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T Tsujimoto

Publications and source records attributed to T Tsujimoto.

At least 37 records · Page 2Linked to original sources
PubMed2000

Assessment of efficiency and safety of adenovirus mediated gene transfer into normal and damaged murine livers.

BACKGROUND: When recombinant adenoviruses are infused directly into the circulation, transgene expression is almost completely restricted to the liver. AIMS: Efficiency and safety of adenovirus mediated gene transfer into damaged livers were examined in mice with liver cirrhosis or fulminant hepatitis. METHODS: Liver cirrhosis and fulminant hepatitis were induced by intraperitoneal administration of thioacetamide and D-galactosamine followed by lipopolysaccharide, respectively. Mice were infused with adenoviruses carrying the Escherichia coli beta-galactosidase gene, lacZ gene, into the tail vein. Transduction efficiency of the lacZ gene was estimated histochemically by X-gal staining and quantitatively using a chemiluminescent assay. Activation of adenovirus specific T cells and development of neutralising antibodies against adenovirus were also examined. RESULTS: Histochemical evaluation revealed that approximately 40%, 80%, and 40% of cells in normal, cirrhotic, and fulminant hepatitis livers, respectively, were stained blue using X-gal staining. Quantitative analyses revealed that levels of lacZ expression in cirrhotic livers were approximately 2.5-fold and sixfold greater than those in normal and fulminant hepatitis livers, respectively. Although transgene expression in fulminant hepatitis livers was significantly lower than that in normal livers, marked levels of transgene expression were achieved even in fulminant hepatitis livers. Significant adverse effects of adenoviruses were not observed in damaged livers. There were no significant differences in cellular or humoral immune responses to adenoviruses among animals with normal, cirrhotic, and fulminant hepatitis livers. CONCLUSIONS: Our results suggest that gene therapy with adenoviruses may be used efficiently and safely, even in patients with severe liver disease.

Adenoviridae↗
PubMed2000

Electrochemotherapy can eradicate established colorectal carcinoma and leaves a systemic protective memory in mice.

Mice bearing subcutaneously established colorectal carcinoma (CRC) were given intratumoral, intravenous or intraperitoneal injection of various doses of bleomycin (BLM), followed by the delivery of direct current, square wave electric pulses to the tumor. Approximately 50% of animals treated with electrochemotherapy with BLM had completely eradicated established CRC tumors. Importantly, it was shown that CRC-specific cytotoxic T lymphocytes were elicited in the spleens of cured animals, resulting in the protection of the rechallenge with CRC. These results indicate that electrochemotherapy with BLM is promising for the treatment of metastatic CRC as well as the original lesion.

Animals↗
PubMed1999

Impairment of inhibitory synaptic transmission in mice lacking synapsin I.

Deletion of the synapsin I genes, encoding one of the major groups of proteins on synaptic vesicles, in mice causes late onset epileptic seizures and enhanced experimental temporal lobe epilepsy. However, mice lacking synapsin I maintain normal excitatory synaptic transmission and modulation but for an enhancement of paired-pulse facilitation. To elucidate the cellular basis for epilepsy in mutants, we examined whether the inhibitory synapses in the hippocampus from mutant mice are intact by electrophysiological and morphological means. In the cultured hippocampal synapses from mutant mice, repeated application of a hypertonic solution significantly suppressed the subsequent transmitter release, associated with an accelerated vesicle replenishing time at the inhibitory synapses, compared with the excitatory synapses. In the mutants, morphologically identifiable synaptic vesicles failed to accumulate after application of a hypertonic solution at the inhibitory preterminals but not at the excitatory preterminals. In the CA3 pyramidal cells in hippocampal slices from mutant mice, inhibitory postsynaptic currents evoked by direct electrical stimulation of the interneuron in the striatum oriens were characterized by reduced quantal content compared with those in wild type. We conclude that synapsin I contributes to the anchoring of synaptic vesicles, thereby minimizing transmitter depletion at the inhibitory synapses. This may explain, at least in part, the epileptic seizures occurring in the synapsin I mutant mice.

Animals↗
PubMed1999

Activation of the insular and opercular regions of the monkey by drinking as revealed by positron emission tomography.

We measured the regional cerebral blood flow (rCBF) of the monkey using positron emission tomography (PET) in the following conditions: self-initiated (SELF), visually-initiated (VISUAL), and color-discriminating go/no-go (GONOGO) hand movement tasks with reward of water, free reward condition (REWARD), and resting states (REST). The insulae, frontal opercula, and mouth sensorimotor area were activated in any of SELF, VISUAL, GONOGO, and REWARD, when compared with REST. These findings suggest that the insular and opercular regions play an important role in the neural processes involved in drinking.

Animals↗
PubMed1999

Hepatocellular carcinoma in an orthotopic mouse model metastasizes intrahepatically in cirrhotic but not in normal liver.

Prognosis of hepatocellular carcinoma (HCC) still remains poor mainly because of intrahepatic metastasis. In the majority of cases, HCC is found in conjunction with liver cirrhosis. It is, therefore, of great importance to investigate the invasive and metastatic behavior of HCC in cirrhotic liver. To examine this, a liver cirrhosis model was produced by injecting thioacetamide i.p. into mice. Murine HCC cells were labeled with the fluorescent carbocyanine dye, DiI, and implanted directly under the capsule of cirrhotic and normal livers of syngeneic mice. DiI-labeled HCC cells in the liver were observed under fluorescent and confocal microscopy. Histological analysis of cirrhotic and normal livers revealed that implanted HCC cells migrated to and invaded the adjacent periportal regions, but not the adjacent centrolobular areas. This characteristic behavior of HCC was more evident in cirrhotic liver than in normal liver. Furthermore, intrahepatic metastasis to unimplanted hepatic lobes was observed in cirrhotic liver as early as 7 days after implantation, while it was not detected in normal liver even 4 weeks later. Thus, an orthotopic animal model for HCC with cirrhosis described here may be suitable for investigating the invasive and metastatic behavior of HCC. Importantly, labeling tumor cells with a fluorescent dye before orthotopic implantation may be a convenient and useful method to investigate the invasive and metastatic behavior of various types of cancer.

Animals↗
PubMed1999

Transient cyclophosphamide treatment before intraportal readministration of an adenoviral vector can induce re-expression of the original gene construct in rat liver.

Although adenovirus is an attractive vehicle for transferring therapeutic genes in vivo, animal studies have indicated that the clinical usefulness of adenoviruses may be limited by their immunogenicity. Although immunosuppressive strategies around the time of initial exposure of adenoviruses have been shown to prevent the formation of neutralizing antibodies and permit the successful readministration of adenoviruses in animals, the practicality of the approaches remains questionable. Because the majority of prospective gene therapy patients have already been infected with wild-type adenoviruses, initial treatment with adenoviruses in humans may correspond to readministration of adenoviruses into animals. It is shown here that although intraportal infusion of adenoviruses carrying a reporter lacZ gene resulted in transient high levels of transgene expression in the rat liver, intraportal readministration of adenoviruses failed to induce detectable levels of transgene expression. Conversely, when animals were treated transiently with cyclophosphamide before the intraportal readministration of adenoviruses, development of neutralizing antibodies and antigen-specific T cell proliferation in response to adenoviral readministration was significantly suppressed and successful re-expression of the transgene was achievable. These results may have important implications for efficacy considerations when adenoviral vectors are employed in clinical settings.

Adenoviridae↗
PubMed1999

Targeted disruption of a melanin biosynthesis gene affects conidial development and UV tolerance in the Japanese pear pathotype of Alternaria alternata.

Structural analysis of the BRM2 gene involved in melanin biosynthesis of the Japanese pear pathotype of Alternaria alternata suggested that this gene encodes 1,3,8-trihydroxynaphthalene reductase. Targeted disruption of the BRM2 gene did not affect pathogenicity, vegetative growth, or the number of conidia produced. Targeted disruption, however, did reduce conidial size and septal number, suggesting that melanin is associated with conidial development. The conidia of brm2 mutant transformants were more sensitive to UV light than those of the wild type, demonstrating that melanin confers UV tolerance.

Alternaria↗
PubMed1998

Activation of the ventral and mesial frontal cortex of the monkey by self-initiated movement tasks as revealed by positron emission tomography.

In order to investigate the neural mechanisms of movement initiation, we measured the regional cerebral blood flow (rCBF) of the monkey during self-initiated and visually-initiated hand movement tasks using positron emission tomography (PET). The orbitofrontal, cingulate, and anteromedial part of the dorsal premotor areas were preferentially activated by the self-initiated hand movement task (SELF). The pre-supplementary motor area and the cingulate motor area were also included in the active foci during the task. In the visually-initiated task (VISUAL), the V1, V2, V3, V3A, and V4 were activated, whereas the activity of the dorsolateral premotor and primary motor areas was not significantly different between the two tasks. These findings suggest that the orbitofrontal and mesial frontal cortices play an important role in the neural processes involved in self-initiation of movement and self-regulation of inner drives.

Animals↗
PubMed1998

Facilitation of the presynaptic calcium current at an auditory synapse in rat brainstem.

1. The presynaptic calcium current (IpCa) was recorded from the calyx of Held in rat brainstem slices using the whole-cell patch clamp technique. 2. Tetanic activation of IpCa by 1 ms depolarizing voltage steps markedly enhanced the amplitude of IpCa. Using a paired pulse protocol, the second (test) response was facilitated with inter-pulse intervals of less than 100 ms. The facilitation was greater at shorter intervals and was maximal (about 20%) at intervals of 5-10 ms. 3. When the test pulse duration was extended, the facilitation was revealed as an increased rate of IpCa activation. From the current-voltage relationship measured at 1 ms from onset, facilitation could be described by a shift in the half-activation voltage of about -4 mV. 4. IpCa facilitation was not attenuated when guanosine-5'-O-(3-thiotriphosphate) (GTPgammaS) or guanosine-5'-O-(2-thiodiphosphate) (GDPbetaS) was included in the patch pipette, suggesting that G-proteins are not involved in this phenomenon. 5. On reducing [Ca2+]o, the magnitude of facilitation diminished proportionally to the amplitude of IpCa. Replacement of [Ca2+]o by Ba2+ or Na+, or buffering of [Ca2+]i with EGTA or BAPTA attenuated IpCa facilitation. 6. We conclude that repetitive presynaptic activity can facilitate the presynaptic Ca2+ current through a Ca2+-dependent mechanism. This mechanism would be complementary to the action of residual Ca2+ on the exocytotic machinery in producing activity-dependent facilitation of synaptic responses.

Animals↗
PubMed1998

G-Protein-coupled modulation of presynaptic calcium currents and transmitter release by a GABAB receptor.

Presynaptic GABAB receptors play a regulatory role in central synaptic transmission. To elucidate their underlying mechanism of action, we have made whole-cell recordings of calcium and potassium currents from a giant presynaptic terminal, the calyx of Held, and EPSCs from its postsynaptic target in the medial nucleus of the trapezoid body of rat brainstem slices. The GABAB receptor agonist baclofen suppressed EPSCs and presynaptic calcium currents but had no effect on voltage-dependent potassium currents. The calcium current-EPSC relationship measured during baclofen application was similar to that observed on reducing [Ca2+]o, suggesting that the presynaptic inhibition generated by baclofen is caused largely by the suppression of presynaptic calcium influx. Presynaptic loading of the GDP analog guanosine-5'-O-(2-thiodiphosphate) (GDPbetaS) abolished the effect of baclofen on both presynaptic calcium currents and EPSCs. The nonhydrolyzable GTP analog guanosine 5'-O-(3-thiotriphosphate) (GTPgammaS) suppressed presynaptic calcium currents and occluded the effect of baclofen on presynaptic calcium currents and EPSCs. Photoactivation of GTPgammaS induced an inward rectifying potassium current at the calyx of Held, whereas baclofen had no such effect. We conclude that presynaptic GABAB receptors suppress transmitter release through G-protein-coupled inhibition of calcium currents.

Animals↗
PubMed1998

Significance of increased plasma adrenomedullin concentration in patients with cirrhosis.

BACKGROUND/AIMS: Adrenomedullin recently discovered in human pheochromocytoma is a potent vasodilatory peptide mainly derived from vascular endothelial and smooth muscle cells. Hyperdynamic circulation, ultimately leading to ascites formation, has been attributed to peripheral vasodilatation in liver cirrhosis. However, little is known about the role of adrenomedullin in this condition. METHODS: Plasma adrenomedullin concentrations were measured by radioimmunoassay after extraction and purification in 28 cirrhotic patients without ascites, 12 cirrhotic patients with ascites and 10 healthy subjects. RESULTS: Plasma adrenomedullin concentrations in cirrhotic patients with ascites (12.7+/-4.5 fmol/ml) were significantly higher than those in cirrhotic patients without ascites (8.2+/-2.3 fmol/ml, p<0.005) and healthy subjects (5.8+/-0.8 fmol/ml, p<0.005). Interestingly, plasma adrenomedullin concentrations were highest in patients with refractory ascites (n=5, 15.8+/-3.0 fmol/ml) and were positively correlated with the Child-Pugh score (r=0.44, p<0.01). Moreover, plasma adrenomedullin concentrations were positively correlated with plasma renin activity (r=0.63, p<0.0001), plasma aldosterone (r=0.60, p<0.0001) and plasma norepinephrine concentrations (r=0.60, p<0.0001), and negatively correlated with creatinine clearance (r=-0.61, p<0.0005) and urinary sodium excretion (r=-0.44, p<0.02). Stepwise multiple regression analysis using certain independent variables, including Pugh's score, vasoactive substances, renal function and hemodynamic parameters, showed that the adjusted R square was highest when plasma renin activity and creatinine clearance (standard coefficient=0.53, -0.49, respectively) were considered (adjusted R square=0.61, p<0.0001). CONCLUSIONS: Plasma adrenomedullin concentrations increased with the progression of liver cirrhosis and were highest in cirrhotic patients with refractory ascites. In addition, elevated adrenomedullin was associated with activation of the renin-angiotensin-aldosterone and sympathetic nervous systems, and with functional renal impairment in cirrhosis. Considering the potent vasodilatory action of adrenomedullin, increased adrenomedullin may participate in the hyperdynamic circulation, ultimately leading to ascites formation, in patients with liver cirrhosis.

Adrenomedullin↗
PubMed1998

Inactivation of presynaptic calcium current contributes to synaptic depression at a fast central synapse.

Voltage-gated calcium channels are well characterized at neuronal somata but less thoroughly understood at the presynaptic terminal where they trigger transmitter release. In order to elucidate how the intrinsic properties of presynaptic calcium channels influence synaptic function, we have made direct recordings of the presynaptic calcium current (I(pCa)) in a brainstem giant synapse called the calyx of Held. The current was pharmacologically classified as P-type and exhibited marked inactivation. The inactivation was largely dependent upon the inward calcium current magnitude rather than the membrane potential, displayed little selectivity between divalent charge carriers (Ca2+, Ba2+ and Sr+), and exhibited slow recovery. Simultaneous pre- and postsynaptic whole-cell recording revealed that I(pCa) inactivation predominantly contributes to posttetanic depression of EPSCs. Thus, because of its slow recovery, I(pCa) inactivation underlies this short-term synaptic plasticity.

Animals↗
PubMed1997

Activation of the prefrontal, occipital and parietal cortices during go/no-go discrimination tasks in the monkey as revealed by positron emission tomography.

The regions of the monkey brain involved in cognito-behavioral tasks were mapped using the H2(15)O positron emission tomography (PET) activation technique. Monkeys performed hand movements in an asymmetrically rewarded go/no-go reaction-time task with discrimination between green (go) and red (no-go) light signals. Regional cerebral blood flow (rCBF) was measured during the go/no-go task and a control task in which only go signals were presented. In the go/no-go task, when compared with the control, a significant increase in rCBF was noted in the following regions: (1) the principal sulci; (2) the anterodorsal frontal pole; (3) the anterior part of the inferior occipital sulcus which appeared to be the V4; and (4) the parieto-occipital region. The increase in the principal sulci may be related to the no-go decision and motor suppression in the area, as interpreted in our previous electrophysiological studies. The present results appear to support electrophysiological observations on visual perception and motor suppression and reflect a mode of their functional integration.

Animals↗
PubMed1997

Circular DNA plasmid in the phytopathogenic fungus Alternaria alternata: its temperature-dependent curing and association with pathogenicity.

We found the presence of plasmid DNA in strain T88-56 of the Japanese pear pathotype of Alternaria alternata, which causes black spot of certain cultivars of Japanese pear by producing host-specific AK-toxin. The plasmid, designated pAAT56, was identified to be an approximately 5.4-kilobase (kb) circular molecule by electron microscopic observation and restriction endonuclease mapping. Southern blot analysis showed that pAAT56 DNA had no homology with either nuclear or mitochondrial DNA. Cultures of strain T88-56 grown at 26 degrees showed markedly reduced plasmid levels relative to those grown at lower temperatures. The strain was completely cured of pAAT56 during growth at 29 degrees. Temperature-dependent curing of pAAT56 was confirmed by using single-protoplast isolates from mycelia grown at 23 degrees, most of which maintained the plasmid, and from mycelia grown at 29 degrees, most of which had lost the plasmid. Northern blot analysis detected the presence of three RNA species (approximately 1.7, 2.7 and 5.4 kb) transcribed from pAAT56. The biological function of pAAT56 was observed using single-protoplast isolates from mycelia that either contained or had been cured of pAAT56. The plasmid-containing isolates tended to be reduced in AK-toxin production and pathogenicity compared with the plasmid-cured isolates.

Alternaria↗
PubMed1997

Chemical restraint of African lions (Panthera leo) with medetomidine-ketamine.

Effects of a combination of medetomidine-ketamine as a chemical restraint and antagonistic effects of atipamezole on this combination were investigated in 5 lions. The medetomidine (47.6-58.4 micrograms/kg) and ketamine (1.9-5.7 mg/kg) combination provided complete immobilization with good analgesia and muscle relaxation in 4 lions, while one lioness was poorly sedated by medetomidine, and additional injections of medetomidine and ketamine were required. The duration of anesthesia seemed to be much longer than one hour in 4 of the lions. Atipamezole, at four times the preceding dose of medetomidine, provided a smooth recovery and animals were able to stand up 17-61 min after its injection. Side effects were limited to vomiting after walking in 3 of 5 lions.

Anesthetics, Dissociative↗
PubMed1996

Presynaptic calcium current modulation by a metabotropic glutamate receptor.

Metabotropic glutamate receptors (mGluRs) regulate transmitter release at mammalian central synapses. However, because of the difficulty of recording from mammalian presynaptic terminals, the mechanism underlying mGluR-mediated presynaptic inhibition is not known. Here, simultaneous recordings from a giant presynaptic terminal, the calyx of Held, and its postsynaptic target in the medial nucleus of the trapezoid body were obtained in rat brainstem slices. Agonists of mGluRs suppressed a high voltage-activated P/Q-type calcium conductance in the presynaptic terminal, thereby inhibiting transmitter release at this glutamatergic synapse. Because several forms of presynaptic modulation and plasticity are mediated by mGluRs, this identification of a target ion channel is a first step toward elucidation of their molecular mechanism.

Aminobutyrates↗