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Biomedical subjects

M Yamada

Publications and source records attributed to M Yamada.

At least 757 records · Page 42Linked to original sources
PubMed1998

[In vivo study on intrathecal use of 5-fluoro-2'-deoxyuridine (FdUrd) in meningeal dissemination of malignant tumor].

FdUrd was evaluated in vivo as a potential agent for intrathecal chemotherapy of meningeal carcinomatosis. Neurotoxicity was examined pathologically in normal mice after 4 consecutive intrathecal injections of FdUrd. Using mice models of meningeal carcinomatosis, antitumor activities were studied by evaluating survival time. Pathological examination showed none of the following abnormal findings: demyelination, degeneration and destruction of ependymal linings. FdUrd also had an effect on meningeal carcinomatosis of mice (203 glioma and MM46 transplantable ascitic mammary cancer). In causing FdUrd to exhibit its efficacy, it is necessary to take into consideration the balance between the activity of two key enzymes, thymidine phosphorylase (TPase) (anabolic enzyme) and thymidine kinase (TK) (metabolic enzyme), in tumor tissues as compared with their activity in normal tissues. TPase activity which results in conversion to 5-FU was much lower in malignant glioma and metastatic brain tumors compared with tumors in other extracranial organs. TPase activity in normal brain was much less than in normal tissues in extracranial organs and its activity in gray matter (cortex) was significantly lower than that in white matter. On the other hand, TK activity in malignant brain tumors was much less than that in extracranial organs, however, its activity in normal brain was almost equal to that in normal tissues in extracranial organs. These data obtained in vivo study showed FdUrd to be a possible agent for intrathecal chemotherapy.

Animals↗
PubMed1998

Fas-mediated apoptosis is involved in the elimination of gene-transduced hepatocytes with E1/E3-deleted adenoviral vectors.

Gene-transduced hepatocytes with E1/E3-deleted adenoviral vectors are eliminated immediately and the expression of transduced genes disappears rapidly following the vector administration. In this report, we analysed the involvement of apoptotic cell death in the elimination of hepatocytes infected with adenoviral vectors. An E1/E3-deleted adenoviral vector expressing Escherichia coli beta-galactosidase (LacZ) was injected via the portal vein into congenitally Fas-deficient mice (lpr), Fas ligand-deficient mice (gld) and their control mice, MRL and C3H. 5-Bromo-4-chloro-3-indolyl-beta-D-galactoside (X-gal) staining of the liver specimens showed that 80-100% of hepatocytes were LacZ positive at 7 days after virus administration, suggesting that most of the hepatocytes received the injected adenoviral vectors. In normal mice, the number of LacZ-positive cells decreased dramatically at 14 and 21 days after transduction and few positive cells were observed at day 28. Beta-galactosidase activity, quantified by the O-nitrophenyl-beta-D-galactopyranoside assay, gave comparable results to X-gal staining. At days 14 or 21, many apoptotic hepatocytes and apoptotic infiltrating cells were detected with the terminal deoxyribonucleotidyl transferase-mediated dUTP-digoxigenin nick end-labelling (TUNEL) in situ apoptosis detection method. This observation suggested that the apoptotic process was associated with the elimination of adenovirus-infected hepatocytes. To test the involvement of the Fas-Fas ligand interaction in this apoptotic process, the period of transgene expression was measured in lpr and gld mice, which had received the same amount of AxCALacZ. X-Gal histochemical analysis detected many LacZ-positive cells in lpr or gld mice liver even at 21 or 28 days after AxCALacZ injection. There were significant differences in the reduction rates of beta-galactosidase activity of liver homogenates between lpr and MRL, or gld and C3H mice. Based on these observations, we conclude that the Fas-mediated apoptotic process is involved in the elimination of hepatocytes infected with E1/E3-deleted adenoviral vectors.

Adenoviridae↗
PubMed1998

[Clinical application of FdUrd to meningeal dissemination of malignant tumors].

A clinical trial of intrathecal chemotherapy with FdUrd was performed in sixteen patients with meningeal dissemination of malignant tumors. Twelve of the sixteen patients responded to intrathecal FdUrd chemotherapy (1-5 micrograms/dose) through an Ommaya reservoir placed in the lateral ventricle: complete response, 2; partial response, 10; progressive disease, 4. Only slight nausea was observed in two patients and dull headache in one patient. No other systemic side effects such as myelosuppression or liver dysfunction were observed in any patients. Moreover, no delayed side effects such as marked brain atrophy or leukoencephalopathy developed during the course of this intrathecal chemotherapy despite over thirty consecutive intrathecal administrations. In conclusion, intrathecal FdUrd proved to be safe and effective for the treatment of meningeal dissemination of malignant tumors.

Adenocarcinoma↗
PubMed1998

[Application of a semi-automatic ROI setting system for brain PET images to animal PET studies].

ProASSIST, a semi-automatic ROI (region of interest) setting system for human brain PET images, has been modified for use with the canine brain, and the performance of the obtained system was evaluated by comparing the operational simplicity for ROI setting and the consistency of ROI values obtained with those by a conventional manual procedure. Namely, we created segment maps for the canine brain by making reference to the coronal section atlas of the canine brain by Lim et al., and incorporated them into the ProASSIST system. For the performance test, CBF (cerebral blood flow) and CMRglc (cerebral metabolic rate in glucose) images in dogs with or without focal cerebral ischemia were used. In ProASSIST, brain contours were defined semiautomatically. In the ROI analysis of the test image, manual modification of the contour was necessary in half cases examined (8/16). However, the operation was rather simple so that the operation time per one brain section was significantly shorter than that in the manual operation. The ROI values determined by the system were comparable with those by the manual procedure, confirming the applicability of the system to these animal studies. The use of the system like the present one would also merit the more objective data acquisition for the quantitative ROI analysis, because no manual procedure except for some specifications of the anatomical features is required for ROI setting.

Animals↗
PubMed1998

Missense mutations in the PAX6 gene in aniridia.

PURPOSE: Aniridia is caused by a mutation of the PAX6 gene. Haploinsufficiency of the gene product is thought to result in the aniridia phenotype, because most mutations thus far detected have been large deletions encompassing the entire gene and nonsense, frameshift, or splice errors that result in premature translational termination on one of the alleles. Only two missense mutations have been detected in aniridia pedigrees, each of which occurs in its paired domain or homeodomain. In this study, four novel missense mutations were found in three aniridia pedigrees. METHODS: Polymerase chain reaction-single-strand conformation polymorphism analysis and sequencing of the PAX6 gene were performed using genomic DNA of three aniridia pedigrees and more than 100 healthy control subjects. RESULTS: Three mutations occurred in the N-terminal subdomain of the paired domain, namely N17S, I29V, and R44Q, the first two of which were detected on the same allele of one patient. The other mutation (Q178H) was in the linking portion of the paired domain and homeodomain. CONCLUSIONS: These missense mutations give rise to haploinsufficiency by another route, because the missense mutations presented here resulted in an aniridia phenotype indistinguishable from that caused by a heterozygous deletion of the entire PAX6 gene.

Adult↗
PubMed1998

Scalp localization using a simple algorithm.

We have developed a simple algorithm for scalp localization for craniotomy. Using a series of CT/MRI data, the point on the scalp closest to the center of the lesion (T) is determined by our program. A plane (plane R) is defined by three points: bilateral auditory meati and T. Point S is the intersection of three planes: the plane R, the scalp surface and the midsagittal plane. The distance from either S or the ipsilateral external auditory meatus to T is measured along the scalp surface on the plane R. The distance from the nasion to S along the scalp surface on the midsagittal plane is also measured. To determine the craniotomy site, these distances are measured directly on a patient's scalp in the operating room. This simple and accurate method for scalp localization could be used by installing our program in conventional CT/MR scanners.

Aged↗
PubMed1998

[Combined scatter and attenuation correction for 201Tl myocardial perfusion SPECT using OS-EM algorithm].

There are two possible ways to obtain scatter-corrected images with the ML-EM (maximum likelihood expectation maximization) algorithm: one is the subtraction of scatter estimate si from projection data pi, and then (pi-si) is used for scatter-corrected projection data (denoted as SC(T)); the other method is the addition of scatter estimate si to the projections calculated from the reconstructed image without performing data subtraction (SC(E)). This paper investigated these two ML-EM algorithms of combined scatter and attenuation correction on 201Tl myocardial perfusion SPECT imaging. Scatter windows were placed one full width at half maximum (FWHM) below and above the photopeak centerline. The scatter fraction in the primary peak was estimated using trapezoidal approximation by the triple energy window method. Phantom and clinical images were reconstructed using 6 iterations of ordered subsets EM algorithm (OS-EM). A cylindrical phantom with a cold-rod insert and a heart/thorax phantom with liver insert were used to evaluate scatter and the attenuation compensation technique. A cylindrical phantom filled with uniform 201Tl solution was used to evaluate statistical noise. The percent root-mean-square uncertainty (%RMSU) was used as a quantitative measure of noise amplification. %RMSU showed that the SC(E) method amplified noise less in comparison with the SC(T) method, however, no significant difference in image quality was observed between the two methods. In conclusion, both the SC(T) and SC(E) methods provided significant and similar improvement in the removal of scatter in 201Tl myocardial perfusion SPECT imaging.

Absorption↗
PubMed1998

Chimeras of Kir6.1 and Kir6.2 reveal structural elements involved in spontaneous opening and unitary conductance of the ATP-sensitive K+ channels.

The ATP-sensitive K+ (KATP) channels, e.g. in heart and pancreatic beta-cells, open spontaneously in the absence of intracellular ATP (ATPi). Their unitary conductance is approximately 80 pS with 150 mM extracellular K+. These features are shared by the K+ channels composed of various sulfonylurea receptors (SURs) and Kir6.2, whereas SUR/Kir6.1 channels have a smaller conductance (approximately 35 pS) and do not open spontaneously in the absence of ATPi. To identify the structural elements in Kir6.0 subunits which determine these properties, we analyzed the properties of functional K+ channels composed of SUR2A, the cardiac type SUR, and various chimeras of Kir6.1 and Kir6.2 heterologously expressed in HEK (human embryonic kidney) 293T cells. The analyses indicate that the extracellular linker domain between the two putative membrane-spanning regions is responsible for the difference in the single channel conductance between SUR2A/Kir6.1 and SUR2A/Kir6.2 channels. The cytosolic N-terminal domain of Kir6.2 was mandatory for spontaneous channel opening in the absence of ATPi, although a part of C-terminus was also involved. These results implicate specific regions of Kir6.0 in the spontaneous opening and the single channel conductance.

Adenosine Triphosphate↗
PubMed1997

BDNF potentiates spontaneous Ca2+ oscillations in cultured hippocampal neurons.

Brain-derived neurotrophic factor (BDNF) is thought to regulate neuronal plasticity in developing and matured neurons, although the molecular mechanisms are less well characterized. We monitored changes in the intracellular calcium (Ca2+) levels induced by BDNF using a fluorescence Ca2+ indicator (Fluo-3) by means of confocal laser microscopy in rat cultured hippocampal neurons. BDNF acutely potentiated spontaneous Ca2+ oscillations in dendrites and also in the soma of several neurons, although it increased intracellular Ca2+ in only selective proportion of resting neurons without Ca2+ oscillations. The potentiation was observed both in the frequency and the amplitude of Ca2+ oscillations, completely blocked by K-252a, and significantly reduced by 2-aminophosphonovaleric acid. These findings suggest that BDNF increases glutamate release and N-methyl-D-aspartate (NMDA) channel-gated Ca2+ influx via TrkB and regulates the frequency and the amplitude of Ca2+ oscillations. BDNF may have the potential to modulate spontaneous Ca2+ oscillations to regulate neuronal plasticity in developing hippocampal neurons.

2-Amino-5-phosphonovalerate↗
PubMed1997

Multiple pathways for SOS-induced mutagenesis in Escherichia coli: an overexpression of dinB/dinP results in strongly enhancing mutagenesis in the absence of any exogenous treatment to damage DNA.

dinP is an Escherichia coli gene recently identified at 5.5 min of the genetic map, whose product shows a similarity in amino acid sequence to the E. coli UmuC protein involved in DNA damage-induced mutagenesis. In this paper we show that the gene is identical to dinB, an SOS gene previously localized near the lac locus at 8 min, the function of which was shown to be required for mutagenesis of nonirradiated lambda phage infecting UV-preirradiated bacterial cells (termed lambdaUTM for lambda untargeted mutagenesis). A newly constructed dinP null mutant exhibited the same defect for lambdaUTM as observed previously with a dinB::Mu mutant, and the defect was complemented by plasmids carrying dinP as the only intact bacterial gene. Furthermore, merely increasing the dinP gene expression, without UV irradiation or any other DNA-damaging treatment, resulted in a strong enhancement of mutagenesis in F'lac plasmids; at most, 800-fold increase in the G6-to-G5 change. The enhanced mutagenesis did not depend on recA, uvrA, or umuDC. Thus, our results establish that E. coli has at least two distinct pathways for SOS-induced mutagenesis: one dependent on umuDC and the other on dinB/P.

Bacterial Proteins↗
PubMed1997

Insulin receptor substrate (IRS)-1 and IRS-2 are tyrosine-phosphorylated and associated with phosphatidylinositol 3-kinase in response to brain-derived neurotrophic factor in cultured cerebral cortical neurons.

Brain-derived neurotrophic factor (BDNF), a member of the neurotrophins, promotes differentiation and survival of various types of neurons in the central nervous system. BDNF binds to and activates the tyrosine kinase receptor, TrkB, initiating intracellular signaling and exerting its effects. Phosphatidylinositol 3-kinase (PI3-K), which has been implicated in promotion of neuronal survival by neurotrophic factors, is a component in the signaling pathway of BDNF. We examined how BDNF activates PI3-K in cultured cerebral cortical neurons. We found that insulin receptor substrate (IRS)-1 and -2 are involved in the BDNF signaling pathway that activates PI3-K. IRS-1 and -2 were tyrosine-phosphorylated and bound to PI3-K in response to BDNF. This BDNF-stimulated signaling via IRS-1 and -2 was inhibited by K-252a, an inhibitor of Trk tyrosine kinase. In addition, signaling via IRS-1 and -2 was markedly sustained as well as the BDNF-induced tyrosine phosphorylation of TrkB. On the other hand, we observed no association of PI3-K with TrkB in response to BDNF. These results indicate that the activation of TrkB by BDNF induces the activation of PI3-K via IRS-1 and -2 rather than by a direct interaction of TrkB with PI3-K in cultured cortical neurons.

Animals↗
PubMed1997

New tester strains of Salmonella typhimurium lacking O6-methylguanine DNA methyltransferases and highly sensitive to mutagenic alkylating agents.

Salmonella typhimurium YG7104 and YG7108 are derivatives of the Ames tester strain TA1535, and have chromosomal deletions of the ogtST gene or both the ogtST and adaST genes, respectively. The ogtST and adaST genes encode O6-methylguanine DNA methyltransferases that are involved in the repair of DNA damage caused by alkylating agents. The sensitivities of these strains to 15 mutagens with different structures were tested and compared with those of the parent strain TA1535. Deletion of ogtST or ogtST plus adaST substantially increased the sensitivity of strain TA1535 to the mutagenicity of alkylating agents, such as N-ethyl-N'-nitro-N-nitrosoguanidine, ethyl methanesulfonate or dimethylnitrosamine (DMN). Preincubation of the chemical with S9 mix and bacteria for 20 min at 37 degrees C before pouring them together on agar plates was not necessary to detect the mutagenicity of DMN when strain YG7104 or YG7108 was used as a tester strain. Introduction of plasmid pKM101 did not enhance but rather decreased the sensitivity of YG7104 and YG7108 to alkylating agents. Since the new strains are highly sensitive only to alkylating agents, they will be useful to detect the mutagenicity with high efficiency and to study the mechanism of mutagenesis induced by environmental alkylating agents.

Alkylating Agents↗
PubMed1997

Dentatorubral pallidoluysian atrophy (DRPLA) protein is cleaved by caspase-3 during apoptosis.

Dentatorubral pallidoluysian atrophy (DRPLA) is an autosomal dominant neurodegenerative disorder. It is associated with an abnormal CAG repeat expansion resulting in formation of a protein with an elongated polyglutamine stretch. However, neither the physiological roles of the DRPLA gene product nor molecular mechanisms of its pathogenesis have yet been elucidated. Here we report that the DRPLA protein is cleaved at a site near the N terminus during apoptosis induced by VP-16, staurosporine, or glucocorticoid. Moreover, the in vitro translated DRPLA protein is cleaved by recombinant caspase-3, a member of the cysteine protease family, which is thought to be a main executioner of apoptosis. Using mutant DRPLA proteins, the cleavage site was identified as 106DSLDG110. The cleavage, however, was not modulated by the length of the polyglutamine stretch. These findings suggest that the DRPLA protein is one of the physiological substrates of caspase-3, and its cleavage may result in structural and biochemical alterations associated with apoptosis.

Amino Acid Sequence↗
PubMed1997

Induction of nitric oxide synthase by lipopolysaccharide and its inhibition by auranofin in RAW 264.7 cells.

In RAW 264.7 cells, a murine macrophage cell line, treatment with lipopolysaccharide (1 to 10 ng/ml) stimulated production of nitric oxide (NO), which was inhibited by L-N(G)-monomethyl-L-arginine acetate, an inhibitor of NO synthase. Auranofin, an orally active chrysotherapeutic agent, also inhibited the lipopolysaccharide-induced NO production in a concentration-dependent manner (0.3 to 3 microM). Other gold salts such as aurothioglucose and aurothiomalate had no effect. Western blot analysis demonstrated that the lipopolysaccharide (10 ng/ml)-induced expression of inducible NO synthase protein was inhibited by auranofin as well as by the protein synthesis inhibitor cycloheximide. In addition, the lipopolysaccharide-induced increase in the level of mRNA for inducible NO synthase was also lowered by auranofin. Furthermore, auranofin showed no direct effect on the conversion of [3H]arginine to [3H]citrulline by the cell lysate. These findings indicate that auranofin inhibits lipopolysaccharide-induced NO production by suppressing the expression of inducible NO synthase.

Animals↗
PubMed1997

Construction of mutants of Salmonella typhimurium deficient in 8-hydroxyguanine DNA glycosylase and their sensitivities to oxidative mutagens and nitro compounds.

8-Hydroxyguanine (8-OH-G) DNA glycosylase is an enzyme involved in repair of oxidative DNA damage, e.g., 8-OH-G in DNA. In order to assess the roles of 8-OH-G in spontaneous and chemically-induced mutagenesis, the mutMST gene encoding 8-OH-G DNA glycosylase of Salmonella typhimurium was disrupted in several Ames tester strains, i.e., S. typhimurium TA1535 (hisG46, uvrB-, rfa), TA1975 (hisG46, uvr+, rfa) and TA102 (hisG428, uvr+, rfa). The spontaneous mutation frequencies were increased 2.4 and 1.6 times, respectively, by the mutMST deletions in strains TA1535 and TA1975, which are spontaneously reverted to His+ by mutations mainly at G:C base pairs. The resulting strains YG3001 (TA1535 delta mutMST) and YG3002 (TA1975 delta mutMST) were 2 to 8 times more sensitive to the mutagenicities of methylene blue plus visible light, neutral red plus visible light and 2-nitrofluorene than the parent strains. The strain YG3002 but not YG3001 was about 30 times more sensitive to the mutagenicity of 4-nitroquinoline N-oxide than the parent strain TA1975. Neither hydrogen peroxide nor phenazine methosulfate was mutagenic in the mutMST-deletion strains as well as in the parent strains. In contrast, the mutMST deletion did not affect the spontaneous mutation frequency of strain TA102, which has an A:T base pair at the critical site for reversion. The sensitivities of strain TA102 to the chemicals were not enhanced by the mutMST deletion except for hydrogen peroxide. These results suggest that 8-OH-G in DNA plays important roles in spontaneous mutagenesis occurring at G:C base pairs in S. typhimurium, and some nitro aromatics such as 4-nitroquinoline N-oxide or 2-nitrofluorene as well as the photosensitizers plus visible light can produce 8-OH-G in DNA, thereby inducing mutations. In the case of 4-nitroquinoline N-oxide, 8-OH-G rather than DNA adducts seems to play major roles in mutagenesis in uvr+ background. The new strains could be useful for the evaluation of the roles of 8-OH-G in mutagenesis in S. typhimurium and permit the efficient detection of some oxidative mutagens in the environment.

4-Nitroquinoline-1-oxide↗
PubMed1997

Evaluation of acrosomal status of bovine spermatozoa using concanavalin a lectin.

We report here that fluorescein isothiocyanate-conjugated concanavalin A (FITC-ConA) specifically labels the acrosomal region of acrosome-reacted bovine spermatozoa. This labeling is found to be useful in evaluating the acrosome status of bovine spermatozoa. When fresh bovine spermatozoa that had been fixed with 4% formaldehyde, smeared on glass slides and then air-dried were stained by FITC-ConA, weak fluorescence was observed on the acrosomal region, although almost all the spermatozoa appeared to be acrosome-intact. However, when fresh sperm suspensions were incubated with FITC-ConA and then mounted on glass slides, no fluorescence was observed on the acrosomal region. Therefore, in the ensuing experiments, both the fixation and the FITC-ConA staining of spermatozoa were done in suspension. When ethanol-treated spermatozoa, whose outer membrane may be permeabilized, were stained with FITC-ConA, the fluorescence was extensively observed on the inner acrosomal region. This fluorescence was inhibited in the presence of 0.2 M D-mannose, a competitive sugar, suggesting that FITC-ConA binds specifically to glycocomponents on the inner acrosomal membrane. We next tried to stain fresh or frozen-thawed spermatozoa from 3 different bulls that had been treated with the calcium ionophore A23187, which is known to induce acrosome reaction of bovine spermatozoa, with FITC-ConA. A significant correlation between the percentage of ConA-labeled spermatozoa and that of rose bengal stained negative ones at various time points during A23187 incubation was achieved. Furthermore, suitability of dual staining to distinguish between physiological acrosome reaction (acrosome-lost and live) and degenerative acrosomal loss (acrosome-lost and dead) using FITC-ConA and Hoechst bis-benzimide 33258 (H258) supravital stain was also confirmed. From these results, it was concluded that the FITC-ConA labeling procedure is a feasible and reliable method for the assessment of physiological acrosome reaction of bovine spermatozoa.

Journal Article↗
PubMed1997

Characterization of the gntT gene encoding a high-affinity gluconate permease in Escherichia coli.

We characterized the gntT gene encoding a high-affinity gluconate permease of Escherichia coli K-12. Primer extension and lacZ-operon fusion analyses revealed that gntT has one strong and two weak promoters, all of which are regulated positively by cAMP-CRP and negatively by GntR. The weak promoters became constitutive when separated from the upstream region including the strong promoter that overlaps a putative GntR-binding sequence. Gluconate-specific uptake activity was observed with cells harboring the gntT plasmid clone, which was enhanced by the presence of gntK encoding gluconate kinase.

Amino Acid Sequence↗