Biomedical subjects

Y Ushio

Publications and source records attributed to Y Ushio.

At least 199 records · Page 11Linked to original sources

PubMed1994

A novel isoform of the neurofibromatosis type-1 mRNA and a switch of isoforms during murine cell differentiation and proliferation.

Four types of cDNAs encoding the GTPase-activating protein-related domain (GRD) of the mouse neurofibromatosis type-1 gene (NF1) have been cloned. One of these isoforms was a newly identified form termed type IV. Analysis of the genomic structure of the mouse NF1-GRD revealed two exons (23A and 23B) between exons 23 and 24, leading to the production of four types of NF1-GRD cDNAs by an alternative splicing mechanism. Amino-acid sequences encoded by NF1-GRD are highly conserved between human and mouse. Analysis of the expression of these transcripts in various tissues of adult mouse revealed that the type-I transcript is predominantly expressed in neural tissues such as brain and spinal cord. Other forms, termed types II, III and IV, are also expressed in various tissues. The type-I and type-II transcripts are expressed equivalently in undifferentiated P19 mouse teratocarcinoma cells, whereas type-I expression becomes predominant during neuronal differentiation by retinoic acid treatment. Expression of type I is also shown to be correlated with cessation of cell proliferation in P19 cells, but not in NIH3T3 cells. These, together with other results, suggest that the four types of NF1-GRD transcripts generated by alternative splicing have some important biological roles in cell differentiation and proliferation.

Alternative Splicing↗

PubMed1994

Increased expression of growth-associated protein GAP-43/B-50 following cerebral hemitransection or striatal ischemic injury in the substantia nigra of adult rats.

The substantia nigra receives massive inhibitory gamma-aminobutyric acid (GABA)-ergic inputs, deafferentation of which is supposed to lead to anterograde transsynaptic regression of the nigral neurons. An immunohistochemical technique was used to examine growth-associated protein GAP-43 expression following cerebral hemitransection or transient middle cerebral artery occlusion (MCAO) in the substantia nigra of adult rats. GAP-43 expression was transiently elevated in the substantia nigra pars reticulata (SNr) neurons, but not in the pars compacta neurons at 3 and 4 days post-hemitransection. Massive striatal ischemic injury produced by transient MCAO also caused an increase in GAP-43 synthesis in the SNr neurons at 3 and 4 days after operation. The present findings raise the possibility that deafferentation of the GABAergic inputs leading to transneuronal regression of the SNr neurons is responsible for the elevated expression of GAP-43 in the substantia nigra of adult rats.

Animals↗

PubMed1994

Neostriatal mosaic and type II Ca2+/calmodulin-dependent protein kinase: an immunohistochemical study on the adult rat striatum.

The present immunohistochemical study is concerned with the expression of type II Ca2+/calmodulin-dependent protein kinase (CaM-kinase II), which is supposed to play an essential role in the intracellular Ca2+ signal transduction, in the striatum of adult rats. CaM-kinase II immunoreactivity was differentially concentrated in irregularly shaped compartments within the nucleus in a mosaic-like fashion. The compartment of heightened CaM-kinase II-immunolabeling corresponded to the extrastriosomal matrix visualized by calbindin-D28k-immunostaining. Light microscopic observation showed neurons immunoreactive for CaM-kinase II to be less densely distributed in the striosomes than in the matrix compartment. The present data suggest that these two striatal compartments may differ in an intracellular Ca(2+)-signaling process associated with protein phosphorylation.

Animals↗

PubMed1994

Activation of Ca2+/calmodulin-dependent protein kinase II and phosphorylation of intermediate filament proteins by stimulation of glutamate receptors in cultured rat cortical astrocytes.

We investigated the activation of Ca2+/calmodulin-dependent protein kinase II (CaM kinase II) via stimulation of glutamate receptors and subsequent phosphorylation of vimentin and glial fibrillary acidic protein (GFAP) in cultured rat cortical astrocytes. The indirect immunofluorescence analysis with the anti-CaM kinase II antibody revealed that the enzyme was detected diffusely in the cytoplasm and more intensely in the nucleus. Glutamate elevated the Ca(2+)-independent activity of CaM kinase II through autophosphorylation, and this response was blocked by both DL-2-amino-3-phosphonopropionate and 6-cyano-7-nitroquinoxaline-2,3-dione, but not by D-2-amino-5-phosphonovalerate. In the experiments using 32P-labeled astrocytes, the phosphorylation of vimentin and GFAP as well as autophosphorylation of CaM kinase II were found to be stimulated after the exposure to glutamate. It was concluded by two-dimensional phosphopeptide analysis that the increased phosphorylation of vimentin and GFAP observed in intact cells were due to the activation of CaM kinase II by glutamate. These results suggest that glutamate can activate CaM kinase II through stimulation of both the metabotropic and non-N-methyl-D-aspartate receptors, and that the concomitant phosphorylation of vimentin and GFAP may in turn regulate the functions of intermediate filament proteins in intact astrocytes.

6-Cyano-7-nitroquinoxaline-2,3-dione↗

PubMed1994

Disruption of the blood-cerebrospinal fluid barrier by transient cerebral ischemia.

The influence of transient cerebral ischemia on blood-brain and blood-cerebrospinal fluid (CSF) barrier permeability was studied sequentially by magnetic resonance imaging (MRI) contrast enhancement using gadolinium-diethylene triamine pentaacetic acid (Gd-DTPA) in rats. The unilateral internal carotid and middle cerebral arteries were transiently occluded by inserting a nylon thread into the carotid artery and removing it following a variable interval of 5 to 60 min. Contrast enhancement of the lateral ventricle on the affected side was seen in the enhanced T1-weighted image at the early stage of reperfusion 6 h after the start of ischemia in most of the rats subjected to 30- and 60-min ischemia, and in 3 of 6 rats in the 15-min ischemia group. Autoradiograms of Gd-[14C]DTPA in rats subjected to 60-min ischemia demonstrated that the tracer strongly accumulated in the choroid plexus, the wall of the lateral ventricle and its surrounding brain tissue. On the other hand, parenchymal enhancement of the striatum was seen only in the 60-min ischemia group and appeared later on Day 1 or Day 7. These results indicate that ventricular enhancement on MRI in this model is caused by disruption of the blood-CSF barrier at the choroid plexus of the lateral ventricle. This is the first reported study to demonstrate blood-CSF barrier disruption by transient ischemia.

Animals↗

PubMed1994

Germ-line and somatic mutations of the APC gene in patients with Turcot syndrome and analysis of APC mutations in brain tumors.

The Turcot syndrome (TS) is a rare, probably autosomal recessive, disorder characterized by development of primary neuroepithelial tumors of the central nervous system (CNS) and numerous adenomatous colorectal polyps. To examine the possible involvement of mutations of the APC gene, which is responsible for familial adenomatous polyposis (FAP), in Turcot syndrome, we examined DNAs from TS patients for alterations in this gene by means of ribonuclease protection analysis. Germ-line APC mutations were detected in each of three unrelated cases of TS, and additional (somatic) mutations were observed in colonic adenomas that had developed in one of these patients. However, no somatic mutations in APC were found among 91 neuroepithelial tumors (medulloblastoma, glioblastoma, astrocytoma, and oligodendroglioma), whether sporadic or associated with TS. These results suggest that the APC gene is associated with pathogenesis of one feature of TS, but that at least one other gene is responsible for the genesis of neuroepithelial tumors in the CNS.

Adenomatous Polyposis Coli↗

PubMed1994

Elevated immunoreactivity for glutamic acid decarboxylase in the rat cerebral cortex following transient middle cerebral artery occlusion.

We investigated the expression of glutamic acid decarboxylase (molecular weight 67,000; GAD67) immunohistochemically in the rat cerebral cortex following transient middle cerebral artery occlusion (MCAO) capable of producing slowly progressive neuronal damage. An increase in GAD67 immunoreactivity was observed in the cerebral cortex ipsilateral to the ischemic insult, most prominent in lamina IV, 3 to 14 days after MCAO. At this stage, light microscopy showed GAD67-positive puncta to be larger and more strongly immunoreactive in the ipsilateral cortex than those in the contralateral side. The elevated expression of GAD67 in the insulted cortex may reflect part of the adaptive functional changes in GABA transmission with slowly progressive cortical ischemic damage.

Animals↗

PubMed1994

Subdivisional ischemic injury of the unilateral striatum causes apomorphine-induced rotational behavior in rats.

Behavioral and histological studies were performed on a reversible ischemia model in rats. At 60 days after unilateral transient middle cerebral artery occlusion for 30 min, the operated rats exhibited the ipsiversive rotational behavior elicited by systemic administration of dopamine receptor agonist apomorphine in a dose-dependent manner. Histologically, the ipsilateral striatum of the rats showed a subdivisional ischemic injury, while the nigral dopaminergic neurons appeared intact. The striatal lesions having a cell type-specific injury were located in the dorsolateral portion of the rostral striatum and in the lateral portion of the caudal part of the nucleus. Thus, the transient cerebral ischemia could successfully produce selective damage of a striatal subdivision, which causes an abnormality in motor controls in response to dopamine receptor stimulation. The present data may provide a part of functional and anatomical basis for understanding the movement disorders associated with basal ganglia dysfunction (e.g., parkinsonism), which may occur in patients with cerebrovascular disorders.

Animals↗

PubMed1994

In vivo induction of the growth associated protein GAP43/B-50 in rat astrocytes following transient middle cerebral artery occlusion.

Immunohistochemistry was used to investigate the induction of growth-associated protein GAP43/B-50 in the astrocytes of rat cerebrum in vivo following ischemic injury produced by 30 min of transient middle cerebral artery occlusion. Three days after operation, GAP43 immunoreactivity first appeared in some astrocytic populations surrounding the infarcted lesion. Induction of GAP43 in those astrocytes persisted for up to 14 days and disappeared at 30 days postoperation. Double-immunofluorescence staining confirmed that the GAP43-immunoreactive astrocytes examined were all positive for glial fibrillary acidic protein. Our present data suggest that certain astrocytes could be induced to synthesize GAP43 in vivo in response to an ischemic insult in adult rats.

Animals↗

PubMed1994

Changes of monosaccharide availability of human hybridoma lead to alteration of biological properties of human monoclonal antibody.

The effect of glucose and other monosaccharide availability in culture medium on production of antibody by human hybridomas has been studied. Human hybridoma cells C5TN produce an anti lung cancer human monoclonal antibody, and the light chain is N-glycosylated at the variable region. When the cell line was grown in the presence of various concentrations of glucose, the antibodies produced changed their antigen-binding activities. Analysis of the light chains produced under these condition revealed that four molecular-mass variant light chains ranging from about 26 to 32 kDa were secreted. The twenty six-kDa species, which corresponds to a non-glycosylated form of the light chain, was recovered after enzymatic removal of all N-linked carbohydrate chains, indicating that the source of the heterogeneity of the light chain is due to the varied glycosylation. When the C5TN cells were cultured in medium containing either fructose, mannose or galactose instead of glucose, galactose elevated the antigen binding activity of the antibody more than the other sugars. These results suggest that change of glucose availability affects the antigen-binding activity of the antibody via the alteration of the glycosylation.

Antibodies, Monoclonal↗

PubMed1994

Neurotoxicity and pharmacokinetics of ventriculolumbar perfusion of methyl 6-[3-(2-chloroethyl)-3-nitrosoureido]-6-deoxy-alpha-D-glucopyranoside (MCNU) in dogs.

Ventriculolumbar perfusion of methyl 6-[3-(2-chloroethyl)-3-nitrosoureido]-6-deoxy-alpha-D-glucopyranoside (MCNU), a water soluble nitrosourea with log P -0.71, may be efficacious in the treatment of subarachnoid dissemination of malignant glioma. We used 2 dogs to study the neurotoxicity and pharmacokinetics of MCNU. MCNU (1 mg), dissolved in 10 ml of artificial CSF, was administered via the right lateral ventricle during a period of 18 to 42 min and the CSF was drained by lumbar puncture. The perfusion was repeated once a week for 10 consecutive weeks. No neurological and systemic symptoms were noted after perfusion. Histological examination of the brain and spinal cord showed local denudation of the ependyma and local subependymal spongy degeneration and gliosis in the lateral ventricle into which MCNU was administered in one dog and local denudation of the ependyma in the other. When administration was over a period of 21 to 38 min, the MCNU concentration in the lumbar CSF peaked at 11.11 to 50.67 micrograms/ml, in 28 to 78 min. The area under the drug concentration-time curve (AUC) was 1152 micrograms x min/ml on average, significantly larger than that of ACNU. The elimination phase followed linear kinetics and the half-time was 41.1 min on average, significantly longer than that of ACNU. These findings suggest that ventriculolumbar perfusion of MCNU may be effective in the treatment of subarachnoid dissemination of malignant glioma notwithstanding some local histological changes.

Animals↗

PubMed1994

Pharmacokinetics of intrathecal 1-(4-amino-2-methyl-5-pyrimidinyl) methyl-3-(2-chloroethyl)-3-nitrosourea hydrochloride in rats.

The pharmacokinetics of intrathecal 1-(4-amino-2-methyl-5-pyrimidinyl) methyl-3-(2-chloroethyl)-3-nitrosourea hydrochloride (ACNU) were studied in female Wistar rats by macroscopical autoradiography using 14C labeled ACNU. In normal rats, ACNU rapidly distributed in the subarachnoid space and ventricles after intracisternal administration. Diffusional transport into the brain tissue was limited to a depth of 1 or 2 mm from the cerebrospinal fluid (CSF) surface of the brain. Clearance of ACNU from the CSF space and brain was relatively fast and the half time of ACNU concentration at the cortical or ventricular surface was 10 min. In rats with leptomeningeal tumor induced by intracisternal inoculation of Walker 256 carcinosarcoma cells, the distribution pattern of ACNU after intracisternal administration was essentially the same as in normal rats until the tumor had grown in the subarachnoid space to form more than 10 or 20 layers of tumor cells. ACNU was distributed in the tumor as well. When the tumor had grown to form masses in the subarachnoid space, ACNU failed to penetrate to more than a depth of 1 or 2 mm from the tumor surface. Our results suggest that intrathecal ACNU administration may have no, or minor side effects on the brain and that it can eliminate floating or thin layered tumor cells in the subarachnoid space but not bulky tumors.

Animals↗

PubMed1994

Expression of PDGF, PDGF-receptor, EGF-receptor and sex hormone receptors on meningioma.

The expression of platelet derived growth factors (PDGF), the PDGF-Receptor (R) (alpha and beta types), epidermal growth factor (EGF)-Receptor (R) and sex hormone (oestrogen and progesterone) receptors was studied in 22 meningiomas. All tumours were PDGF-R beta type positive and 21 (95%) were PDGF positive. Only 2 (9%) were PDGF-R alpha type positive, 13 (59%) were EGF-R positive. The expression of these proteins was not related to the histological type or the malignancy of the meningiomas although the expression of PDGF and PDGF-R beta tended to be stronger in malignant meningiomas. Oestrogen and progesterone receptor protein were examined in 19 patients (10 females and 9 males). None of the meningioma cells revealed oestrogen receptor protein while 17 (89%) of the 19 meningiomas were positive for progesterone receptor protein. The expression of progesterone receptor was not related to histological type or malignancy. Our studies suggest that the autocrine system, through PDGF and PDGF-R type beta, may play an important role in the tumourigenicity of meningiomas. EGF-R was present in almost half and progesterone receptor in most of the meningiomas. There was no correlation between the expression of either PDGF, PDGF-R or EGF-R and the expression of progesterone receptor protein.

Adult↗

PubMed1994

Mycotic aneurysm accompanied by aspergillotic granuloma: a case report.

Solid granulomas and aneurysms caused by Aspergillus of the central nervous system are rare. A fungal aneurysm is usually situated proximally on the intracranial vessels and is often fatal. We report a case in which a ruptured aneurysm arose from the distal portion of the left posterior inferior cerebellar artery (PICA) in the course of treatment for chronic meningitis forming a granuloma at the left pyramis. After biopsy of the granuloma, he presented subarachnoid hemorrhage caused by a saccular PICA aneurysm. Histologic examination of the aneurysmal wall and the granuloma revealed infection with Aspergillus hyphae. The literature regarding the management of aspergillotic aneurysm is reviewed.

Aneurysm, Infected↗

PubMed1994

Isolated oculomotor nerve palsy following midbrain infarction.

A 62 year-old male patient presented with isolated oculomotor nerve palsy following a small infarction of the medial ventral midbrain, documented by magnetic resonance imaging. There was a 12-year history of hypertension, but no diabetes mellitus. Angiography revealed atherosclerosis of the paramedian mesencephalic arteries. Magnetic resonance imaging may be useful in patients with small brain stem infarctions.

Cerebral Angiography↗

PubMed1994

Cellular localization of type II Ca2+/calmodulin-dependent protein kinase in the rat basal ganglia and intrastriatal grafts derived from fetal striatal primordia, in comparison with that of Ca2+/calmodulin-regulated protein phosphatase, calcineurin.

We investigated immunohistochemically the cellular localization of multifunctional type II Ca2+/calmodulin-dependent protein kinase in the rat basal ganglia and intrastriatal grafts derived from fetal striatal primordia, in comparison with that of calcineurin, a reliable marker for striatal medium-sized spinous neurons. The type II Ca2+/calmodulin-dependent protein kinase-positive neurons were of medium size, with a mean diameter of 16.1 +/- microns (average +/- S.D., n = 72, range 13.6-18.3 microns) and comprised approximately 70% of the total neuronal population in the striatum. Light microscopy showed that the type II Ca2+/calmodulin-dependent protein kinase-positive cells had round, triangular or polygonal cell bodies with relatively little cytoplasm. Analysis of serial sections showed that type II Ca2+/calmodulin-dependent protein kinase and calcineurin immunoreactivities were co-localized in the striatal neurons examined with a similar distribution pattern. Type II Ca2+/calmodulin-dependent protein kinase-positive cells were always immunoreactive for calcineurin and cells negative for type II Ca2+/calmodulin-dependent protein kinase showed no apparent calcineurin immunoreactivity. Type II Ca2+/calmodulin-dependent protein kinase-positive nerve fibers in the globus pallidus and substantia nigra almost disappeared following striatal ischemic injury produced by transient middle cerebral artery occlusion and cerebral hemitransection, respectively, suggesting that these immunopositive fibers were striatal projections. Thus, most type II Ca2+/calmodulin-dependent protein kinase-positive neurons in the rat striatum are considered to be of the medium-sized spinous type. Type II Ca2+/calmodulin-dependent protein kinase or calcineurin immunoreactivity was also observed in a large number of neurons in transplants derived from fetal striatal primordia grafted into striatal ischemic lesions. In addition, type II Ca2+/calmodulin-dependent protein kinase- or calcineurin-immunoreactive nerve fibers appeared in the deafferented globus pallidus of the host rats, suggesting that the striatopallidal pathway was reformed by striatal projection neurons of the transplants. This finding may also indicate that Ca2+/calmodulin-regulated enzymes are useful for tracing striatal projection fibers as endogenous marker proteins.

Animals↗

PubMed1994

Occurrence of heat shock response in deafferented neurons in the substantia nigra of rats.

The substantia nigra is innervated by massive inhibitory GABAergic projections from the striatum and globus pallidus, deafferentation of which is supposed to lead to anterograde trans-synaptic degeneration of the nigral neurons. An immunohistochemical method was used to examine the induction of 72,000 mol. wt heat shock protein in the substantia nigra following cerebral hemitransection or transient middle cerebral artery occlusion. At three and four days post-transection, strong immunoreactivity for 72,000 mol. wt heat shock protein was found in the ipsilateral substantia nigra pars reticulata. Light microscopic observation revealed a number of pars reticulata neurons showing strong immunoreactivity for 72,000 mol. wt heat shock protein in their perikarya and proximal processes. In addition, Golgi-like stained neurons with dystrophic features were occasionally observed in the ipsilateral substantia nigra pars reticulata. The immunoreactivity for 72,000 mol. wt heat shock protein in the ipsilateral pars reticulata gradually declined and almost disappeared by 15 days after transection. No apparent induction of 72,000 mol. wt heat shock protein was found in the substantia nigra pars compacta throughout the time period examined. Massive striatal ischemic injury produced by transient middle cerebral artery occlusion also induced expression of 72,000 mol. wt heat shock protein in the pars reticulata neurons three and four days postoperatively. These findings suggest that deafferentation of the striatal or striatopallidal inputs per se is a harmful stress for the substantia nigra pars reticulata neurons, inducing 72,000 mol. wt heat shock protein synthesis. The present data may contribute to our understanding of the molecular basis of the pathomechanism of the transneuronal regression of substantia nigra pars reticulata neurons, which may occur after removal of inhibitory GABAergic inputs.

Afferent Pathways↗

PubMed1994

Microglial response to transient focal cerebral ischemia: an immunocytochemical study on the rat cerebral cortex using anti-phosphotyrosine antibody.

Microglial response to transient focal ischemia was examined using an immunohistochemical method with a monoclonal antibody to phosphotyrosine (P-Tyr). For this purpose, a rat model of reversible middle cerebral artery occlusion for 1 h was used. Compared with results in the noninsulted hemisphere, there was a significant increase in P-Tyr immunolabeling of the microglia in the insulted cerebral cortex 3 h postreperfusion. This microglial reaction progressed up to 24 h after ischemic insult. In the affected cerebral cortex, morphological changes of the microglial positive for P-Tyr were also observed, with shortened and thickened processes, enlarged cell bodies, and ameboid features. Cell density analysis did not show any apparent change in number of P-Tyr-positive microglia in the insulted cortex at 6, 12, and 24 h after reperfusion, suggesting that the cells with increased P-Tyr immunoreactivity were resident microglia. The present findings suggest that signal transduction mediated by tyrosine phosphorylation is involved in the microglial response to ischemic injury in the rat cerebral cortex.

Animals↗