Biomedical subjects

Moo Ho Won

Publications and source records attributed to Moo Ho Won.

At least 37 records · Page 2Linked to original sources

PubMed2006

Immunohistochemical studies of human ribosomal protein S3 (rpS3).

The human ribosomal protein S3 (rpS3) was expressed in E. coli using the pET-15b vector and the monoclonal antibodies (mAbs) were produced and characterized. A total of five hybridoma cell lines were established and the antibodies recognized a single band of molecular weight of 33 kDa on immunoblot with purified rpS3. When the purified rpS3 was incubated with the mAbs, the UV endonuclease activity of rpS3 was inhibited up to a maximum of 49%. The binding affinity of mAbs to rpS3 determined by using a biosensor technology showed that they have similar binding affinities. Using the anti-rpS3 antibodies as probes, we investigated the cross-reactivities of various other mammalian brain tissues and cell lines, including human. The immunoreactive bands on Western blots appeared to be the same molecular mass of 33 kDa in all animal species tested. They also appear to be extensively cross-reactive among different organs in rat. These results demonstrated that only one type of immunologically similar rpS3 protein is present in all of the mammalian brain tissues including human. Furthermore, these antibodies were successfully applied in immunohistochemistry in order to detect rpS3 in the gerbil brain tissues. Among the various regions in the brain tissues, the rpS3 positive neurons were predominantly observed in the ependymal cells, hippocampus and stantia nigra pars compacta. The different distributions of rpS3 in brain tissues reply that rpS3 protein may play an important second function in the neuronal cells.

Animals↗

PubMed2006

Differential effects and changes of ceruloplasmin in the hippocampal CA1 region between adult and aged gerbils after transient cerebral ischemia.

In this study, we examined the differential effects and changes of ceruloplasmin between adult and aged gerbil hippocampus after transient forebrain ischemia. Ceruloplasmin in the hippocampal CA1 region of adult and aged gerbils was significantly changed after ischemia/reperfusion. Whereas, it was not significantly changed in the CA2/3 region compared to the CA1 region after ischemia. Ceruloplasmin immunoreactivity and its protein level in aged gerbil CA1 region were higher than those in adult gerbil CA1 region. Ceruloplasmin in the CA1 region was highest in adult gerbils and aged gerbils at 24h and 12h after transient ischemia, respectively. At these time points, strong ceruloplasmin immunoreactivity was observed in CA1 pyramidal cells. Thereafter, ceruloplasmin was decreased with time after ischemia. Four days after ischemia/reperfusion, ceruloplasmin immunoreactivity in both adult and aged gerbils was expressed in astrocytes in the CA1 region. Ceruloplasmin treatment in adult ischemic gerbils showed strong protective effect against ischemic damage in CA1 pyramidal cells compared to that in aged ischemic gerbils. We conclude that ceruloplasmin early increases in the aged gerbil CA1 region compared to that of the adult gerbil CA1 region may be associated with the earlier induction of reactive oxygen species, and ceruloplasmin shows strong neuroprotective effects in adults compared to those in aged gerbils.

Age Factors↗

PubMed2006

Mineralocorticoid and glucocorticoid receptor expressions in astrocytes and microglia in the gerbil hippocampal CA1 region after ischemic insult.

In the present study, we observed expression and changes of mineralocorticoid receptor (MR) and glucocorticoid receptor (GR) in the gerbil hippocampal CA1 region, but not in the CA2/3 region, after 5 min of transient forebrain ischemia. In blood, corticosterone levels were increased biphasically at 30 min and 12 h after ischemia/reperfusion, and thereafter its levels were decreased. In the sham-operated group, MR and GR immunoreactivities were weakly detected in the CA1 region. By 3 days after ischemia, MR and GR were not significantly altered in the CA1 region: at 12 h after ischemia, GR was expressed in a few neurons in the CA1 region, whereas MR was not expressed in any neurons after ischemic insult. From 4 days after ischemia, MR and GR immunoreactivities were detected in astrocytes and microglia in the CA1 region, and at 7 days after ischemia, MR and GR immunoreactivities peaked in the hippocampal CA1 region. At this time, 55% of astrocytes and 30% of microglia showed MR immunoreactivity, and 20% of astrocytes and 40% of microglia showed GR immunoreactivity. Western blot analyses showed that the pattern of changes in MR and GR protein levels was similar to the immunohistochemical changes observed after transient forebrain ischemia. From 4 days after ischemia, MR and GR protein levels were increased time-dependently after ischemia. In conclusion, enhanced MR and GR expressions in astrocytes and microglia were detected in the hippocampal CA1 region 4-7 days after ischemia/reperfusion. At this time, GR immunoreactivity was abundant in microglia, whereas MR immunoreactivity was prominent in astrocytes. The specific distribution of corticosteroid receptors in the astrocytes and microglia may be associated with the differences of MR and GR functions against ischemic damage.

Animals↗

PubMed2006

Tat-mediated protein transduction of human brain pyridoxine-5-P oxidase into PC12 cells.

Pyridoxine-5-P oxidase catalyses the terminal step in the biosynthesis of pyridoxal-5-P, the biologically active form of vitamin B6 which acts as an essential cofactor. Here, a human brain pyridoxine-5-P oxidase gene was fused with a gene fragment encoding the HIV-1 Tat protein transduction domain (RKKRRQRRR) in a bacterial expression vector to produce a genetic in-frame Tat-pyridoxine-5-P oxidase fusion protein. Expressed and purified Tat-pyridoxine-5-P oxidase fusion protein transduced efficiently into PC12 cells in a time- and dose-dependent manner when added exogenously to culture media. Once inside the cells, the transduced Tat-pyridoxine-5-P oxidase protein showed catalytic activity and was stable for 48 h. Moreover, the formation of pyridoxal-5-P was increased by adding exogenous Tat-pyridoxine-5-P oxidase to media pre-treated with the vitamin B6 precursor pyridoxine. In addition, the intracellular concentration of pyridoxal-5-P was markedly increased when Tat-pyridoxal kinase was transduced together with Tat-pyridoxine-5-P oxidase into cells.These results suggest that the transduction of Tat-pyridoxine-5-P oxidase fusion protein presents a means of regulating the level of pyridoxal-5-P and of replenishing this enzyme in various neurological disorders related to vitamin B6.

Animals↗

PubMed2006

Rip immunoreactivity significantly decreases in the stratum oriens of hippocampal CA1 region after transient forebrain ischemia in gerbils.

In the present study, we observed ischemia-related changes in Rip recognizing the promyelinating and myelinating oligodendrocytes in the hippocampus proper after 5 min of transient forebrain ischemia in gerbils. Rip immunoreactivity was significantly altered in the hippocampal CA1 region but not in the CA2/3 region after ischemic insult. In the sham-operated group, Rip immunoreactivity was shown in the cell bodies and processes of oligodendrocytes in all layers of the hippocampus proper. From 15 min to 2 days after ischemic insult, Rip immunoreactivity was similar to that of sham-operated group. Three days after ischemic insult, Rip-immunoreactive processes were tangled in the stratum oriens of the CA1 region, and Rip protein level decreased from this time after ischemia. Thereafter, Rip immunoreactivity was decreased time dependently in the CA1 region. Seven days after ischemic insult, Rip-immunoreactive processes were tangled and densely detected in the stratum oriens adjacent to the stratum pyramidale. In brief, these results indicate that the significant decrease of Rip immunoreactivity in processes in the stratum oriens of the hippocampal CA1 region occurs at late time after ischemia, and this decrease in Rip immunoreactivity may be associated with delayed neuronal death of CA1 pyramidal cells.

Animals↗

PubMed2006

Altered expression of K+ -Cl- cotransporters affects fast paired-pulse inhibition during GABA receptor activation in the gerbil hippocampus.

K+ -Cl- cotransporter (KCC) plays an important role in maintaining neuronal activity. However, the effect of seizure activity or pharmacological manipulation of GABAergic transmission on KCC expression remains to be clarified. Therefore, the present study was performed to investigate whether seizure activity or GABA receptor agonist treatment changes KCC expression in the gerbil hippocampus. Furthermore, the effect of blockade of KCC on inhibitory transmission in the dentate gyrus was identified following applications of GABA receptor agonists. The distribution of KCC immunoreactivity in the hippocampus was similarly detected between seizure-resistant (SR) and seizure-sensitive (SS) gerbils. Baclofen (a GABAB receptor agonist) treatment markedly increased KCC expression in the gerbil hippocampus. Baclofen treatment significantly reduced paired-pulse inhibition in the dentate gyrus. Furosemide (a KCC inhibitor) treatment amplified the effect of baclofen on paired-pulse responses. In contrast, muscimol (a GABAA receptor agonist) treatment reduced KCC expression. Enhanced paired-pulse inhibition by muscimol treatment was not affected by furosemide treatment. These findings suggest that seizure activity in the gerbil may not affect KCC expression in the hippocampus. In addition, altered KCC immunoreactivity induced by baclofen or muscimol may play an important role in maintaining or regulating inhibitory transmission during GABA receptor activation.

Animals↗

PubMed2006

Ischemia-related changes of glial-derived neurotrophic factor and phosphatidylinositol 3-kinase in the hippocampus: their possible correlation in astrocytes.

In the present study, we observed the changes of endogenous expression of glial-cell-line-derived neurotrophic factor (GDNF) and phosphatidylinositol 3-kinase (PI-3 kinase) in the gerbil hippocampus after transient forebrain ischemia and investigated the correlation between GDNF and PI-3 kinase in the ischemic hippocampus. In the sham-operated group, GDNF and PI-3 kinase immunoreactivity was not found in any cells in the hippocampal CA1 region. GDNF, not PI-3 kinase, immunoreactivity was expressed in non-pyramidal cells in the CA1 region at 6 h after ischemic insult. At 12-24 h after ischemia, GDNF and PI-3 kinase immunoreactivity in the CA1 region was similar to that of the sham-operated group. From 2 days after ischemic insult, GDNF- and PI-3-kinase-immunoreactive astrocytes were detected in the CA1 region, and GDNF and PI-3 kinase immunoreactivity in astrocytes was highest in the CA1 region 4 days after ischemic insult. Moreover, at this time point, GDNF and PI-3 kinase were co-localized in some astrocytes. Western blotting showed that ischemia-related changes of GDNF and PI-3 kinase protein levels were similar to the immunohistochemical changes after ischemia. These results suggest that GDNF and PI-3 kinase may be related to delayed neuronal death and that GDNF and PI-3 kinase may be involved in activation of astrocytes.

Animals↗

PubMed2006

Epileptogenic roles of astroglial death and regeneration in the dentate gyrus of experimental temporal lobe epilepsy.

Recent studies have demonstrated that blockade of neuronal death in the hippocampus cannot prevent epileptogenesis in various epileptic models. These reports indicate that neurodegeneration alone is insufficient to cause epilepsy, and that the role of astrocytes in epileptogenesis should be reconsidered. Therefore, the present study was designed to elucidate whether altered morphological organization or the functionalities of astrocytes induced by status epilepticus (SE) is responsible for epileptogenesis. Glial responses (reactive microgliosis followed by astroglial death) in the dentate gyrus induced by pilocarpine-induced SE were found to precede neuronal damage and these alterations were closely related to abnormal neurotransmission related to altered vesicular glutamate and GABA transporter expressions, and mossy fiber sprouting in the dentate gyrus. In addition, newly generated astrocytes showed down-regulated expressions of glutamine synthase, glutamate dehydrogenase, and glial GABA transporter. Taken together, our findings suggest that glial responses after SE may contribute to epileptogenesis and the acquisition of the properties of the epileptic hippocampus. Thus, we believe that it is worth considering new therapeutic approaches to epileptogenesis involving targeting the inactivation of microglia and protecting against astroglial loss.

Animals↗

PubMed2006

Transient ischemia-induced expression and changes of tyrosine kinase A in the hippocampal dentate gyrus of the gerbil.

The present study examined ischemia-related changes in tyrosine kinase A (trkA) immunoreactivity and its protein content in the dentate gyrus after 5 min of transient forebrain ischemia in gerbils. One day after ischemic insult, cresyl violet-positive polymorphic cells showed ischemic degeneration. The ischemia-induced changes in trkA immunoreactivity were found in the polymorphic layer (PL) and granule cell layer (GCL) of the dentate gyrus. In the sham-operated group, trkA immunoreactivity in the dentate gyrus was very weak. From 30 min after ischemia, trkA immunoreactivity was increased in the dentate gyrus and peaked in the dentate gyrus at 12 h after ischemia-reperfusion. Thereafter, trkA immunoreactivity was decreased time-dependently after ischemia-reperfusion. Four days after ischemic insult, trkA immunoreactivity was similar to that of the sham-operated group. In addition, it was found that ischemia-related changes in trkA protein content were similar to the immunohistochemical changes. These results suggest that the chronological changes of trkA in the dentate gyrus after transient forebrain ischemia may be associated with ischemic damage in polymorphic cells of the dentate gyrus.

Analysis of Variance↗

PubMed2006

Calbindin D-28k immunoreactivity increases in the hippocampus after long-term treatment of soy isoflavones in middle-aged ovariectomized and male rats.

This article investigates the long-term effects of soybean isoflavones (ISO) on the changes of calbindin D-28k (CB) immunoreactivity in the hippocampus in middle-aged ovariectomized female rats as well as middle-aged control female and male rats to identify any correlation between calcium and phytoestrogens. In the CA1 region, CB immunoreactivity in the ovariectomized females was similar to that of the control females, whereas CB immunoreactivity in the males was significantly lower than that of the control females. In the dentate gyrus, CB immunoreactivity in the ovariectomized females and males was significantly lower than that of the control females. CB immunoreactivity in all groups was increased dose-dependently after ISO treatment in the CA1 region and dentate gyrus. This result suggests that ISO treatment enhances the expression of CB immunoreactivity in the hippocampus in the middle-aged rats.

Age Factors↗

PubMed2006

trans-10,cis-12 conjugated linoleic acid inhibits the G1-S cell cycle progression in DU145 human prostate carcinoma cells.

Conjugated linoleic acid (CLA) is a group of positional and geometric isomers of linoleic acid, and has evidenced anti-cancer activities in experimental animal cancer models and in vitro studies. The two predominant isomers of CLA are cis-9,trans-11 CLA (c9t11) and trans-10,cis-12 CLA (t10c12). The present study was performed to study the effect of the individual CLA isomers on DU145 cell growth. The cells were incubated in serum-free medium with different concentrations of the fatty acids. Treatment of cells with t10c12 (at 2.5-10 micromol/L) resulted in a dose-dependent reduction in the numbers of viable cells, whereas c9t11 CLA at a concentration of 5 micromol/L slightly increased viable cell numbers at 3 days (P < .05). DNA flow cytometric analysis revealed that the treatment of DU145 cells with t10c12 for 24 hours induced a small but significant increase in the number of cells in the G1 phase, accompanied by a complementary decrease in cells in the S phase. c9t, however, had no effect on cell cycle progression. To determine the molecular mechanisms underlying t10c12-induced G1 arrest, the levels of cell cycle regulatory proteins were estimated by western blot analyses. t10c12 induced a marked increase in p21(CIP1/WAF1) protein levels in a dose-dependent manner. p27(KIP1) was not affected by t10c12. t10c12 moderately decreased cyclin A and cyclin D1 protein levels (P > .05). However, t10c12 did not affect the expression of cyclin-dependent kinase (CDK) 2, CDK4, or cyclin E. t10c12 increased p21(CIP1/WAF1) bound to CDK2 and attenuated CDK2 activity. These results indicate that t10c12-induced p21(CIP1/WAF1) binds to CDK, and inhibits the activity of this enzyme, which results in the observed decrease in the G1-S progression in DU145 cells.

Blotting, Western↗

PubMed2006

Berberry extract reduces neuronal damage and N-Methyl-D-aspartate receptor 1 immunoreactivity in the gerbil hippocampus after transient forebrain ischemia.

In the present study, we studied the neuroprotective effects of berberry extract (BE) against ischemic damage and the temporal and spatial alterations of N-methyl-D-aspartate receptor type 1 (NR1) and NR2A/2B immunoreactivities in the gerbil hippocampal CA1 region after transient ischemia to examine anti-ischemic effects and its role in transient forebrain ischemia. In the vehicle-treated group, the percentage of cresyl violet positive pyramidal cells in the CA1 region was about 11.4% compared to the sham-operated group 4 d after ischemic insult. BE showed neuroprotective effects against ischemic damage after ischemia-reperfusion. In the BE-treated groups, about 60-75% of CA1 pyramidal cells were stained with cresyl violet 4 d after ischemic insult. We observed the percentage of berberine (7.45+0.85 mg/g in BE) by HPLC, which is active ingredient of BE. NR1 immunoreactivity in the stratum pyramidale of the CA1 region in the vehicle-treated group was significantly increased at 30 min after transient forebrain ischemia, while at this time the NR1 immunoreactivity in the BE-treated groups was significantly low compared to the vehicle-treated group. The pattern of NR2A/B immunoreactivity in the stratum pyramidale of the BE-treated group and its protein levels were similar to that in the vehicle-treated group after ischemic insult. These results suggest that BE has potent neuroprotective effects against ischemic damage via the reduction of NR1 activity.

Animals↗

PubMed2006

Age-related changes in calretinin-immunoreactive periglomerular cells in the rat main olfactory bulb.

The changes of calretinin (CR)-immunoreactive periglomerular cells in the glomerular layer of the main olfactory bulb (MOB) were investigated in rats differing ages from postnatal month 1 (PM 1) to PM 24. The number of cresyl violet-positive periglomerular cells was similar between PM 1 and PM 12, but they decreased slightly in the PM 24 group. The size of CR-immunoreactive periglomerular cells in the glomerular layer increased with age, while their numbers did not change significantly in the PM 6-PM 24 groups. In the PM 24 group, numbers of CR-positive periglomerular cell bodies and their processes decreased, while the size of CR-positive cell bodies in the glomeruli was larger than that of the previous groups. These results suggest that CR-immunoreactive periglomerular cells in the rat MOB are well-developed in the PM 6 group, and that periglomerular cells in the PM 24 group show poor CR-immunoreactivity compared to those in the PM 6 group.

Aging↗

PubMed2006

Study on the ophthalmic diseases in ICR mice and BALB/c mice.

In pharmaceutical companies and research institutes, many toxicity tests are performed with laboratory animals. This study was performed to produce reference data for eye toxicity tests and to investigate the ophthalmic diseases of 408 ICR mice and 119 BALB/c mice, which are commonly used as subjects in toxicity tests. The experimental animals without clinical disorders were selected regardless of sex. The ophthalmic diseases were examined by using special ophthalmic instruments: direct ophthalmoscope, indirect ophthalmoscope, slit-lamp biomicroscope and focal illuminator. The most prevalent ocular variation within normal limits was hyaloid vessel remnant (ICR mice, 28.2%; BALB/c mice, 31.9%) and the incidence gradually decreased with age. The ocular diseases found in ICR mice were retinal degeneration (9.8%), corneal scar (4.2%), focal cataract (2.2%), anisocoria (1.2%), corneal ulcer (0.2%) and uveitis (0.2%). In BALB/c mice, corneal scar (9.2%), focal cataract (1.7%) and corneal ulcer (0.8%) were the ocular diseases found.

Animals↗

PubMed2005

Transduced human copper chaperone for Cu,Zn-SOD (PEP-1-CCS) protects against neuronal cell death.

Reactive oxygen species (ROS) contribute to the development of various human diseases. Cu,Zn-superoxide dismutase (SOD) is one of the major means by which cells counteract the deleterious effects of ROS. SOD activity is dependent upon bound copper ions supplied by its partner metallochaperone protein, copper chaperone for SOD (CCS). In the present study, we investigated the protective effects of PEP-1-CCS against neuronal cell death and ischemic insults. When PEP-1-CCS was added to the culture medium of neuronal cells, it rapidly entered the cells and protected them against paraquat-induced cell death. Moreover, transduced PEP-1-CCS markedly increased endogenous SOD activity in the cells. Immunohistochemical analysis revealed that it prevented neuronal cell death in the hippocampus in response to transient forebrain ischemia. These results suggest that CCS is essential to activate SOD, and that transduction of PEP-1-CCS provides a potential strategy for therapeutic delivery in various human diseases including stroke related to SOD or ROS.

Animals↗

PubMed2005

Human brain pyridoxal-5'-phosphate phosphatase: production and characterization of monoclonal antibodies.

We cloned and expressed human pyridoxal-5\'-phosphate (PLP) phosphatase, the coenzymatically active form of vitamin B6, in Escherichia coli using pET15b vector. Monoclonal antibodies (mAb) were generated against purified human brain PLP phosphatase in mice, and four antibodies recognizing different epitopes were obtained, one of which inhibited PLP phosphatase. The binding affinities of these four mAbs to PLP phosphatase, as determined using biosensor technology, showed that they had similar binding affinities. Using the anti-PLP phosphatase antibodies as probes, we investigated their cross-reactivities in various mammalian and human tissues and cell lines. The immunoreactive bands obtained on Western blots had molecular masses of ca. 33 kDa. Similarly fractionated extracts of several mammalian cell lines all produced a single band of molecular mass 33 kDa. We believe that these PLP phosphatase mAbs could be used as valuable immunodiagnostic reagents for the detection, identification, and characterization of various neurological diseases related to vitamin B6 abnormalities.

Animals↗

PubMed2005

The effect of P2X receptor activity on GABAA receptor-mediated inhibition in the gerbil hippocampus.

In the present study, to elucidate the effect of altered P(2)X receptor transmission on GABA(A) receptor expression and its transmission, we studied the morphological and electrophysiological responses of GABA(A) receptor in the gerbil hippocampus following P(2)X receptor antagonist/agonist treatment. Suramin or pyridoxal phosphate-6-azophenyl-2',4'-disulfonic acid (PPADS) treatment did not affect GABA(A) receptor immunoreactivities and paired-pulse responses in the gerbil hippocampus. In addition, ATP treatment did not significantly affect population spike amplitude ratios and EPSP slope ratios in the gerbil dentate gyrus. Co-application, but not pretreatment, of PPADS or suramin enhanced the effect of muscimol on paired-pulse inhibition in the dentate gyrus. In contrast, co-application of ATP reduced the effect of muscimol in the dentate gyrus. These findings indicate that the blockade of P(2)X receptor did not affect GABA(A) receptor immunoreactivities, and P(2)X receptor may modulate GABA(A) receptor-mediated inhibition when in co-activation with GABA(A) receptor. Therefore, our findings suggest that the relationship between GABA(A) receptor and P(2)X receptor may not be reciprocal, although GABA(A) receptor activity affects P(2)X receptor functionality and its expression.

Adenosine Triphosphate↗

PubMed2005

The immunoreactivity and activity of adenylate cyclase type I are changed in the hippocampal CA1 region after transient forebrain ischemia in gerbils.

Adenylate cyclase (AC) has a specific sensitivity to Ca2+/calmodulin. AC-I, one of the mediator of learning and memory, plays an important role in signal transduction underlying learning and memory function. In the present study, we found ischemia-related changes of AC-I in the hippocampal CA1 region, but not in the CA2/3 region, after 5 min of transient forebrain ischemia in gerbils. In the sham-operated group, AC-I immunoreactive neurons were detected in pyramidal and non-pyramidal cells in the hippocampus proper. AC-I immunoreactivity was significantly increased at 3 h in the CA1 region after ischemic insult. Thereafter, AC-I immunoreactivity was gradually decreased. Four days after ischemic insult, AC-I-immunoreactive CA1 pyramidal cells in the stratum pyramidale were very few due to delayed neuronal death. The results of Western blot analysis showed that changes of AC-I protein contents were similar to immunohistochemical data after ischemic insult. Gpp(NH)p-dependent AC-I activity in hippocampal CA1 region was not changed in all groups, while Ca2+/calmodulin-dependent AC-I activity in hippocampal CA1 region was significantly decreased 24 h after ischemia-reperfusion. These results suggest that the decrease of AC-I activity may be associated with impairment of neurodevelopment and neuroplasticity including learning and memory although the AC-I immunoreactivity was maintained 24 h postischemic group compared to that of the sham-operated group.

Adenylyl Cyclases↗