Biomedical subjects

T Shirakawa

Publications and source records attributed to T Shirakawa.

At least 37 records · Page 2Linked to original sources

PubMed2001

Drug-resistant human bladder-cancer cells are more sensitive to adenovirus-mediated wild-type p53 gene therapy compared to drug-sensitive cells.

We investigated the therapeutic potential and molecular mechanism of adenovirus-mediated wt p53 gene therapy for drug-resistant human bladder cancers. KK47, a human bladder-cancer cell line, along with the drug-resistant sublines KK47/DDP10, KK47/DDP20 (cisplatin-resistant) and KK47/ADM (doxorubicin-resistant) were used for the experiments. All 4 KK47 cell lines had genetically normal p53 genes. Using an in vitro cytotoxicity assay, the drug-resistant cell lines were more sensitive to Ad-CMV-p53 cell killing than the KK47 parental cell line. Ad-CMV-p53 induced higher levels of p53 protein and mRNA in the drug-resistant cell lines than in the parental cell line and, consequently, higher levels of p21 and Bax mRNA, which resulted in higher percentages of G(1) cell-cycle arrest and apoptosis. The higher efficiencies of adenoviral gene transfer in the drug-resistant cell lines were confirmed by X-gal staining after infection with Ad-CMV-beta-gal. In conclusion, adenovirus-mediated wt p53 gene therapy was more effective in the drug-resistant bladder-cancer cell lines than in the drug-sensitive bladder-cancer cell line.

Adenoviridae↗

PubMed2001

Suppression of the transcriptional activity and DNA binding of nuclear factor-kappa B by a paraneoplastic cerebellar degeneration-associated antigen.

Paraneoplastic cerebellar degeneration (PCD) associated with gynecological malignancies is a disorder in which an autoimmune mechanism has been suggested, and both antibody- and cell-mediated immune responses exist against pcd17/cdr2, a neural protein expressed in cerebellar Purkinje neurons and brainstem neurons. In this report, we describe that pcd17 can suppress the basal or activated NF-kappaB-dependent transcriptional activity in a co-transfection study. The DNA binding of constitutive NF-kappaB complexes decreased in the nucleus of TNF-alpha-stimulated neuroblastoma cells, though pcd17 does not bind to classical NF-kappaB consensus site. These data indicate that pcd17 is a potential repressor for NF-kappaB-dependent gene transcription in neurons.

Animals↗

PubMed2001

Circadian rhythm of nitric oxide production in the dorsal region of the suprachiasmatic nucleus in rats.

Extracellular concentration of nitrite (NO2-), an oxidized product of nitric oxide (NO), was measured consecutively in the dorsal region of the rat suprachiasmatic nucleus (SCN) by means of in vivo microdialysis. The NO2- concentrations in the dialysates showed robust circadian rhythm under a 12:12 h light/dark cycle and were higher during the dark phase than during the light phase. When the rats were transferred to constant darkness, the 24 h rhythm of NO2- persisted without damping the amplitude. The NO2- level was significantly lowered by an injection of NO synthase inhibitor (NG-monomethyl-L-arginine, 10 mg/kg i.p.). These findings indicate that the daily fluctuation of NO2- in the dorsal region of the SCN, which represents endogenous rhythm of NO, is regulated independently of photic inputs into the SCN and may be related to the circadian clock functions.

Animals↗

PubMed2001

Fibroblast growth factor-binding protein expression changes with disease progression in clinical and experimental human squamous epithelium.

Basic fibroblast growth factor (bFGF) is synthesized by a wide variety of normal and malignant cells. However, bFGF cannot exert its effects unless it gets outside of the cell. Since it lacks a signal sequence to direct secretion, the method by which cells release it remains unclear. A 17 kDa secreted binding protein for bFGF (FGF-BP, HBp-17) is expressed at high levels in squamous cell carcinoma (SCC) and transformed keratinocytes and may act as a chaperone to transport bFGF outside of the cell. In our study, FGF-BP mRNA expression in normal keratinocytes was higher than in 5/5 SCCs. Using a new monoclonal antibody, we demonstrate that FGF-BP can dimerize. Immunoassays demonstrate that normal keratinocytes have a higher level of FGF-BP than SCCs. In normal human squamous epithelium, we observed diffuse, moderate to intense cytoplasmic and membranous expression of FGF-BP. Expression decreased and became focal with disease progression to invasive cancer. Injection of immortalized but non-tumorigenic HaCaT cells transduced with FGF-BP into normal human skin xenografts failed to result in tumors. Transfection of FGF-BP into the SCCs Det 562 and FaDu did not promote tumor growth more than controls, and peri-tumoral microvessel density was lower in FGF-BP-transfected than in control tumors. Taken together, these data suggest that FGF-BP expression in squamous epithelium does not play an important role in progression to invasive carcinoma.

Antibodies, Monoclonal↗

PubMed2001

Induction of major histocompatibility complex class I molecules on human neuroblastoma line cells by a flavoid antioxidant.

Human major histocompatibility complex (MHC) class I expression is usually suppressed in neuronal cells and neuroblastoma cells. In the present study, we analyzed the effect of a flavonoid antioxidant, silymarin, on the induction of MHC class I molecules in human neuroblastoma line cells. Treatment of neuroblastoma cells with silymarin resulted in the expression of MHC class I molecules. Silymarin treatment enhanced the transcriptional activity of the reporter construct containing MHC class I promoter truncated within -428 bp of transcription initiation, but not the construct containing the promoter truncated within -284 bp. Because an E-box element is located between -428 and -285 bp of the transcription initiation, results suggest that silymarin acts on the enhancer activity of the E-box in the MHC class I promoter. Our findings indicate that silymarin induces the transcriptional factors to enhance the MHC class I promoter through the class I E-box element.

Antioxidants↗

PubMed2001

Induction of cytotoxic T lymphocytes specific for paraneoplastic cerebellar degeneration-associated antigen in vivo by DNA immunization.

Paraneoplastic cerebellar degeneration associated with gynecological and breast malignancies (PCD) is known to develop autoantibodies and autoreactive cytotoxic T lymphocytes (CTLs) specific for a cytoplasmic protein of Purkinje cells PCD17/cdr2, in the blood of patients. The functional roles of these antibodies and CTLs in the pathogenesis of PCD are unknown. Induction of immune response to this antigen in experimental animals should be useful to clarify the immune mechanisms in PCD patients. We immunized Balb/c mice with naked DNA, which is encoded human PCD17 neural protein in an eukaryotic expression plasmid under control of CMV promoter, and explored whether or not humoral and cell-mediated immune responses against PCD17 could be induced in vivo. We show that DNA immunization with naked pcd17 cDNA could induce autoantibodies against the cytoplasmic protein of Purkinje cells and CTLs could lyse syngenic myeloma cells pulsed with H-2K-restricted PCD17 peptide. In spite of the generation of anti-Purkinje cell antibodies and PCD17-specific CTLs in vivo, neither clinical nor pathological changes consistent with significant cerebellar degeneration have been detected.

Animals↗

PubMed2001

Polymorphisms of renin-angiotensin system genes in childhood IgA nephropathy.

We investigated whether polymorphisms of the renin-angiotensin system genes are involved in IgA nephropathy in Japanese children. We identified the M235T polymorphism of the angiotensinogen (AGT) gene, the I/D polymorphisms of the angiotensin-converting enzyme (ACE) gene, and the A1166C polymorphism of the angiotensin II type 1 receptor gene in 95 Japanese children with IgA nephropathy and 99 healthy Japanese adults. There were no differences in the genotype and allele frequencies of these genes between patients with IgA nephropathy and controls. Urinary protein excretion at the time of biopsy was significantly greater in patients with the TT genotype of the AGT gene than in those with the MM/MT genotypes of the AGT gene (1.32 +/- 1.42 versus 0.75 +/- 0.78 g/day; P = 0.01) and in patients with the ID/DD genotypes of the ACE gene than in those with the II genotype of the ACE gene (1.45 +/- 1.50 versus 0.63 +/- 0.56 g/day; P = 0.001). Thus, the TT genotype of the AGT gene and the ID/DD genotype of the ACE gene are associated with increased severity of proteinuria, suggesting that AGT and ACE gene polymorphisms may play a significant role in the progression of IgA nephropathy in Japanese children.

Adolescent↗

PubMed2001

Y-chromosome microdeletion and phenotype in cytogenetically normal men with idiopathic azoospermia.

OBJECTIVE: To determine the prevalence of microdeletions of the long arm of chromosome Y within the AZFa, AZFb, and AZFc subregions in patients with idiopathic azoospermia, and then correlate the microdeletions with clinical phenotypes to determine the most important subregion for screening. DESIGN: Controlled clinical study. SETTING: Male infertility clinic, Kobe University Hospital. PATIENT(S): Among 89 consecutive azoospermic patients, those whose infertility was related to known hereditary, endocrine, or obstructive causes or a cytogenetic abnormality were excluded; 54 remaining patients were studied using a polymerase chain reaction (PCR). Of these patients, 33 had Sertoli cell only syndrome, 10 had maturation arrest, and 11 had hypospermatogenesis. INTERVENTION(S): Blood and semen samples and testicular biopsies were obtained from all of the participants. MAIN OUTCOME MEASURE(S): We performed semen analysis, polymerase chain amplification of 28 DNA loci on the long arm of the Y chromosome involving the DAZ (deleted in azoospermia), and measured the plasma FSH, LH, testosterone, prolactin, and estradiol levels. RESULT(S): Microdeletions were detected in 14 of the 54 patients (nine with Sertoli cell only, three with maturation arrest, and two with hypospermatogenesis). Most microdeletions involved AZFb or AZFc. Patients with hypospermatogenesis or maturation arrest showed deletion only in AZFc. The DAZ gene was deleted in four patients with Sertoli cell only and one patient with maturation arrest. The RBM gene was deleted in two patients with Sertoli cell only who had particularly large deletions, but in no patients with arrest or hypospermatogenesis. CONCLUSION(S): Cytogenetically azoospermic patients should be examined for microdeletions before undertaking assisted reproduction. AZFc may be the most important subregion to screen. In addition, intact AZFa and AZFb subregions may be important for the presence of germ cells.

Adult↗

PubMed2001

Regional pacemakers composed of multiple oscillator neurons in the rat suprachiasmatic nucleus.

Regional specificities of the dorsal and ventral regions of the suprachiasmatic nucleus (SCN) were examined to elucidate the structure of multioscillator circadian organization. The circadian rhythms of arginine vasopressin (AVP) and vasoactive intestinal polypeptide (VIP) release, and of electrical activity of individual neurons were measured in an organotypic, static slice culture of the SCN obtained from neonatal rats. Five days after the start of culture, robust circadian rhythms were detected in AVP release with a peak located consistently at the middle of the original light phase, while the 24 h profiles of VIP release were either arrhythmic or rhythmic. In the latter case, a phase delay of 5-7 h was observed in the circadian peak from the AVP rhythm. Multi-channel, extracellular recording revealed that 51 (76.1%) out of 67 firing neurons, examined in the SCN, showed circadian rhythms in their firing rate. The percentage of rhythmic neurons was significantly larger in the dorsal (86.8%) than in the ventral (62.1%) region of the SCN, where the AVP and VIP containing neurons predominate, respectively. Twenty-seven percent of the firing rhythms were almost antiphasic from the majority of rhythms. There was no regional specificity in the distribution of the antiphasic rhythm. These findings, that the dorsal and ventral regions of the SCN both contain circadian pacemakers with different properties that regulate the AVP and VIP release separately, is probably due to differences in the number and, hence, the coupling strength of oscillating neurons.

Action Potentials↗

PubMed2001

Multiple oscillators in the suprachiasmatic nucleus.

The suprachiasmatic nucleus (SCN) of the hypothalamus is the site of the pacemaker that controls circadian rhythms of a variety of physiological functions. Data strongly indicate the majority of the SCN neurons express self-sustaining oscillations that can be detected as rhythms in the spontaneous firing of individual neurons. The period of single SCN neurons in a dissociated cell culture is dispersed in a wide range (from 20h to 28h in rats), but that of the locomotor rhythm is close to 24h, suggesting individual oscillators are coupled to generate an averaged circadian period in the nucleus. Electrical coupling via gap junctions, glial regulation, calcium spikes, ephaptic interactions. extracellular ion flux, and diffusible substances have been discussed as possible mechanisms that mediate the interneuronal rhythm synchrony. Recently, GABA (gamma-aminobutyric acid), a major neurotransmitter in the SCN, was reported to regulate cellular communication and to synchronize rhythms through GABA(A) receptors. At present, subsequent intracellular processes that are able to reset the genetic loop of oscillations are unknown. There may be diverse mechanisms for integrating the multiple circadian oscillators in the SCN. This article reviews the knowledge about the various circadian oscillations intrinsic to the SCN, with particular focus on the intercellular signaling of coupled oscillators.

Animals↗

PubMed2001

Gene therapy for bladder cancer using adenoviral vector.

BACKGROUND AND PURPOSE: Bladder cancer is common. Current treatment for patients with superficial bladder cancer involves transurethral resection followed by adjuvant bacillus Calmette-Guérin (BCG) administration. Adjuvant BCG has been reported to be effective in 38% to 68% of patients; however, more than 30% of patients do not respond. Because p53 mutations are common among superficial bladder cancers, we tested the feasibility of using p53 as a gene therapy agent for targeting superficial tumors, which are easily accessible using an intravesical approach. MATERIALS AND METHODS: Wild-type p53 was transduced into various human and murine bladder cancer cell lines (HTB9, KU-1, and MBT-2) using a recombinant adenoviral vector (Ad5CMV-p53) in vitro. Also, subcutaneous tumors were established and then treated with intratumoral injection of Ad5CMV-p53 or control viruses. RESULTS: In vitro assays revealed significant growth suppression of target cells by Ad5CMV-p53 in comparison with those receiving the control Ad5-CMV-PA vector or untreated control cells. In vivo studies using subcutaneous bladder tumor models established in syngeneic mice demonstrated that the rate of tumor growth and volume was reduced to a greater extent by 14 days of intratumoral injection of Ad5CMV-p53 rather than Ad5CMV-PA. Furthermore, the survival of host animals bearing tumors that were infected with Ad5CMV-p53 was significantly longer than that of the control group treated with Ad5CMV-PA (P < 0.01). CONCLUSION: Our data suggest that Ad5CMV-p53 is effective in suppressing bladder cancer growth and improving host survival.

Adenoviridae↗

PubMed2001

Genetic polymorphisms of CC chemokine receptor 3 in Japanese and British asthmatics.

Whole genome scan analyses have revealed that chromosomal region 3p21-24, which contains a gene cluster of CC chemokine receptors such as CCR3, is possibly linked to asthma. Because CCR3 ligands play a pivotal role in the selective recruitment and activation of inflammatory cells in the asthmatic airway, the authors examined whether there is any association between asthma and the CCR3 gene polymorphisms. Three polymorphisms were identified using the single stranded conformational polymorphism method in Japanese (Asian) and British (Caucasian) subjects; one silent mutation T51C and two missense mutations G824A and T971C. These polymorphisms were examined in 391 Japanese subjects (210 asthmatics and 181 nonasthmatic controls) and 234 British subjects (142 asthmatics and 92 nonasthmatic controls). Asthma diagnosis was based on episodic symptoms, documented wheeze, and the presence of reversible airflow limitation. CCR3 T51C demonstrated a significant association with the diagnosis of asthma in the British population (odds ratio 2.35, p<0.01), but not in the Japanese population. Multiple logistic regression analysis also showed that CCR3 T51C was associated with asthma (odds ratio 2.83, p < 0.02), independent of atopic phenotypes such as high levels of total or house dust mite-specific immunoglobulin-E in serum. In conclusion, a significant association between asthma and CCR3 T51C polymorphism localized on chromosome 3p21 was found.

Alleles↗

PubMed2001

Color/power Doppler sonographic differential diagnosis of superficial lymphadenopathy: metastasis, malignant lymphoma, and benign process.

OBJECTIVE: Differentiation of lymph nodes as metastasis, malignant lymphoma, and benign lesions by gray scale and color/power Doppler ultrasonography has been reported. In this study we sought to determine patterns of abnormal vascularity, the relationship between diagnostic criteria and node size, and which vessel in the node was best suited for Doppler spectral analysis. Extrahilar vessels, defined as vessels entering through the lymphatic surface and not through the hilus, were also investigated. METHODS: One hundred twenty-nine nodes were examined by color/power Doppler ultrasonography. RESULTS: Extrahilar vessels were seen in 85.4% of metastatic, 40.5% of lymphomatous, and 7.7% of benign nodes regardless of node size. The mean pulsatility index and resistive index values of metastatic nodes were higher than those of benign nodes regardless of their size. Those of lymphoma were intermediate compared with the others. CONCLUSIONS: A pulsatility index greater then 1.3 and a resistive index greater than 0.72 suggested malignancy as measured from any vessel.

Breast Neoplasms↗

PubMed2001

[A case of post-tuberculosis chronic empyema with effective low-dose, long-term clarithromycin administration].

A 78-year-old woman with post-tuberculosis chronic empyema who underwent unsuccessful decortication in July 1993, was admitted to our hospital in November 1994 because of M. tuberculosis in her sputum. One month after anti-tuberculosis treatment, the M. tuberculosis disappeared from the sputum, but a pleurocutaneous fistula that exuded abundant bloody pus containing Pseudomonas aeruginosa persisted in the area of chronic empyema. In July 1997, 200 mg of clarithromycin (CAM) was administered daily; 21 months later, the pus and pleurocutaneous fistula had disappeared. Post-tuberculosis chronic empyema with a fistula is generally an intractable condition that necessitates surgery. In this case, the empyema did not improve during the 4 years after the surgery. However, low-dose, long-term CAM administration brought about a disappearance of pus from the fistula and closure of the empyema spaces.

Aged↗

PubMed2001

[Prospects for molecular research in urological oncology: gene therapy].

The recent great advances in genetic engineering are now making possible the identification and isolation of the trigger genes of many hereditary illnesses, and the clarification of the relevant molecular mechanisms. The idea that if the genetic abnormalities responsible for illness could be established at a DNA level, treatment at the genetic level repairing damaged genes or supplying absent ones would also be possible was the incentive for the recent boom in gene therapy. Clinical research into gene therapy began in 1990 and currently over 3,000 patient cases are being studied. Some 70% of these are cancer patients. This is not simply because such patients are relatively numerous, but is also a sign of the wish, held earnestly by many researchers and clinicians as well as cancer patients and their families, to at last overcome this intractable disease. Gene therapy, so far conducted mainly in the United States, has hitherto not lived up to initial expectations in its concrete results. The reason for this results mainly in technical factors, such as the rate of success in implanting genes into target cells, the rate of successful expression of the implanted genes, and the successful achievement of specific expression at the target site. Gene therapy in the form of clinical research into renal cancer and lung cancer is now under way in Japan. It is too early at this stage to evaluate this work, but the present paper takes this opportunity to give an outline of gene therapy, and to examine its current state, future prospects and problem areas with particular reference to cancer.

Bone Neoplasms↗

PubMed2001

Month of birth, atopic disease, and atopic sensitization.

BACKGROUND: Japanese cedar (Cryptmeria japonica; CJ) pollen and house dust mites are the two important aeroallergens in Japan. However, no epidemiological survey has been performed in Japan to investigate the relationship between month of birth and manifestations of allergic disease and sensitization. OBJECTIVE: This study evaluates the correlation between month of birth and sensitization to aeroallergens or the occurrence of allergic disease on 755 Japanese school children aged 12-13 years. METHODS: The personal history of atopic disease (bronchial asthma, allergic rhinitis, eczema, and allergic conjunctivitis) as recorded by questionnaires was investigated in relation to total serum IgE and specific IgE toward house dust mites and CJ pollen. RESULTS: Positive specific IgE toward house dust mites was significantly less prevalent in the children born between January and March than those born during the rest of the year (p < 0.01). Positive specific IgE toward CJ pollen was significantly more prevalent in the children born between December and January than those born during the rest of the year (p < 0.05). High total IgE was less prevalent in the children born between February and April than in children born during the rest of the year (p = 0.05). The prevalence of bronchial asthma was 26.2% among children born between November and December, compared with a ratio of 17.3% among children born during the rest of the year (p < 0.05). A significantly higher proportion of the children with allergic rhinitis was born between August and October than during the rest of the year (p < 0.05). The prevalence of allergic conjunctivitis was 15.8% among the children who were born between December and January, compared with 9.1% among children born during the rest of the year (p < 0.01). No relationship between prevalence of eczema and season of birth was found. CONCLUSION: Month of birth appears to influence the risk in the development of allergic sensitization and atopic diseases. The findings concerning higher CJ pollen sensitization in children born in the months that proceed the CJ pollen seasons are as evident as the house-dust-mite-related findings.

Adolescent↗

PubMed2000

Activation of thrombosis and fibrinolysis following brain infarction.

To clarify the sequence of alterations in the thrombotic and fibrinolytic systems after acute brain infarction, we prospectively examined sequential changes in coagulatory markers in 38 patients suffering from cardioembolic infarcts (CEI), 41 patients with atherothrombotic infarcts (ATI), 58 patients with lacunar infarcts (LI), and 32 age-matched controls. The plasma level of thrombin-antithrombin III complex (TAT), fibrinopeptide A (FpA), D-dimer, fibrin degradation products-E (FDP-E), fibrinogen, alpha2-plasmin inhibitor-plasmin complex (PIC), and percent activity of antithrombin III (AT-III) were measured within 48 h, at 1 week, and at 3 weeks after the stroke onset. Significantly elevated levels of TAT and FpA, which are both markers of thrombin formation, were observed in CEI patients, and these elevated levels were associated with increasing D-dimer levels for 3 weeks (P<0.0001). D-Dimer in CEI patients was significantly elevated compared to control, LI and ATI levels within 48 h (P<0.001). Percent activity of AT-III was significantly decreased in CEI patients for 3 weeks compared to this activity in controls, LI and ATI (P<0.001). TAT and FpA also increased significantly within 48 h in ATI subjects and declined thereafter. A significant elevation of FDP-E (P<0.001) and D-dimer (P<0.05, P<0.01) was detected in parallel with increasing fibrinogen for 3 weeks. However, there was no significant depletion of percent activity of AT-III in ATI. In LI subjects, no significant elevation of TAT, D-dimer or FDP-E were observed within 1 week. PIC increased significantly in three subtypes of brain infarcts, but did not differ significantly among the three subtypes for 3 weeks. An accurate assessment of sequential alterations in thrombotic and fibrinolytic markers in the acute stage of brain infarct should contribute to the clinical diagnosis of brain infarct subtype. Alterations in these markers in response to activation of the coagulatory system are attributable to the different pathogenesis of ischemic stroke.

Aged↗

PubMed2000

Synaptic communication of cellular oscillations in the rat suprachiasmatic neurons.

Circadian firing rhythms of cultured rat suprachiasmatic nucleus were measured simultaneously from 4-8 neurons by using a multi-electrode dish and neuronal interactions were examined by a cross-correlation analysis of spontaneous action potentials. Functional connections were detected in the neuron pairs showing synchronized circadian firing rhythms, and when the connections were lost, firing rhythms were desynchronized. After the prolonged treatment with tetrodotoxin, cross-correlation and circadian rhythm synchronization were abolished concomitantly in most neuron pairs. Cellular mechanisms involving Na(+)-channel dependent communication are responsible for the synchronization of the circadian rhythms in individual suprachiasmatic nucleus (SCN) neurons.

Action Potentials↗