacteria grown at 22 degrees C, but transmission electron microscopy did not reveal the presence of fimbrial structures on the bacterial cell surface. Outer-membrane proteins (OMPs) extracted from isolate Ae398, grown at 22 degrees C and 37 degrees C, showed similar SDS-PAGE protein profiles. Most proteins were < 60 kDa. A major 43-kDa protein was seen only in the boiled OMP extract. The biotinylated 43-kDa protein bound specifically to HEp-2 cells. Microbeads coated with the 43-kDa protein were also adherent to HEp-2 cells, and anti-43-kDa protein antibody blocked adherence of 43-kDa protein-coated latex beads. These data suggest that the 43-kDa OMP functions as an adhesin in A. caviae.

Adhesins, Bacterial↗

PubMed2001

[Tumor markers for hepatocellular carcinoma].

The diagnosis of hepatocellular carcinoma (HCC) is based mainly on serological tumor markers, such as alpha-fetoprotein, L3% fraction thereof and PIVKA-II, and imaging modalities. These are not correlated but are complementary. Hence, a combination designed to take advantage of the characteristics of each needs to be worked out. First, it is necessary to identify the patients at high risk for developing HCC, such as those with chronic hepatitis or liver cirrhosis, and in the follow-up conduct regular check-ups for serological tumor markers. Those testing positive for any marker are at the highest risk for developing HCC, even when imaging fails to disclose any space-occupying lesions. Following these high-risk patients, in concert with imaging, enables accurate evaluation of the efficacy of therapies for HCC. Since serological tumor markers can signal the development of HCC earlier than any other laboratory test, they offer an excellent means of identifying relapsing HCC. Equally important in the management of patients with HCC are biological indicators for malignancy, selection of therapeutic interventions and prediction of the outcome.

Biomarkers↗

PubMed2000

Immediate inhibition by oral l-ephedrine of passive cutaneous anaphylaxis of rats: indirect inhibition of anaphylactic chemical mediator release from the mast cell.

OBJECTIVE AND DESIGN: We previously reported that oral l-ephedrine showed extraordinarily rapid inhibition of 48-h passive cutaneous anaphylaxis (PCA) in rats. In the present study, in vivo and in vitro experiments were performed to elucidate a possible mechanism for the inhibition. MATERIALS: Rat antiserum was prepared with dinitrophenylated Ascaris suum extract + Bordetella pertussis. TREATMENT: Wistar rats were passively skin-sensitised, actively sensitised or non-sensitised. l-Ephedrine immediately before provocations was orally or intravenously administered in in vivo experiments. In in vitro experiments, the drug was added at various time and concentrations before the challenge. METHODS: The intensity of PCA was assessed by dye leakage method. Histamine and serotonin released in vitro or retained in the skin in vivo by anaphylaxis were assayed fluorometrically. RESULTS: Oral l-ephedrine rapidly inhibited the PCA by inhibiting the release of histamine and serotonin from the reaction site, whereas anaphylactic histamine and serotonin releases from skin fragments were not affected by the drug. Furthermore, the orally administered drug influenced neither the histamine- nor serotonin-induced cutaneous vascular permeability. CONCLUSIONS: These results were strongly indicative that the prompt suppression of the PCA by oral l-ephedrine was not exerted following the drug was absorbed from the gastrointestinal tract. Thus, the result may be from an indirect inhibition of chemical mediator release, possibly through an unidentified stimulation of the nervous system, but not from the inhibition of chemical mediator release by the direct interaction of drug to mast cells and not from the decreased vascular permeability.

Administration, Oral↗

PubMed2000

Oral beta-stimulants can inhibit passive cutaneous anaphylaxis in rats through an indirect inhibitory mechanism: possible involvement of afferent and efferent nervous system via gastric beta2-adrenoceptor stimulation.

OBJECTIVE AND DESIGN: We previously demonstrated that oral l-ephedrine exerts an extremely rapid (within 20 s) inhibition of 48-h passive cutaneous anaphylaxis reaction (PCA) in rats by a possibly unidentified mode of action. In the present experiments, we elucidated the mechanism of the PCA inhibition by l-ephedrine using adrenoceptor agonists and antagonists. MATERIALS: Rat antiserum was prepared with dinitrophenylated Ascaris suum extract + Bordetella pertussis. TREATMENT: Passively skin-sensitised Wistar rats were mainly used. l-Ephedrine, and adrenoceptor agonists and antagonists were orally administered immediately before PCA provocation. Catecholamine depleting (6-hydroxydopamine, 6-OHDA), amine depleting (reserpine) or ganglion blocking (hexamethonium) agent was intraperitoneally or intravenously administered before the provocation. METHODS: The effects of the drugs on PCA were assessed by inhibition of the dye leakage. RESULTS: beta-(propranolol) and beta2-(butoxamine) blocking agents reduced the inhibition of PCA by l-ephedrine, while the inhibition was not altered by either an a-blocking agent (phentolamine) or a beta1-(atenolol) selective antagonist. On the other hand, beta-(isoproterenol) and beta2-selective (salbutamol) agonists showed extremely rapid inhibition of PCA. However, the beta-selective agonist (dobutamine) had no effect on the reaction. The pretreatment with hexamethonium, reserpine or 6-OH-DA substantially attenuated the inhibitory effect of l-ephedrine on PCA. CONCLUSIONS: The results strongly suggest that beta2-adrenoceptors locate in the stomach and that their receptor excitement finally may lead to the inhibition of PCA via the stimulation of the central and peripheral nervous systems.

Adrenergic beta-2 Receptor Agonists↗

PubMed2000

Simultaneous production of granulocyte colony-stimulating factor and parathyroid hormone-related protein in bladder cancer.

A case of bladder cancer with simultaneous production of granulocyte colony-stimulating factor (G-CSF) and parathyroid hormone-related protein (PTHrP) is reported. An 81-year-old male patient was admitted to the Saitama Medical School for treatment of gross hematuria, leukocytosis and hypercalcemia and diagnosed as having advanced bladder cancer. Immediately after a cystectomy was carried out, his white cell count and serum calcium levels returned to normal. However, the tumors recurred locally and the recurrence was accompanied by an increase in the serum G-CSF and PTHrP levels with a recurrent elevation of white cell count and the serum calcium level. The production of G-CSF and PTHrP in the tumor cells was confirmed by immunohistochemistry.

Aged↗

PubMed2000

Hypercalcemia in a patient with renal cell carcinoma producing parathyroid hormone-related protein and interleukin-6.

A patient with renal cell carcinoma who developed humoral hypercalcemia of malignancy is reported. A 52-year-old male patient was diagnosed with renal cell carcinoma and multiple lung metastases. A cell line isolated from the surgical specimen exhibited continuous production of parathyroid hormone-related protein (PTHrP) in vitro. The production of PTHrP from the cancer cells was confirmed by RT-PCR and immunoradiometric assay. The serum calcium level was not enhanced, whereas the lung lesion was developing and producing interleukin-6, a possible modulator of osteoclastic resorption. Hypercalcemia was induced when the PTHrP concentration increased up to 3.3 pmol/L.

Carcinoma, Renal Cell↗

PubMed2000

Detection of Porphyromonas endodontalis in infected root canals by 16S rRNA gene-directed polymerase chain reaction.

Porphyromonas endodontalis has been isolated from the endodontic infections mainly in symptomatic teeth. This study evaluated the occurrence of P. endodontalis in both symptomatic and asymptomatic endodontic infections using 16S rRNA gene-directed polymerase chain reaction. P. endodontalis was detected in 39.5% of the cases (17 of 43 teeth). It was present in 4 of the 6 cases with acute periradicular abscess (66.7%) and in 13 of the 37 other cases (35.1%). The presence of P. endodontalis was associated with an asymptomatic periradicular lesion in 6 cases (25%) and in 10 teeth with tenderness to percussion (52.6%). P. endodontalis was also found in one asymptomatic case without evidence of periradicular pathosis. Our results indicated that, although P. endodontalis is commonly detected in symptomatic cases, it can be present in asymptomatic root canal infections. Further studies should determine if this bacterial species is really an important endodontopathogen.

Adult↗

PubMed1999

Patch clamp studies on V-type ATPase of vacuolar membrane of haploid Saccharomyces cerevisiae. Preparation and utilization of a giant cell containing a giant vacuole.

A method for obtaining giant protoplasts of Escherichia coli (the spheroplast incubation (SI) method: Kuroda et al. (Kuroda, T., Okuda, N., Saitoh, N., Hiyama, T., Terasaki, Y., Anazawa, H., Hirata, A., Mogi, T., Kusaka, I., Tsuchiya, T., and Yabe, I. (1998) J. Biol. Chem. 273, 16897-16904) was adapted to haploid cells of Saccharomyces cerevisiae. The yeast cell grew to become as large as 20 micrometer in diameter and to contain an oversized vacuole inside. A patch clamp technique in the whole cell/vacuole recording mode was applied for the vacuole isolated by osmotic shock. At zero membrane potential, ATP induced a strong current (as high as 100 pA; specific activity, 0.1 pA/micrometer(2)) toward the inside of the vacuole. Bafilomycin A(1,) a specific inhibitor of the V-type ATPase, strongly inhibited the activity (K(i) = 10 nM). Complete inhibition at higher concentrations indicated that any other ATP-driven transport systems were not expressed under the present incubation conditions. This current was not observed in the vacuoles prepared from a mutant that disrupted a catalytic subunit of the V-type ATPase (RH105(Deltavma1::TRP)). The K(m) value for the ATP dose response of the current was 159 microM and the H(+)/ATP ratio estimated from the reversible potential of the V-I curve was 3.5 +/- 0.3. These values agreed well with those previously estimated by measuring the V-type ATPase activity biochemically. This method can potentially be applied to any type of ion channel, ion pump, and ion transporter in S. cerevisiae, and can also be used to investigate gene functions in various organisms by using yeast cells as hosts for homologous and heterogeneous expression systems.

Anti-Bacterial Agents↗

PubMed1999

Suppression of default apoptosis in androgen-dependent cells by testosterone-mediated bcl-2 expression.

BACKGROUND: The significance of apoptosis with regard to the development and progression of androgen-dependent cells has not been clearly understood. In the present study we investigated the expression of the bcl-2 proto-oncogene after androgen deprivation and its role in cell growth in an androgen-dependent cell line. METHODS: We used SC2G, an androgen-dependent mouse mammary carcinoma cell line cloned from Shionogi carcinoma 115 (SC115). The expression of bcl-2 mRNA and protein in SC2G cells was measured by reverse transcription-polymerase chain reaction and western blotting, respectively. We also investigated the effects of antisense oligodeoxynucleotides (ODN) complementary to strategic sites in the mouse bcl-2 gene in SC2G cells. RESULTS: When SC2G cells were cultured in serum-free medium, the number of viable cells was significantly larger among cells with testosterone than those without testosterone after 3 days. Apoptosis was demonstrated in approximately 30% of positive-staining nuclei in SC2G cells cultured in testosterone-free medium. The levels of bcl-2 mRNA and protein in SC2G cells started to decrease after testosterone withdrawal. The cell density of SC2G cells decreased after 4 days culture with antisense ODN when compared with cells cultured in the presence of sense control. CONCLUSIONS: These data indicate that bcl-2 proto-oncogene inhibits the self-programmed apoptosis of androgen-dependent cells, suggesting the possibility of an antisense therapy for hormone-refractory prostate cancer, which is reported to express high levels of Bcl-2 protein.

Animals↗

PubMed1998

TAP polymorphism is not associated with ankylosing spondylitis and complications with acute anterior uveitis in HLA-B27-positive Japanese.

HLA-B27 is associated with the etiology of ankylosing spondylitis (AS) and acute anterior uveitis (AAU). Transporter associated with antigen processing (TAP) 1 and TAP2 polymorphism influences the range of peptide presented by HLA class I molecules. In this report, contribution of TAP polymorphism to the susceptibility to AS and AAU was studied in HLA-B27-positive Japanese individuals. Patients were classified into three groups: 16 AS patients, 14 AAU patients and 22 patients with both AS and AAU. Twelve HLA-B27-positive healthy individuals were included as a control. TAP polymorphism was detected by PCR-RFLP methods. Significant differences in frequencies of TAP1 alleles were not found between patient groups. None of the TAP2 frequencies showed increased or decreased frequencies compared with HLA-B27-positive healthy controls. In comparison with a random Japanese control, TAP2D allele frequency was significantly increased in the AAU group, but failed to reach a significant level in a group consisting of the AAU-only patients and the patients with both AS and AAU. All of the patient groups were noted to have a significantly increased prevalence of the TAP2H allele as compared to random controls; however, the higher frequency of this allele was detected in HLA-B27 healthy controls as well. These observations suggest a linkage disequilibrium between TAP2D, TAP2H and HLA-B27 in Japanese.

ATP Binding Cassette Transporter, Subfamily B, Mem↗

PubMed1998

[Anti-proliferative effects of heating on the human prostatic carcinoma cells in culture].

Prostatic cancers are well-known to be sensitive to heat stress. However, the mechanism by which the cancer cells are killed by high temperature remains poorly understood. The present study was undertaken to determine the anti-proliferative effects of heat stress on the prostatic cancer cells in culture. Heat shock at 43 degrees C inhibited the cell growth of three different prostatic cell lines. Flow cytometrical analysis using BrdU and PI showed a decrease in the proportion of cells in an S phase, accompanied by cell accumulation in G1 and G2, in both JCA-1 and PC-3 but not in LNcap. Both JCA-1 and PC-3 presented a strong expression of hsp70 at 37 degrees C. The heat shock caused apparent enhancement of the expression of hsp70 through the cell cycle. A treatment at 43 degrees C for 8 hours resulted in not only an apparent increment of positive hsp70 cells, but cells with subdiploid DNA content in LNcap. Flow cytometrical analysis by FITC-labeled Annexin V showed increment of apoptotic cells at 43 degrees C for 8 hours in LNcap cells. The results suggest that apoptosis is an important pathway of heat-induced killing of these cells. In conclusion, the cell growth of prostatic cancers may be affected by the temperature through relationship of the cell cycle and hsp70.

Apoptosis↗

PubMed1997

Sensitive high-performance liquid chromatographic method with fluorometric detection for the determination of heparin and heparan sulfate in biological samples: application to human urinary heparan sulfate.

A sensitive high-performance liquid chromatographic method for the determination of unsaturated disaccharides produced from heparin and heparan sulfate is described. Heparan sulfate was depolymerized using a combination of heparin lyase I (EC 4.2.2.7), heparin lyase II and heparin lyase III (EC 4.2.2.8). Seven unsaturated disaccharides were separated under isocratic conditions within 25 min using acetonitrile-H2O-0.2 M sodium phosphate buffer (pH 7.0)-3.0 M ammonium chloride (32:10:1:1) and were monitored by fluorescence detection using 2-cyanoacetamide as a post-column derivatizing reagent. As little as 2 pmol of a disaccharide could be detected with excitation at 346 nm and emission at 410 nm. This method was applied to the analysis of normal human urine. It was revealed that the concentration of normal human urinary heparan sulfate is 1.53+/-0.36 mg/mg creatinine (n=4).

Adult↗

PubMed1997

VMA11 and VMA16 encode second and third proteolipid subunits of the Saccharomyces cerevisiae vacuolar membrane H+-ATPase.

The vacuolar membrane H+-ATPase (V-ATPase) of the yeast Saccharomyces cerevisiae is composed of peripheral catalytic (V1) and integral membrane (V0) domains. The 17-kDa proteolipid subunit (VMA3 gene product; Vma3p) is predicted to constitute at least part of the proton translocating pore of V0. Recently, two VMA3 homologues, VMA11 and VMA16 (PPA1), have been identified in yeast, and VMA11 has been shown to be required for the V-ATPase activity. Cells disrupted for the VMA16 gene displayed the same phenotypes as those lacking either Vma3p or Vma11p; the mutant cells lost V-ATPase activity and failed to assemble V-ATPase subunits onto the vacuolar membrane. Epitope-tagged Vma11p and Vma16p were detected on the vacuolar membrane by immunofluorescence microscopy. Density gradient fractionation of the solubilized vacuolar proteins demonstrated that the tagged proteins copurified with the V-ATPase complex. We conclude that Vma11p and Vma16p are essential subunits of the V-ATPase. Vma3p contains a conserved glutamic acid residue (Glu137) whose carboxyl side chain is predicted to be important for proton transport activity. Mutational analysis of Vma11p and Vma16p revealed that both proteins contain a glutamic acid residue (Vma11p Glu145 and Vma16p Glu108) functionally similar to Vma3p Glu137. These residues could only be functionally substituted by an aspartic acid residue, because other mutations we examined inactivated the enzyme activity. Assembly and vacuolar targeting of the enzyme complex was not inhibited by these mutations. These results suggest that the three proteolipid subunits have similar but not redundant functions, each of which is most likely involved in proton transport activity of the enzyme complex. Yeast cells contain V0 and V1 subcomplexes in the vacuolar membrane and in the cytosol, respectively, that can be assembled into the active V0V1 complex in vivo. Surprisingly, loss-of-function mutations of either Vma11p Glu145 or Vma16p Glu108 resulted in a higher degree of assembly of the V1 subunits onto the V0 subcomplex in the vacuolar membrane.

Amino Acid Sequence↗

PubMed1997

A novel interferon-inducible gene expressed during myeloid differentiation.

The acute promyelocytic leukemia cell line, NB4, can be induced to differentiate to mature granulocytes by retinoic acid treatment. A novel retinoic acid-inducible cDNA clone, designated RI58, was isolated from a cDNA library constructed from retinoic acid-treated NB4 cells by differential hybridization. RI58 cDNA encodes a protein of 58kDa which has a similarity in its amino acids sequence to interferon (IFN)-inducible proteins. In addition, RI58 was induced by recombinant human IFN-alpha (rhIFN-alpha) in NB4 cells. RI58 was detectable within 4 hours post-stimulation with rhIFN-alpha, while it took as long as 1day after retinoic acid stimulation. Culture supernatant from retinoic acid-treated NB4 cells also induced RI58 expression similarly as rhIFN-alpha. This activity in culture supernatant was inhibited by anti-leukocyte IFN antiserum which showed specific reactivity to rhIFN-alpha. These results indicate that RI58 is induced by retinoic acid stimulation through autocrinally secreted IFN-alpha from NB4 cells. In the retinoic acid-treated NB4 cells, the expression of RI58 was increased along the process of differentiation. On the other hand, it was expressed constitutively in untreated non-hematopoietic cell lines and mature hematopoietic cell lines.

Amino Acid Sequence↗