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T Ono

Publications and source records attributed to T Ono.

At least 559 records · Page 31Linked to original sources
PubMed1995

Molecular cloning and expression of rat squalene epoxidase.

Squalene epoxidase (SE) (EC 1.14.99.7) catalyzes the first oxygenation step in sterol biosynthesis and is suggested to be one of the rate-limiting enzymes in this pathway. Rat SE cDNA was isolated by selecting yeast transformants expressing rat cDNA in the presence of transformants expressing rat cDNA in the presence of terbinafine, an inhibitor specific for fungal SE. The expression of rat SE in the isolated terbinafine-resistant clone was confirmed by its survival in the presence of either terbinafine or an inhibitor specific for mammalian SE, NB-598, but not in the presence of both terbinafine and NB-598. Rat SE polypeptide deduced from the nucleotide sequence contains 573 amino acids, and its molecular weight is 63,950 Da. The amino acid sequence reveals one potential transmembrane domain, a hydrophobic segment (Leu27 to Tyr43) in the NH2-terminal region. This region also contains a beta 1-alpha A-beta 2 motif, which is the consensus sequence for an FAD binding domain, suggesting that SE is a flavoenzyme. This deduced rat SE sequence is 30.2% identical to the ERG 1 gene, which encodes SE from an allylamine-resistant Saccharomyces cerevisiae mutant. Expression of a full-length rat SE protein in Escherichia coli confirms this polypeptide as a functional SE. This is the first report of the molecular cloning of mammalian SE.

Amino Acid Sequence↗
PubMed1995

Repression of transient expression by DNA methylation in transcribed regions of reporter genes introduced into cultured human cells.

We developed a convenient method to methylate all CpG dinucleotides in both strands in a selected region of a plasmid, and investigated the effect of DNA methylation in the transcribed regions of reporter genes on the transient expression in HeLa cells. In a construct containing the chloramphenicol acetyltransferase (CAT) gene linked to the SV40 early promoter, methylation in the CAT structural gene repressed CAT activity. Methylation in the transcribed region of the Escherichia coli lacZ gene driven by the human cytomegalovirus immediate early promoter also inhibited expression of beta-galactosidase activity. These results suggest that methylation in the transcribed region as well as promoter methylation may affect transcription.

Chloramphenicol O-Acetyltransferase↗
PubMed1995

Immunohistochemical localization of two types of fatty acid-binding proteins in rat ovaries during postnatal development and in immature rat ovaries treated with gonadotropins.

BACKGROUND: The ovary of adult rats expresses two types of cytoplasmic fatty acid binding proteins (FABP), i.e., heart FABP (H-FABP) and intestinal 15 kDa protein (I-15P). We studied immunohistochemically the cellular localizations of these FABPs in the ovaries of rats at various postnatal ages and in the ovaries of immature (3-week-old) rats treated with pregnant mare serum gonadotropin (PMSG) and human chorionic gonadotropin (HCG). METHODS: The cryosections of ovaries were incubated with polyclonal antibodies against H-FABP and I-15P, and the immunoreactions were visualized at both light and electron microscopic levels. RESULTS: The immunoreactivity for H-FABP occurred temporarily in the follicular epithelial (granulosa) cells from 3 days to 2 weeks post partum, and then was localized exclusively to the theca/interstitial gland cells from 2 weeks to adulthood. In contrast, the immunoreactivity for I-15P appeared temporarily in a small subset of theca/interstitial gland cells from 2 to 3 weeks, disappeared at 4 weeks, and was localized exclusively to the corpus luteum cells after the onset of ovulation in the animal around 5 weeks. In the immature rat ovaries induced to ovulate by treatment with gonadotropins, I-15P-immunoreactive cells were first recognized in the luteinized granulosa layer of large preovulatory follicles, and increased in number progressively in the developing corpora lutea after the ovulation. CONCLUSIONS: Two types of FABPs are expressed in distinct steroid-producing cell types of rat ovary, and their expressions seem to be regulated in coincidence with the expressions of respective steroid hormones. These results suggest that FABPs play specific roles in the ovarian hormone synthesis.

Aging↗
PubMed1995

Recognition and neural plasticity responding to deficient nutrient intake scanned by a functional MRI in the brain of rats with L-lysine deficiency.

Each L-amino acid (AA) in plasma and brain remains unchanged while normal diet is available. Once L-lysine (Lys) deficient diet was offered to rats, Lys in plasma and brain declined, and anorexia occurred. When solutions of AAs were offered, they selected the Lys solution, and their food intake and growth normalized. The single neuron activity in the lateral hypothalamic area of these rats suggested that neural plasticity occurred, specifically responding to Lys, both by iontophoretic application and during ingestion of AA. The recognition site for deficient nutrient intake in the brain of rats with Lys deficiency was identified by non-invasive magnetic resonance imaging (MRI 4.7 tesla, 40 cm bore in diameter) developed to monitor changes in cerebral blood flow and oxygenation in rat brain. Wistar strain young male rats fed with Lys deficient diet for 4 days, were adapted to be settled in the center of the bore. When they received a Lys injection intraperitoneally (0.2 M, 10 mL/kg), a signal intensity decrease in the medial and lateral hypothalamus appeared 30 minutes later in T2 weighted images, reflecting increased oxygenation which lasted for 30 minutes, and then gradually recovered. These changes never occurred in any other areas of the brain of rats with Lys deficiency, i.e., the thalamus, the cortex, the hippocampus, etc. There were no changes in the signal intensity with control injection of saline. In addition, oxygen consumption in the brain of rats without Lys deficiency was not altered by intraperitoneal Lys injection. The present results suggest that in essential AA deficiency, the medial and lateral hypothalamus may play important roles in recognition responses to particular deficient nutrients in order to maintain homeostasis.

Amino Acids↗
PubMed1995

Effects of presence of an egg and calcium deposition in the shell gland on levels of messenger ribonucleic acid of CaBP-D28K and of vitamin D3 receptor in the shell gland of the laying hen.

Levels of calbindin-D28K (CaBP-D28K) mRNA and vitamin D3, 1 alpha, 25(OH)2D3, receptor (VDR) mRNA in the intestine and shell gland were measured in the laying hen during the ovulatory cycle by Northern and slot blot analyses. In addition, effects of presence of an egg and calcium deposition in the shell gland on the levels of both mRNAs were studied either by delay in oviposition (retaining the egg in the shell gland beyond the expected time of oviposition) or premature oviposition (emptying the shell gland and arrest of calcium deposition). During the ovulatory cycle mRNA levels of CaBP-D28K and of VDR remained relatively constant in the intestine. In contrast, both mRNA levels of the shell gland were low when there was an egg in the infundibulum, magnum, and isthmus, but significantly increased when there was an egg in the shell gland during shell formation. The im injection of indomethacin 3 hr before expected ovulation delayed oviposition but increased neither shell thickness nor mRNA levels of CaBP-D28K and of VDR. On the other hand, premature oviposition reduced mRNA levels of both CaBP-D28K and VDR in the shell gland. These results suggest that the presence of an egg and calcium deposition in the shell gland may be a stimulatory factor for synthesis and accumulation of CaBP-D28K mRNA and VDR mRNA in association with calcification.

Animals↗
PubMed1995

Biochemical and immunohistochemical analysis of cathepsins B, H, L and D in human melanocytic tumours.

We carried out biochemical and immunohistochemical analyses of cathepsins B, H, L and D in human melanocytic tumours using monospecific antibodies against rat cathepsins. In Western blot analysis, anti-rat cathepsin antibodies reacted with the cathepsins from normal human tissues and human malignant melanoma. However, the molecular profiles of the cathepsins from human melanoma were slightly different from those of the rat cathepsins, suggesting a distinct intracellular processing mechanism for cathepsins in human melanoma. Although cathepsins B, H, L and D were expressed in primary and metastatic melanomas and pigmented naevi immunohistochemically, the intensity of staining in metastatic melanomas was stronger than in primary melanomas and pigmented naevi. These findings suggest that anti-rat cathepsin antibodies may be useful in biochemical and/or immunohistochemical analysis of human melanocytic tumours.

Animals↗
PubMed1995

Clinical relevance of cathepsin B-like enzyme activity and cysteine proteinase inhibitor in melanocytic tumours.

We studied the relationship between cathepsin B (CB)-like enzyme activities and cysteine proteinase inhibitor (CPI) activities in lesions of human pigmented naevi (PN), primary melanomas (PM) and metastases/metastatic melanomas (MM). The CB-like enzyme activities in PM and MM were 2.5 and 6.8 times higher than in PN, respectively. The CB-like/CPI ratios in PM and MM were 1.9 and 11.6 times higher than in PN, respectively. CPI had the highest activity in PM and the lowest activity in MM. The disease-free interval of patients with a high CB-like enzyme activity (> or = 80 U/mg protein) and/or a high CB-like/CPI ratio (> or = 7) was shorter than that of patients with a low CB-like enzyme activity (< 80 U/mg protein) and/or a low CB-like/CPI ratio (< 7). Analysis of CB-like enzyme activity and/or the CB-like/CPI ratio may be useful in predicting the prognosis of human malignant melanoma.

Adult↗
PubMed1995

Spectra of superoxide-induced mutations in the lacI gene of a wild-type and a mutM strain of Escherichia coli K-12.

We have analyzed the spectra of superoxide-induced mutations in the chromosomal lacI gene of a wild-type and a mutM strain of Escherichia coli K-12. The mutM strain is known to be deficient in removing 8-hydroxyguanine from DNA. An intracellular superoxide-generating agent, menadione, was used to cause the mutation. Analysis of the mutated DNA showed marked differences between the mutants from the wild type and those from the mutM strain. In the mutants from the wild type, all possible base-pair substitutions were present and their proportions were similar to each other, whereas in those from the mutM bacteria there was a 90% bias in favor of transversion. Furthermore, in the mutM strain GC-to-CG transversion rather than GC-to-TA was predominantly induced. 64% of the GC-to-CG transversions in the mutM strain occurred at the site of (CT/GC)GGC (mutated base underlined). The favorable mutation site, CTGGC, was the same as that of the UV- and sunlight-induced mutations previously reported: the mutations observed there were also G-to-C transversions. We speculate from these results that the superoxide in the cells may lead to production of a modified guanine that can pair with guanine and is subject to removal by the MutM protein.

Base Sequence↗
PubMed1995

Effect of tauroursodeoxycholic acid on bile flow and calcium excretion in ischemia-reperfusion injury of rat livers.

BACKGROUND/AIMS: Tauroursodeoxycholic acid is known to have a hepatoprotective action in cholestatic disorders. We evaluated whether oral pretreatment with tauroursodeoxycholic acid could protect the liver from ischemia-reperfusion injury, with particular regard to its effect on bile flow and biliary calcium excretion. METHODS: A 1-hour in vivo ischemia-reperfusion model of 70% of the lobes of rat liver was used. Animals were divided into six groups (each group; n = 8); a non-ischemia sham group (CS), a control group without bile acids (CON), and 4 bile acid groups; 10 mg/kg and 50 mg/kg (U10, U50), taurocholic acid 10 mg/kg (CA10) and tauroursodeoxycholic acid 10 mg/kg (CD10). Bile acids were given orally for 7 days before operation. RESULTS: Three hours after reperfusion, oral bile acid pretreatment failed to reduce the hepatic ischemia-reperfusion injury biochemically, but histological improvement was observed in the tauroursodeoxycholic acid groups. After reperfusion, tauroursodeoxycholic acid significantly increased bile flow from the ischemic liver, and also significantly increased serum calcium concentration. Although tauroursodeoxycholic acid did not change biliary calcium concentration, it significantly enhanced total biliary calcium output during reperfusion. CONCLUSION: Thus, tauroursodeoxycholic acid inhibited tissue calcium accumulation and enhanced sinusoidal and biliary calcium output during hepatic ischemia-reperfusion. However, it is still unclear if calcium mobilization is part of the protective mechanisms of tauroursodeoxycholic acid in ischemia-reperfusion injury of the liver.

Analysis of Variance↗
PubMed1995

Purification and characterization of cytoplasmic dynein of rabbit liver.

Cytoplasmic dynein is a microtubule-dependent motor protein, which plays a role in intracellular transport. However, there have been few studies regarding the role of cytoplasmic dynein in the liver. Purification of cytoplasmic dynein from rabbit liver took advantage of the affinity of microtubule-dependent motor proteins for microtubules. Purified dynein contained heavy chain (450 kDa), intermediate chain (75 kDa), light chains (45-58 kDa) and dynactin (150 kDa). The subunit composition was consistent with previously reported data on brain cytoplasmic dynein. Microtubules prepared from bovine brain were driven by purified cytoplasmic dynein from rabbit liver, and movements of microtubules were visualized by video-enhanced differential interference contrast microscopy. The mean velocity of the motile microtubules was 1.09 +/- 0.13 microns/s. Our study provides evidence of rapid intracellular transport in hepatocytes controlled by cytoplasmic dynein.

Animals↗
PubMed1995

Dephosphorylation of abnormal sites of tau factor by protein phosphatases and its implication for Alzheimer's disease.

The abnormally phosphorylated forms of tau factor are major constituents of neurofibrillary tangles in Alzheimer's disease brain. In order to investigate protein phosphatases which are related to dephosphorylation of abnormal phosphorylation sites, we examined the dephosphorylation of tau factor phosphorylated by three proline-directed type protein kinases. Tau factor phosphorylated by cdc2 kinase and tau protein kinase II was dephosphorylated by the holoenzyme of protein phosphatase 2A and calcineurin, while either the catalytic subunit of protein phosphatase 2A or protein phosphatase 2C could not catalyze the dephosphorylation. From the kinetic analysis, we concluded that tau factors phosphorylated by the protein kinases serve as good substrates for protein phosphatase 2A and calcineurin. On the other hand, tau factor phosphorylated by glycogen synthase kinase 3 alpha was dephosphorylated by the catalytic subunit of protein phosphatases 2A as well as the holoenzyme of protein phosphatase 2A and calcineurin. It has been reported that serines 199, 202 and 396 according to the numbering of the longest human tau isoform are among the major abnormal phosphorylation sites of tau factor. We synthesized two phosphopeptides which contained phosphoserines 199 and 202 or phosphoserine 396 and prepared the polyclonal antibodies specific for the phosphopeptides. Using these antibodies, we confirmed that the holoenzyme of protein phosphatase 2A and calcineurin could dephosphorylate phosphoserines 199, 202 and 396 in tau factor. The catalytic subunit of protein phosphatase 2A could dephosphorylate phosphoserine 396 but not phosphoserines 199 and 202. Neurofibrillary tangles in Alzheimer's disease brain were immunostained with both antibodies but the normal neurons in the normal aged brains were not. The results suggest that protein phosphatase 2A and calcineurin can be involved in the dephosphorylation of abnormal phosphorylation sites in tau factor and that the dephosphorylation of phosphoserine 396 is differently regulated from phosphoserines 199 and 202.

Aged↗
PubMed1995

Amygdala role in conditioned associative learning.

Amygdala role in emotion was reviewed in reference to recent amygdala lesion studies and neuronal responses in the rat amygdala to conditioned stimuli. Extensive lesion studies suggest that the amygdala is crucial in various kinds of motivated and emotional behavior, and related autonomic responses. These amygdala functions critically depend on learning and memory. Amygdala lesions, both before and after training of conditioned associative learning, impaired emotional expression without simple sensory-motor deficits. Pharmacological experiments indicated neurotransmission in the amygdala is mediated through NMDA and AMPA receptors. These results strongly suggest the amygdala involvement in acquiring and storing associative memory (i.e. stimulus-affect association), by which animals recognize and evaluate the biological significance of a stimulus. This information is then transferred to the brainstem executing system. In the neurophysiological experiments, there were topographic distributions of sensory-responsive neurons within the amygdala, which were well correlated to anatomical data. The responses of rat amygdala neurons changed plastically during learning. Furthermore, more sensory-responsive neurons were encountered in the amygdala of rats trained to associate the sensory stimuli with a reinforcement than in the amygdala of rats that were not trained. In trained rats, multimodal neurons that responded to conditioned and unconditioned stimuli were frequently found in the basolateral and central nuclei of the amygdala. The results suggest that basolateral and central nuclei are foci where various sensory modalities converge, and which might perform critical functions in acquiring and storing long-term associative memory to link between sensory information and affective significance.

Amygdala↗
PubMed1995

Neuronal responsiveness to various sensory stimuli, and associative learning in the rat amygdala.

Neuronal activities were recorded from the amygdala and amygdalostriatal transition area of behaving rats during discrimination of conditioned auditory, visual, olfactory, and somatosensory stimuli associated with positive and/or negative reinforcements. Neurons were also tested with taste solution and various sensory stimuli that were not associated with reinforcement. Of the 1195 neurons tested, 475 responded to one or more sensory stimuli. Of these, 256 neurons responded exclusively to a unimodal sensory stimulus, 128 to multimodal sensory stimuli, and the remaining 91 could not be classified. Distribution of unimodal neurons was correlated with anatomical projections to the amygdala from sensory thalamus or sensory cortices. Multimodal neurons were located mainly in the basolateral and central nuclei of the amgydala. Response latencies of neurons in the basolateral nucleus were longer than those in other nuclei and neurons in the central nucleus had both short and long latencies. Neurons responsive to a given stimulus were more frequently encountered in the amygdalas of the trained rats than in those of the rats not trained to associate that stimulus with a reinforcement. Multimodal neurons that responded to conditioned and/or unconditioned stimuli used in the associative learned tasks were concentrated in the basolateral and central nuclei. The results indicate that some amygdalar neurons receive exclusive single sensory information, and the others receive information from two or more sensory inputs. Considering the long latencies and multimodal responsiveness, the basolateral and central nuclei of the amygdala might be foci where various kinds of sensory information converge. It is also suggested that the basolateral and central nuclei of the amygdala have critical roles in associative learning to relate sensory information to reinforcement or affective significance.

Acoustic Stimulation↗
PubMed1995

Generators of somatosensory evoked potentials investigated by dipole tracing in the monkey.

Generators of somatosensory evoked potentials, elicited by electrical stimulation of the median nerve in anaesthetized monkeys (Macaca fuscata), were investigated by submitting a three-dimensional reconstructed brain model to dipole tracing, which can equate surface potential distributions to an approximate corresponding equivalent dipole. The following components of the somatosensory evoked potentials were simultaneously recorded from 21-27 epidural electrodes: P7 (the letter indicates positive or negative polarity; the number indicates the approximate latency of the peak in ms) was recorded widely from various locations on both the left and right hemispheres, P10 was recorded near the anterior side of the central sulcus contralateral to the stimulation side, N10 was recorded near the posterior side of the contralateral central sulcus, P12 was recorded on both sides of the contralateral central sulcus, and P18 was recorded posterior to the contralateral central sulcus. Current source generators (dipoles) of each component of somatosensory evoked potentials were localized by dipole tracing: a dipole for P7 was located in the thalamus contralateral to the stimulation side; a dipole for P10 and N10 in the posterior wall of the contralateral central sulcus (area 3b); a dipole for P12 in the contralateral post central gyrus (areas 1 and 2); and a dipole for P18 in the anterior wall of the contralateral intraparietal sulcus (area 5). The locations and latencies of dipoles that generated cortical components of somatosensory evoked potentials, estimated by dipole tracing, were confirmed by direct cortical surface recording from a 16-25 electrode array placed directly on the cortical surface; and multiple unit recording from the anterior and posterior parietal cortices. After excision of area 5, P18 and N18 were abolished, whereas P10, N10, and P12 were not affected. The results suggest that dipoles for somatosensory evoked potentials progressed from the thalamus to area 5 via the primary somatosensory area. This progress is consistent with the hierarchical sequence of somatosensory information processing.

Anesthesia↗
PubMed1995

Physiological results of monkey brain ischemia, and protection by a calcium blocker.

Physiological and histological investigation was undertaken to examine dynamic and metabolic changes due to transient ischemic insult of the monkey brain with and without postischemic treatment by the calcium entry blocker, NC-1100 (1 mg/kg, IV). Monkeys were subjected to temporary occlusion of the eight major arteries: bilateral common carotid, internal and external carotid, and vertebral arteries. Blood flow was restored after 5-, 10-, 13-, and 15-min ischemia in different monkeys. The amplitudes of extradural, cortical, and hippocampal electroencephalograms decreased severely within 1-6 min after beginning occlusion. Complete recovery of these electroencephalograms required more than 1 h. During ischemia, significant change was obvious in arterial glucose, and systolic, diastolic, and mean blood pressure, all of which increased. There were no significant physiological differences between the untreated and NC-1100-treated groups, except decreased diastolic blood pressure and slightly lower postischemic heart rate in the treated group. These small differences might be accounted for by the effect of the calcium blocker. Ten to 15 minutes ischemia caused cell changes, including cell death, which were confined almost exclusively to the CA1 subfield of untreated hippocampi examined the fifth day after occlusion. However, no ischemia-induced cell change was observed in the CA1 subfield of hippocampi subjected to 10 to 15 min ischemia in the NC-1100-treated group. It was concluded that a calcium entry blocker can protect neurons from mild ischemia-induced injury and might ameliorate morphological damage and functional impairment of the brain due to ischemia in patients who suffer transient anoxic or hypoxic injury. The present physiological data should contribute to their clinical treatment.

Analysis of Variance↗
PubMed1995

Ischemic neuronal damage specific to monkey hippocampus: histological investigation.

We previously reported lesions confined specifically to the hippocampus when produced by occluding eight vessels (the bilateral vertebral, common, internal, and external carotid arteries), which supply blood to the brain. However, histopathological changes in the primate brain, caused by ischemic injury, have not previously been thoroughly investigated. In the present study, macaque monkeys were subjected to 5-18-min ischemia by occluding the eight vessels. After the brains were perfused and fixed 5 days after the occlusion, all regions were histologically investigated for ischemic cell changes. Ischemia for 5 min produced no ischemic cell change. Ischemia for 10-15 min produced cell death limited to the deeper portion of the pyramidal cell layer of the CA1 subfield in the hippocampus. In most monkeys, no cell death was observed in any brain region outside of the hippocampus after ischemia for up to 15 min. Ischemia for 18 min produced more widespread cell death in the CA1 subfield of the hippocampus, and cell death was no longer confined to the hippocampus, but was observed in layers III, V, and VI of the neocortices, the striatum, and some other regions. Brains that were perfused and fixed 1 year after 15-min ischemic insult revealed no ischemic cell morphological change in any region, but the number of pyramidal cells in the CA1 subfield was decreased to about half. The results indicate that the CA1 subfield of the monkey hippocampus is the precise region of the brain most susceptible to ischemic insult in the primate forebrain, and after a critical time (15-min ischemia in this procedure) ischemic cell changes occur suddenly and extensively. Ischemia due to occlusion of eight arteries for 10-15 min could produce a model of human amnesia caused by transient ischemic insult.

Amnesia↗
PubMed1995

Spontaneous mutant frequency of lacZ gene in spleen of transgenic mouse increases with age.

Spontaneous mutant frequency of lacZ gene in spleen of transgenic MutaTM mouse was examined at different ages. It was (3.2 +/- 1.3 (SD)) x 10(-5) at newborn and increased almost linearly with age up to (8.3 +/- 1.8) x 10(-5) at one year. Since the mutation of the gene is not likely to be subject to selection in vivo, the data support the idea that spontaneous mutation takes place throughout aging process and accumulates with age if not selected out by cell death.

Aging↗