Biomedical subjects

H Watanabe

Publications and source records attributed to H Watanabe.

At least 163 records · Page 9Linked to original sources

PubMed2001

Number and position of mutations in the interferon (IFN) sensitivity-determining region of the gene for nonstructural protein 5A correlate with IFN efficacy in hepatitis C virus genotype 1b infection.

To explore the relationship between responses to interferon (IFN) and the mutation patterns in the IFN sensitivity-determining region (ISDR; amino acid positions 2209-2248) in the NS5A gene of hepatitis C virus genotype 1b, a cohort of 334 patients was analyzed. The number of mutations in the ISDR was higher in patients with sustained response (SR) than in patients with transient or no response (P<.001). Patients with viruses mutated at positions 2209 (P=.02), 2216 (P=.01), or 2227 (P=.02) more frequently experienced SR than did those without these mutations. Mutation occurred most frequently at position 2218, where the presence of cysteine was significantly associated with SR. Thus, the mutation pattern in the ISDR affects the virologic response to IFN and reflects different influences on the function of the NS5A protein. ISDR sequence analysis would allow the prediction of clinical IFN efficacy in individual patients.

Amino Acid Sequence↗

PubMed2001

Brain 5alpha-dihydroprogesterone and allopregnanolone synthesis in a mouse model of protracted social isolation.

Allopregnanolone (ALLO), is a brain endogenous neurosteroid that binds with high affinity to gamma-aminobutyric acid type A (GABA(A)) receptors and positively modulates the action of GABA at these receptors. Unlike ALLO, 5alpha-dihydroprogesterone (5alpha-DHP) binds with high affinity to intracellular progesterone receptors that regulate DNA transcription. To investigate the physiological roles of ALLO and 5alpha-DHP synthesized in brain, we have adopted a mouse model involving protracted social isolation. In the frontal cortex of mice, socially isolated for 6 weeks, both neurosteroids were decreased by approximately 50%. After administration of (17beta)-17-(bis-1-methyl amino carbonyl) androstane-3,5-diene-3-carboxylic acid (SKF105,111), an inhibitor of the enzyme (5alpha-reductase Type I and II) that converts progesterone into 5alpha-DHP, the ALLO and 5alpha-DHP content of frontal cortex of both group-housed and socially isolated mice decreased exponentially to 10%-20% of control values in about 30 min. The fractional rate constants (k h(-1)) of ALLO and 5alpha-DHP decline multiplied by the ALLO and 5alpha-DHP concentrations at any given steady-state estimate the rate of synthesis required to maintain that steady state. After 6 weeks of social isolation, ALLO and 5alpha-DHP biosynthesis rates were decreased to 30% of the values calculated in group-housed mice. Moreover, in socially isolated mice, the expression of 5alpha-reductase Type I mRNA and protein was approximately 50% lower than in group-housed mice whereas 3alpha-hydroxysteroid oxidoreductase mRNA expression was equal in the two groups. Protracted social isolation in mice may provide a model to investigate whether 5alpha-DHP by a genomic action, and ALLO by a nongenomic mechanism down-regulate the action of drugs acting as agonists, partial agonists, or positive allosteric modulators of the benzodiazepine recognition sites expressed by GABA(A) receptors.

Animals↗

PubMed2001

Dynamic changes in subnuclear NP95 location during the cell cycle and its spatial relationship with DNA replication foci.

We determined the expression and subcellular localization of nuclear protein NP95 during the cell cycle in mouse 3T3 cells. The levels of NP95 mRNA and protein were extremely low in quiescent cells; however, stimulation with 10% serum increased their expressions in a time course similar to that of the late growth-regulated gene proliferating cell nuclear antigen (PCNA). Subnuclear location of NP95 dynamically changed during the cell cycle. Double immunostaining for NP95 and chromatin-bound PCNA, a marker of DNA replication sites, revealed that NP95 was almost exclusively colocalized with chromatin-bound PCNA throughout the nucleus in early S phase and partly in mid-S phase. Distinct localization of the two proteins, however, became evident in mid-S phase, and thereafter, many chromatin-bound PCNA foci not carrying NP95 foci could be detected. In G2 phase, nodular NP95 foci were still identified without any chromatin-bound PCNA foci. Chromatin-bound PCNA was observed as a pre-DNA replication complex at the G1/S boundary synchronized by hydroxyurea treatment, while NP95 was detected in nucleolar regions as unique large foci. There was no significant redistribution of NP95 foci shortly after DNA damage by gamma-irradiation. Nodular NP95 foci characteristically seen in G2 phase were also detected in G2-arrested cells following gamma-irradiation. Taken together, our results indicate that NP95 is assigned to a late growth-regulated gene and suggest that NP95 does not take a direct part in DNA replication as part of the DNA synthesizing machinery, like PCNA, but is presumably involved in other DNA replication-linked nuclear events.

3T3 Cells↗

PubMed2001

Transcriptional cross-talk between Smad, ERK1/2, and p38 mitogen-activated protein kinase pathways regulates transforming growth factor-beta-induced aggrecan gene expression in chondrogenic ATDC5 cells.

In chondrogenesis, members of the transforming growth factor-beta (TGF-beta) superfamily play critical roles by inducing gene expression of cartilage-specific molecules. By using a chondrogenic cell line, ATDC5, we investigated the TGF-beta-mediated signaling pathways involved in expression of the aggrecan gene (Agc). At confluency, TGF-beta induced Agc expression within 3 h, and cycloheximide blocked this induction, indicating that de novo protein synthesis is essential for this response. At this stage, TGF-beta induced rapid, transient phosphorylation of Smad2, extracellular signal-activated kinase 1/2 (ERK1/2), and p38 mitogen-activated protein kinase (MAPK). Inhibition of the Smad pathways by transfection with a dominant negative Smad4 construct significantly reduced TGF-beta-induced Agc expression, indicating that Smad signaling is essential for this response. Furthermore, an inhibitor of the ERK1/2 pathway, U0126, or inhibitors of the p38 MAPK pathway, SB203580 and SKF86002, repressed TGF-beta-induced Agc expression in a dose-dependent manner, indicating that ERK1/2 or p38 MAPK activation is also required for TGF-beta-induced Agc expression in confluent ATDC5 cells. In differentiated ATDC5 cells, persistently high basal levels of ERK1/2 and p38 MAPK phosphorylation correlated with elevated basal Agc expression, which was inhibited by incubation with inhibitors of these pathways. Whereas Smad2 was rapidly phosphorylated by TGF-beta and involved in the initial activation of Agc expression in confluent cells, Smad2 activation was not required for maintaining the high level of Agc expression. Taken together, these results suggest an important role for transcriptional cross-talk between Smad and MAPK pathways in expression of early chondrocytic phenotypes and identify important changes in the regulation of Agc expression following chondrocyte differentiation.

Aggrecans↗

PubMed2001

Association of intermediate T cell receptor cells, mainly their NK1.1(-) subset, with protection from malaria.

Mice were infected with Plasmodium (P.) yoelii blood-stage parasites. Both the liver and spleen were the sites of inflammation during malarial infection at the beginning of day 7. The major expanding cells were found to be NK1.1(-) intermediate alphabetaTCR (alphabetaTCR(int)) in the liver and spleen, although the population of NK1.1(+) alphabetaTCR(int) cells remained constant or slightly increased. These TCR(int) cells are of extrathymic origin or are generated by an alternative intrathymic pathway and are distinguished from conventional T cells of thymic origin. During malarial infection, the population of conventional T cells did not increase at all. TCR(int) cells purified from the liver of mice which had recovered from P. yoelii infection protected mice from malaria when they were transferred into 6.5-Gy-irradiated mice. Interestingly, the immunity against malaria seemed to disappear as a function of time after recovery, namely, mice which had recovered from malaria 1 year previously again became susceptible to malarial infection. The present results suggest that TCR(int) cells are intimately associated with protection against malarial infection and, therefore, that mice which had recovered from malaria 1 year previously lost such immunity.

Animals↗

PubMed2001

Neutrophil depletion exacerbates experimental Chagas' disease in BALB/c, but protects C57BL/6 mice through modulating the Th1/Th2 dichotomy in different directions.

To elucidate the roles of neutrophils in experimental Chagas' disease, we depleted the peripheral neutrophils in BALB/c and C57BL/6 mice with a monoclonal antibody 1 day before Trypanosoma cruzi infection. Neutrophil depletion in BALB/ c mice resulted in exacerbation of the disease and decreased expression of mRNA for Th1 cytokines, including IL-2 and IFN-gamma, IL-12p40 and TNF-alpha in their spleens after the infection, while a Th2 cytokine, IL-10, increased especially 1 day after infection. Neutrophils from infected BALB / c mice expressed mRNA for IL-12p40, IFN-gamma, TNF-alpha and Th1 chemoattractive chemokines, monokine induced by IFN-gamma (MIG) and macrophage inflammatory protein-1alpha (MIP-1alpha ). In contrast, in C57BL/6 mice neutrophil depletion induced resistance to the disease and enhanced the expression of the above Th1 cytokines, although IL-10 mRNA in neutrophil-depleted C57BL/6 mice was also higher than in control mice. Neutrophils from C57BL/6 mice did not express IL-12p40, IFN-gamma and MIG but expressed TNF-alpha, MIP-1alpha and IL-10. Therefore, neutrophils may play opposite roles in these two strains of mice with respect to protection versus exacerbation of T. cruzi infection, possibly through modulating the Th1/Th2 dichotomy in different directions.

Animals↗

PubMed2001

Abundance of unconventional CD8(+) natural killer T cells in the large intestine.

Natural killer T (NKT) cells are mainly present in the liver and thymus, and the majority of these T cells express either a CD4(+) or a double-negative (DN) CD4(-)8(-) phenotype. In the present study, we examined whether such NKT cells were present in the intestine. NKT cells were rare in all sites of the small intestine, including an intraepithelial site. However, a considerable number of NKT cells were found at an intraepithelial site in the large intestine. This result was confirmed by both immunofluorescence and immunohistochemistry. In contrast to conventional NKT cells, NKT cells in the large intestine were CD8(+) or DN CD4(-)8(-). In the case of conventional NKT cells, their existence is known to depend on non-classical MHC class I-like antigens (i. e. CD1d) but not on classical MHC class I antigens. However, the NKT cells in the large intestine were independent of the presence of both CD1d and classical MHC class I antigens. These results were obtained using knockout mice lacking the corresponding genes and molecules. NKT cells in the large intestine were mainly alpha betaTCR(+) (> 75 %) but did not use an invariant chain of Valpha14Jalpha281, which is preferentially used by conventional NKT cells. These NKT cells did not bias the TCR-Vbeta usage toward Vbeta8. These findings suggest that the large intestine is a site in which unconventional NKT cells carrying the CD8(+) phenotype (or DN CD4(-)8(-)) are abundant and that these cells are independent of MHC and MHC-like antigens.

Animals↗

PubMed2001

Capillary electrochromatography using fibers as stationary phases.

Fiber-packed capillary columns have been evaluated in chromatographic performance in capillary electrochromatography (CEC). The change of electroosmotic flow (EOF) velocity and selectivity using different kinds of fiber materials was examined. Although the EOF velocity among the different fiber packed columns was almost the same, retention of parabens was larger on the Kevlar-packed column than on the Zylon-packed one, and was larger on the as-span-type fiber-packed column than on the high-modulus-type packed one. Using 200 microm ID x 5 cm Kevlar packed column combined with a 100 microm ID x 20 cm precolumn capillary and a 530 microm ID x 45 cm postcolumn capillary, the separation of three parabens within 30 s was achieved. Other compounds were also separated in a few minutes by the fiber-packed CEC method.

Electrophoresis, Capillary↗

PubMed2001

Novel asymmetric oxy-michael addition reaction of the chiral ketones to the achiral gamma--or delta-hydroxy-alpha,beta-unsaturated carbonyl compounds.

A novel asymmetric oxy-Michael addition reaction was developed. In the presence of a catalytic amount of base, chiral ketones 1 and 2, derived from D-glucose and D-fructose, respectively, reacted with omega-hydroxy enones or enoates 3a-e, 17 and 21 to form the hemiacetal-derived alkoxide which underwent stereoselective intramolecular Michael addition to give cyclic acetals. Although the stereoselectivities in the formation of the five-membered acetal rings were modest, six-membered ring formation proceeded with high stereoselectivity and the utility of the reaction was demonstrated by a simple syntheses of natural products.

Biological Factors↗

PubMed2001

Suppressive effects of thyroxine on glucocorticoid (gc)-induced metabolic changes and cataract formation on developing chick embryos.

To prevent cataracts induced by glucocorticoids (GC) as a systemic disease, the suppression of oxidative stress caused by GC in the hepatic metabolism is of significant interest. In this study, to elucidate the formative mechanism of GC-induced cataracts, we examined the preventive effect and then analysed the mechanisms of thyroxine on GC-induced cataract formation. Fifteen day old chick embryos were administered with 0.25 micromol hydrocortisone succinate sodium (HC), and then 12-30 nmol of thyroxine 4 hr after HC administration. At the indicated time after HC treatment, we examined the incidence of cataract formation, the levels of serum glucose and lipids, lenticular and hepatic glutathione (GSH), and lipid peroxide (LPO) in the lens, blood and liver. Almost all lenses (96%) removed 48 hr after HC administration were opaque. Thyroxine prevented HC-induced cataract formation effectively, and suppressed the elevations of serum glucose and LPO in the lens, blood and liver. The treatment prevented the decreased lenticular GSH level at 48 hr, but the hepatic GSH level at 24 hr remained lowered in contrast to the results of previous studies using insulin. Moreover, thyroxine did not decrease the elevated serum lipid level (triglyceride and non-esterified fatty acid) caused by HC. Under thyroxine treatment, in constant to insulin, acceleration of GSH-GSSG cycle rather than GSH de novo synthesis keeps a certain level of hepatic GSH necessary for diminishing the elevation of LPO as a risk factor of GC-induced cataract formation. The regulation of metabolic changes ensured the maintenance of hepatic GSH level, which is necessary to reduce oxidative stress produced by GC and to protect the lens from oxidative stress leading to opacification.

Animals↗

PubMed2001

Developmental effects of estrogenic agents on mice, fish, and frogs: a mini-review.

Laboratory experiments have demonstrated that exposure of rodents to sex hormones during prenatal or early postnatal life can cause permanent and irreversible alterations of the endocrine and reproductive organs, such as ovary, fallopian tube, uterus, cervix, vagina, and mammary gland in females; and testis, epididymis, prostate, and seminal vesicle in males; as well as non reproductive organs including bones and muscle and immune and nervous systems in both sexes. Early development of Xenopus laevis into the tadpole and Fundulus heteroclitus goes through a rapid cell division, gastrulation, neurulation, and organogenesis within 1 week after fertilization. The developing embryo is very fragile and sensitive to estrogenic agents. Thus, these animals can be used as a suitable model for examining the effect of endocrine disruptors (hormonally active agents) on the development of aquatic living beings, which are most likely to be exposed to the compounds.

Animals↗

PubMed2001

A flagellar gene fliZ regulates the expression of invasion genes and virulence phenotype in Salmonella enterica serovar Typhimurium.

We previously reported that the fliZ gene encodes a positive regulatory factor for the class 2 flagellar operons in Salmonella enterica serovar Typhimurium. In this study, we found that the fliZ mutation reduced not only the amounts of excreted flagellar proteins, but also those of several secreted invasion proteins encoded by the genes within Salmonella pathogenicity island 1. Using the lacZ gene fused to a subset of virulence-associated genes, we show that this downregulation was caused by a decreased transcription of the hilA gene, which encodes a positive regulator for the invasion genes. We further show that the fliZ mutation reduced invasion ability of S. enterica serovar Typhimurium to HEp-2 cells. Consistent with these results, orally challenged cells of the fliZ mutant show an attenuated virulence phenotype in a mouse typhoid model. These results indicate that the fliZ gene product positively regulates the invasion genes and is necessary for expression of full virulence.

Animals↗

PubMed2001

MR-based statistical atlas of the Göttingen minipig brain.

Thedomestic pig is increasingly being used as an experimental model for brain imaging studies with positron emission tomography (PET). The recording of radiotracer uptake by PET gives functional and physiological information, but with poor spatial resolution. To date, anatomical regions of interest in pig brain have been defined in MR images obtained for each individual animal, because of the lack of a standard stereotaxic coordinate system for the pig brain. In order to define a stereotaxic coordinate system, we coregistered T1-weighted MR images from 22 male Göttingen minipigs and obtained a statistically defined surface rendering of the average minipig brain in which stereotaxic zero is defined by the position of the pineal gland. The average brain is now used as a target for registration of dynamic PET data, so that time-activity curves can be extracted from standard volumes of interest. In order to define these volumes, MR images from each individual pig were manually segmented into a total of 34 brain structures, including cortical regions, white matter, caudate and putamen, ventricular system, and cerebellum. The mean volumes of these structures had variances in the range of 10-20%. The 34 brain volumes were transformed into the common coordinate system and then used to generate surface renderings with probabilistic threshold greater than 50%. This probabilistic threshold gave nearly quantitative recovery of the mean volumes in native space. The probabilistic volumes in stereotaxic space are now being used to extract time-radioactivity curves from dynamic PET recordings.

Animals↗

PubMed2001

Systematic separation and purification of elastase, gelatinase (matrix metalloproteinase 9), and collagenase (matrix metalloproteinase 8) from polymorphonuclear leukocytes in dialyzers previously used by patients with renal failure.

We developed a simple and effective method for the systematic separation and purification of human polymorphonuclear leukocyte (PMN) proteinases, elastase, gelatinase (matrix metalloproteinase 9, type IV collagenase), and collagenase (matrix metalloproteinase 8), derived from the extracts of hollow fiber dialyzers that had been utilized in the treatment of patients with renal failure. The fraction containing elastase was grossly separated from that containing gelatinase and collagenase by heparin-Sepharose chromatography and purified in an aprotinin column. The remaining two enzymes were then separated using the gelatin-Sepharose column after gel chromatography following ammonium sulfate precipitation. Gelatinase and collagenase were further purified by gelatin-Sepharose chromatography as a latent form and by collagen-Sepharose chromatography as an activated form. This novel method offers procedural advantages over existing methods that separate PMNs from the whole blood of volunteers for experimental research purposes.

Animals↗

PubMed2001

Animal cellulases.

Previous dogma has maintained that cellulose, ingested by xylophagous or herbivorous animals, is digested by cellulolytic symbiotes. The first evidence in conflict with this contention involved the demonstration of cellulolytic activities in symbiote-free secreting organs (e.g., the salivary glands of termites) or defaunated guts. Following these demonstrations, possible endogenous cellulase components were purified from several cellulose-digesting invertebrates, but this research did little to change the general view concerning animal cellulose digestion. Thanks to recent developments in molecular biology, the existence of cellulases of animal origin has been firmly established. To date, cellulase genes have been reported from arthropods (insects and a crayfish) and nematodes. This paper describes and discusses the presence and nature of endogenous cellulases in higher animals.

Amino Acid Sequence↗

PubMed2001

Mass screening program for prostatic cancer in Japan.

Because the idea of "mass screening" in Japan is based upon the concept of "groupism", a fundamental feature of Japanese society, it is very different from the idea of "screening" in Western countries, which arises from an "individualistic" concept. The prevalence of prostatic cancer in Japan remains lower than that in Western countries but it has recently shown a great increase. In this situation, a mass screening program for prostatic cancer, using prostate-specific antigen (PSA) for the primary study, is thought to be promising in Japan. To exclude false-positive cases in the "gray zone" of PSA values, transrectal power Doppler sonography is remarkably effective.

Biopsy↗

PubMed2001

Low-intensity fetal lungs on MRI may suggest the diagnosis of pulmonary hypoplasia.

BACKGROUND: Pulmonary hypoplasia is a common cause of neonatal death. Despite the recent advances in prenatal diagnosis with US, the diagnosis of pulmonary hypoplasia is difficult. The recent application of fast MR imaging may provide additional valuable information. OBJECTIVE: To evaluate pulmonary hypoplasia in the fetus with MRI. MATERIALS AND METHODS: The subjects comprised 23 fetuses (18-40 weeks' gestation), including major anomalies diagnosed on fetal ultrasonography (n = 20), maternal abnormality (n = 2) and one normal twin. MRI was performed with a 1.5-T magnet and half-Fourier acquisition single-shot turbo spin-echo (HASTE) sequences. MR images were interpreted by three radiologists with special attention to the intensity of the lungs. The lung-to-liver intensity ratio was calculated by means of region-of-interest (ROI) analysis. The diagnosis of pulmonary hypoplasia depended on clinical, surgical and autopsy findings. RESULTS: All fetuses with normal pulmonary development showed high intensity in the lung except for one fetus at 24 weeks' gestational age. All fetuses with pulmonary hypoplasia showed lung of low intensity. CONCLUSIONS: Low-intensity fetal lung on MRI imaging indicates pulmonary hypoplasia after 26 weeks' gestation.

Female↗

PubMed2001

Schwannoma of the extremities: the role of PET in preoperative planning.

The aim of this study was to determine the relative utility of various preoperative diagnostic imaging modalities for the evaluation of benign schwannoma, including positron emission tomography (PET) utilising fluorine-18 fluoro-2-deoxy-D-glucose (FDG) and fluorine- 18 alpha-methyl tyrosine (FMT). computed tomography (CT), magnetic resonance imaging (MRI) and digital subtraction angiography (DSA). We retrospectively reviewed imaging findings in 22 patients with 25 histopathologically documented benign schwannomas of the extremities. Pre-operative imaging included: FDG-PET (n=22), FMT-PET (n=17), MRI (n=25), CT (n=16) and DSA (n=17). All 22 lesions examined by PET with FDG and/or FMT showed accumulation. The standardised uptake values (SUVs) for FDG-PET for the 22 examined tumours ranged from 0.33 to 3.7, and eight of them (36.4%) were assessed as malignant on the basis of their uptake. The SUVs for FMT ranged from 0.44 to 1.47, and 15 out of the 17 evaluated (88.2%) showed values indicating the tumour to be benign. CT demonstrated variable attenuation and contrast enhancement. MRI signal characteristics were relatively consistent: iso-signal or darker than skeletal muscle on T1-weighted and isosignal or brighter than subcutaneous fat on T2-weighted images. The venous tumour staining depicted on DSA was found to be significantly correlated with FDG accumulation. All tumours but one were treated by surgical enucleation. One tumour suspected to be malignant on the basis of imaging findings was treated with primary wide resection. Although CT, MRI and PET studies are all useful for the detection and localisation of schwannoma, our findings suggest that, among the imaging modalities studied, FMT-PET may be the most reliable technique for the differentiation of benign schwannoma from malignancy.

Adult↗