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Biomedical subjects

S Shimohama

Publications and source records attributed to S Shimohama.

At least 127 records · Page 7Linked to original sources
PubMed1995

Glutamate-induced antigenic changes of phospholipase C-delta in cultured cortical neurons.

Phosphoinositide-specific phospholipase C (PLC) is a key enzyme in signal transduction. It was previously demonstrated that an antibody to an isozyme of PLC, PLC-delta, produces intense staining of neurofibrillary tangles (NFT), the neurites surrounding senile plaque (SP) cores and neuropil threads in the brains of patients with Alzheimer's disease (AD). Although the etiology of neuronal degeneration in AD is still to be defined, excitotoxic glutamate might be a candidate. In the present study, an anti-PLC-delta antibody was used to examine the influence of glutamate on PLC-delta immunoreactivity in cultured rat cortical neurons. Exposure to glutamate caused the death of cultured cortical neurons and exhibited increased immunostaining with the anti-PLC-delta antibody. Subtoxic doses of glutamate also increased PLC-delta immunoreactivity in a dose-dependent manner. Both glutamate-induced neuronal degeneration and the increases in PLC-delta immunoreactivity were prevented by removal of extracellular Ca2+ or the application of an N-methyl-D-aspartate (NMDA) receptor antagonist, MK-801. The glutamate-induced increase in PLC-delta immunoreactivity was also prevented by N omega-nitro-L-arginine, a nitric oxide (NO) synthase inhibitor. These results suggest that NO formation secondary to Ca2+ influx by NMDA receptor activation leads to similar modifications of PLC-delta to those seen in AD.

Animals↗
PubMed1995

Characterization of the association of phospholipase C-delta with Alzheimer neurofibrillary tangles.

Phosphoinositide-specific phospholipase C (PLC) is a key enzyme in signal transduction. We have previously demonstrated that an antibody to the PLC isozyme, PLC-delta, intensely stained neurofibrillary tangles (NFT) in the brain tissue of AD patients [Am. J. Pathol., 139 (1991) 737-742]. To clarify the crucial involvement of abnormal PLC-delta accumulation contributing to the formation of NFT, we performed light and electron microscopic immunocytochemistry. To determine PLC-delta's association with NFT, its resistance to solubilization was also studied. Anti-PLC-delta antibody marked the same NFT-bearing neurons containing tau immunoreactivity with tau more clearly on NFT filaments and PLC-delta covering it superficially at the light microscope level. The double stained preparations with anti-PLC-delta antibody and bFGF binding suggested that PLC-delta is an intracellular marker and is not retained after neuronal death. Employing immunoperoxidase and immunogold electron microscopic immunocytochemistry, we found that the antibody to PLC-delta reacts mostly with amorphous granular materials, and occasionally with some abnormal filaments within NFT. Nevertheless, PLC-delta in NFT was resistant to removal by high salt or ionic detergent, indicating it is an integral NFT component. These results indicate that antigenic determinants unique to PLC-delta are mainly present intraneuronally on the amorphous granular components of NFT as well as the abnormal filaments, suggesting PLC-delta's interactions and possible role in the formation of intraneuronal filamentous inclusions in AD.

Aged↗
PubMed1995

Patch sensor detection of glutamate release evoked by a single electrical shock.

We tried to detect minimal stimulation-induced glutamate overflow from the surface of a hippocampal slice using an outside-out patch electrode excised from pyramidal cell membranes. The amplitude of the stimulation-induced patch current was dependent on the distance between the slice surface and the tip of patch sensor. The current-voltage relations of the stimulation-induced patch current were similar to those of the current evoked puff by application of L-glutamate to the patch. This indicates that the stimulation-induced patch current was produced by glutamate released from presynaptic terminals, and thus this technique may be useful in the study of transmitter release evoked by minimal electrical stimulation in brain slices.

Action Potentials↗
PubMed1995

Alteration of phospholipase C-delta protein level and specific activity in Alzheimer's disease.

Phosphoinositide-specific phospholipase C (PLC) is a key enzyme in signal transduction. We have previously demonstrated that an antibody to an isozyme of PLC, PLC-delta, produced intense staining of neurofibrillary tangles in the brains of patients with Alzheimer's disease. In the present study, we investigated the protein level and activity of this enzyme in control and Alzheimer brains. Western blot analysis using a specific antibody for PLC-delta showed that the concentration of PLC-delta protein was significantly higher in the cytosolic fraction of Alzheimer disease cortical tissue than in control brains. The activity of PLC-delta, which hydrolyzes phosphatidylinositol, was also investigated, and we found that PLC-delta activity was not significantly different in the Alzheimer and control cytosolic fractions. These results indicate that the specific activity of PLC-delta is decreased in Alzheimer brains and suggest that inactivation of PLC-delta might be related to the pathophysiology of this disease.

Aged↗
PubMed1995

Signal transduction mechanisms in Alzheimer disease.

The present article focuses on our studies on the metabolism of the inositol phospholipids in Alzheimer disease (AD). The phospholipase C (PLC) isozyme, PLC-delta 1, was abnormally accumulated in neurofibrillary tangles (NFT), the neurites surrounding senile plaque cores, and neuropil threads in AD brains. Anti-PLC-delta 1 antibody marked the same NFT-bearing neurons containing tau immunoreactivity. Electron microscopic immunocytochemistry revealed that antigenic determinants unique to PLC-delta 1 are mainly present intraneuronally on the amorphous granular components of NFT as well as the abnormal filaments. Although the concentration of PLC-delta 1 protein was significantly higher in the cytosolic fraction of AD cortical tissue than in control brains, the specific activity of PLC-delta 1 is decreased in AD brains. The amounts of PLC-beta 1 and -gamma 1 and type beta protein kinase C were significantly reduced in the membranous fraction of the AD temporal cortical tissues compared with controls. The PLC-delta 1 abnormality was also present in nonneuronal tissues as well as the brains of patients with AD. It was revealed that nitric oxide (NO) formation secondary to Ca2+ influx by N-methyl-D-aspartate (NMDA) receptor activation leads to modifications of PLC-delta 1 similar to those seen in AD brains. These results suggest that altered Ca2+ homeostasis, occurring as a consequence of aberrant phosphoinositide metabolism, may be related to key features of AD such as neurofibrillary degeneration, aberrant amyloid deposits, and neuronal death.

Alzheimer Disease↗
PubMed1995

Differential involvement of phospholipase C isozymes in Alzheimer's disease.

Phosphoinositide-specific phospholipase C (PLC) is a key enzyme in signal transduction. We demonstrated that an antibody to an isozyme of PLC, PLC-delta 1, produced intense staining of neurofibrillary tangles (NFT) in the brains of patients with Alzheimer's disease (AD). Western blot analysis using a specific antibody for PLC-delta 1 showed that the concentration of PLC-delta 1 protein was significantly higher in the cytosolic fraction of AD cortical tissues than in controls. The activity of PLC-delta 1, which hydrolyzes phosphatidylinositol, was not significantly different in the Alzheimer and control cytosolic fraction, indicating that the specific activity of PLC-delta 1 is decreased in Alzheimer brains. Western blot analysis also showed that the concentration of PLC-delta 1 and PLC-gamma 1 was significantly lower in the membraneous fraction of AD cortical tissues than in controls. These results suggest a differential involvement of PLC isozymes in AD.

Aged↗
PubMed1995

Effects of brain-derived neurotrophic factor on 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-induced parkinsonism in monkeys.

The effects of intrathecal infusion of brain-derived neurotrophic factor (BDNF) were examined in a 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-induced parkinsonian model in monkeys. Nine Japanese monkeys were divided randomly into three groups, an untreated control (n = 3), a BDNF group (n = 3), and a non-BDNF group (n = 3). Animals in the BDNF group received continuous intrathecal infusion of 10 ml of cell culture medium containing 10 micrograms of BDNF protein; the non-BDNF group received intrathecal infusion of the same culture medium without BDNF. To induce parkinsonian syndromes, a total of 1 mg/kg 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine was administered intravenously to each monkey in both the BDNF and non-BDNF groups. The neurological signs in the monkeys were monitored for 2 weeks and were scored according to the monkey parkinsonism rating scale; histological changes in the substantia nigra were evaluated after the 2-week observation period. The BDNF-treated animals remained asymptomatic during the 1st week and showed mild parkinsonism during the 2nd week, whereas the non-BDNF group showed typical parkinsonian syndrome during the 1st week, with deterioration in the 2nd week. Histological damage in the substantia nigra correlated well with the clinical features. Severe neuronal cell loss in the substantia nigra was observed in animals with severe parkinsonism (those in the non-BDNF group), whereas significantly less damage was observed in this region in the BDNF group.(ABSTRACT TRUNCATED AT 250 WORDS)

1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine↗
PubMed1995

[Protein kinase C].

Protein kinase C (PKC) is a phospholipid-dependent protein kinase that plays important roles in various cellular functions. At least 11 PKC isozymes are known, and structural differences classify these isozymes into three groups, conventional PKC (cPKC), novel PKC (nPKC) and atypical PKC (aPKC). PKC isozymes exhibit differential tissue expressions and enzymatic properties. PKC may also play some roles in pathologic conditions. Several studies have shown the crucial roles of PKC isozymes in the transformation and growth of tumor cells, although these studies failed to show consistent results yet. In Alzheimer's disease, several studies revealed the special and systemic involvement of PKC beta isozyme.

Alzheimer Disease↗
PubMed1995

Reduction of platelet phospholipase C activity in patients with Alzheimer disease.

Phosphoinositide-specific phospholipase C (PLC) is a key enzyme in signal transduction. We have previously demonstrated that a PLC isozyme is abnormally accumulated in the brain tissue in Alzheimer's disease (AD). AD has been suggested to be a systemic disease in which the expression of abnormalities is most prominent in neuronal tissues. In a recent study, we have revealed the increase of the cytosolic protein kinase C (PKC) concentration in platelets of AD patients, suggesting the change of PLC, which is upstream to PKC in phosphoinositide metabolism. In this study, we examined phosphatidylinositol-specific PLC activity in platelets from patients with AD and age-matched controls by measuring the formation of radioactive inositol phosphate. The PLC activity was significantly lower in the AD platelets than in the controls. These findings suggest that aberrant phosphoinositide metabolism is present in nonneuronal tissues as well as the brain in AD.

Aged↗
PubMed1994

The endogenous substrate of low molecular weight acid phosphatase in the brain is an epidermal growth factor receptor.

We have recently reported that low molecular weight (LMW) acid phosphatase, which is supposed to possess phosphotyrosine protein phosphatase activity, showed a significant decrease of activity in Alzheimer brains compared to control brains [Ann. Neurol., 33 (1993) 616-621]. In the present study, we investigated the endogenous substrate of LMW acid phosphatase in the brain. LMW acid phosphatase was purified from bovine brain, and the enzyme was obtained with both a high specific activity and a good yield. The bovine brain enzyme was a monomer with a molecular mass of 17 kDa. We used a specific monoclonal anti-phosphotyrosine antibody to detect phosphotyrosine protein in rat brain extracts. The LMW acid phosphatase from bovine brain dephosphorylated a M(r) 170 kDa phosphotyrosine protein in rat brain extracts. This M(r) 170 kDa protein was considered to be the epidermal growth factor (EGF) receptor using a specific antibody. These results suggest that LMW acid phosphatase in the brain may regulate EGF receptor-dependent transmembrane signalling by dephosphorylating the phosphorylated receptor.

Acid Phosphatase↗
PubMed1994

Nicotine-induced protection of cultured cortical neurons against N-methyl-D-aspartate receptor-mediated glutamate cytotoxicity.

The effects of nicotine on glutamate-induced cytotoxicity were examined using primary cultures of rat cortical neurons. The cell viability was significantly reduced when cultures were briefly exposed to glutamate or N-methyl-D-aspartate (NMDA) then incubated with normal medium for 1 h. A 1-h exposure of the cultures to kainate or alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA) reduced cell viability. Incubating cultures with nicotine for 1-24 h protected cortical neurons against glutamate cytotoxicity. Maximum protection against glutamate cytotoxicity was induced with a 2-h nicotine incubation. Exposure to nicotine for up to 2 h did not affect cell viability by itself although cell viability was reduced in a time-dependent manner when the exposure exceeded 4 h. Neuroprotection by nicotine was dependent on both the concentration and incubation period. Nicotine reduced the NMDA cytotoxicity but did not attenuate that of kainate and AMPA. The neuroprotective effects of nicotine against glutamate cytotoxicity were antagonized by mecamylamine and hexamethonium but not by atropine. These results indicate that nicotinic receptor stimulation induces neuroprotection against glutamate cytotoxicity mediated by NMDA receptors.

Animals↗
PubMed1994

Different modulation of protein kinase C isozymes in dibutyryl cyclic AMP-differentiated PC12h cells.

PC12h cells can be differentiated into sympathetic neuron-like cells by various agents, including nerve growth factor, basic fibroblast growth factor, cyclic AMP analogues, and protein kinase C (PKC) activators. To study the involvement of PKC in the process of PC12h cell differentiation by cyclic AMP treatment, PKC isozymes (alpha, beta I, beta II, and gamma) were analyzed using column chromatography and immunoblotting. Two PKC isozymes, PKC(alpha) and PKC(beta II), were predominantly detected in PC12h cells. When stimulated by dibutyryl cyclic AMP, PKC(alpha) levels declined in the cytosolic fraction of the cells, whereas PKC(beta II) levels increased. Increased PKC(beta II) levels were also detected in the particulate fraction, whereas particulate PKC(alpha) levels did not change. The total PKC activity decreased in the cytosolic fraction following cyclic AMP stimulation of PC12h cells, whereas it stayed constant in the particulate fraction. Fractionation on a hydroxyapatite column showed a decreased level of PKC(alpha) activity and a transient increase followed by a decreased level of PKC(beta II) activity. This discrepancy between increased PKC(beta II) immunoreactivity and reduced PKC(beta II) activity suggested the presence of nonactivatable PKC(beta II) in cyclic AMP-treated PC12h extract. These findings indicate that PKC(alpha) and PKC(beta II) are differentially regulated during the differentiation of PC12h cells. In addition, the differentiation of PC12h cells triggered by cyclic AMP seems to involve characteristic alterations of PKC isozymes.

Animals↗
PubMed1994

Apolipoprotein E polymorphism in Japanese patients with Alzheimer's disease or vascular dementia.

Apolipoprotein E (ApoE) plays a key part in lipid metabolism both in the liver, and in the CNS. To clarify the association of ApoE polymorphism with Alzheimer's disease and vascular dementia in Japan, 13 patients with early onset (age > or = 65) sporadic Alzheimer's disease, 40 patients with late onset (age < or = 65) sporadic Alzheimer's disease, 19 patients with vascular dementia, and 49 non-demented control subjects were analysed. The results showed a significantly increased frequency of the epsilon 4 allele in the patients with late onset sporadic Alzheimer's disease (0.25), but not in the patients with early onset sporadic Alzheimer's disease (0.04) or in the patients with vascular dementia (0.13) compared with controls (0.09). The raised frequency of the epsilon 4 allele in the patients with late onset sporadic Alzheimer's disease was of a lower magnitude than that in United States and Canadian studies. This may in part be due to a lower epsilon 4 frequency in the normal Japanese population and reflect the lower morbidity from Alzheimer's disease in Japan.

Adult↗
PubMed1993

Protective effect of nerve growth factor against glutamate-induced neurotoxicity in cultured cortical neurons.

The effect of recombinant human nerve growth factor (hNGF) and mouse NGF on cultured rat cortical neurons was examined. The DNA fragment coding the human NGF gene was isolated and inserted downstream from the SV40 promoter in a plasmid containing the dihydrofolate reductase cDNA, and this plasmid was introduced into Chinese hamster ovary (CHO) cells to establish cells producing recombinant hNGF. The recombinant hNGF protein secreted by CHO cells was confirmed to be biologically active in an assay using PC12 cells. Brief exposure of cortical cells to glutamate followed by incubation with glutamate-free medium reduced cell viability by 60-70% when compared with the control culture. Simultaneous addition of recombinant hNGF or mouse NGF to rat cortical cultures with glutamate did not affect this reduction of cell viability. However, 24 h pretreatment of rat cortical cultures with recombinant hNGF or mouse NGF resulted in a significant reduction of glutamate-induced neuronal damage. Mouse NGF also protected cortical neurons against N-methyl-D-aspartate (NMDA)- and kainate-induced neuronal damage. These findings suggest that NGF can protect cortical neurons against glutamate-induced neurotoxicity.

Animals↗
PubMed1993

Brain-derived neurotrophic factor pretreatment exerts a partially protective effect against glutamate-induced neurotoxicity in cultured rat cortical neurons.

The effect of recombinant brain-derived neurotrophic factor (BDNF) on cultured rat cortical neurons was examined. Brief exposure of cortical neurons to glutamate followed by incubation with glutamate-free medium reduced cell viability by 60-70% when compared with the control value. Simultaneous addition of recombinant BDNF to rat cortical cultures with glutamate did not affect this reduction of cell viability. However, 24 h pretreatment of rat cortical cultures with recombinant BDNF resulted in a significant reduction of glutamate-induced neuronal damage. These findings suggest that BDNF can protect cortical neurons against glutamate-induced neurotoxicity.

Animals↗
PubMed1993

Abnormal accumulation of phospholipase C-delta in filamentous inclusions of human neurodegenerative diseases.

We have previously demonstrated that an antibody to phosphoinositide-specific phospholipase C (PLC) isozyme, PLC-delta, intensely stained neurofibrillary tangles (NFT) in the brain tissue of Alzheimer's disease (AD). This study was performed to determine if abnormal PLC-delta accumulation might be present in the filamentous inclusions of other neurodegenerative diseases. We found that the anti-PLC-delta antibody stained neuronal inclusions of Pick's disease, progressive supranuclear palsy and diffuse Lewy body disease while the inclusions of idiopathic Parkinson's disease lacked PLC-delta accumulation. These results suggest a possible role for PLC-delta interaction in the formation of intraneuronal filamentous inclusions in human neurodegenerative diseases.

Aged↗
PubMed1993

Aberrant phosphoinositide metabolism in Alzheimer's disease.

Since phosphoinositide-specific phospholipase C (PLC) is one of the key molecules in signal transduction, its involvement was assessed in Alzheimer's disease (AD). The phosphatidyl-inositol (PI)-specific PLC activity in the Alzheimer cytosolic and particulate fractions was not significantly different from that in the control fractions. The PI-specific PLC activity as a function of the free Ca2+ concentration was also similar between control and Alzheimer brains. These results suggest that the PI-specific PLC activity is not altered in AD. Immunostaining of a specific antibody against the PLC isozyme, PLC-delta, demonstrated that this enzyme was abnormally accumulated in neurofibrillary tangles (NFT), the neurites surrounding senile plaque (SP) cores, and neuropil threads in AD brains. Western blot analysis confirmed that PLC-delta was concentrated in the paired helical filament (PHF)-rich fraction of AD brains. PLC-delta marked the same neurons containing tau immunoreactivity and yet tau and PLC-delta often marked different structures within the same neuron, with tau more clearly on NFT and PLC-delta covering it superficially. The double stain with PLC-delta and basic fibroblast growth factor (bFGF) binding suggest that PLC-delta is an intracellular marker, showing little overlap with bFGF binding, an extracellular marker. All of this was consistent with the electron microscopy, with PLC-delta being NFT associated. Antibodies to other PLC isozymes did not produce positive immunostaining of these pathologic structures. Moreover, diffuse and amorphous deposits of PLC-delta were found to precede the accumulation of fibrillary deposits. These results suggest that PLC-delta accumulation plays a possible role in the formation of intraneuronal inclusions in AD.

Alzheimer Disease↗