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Biomedical subjects

M J Kim

Publications and source records attributed to M J Kim.

At least 109 records · Page 6Linked to original sources
PubMed2000

Immunohistochemical study on the distribution of Bcl-2 and Bax in the central nervous system of the transgenic mice expressing a human Cu/Zn SOD mutation.

In the present study, we performed immunohistochemical studies to investigate the changes of Bcl-2 and Bax in the central nervous system of the transgenic mice expressing a human Cu/Zn SOD mutation. In contrast to the controls, a high density of Bcl-2-IR astrocytes were detected all around the gray matter of the spinal cord of the mutant transgenic mice. Bcl-2-IR astrocytes were also detected in the cerebellum and brainstem of transgenic mice. Specific immunoreactivity for Bax was seen in the spinal cord and brainstem of transgenic mice. Immunostaining for Bax was identified only in neurons and not in glial cells. Our present study demonstrated the distribution of Bcl-2 and Bax in detail using immunohistochemical methods through the central nervous system of the transgenic mice, for the first time.

Animals↗
PubMed2000

Structural analysis of multifunctional peptide motifs in human bifunctional tRNA synthetase: identification of RNA-binding residues and functional implications for tandem repeats.

Human bifunctional glutamyl-prolyl-tRNA synthetase (EPRS) contains three tandem repeats linking the two catalytic domains. These repeated motifs have been shown to be involved in protein-protein and protein-nucleic acid interactions. The single copy of the homologous motifs has also been found in several different aminoacyl-tRNA synthetases. The solution structure of repeat 1 (EPRS-R1) and the secondary structure of the whole appended domain containing three repeated motifs in EPRS (EPRS-R123) was determined by nuclear magnetic resonance (NMR) spectroscopy. EPRS-R1 consists of two helices (residues 679-699 and 702-721) arranged in a helix-turn-helix, which is similar to other RNA binding proteins and the j-domain of DnaJ, and EPRS-R123 is composed of three helix-turn-helix motifs linked by an unstructured loop. When tRNA is bound to the appended domain, chemical shifts of several residues in each repeat are perturbed. However, the perturbed residues in each repeat are not the same although they are in the same binding surface, suggesting that each repeat in the appended domain is dynamically arranged to maximize contacts with tRNA. The affinity of tRNA to the three-repeated motif was much higher than to the single motif. These results indicate that each of the repeated motifs has a weak intrinsic affinity for tRNA, but the repetition of the motifs may be required to enhance binding affinity. Thus, the results of this work gave information on the RNA-binding mode of the multifunctional peptide motif attached to different ARSs and the functional reason for the repetition of this motif.

Amino Acid Motifs↗
PubMed2000

Immunocytochemical study on the distribution of p53 in the hippocampus and cerebellum of the aged rat.

A role for p53-mediated modulation of neuronal viability has been suggested by the finding that p53 expression is increased in damaged neurons in models of ischemia and epilepsy. P53 gene upregulation precedes apoptosis in many cell types, and a potential role for this molecule in apoptosis of neurons has already been demonstrated in Alzheimer's disease. Recent studies suggest that p53-associated apoptosis may be a common mechanism of cell loss in several important neurodegenerative diseases. In the present study, we examined changes in p53-immunoreactive (IR) neurons in the brains of aged rats for the first time employing immunocytochemical and in situ hybridization methods. P53-IR neurons were found in the CA1 region of hippocampus, septal region and cerebellum in the aged rats, but there was no p53-IR cell in the brains of adult rats. In the hippocampus of the aged rat, p53-IR cells predominated in the stratum oriens and pyramidal layers, while the molecular layer contained relatively few p53-IR cells. The most prominent population of immunoreactive labeling in cerebellar cortex was localised within the cell bodies of Purkinje cells and dendrites in molecular layers. Upregulation of p53 in the Purkinje cells observed in this study suggests that significant loss of Purkinje cells with aging may be regulated with several apoptosis-controlling factors including p53 and oxidative stress mechanism. Further investigations are required to establish whether direct functional relations exist between p53 and the apoptotic neuronal death in normal aging or Alzheimer brains.

Aging↗
PubMed2000

Immunohistochemical study on the distribution of the type I and type II voltage-gated sodium channels in the gerbil cerebellum.

In this study, we investigated the distribution of the type I and type II Na+ channels in the gerbil cerebellum by immunohistochemistry. Strong uniform staining for type I was observed in the granular layer, whereas there was little evidence of concentrated labeling in the cell bodies and processes of Purkinje cells. The most intense staining for type II was observed in the cell bodies and dendrites of Purkinje cells, with a strong signal in the molecular layer. This localization study has shown clearly that the type I and type II Na+ channel subunits have differential distribution in the gerbil cerebellum, for the first time. The present study may provide useful data for the future investigations to understand the roles of voltage-gated sodium channels in neurological pathways.

Animals↗
PubMed2000

Heat shock protein 90 mediates protein-protein interactions between human aminoacyl-tRNA synthetases.

Heat shock protein 90 (hsp90) is a molecular chaperone responsible for protein folding and maturation in vivo. Interaction of hsp90 with human glutamyl-prolyl-tRNA synthetase (EPRS) was found by genetic screening, co-immunoprecipitation, and in vitro binding experiments. This interaction was sensitive to the hsp90 inhibitor, geldanamycin, and also ATP, suggesting that the chaperone activity of hsp90 is required for interaction with EPRS. Interaction of EPRS with hsp90 was targeted to the region of three tandem repeats linking the two catalytic domains of EPRS that is also responsible for the interaction with isoleucyl-tRNA synthetase (IRS). Interaction of EPRS and IRS also depended on the activity of hsp90, implying that their association was mediated by hsp90. EPRS and IRS form a macromolecular protein complex with at least six other tRNA synthetases and three cofactors. hsp90 preferentially binds to most of the complex-forming enzymes rather than those that are not found in the complex. In addition, inactivation of hsp90 interfered with the in vivo incorporation of the nascent aminoacyl-tRNA synthetases into the multi-ARS complex. Thus, hsp90 appears to mediate protein-protein interactions of mammalian tRNA synthetases.

Adenosine Triphosphate↗
PubMed2000

The mechanism of phospholipase C-gamma1 regulation.

Phospholipase C (PLC)1 hydrolyzes phosphatidylinositol 4,5-bisphosphate to generate the second messengers, inositol 1,4,5-trisphosphate (IP3) and diacylglycerol (DAG). IP3 induces a transient increase in intracellular free Ca2+, while DAG directly activates protein kinase C. Upon stimulation of cells with growth factors, PLC-gamma1 is activated upon their association with and phosphorylation by receptor and non-receptor tyrosine kinases. In this review, we will focus on the activation mechanism and regulatory function of PLC-gamma1.

Cell Division↗
PubMed2000

Immunological characterization of 130 kDa phospholipase C-beta 4 isozyme in rat cerebellar Purkinje cells.

We have developed a monoclonal antibody, beta4-22-9 that recognizes an epitope on pleckstrin homology (PH) domain (1-136 amino acids) located at the amino terminus of rat 130 kDa phospholipase C-beta4 (PLC-beta4). Tissue distribution of 130 kDa PLC-beta4 was determined with this monoclonal antibody. One hundred and thirty kiloDalton PLC-beta4 was exclusively expressed in the nervous tissues including cerebellum and cerebrum, especially at very high levels in cerebellum. The expression of 130 kDa PLC-beta4 in cerebellum was gradually increased from 5 days after birth to adulthood. In the adult cerebellum, especially high level of 130 kDa PLC-beta4 was localized on the periphery of Purkinje cells in the patch-like pattern. These data implies that 130 kDa PLC-beta4 may a role in cerebellar Purkinje cells.

Animals↗
PubMed2000

Differential expression of nicotinamide adenine dinucleotide phosphate-diaphorase in hypothalamic areas of obese Zucker rats.

Several studies have suggested that the activity of nitric oxide synthase (NOS) may be involved in the regulation of food intake in the genetically obese Zucker rats. In the present study, we investigated the expression of NOS in various hypothalamic regions of obese and lean Zucker rats using nicotinamide adenine dinucleotide phosphate (NADPH)-diaphorase histochemistry. Obese Zucker rats showed significantly lower staining intensities of NADPH-diaphorase-positive neurons in the paraventricular nucleus (PVN), lateral hypothalamic area (LHA) and ventromedial hypothalamic nucleus (VMH) than lean Zucker rats did. The differences in staining intensities between obese and lean Zucker rats were large in both the PVN and LHA, but such differences were relatively small in the VMH.

Animals↗
PubMed2000

Immunohistochemical study on the distribution of six members of the Kv1 channel subunits in the rat basal ganglia.

The differential expression of specialized voltage-gated potassium (Kv) channel subtypes in the nervous system probably reflects the wide range of functions. Although there have been previous reports in the cellular and subcellular localizations of various Kv mRNAs and proteins, the comprehensive study described here is the first in which the expression of six Kv1 channel subunits have been directly compared in the rat basal ganglia. In the present study, we have found that staining patterns of the six Kv1 channel subunits overlap in some areas of the basal ganglia, but each has a unique pattern of expression. It was noted that Kv 1.4 subunit had a strikingly high level of expression in the globus pallidus compared to the caudate-putamen. This distinct distribution formed the clear demarcations between caudate-putamen and globus pallidus. The dot-like staining pattern of Kv1 subunits was observed through the accumbens nucleus. Strong staining for Kv1.4 was observed in the cerebral peduncle, not in the subthalamic nucleus. In the substantia nigra, immunoreactivity for Kv1.4 subunit was prominent in the pars reticulata of the substantia nigra. The staining intensity for Kv1.2 was high in the pars compacta of the substantia nigra. Our immunohistochemical results may support the notion that the formation of heteromultimeric Kv channels possibly represents an important contribution to the generation of Kv channel diversity in the brain, especially in the basal ganglia.

Animals↗
PubMed2000

Identification and partial characterization of a novel hemolysin from Leptospira interrogans serovar lai.

It has been suggested that leptospiral hemolysins are important in the virulence and pathogenesis of leptospirosis. We have isolated an Escherichia coli clone carrying the 7.8kb DNA insert from a genomic library of Leptospira interrogans serovar lai by plaque hybridization using a sequence derived from the sphingomyelinase C gene (sphA) of L. borgpetersenii. The clone showed a clear beta-hemolytic zone on sheep blood agar and high hemolytic activities on both human and sheep erythrocytes in liquid assays. The clone carried at least two genes responsible for the hemolytic activities, encoded by two open reading frames of 1662 and 816 nucleotides, which are named sphH and hap-1 (hemolysis associated protein-1), respectively. The SphH showed 75% homology to the SphA at the amino acid level, and the Hap-1 showed no significant homology in major databases. Interestingly, however, E. coli cells harboring sphH did not show sphingomyelinase or phospholipase activities. Moreover, SphH-mediated hemolysis was osmotically protected by polyethylene glycol 5000, suggesting that the hemolysis is likely to be caused by pore formation on the membrane. The SphH was successfully expressed in E. coli as a histidine (His)-SphH fusion protein. Both sphH and hap-1 were highly conserved among the Leptospira species, except for the absence of sphH in non-pathogenic L. biflexa serovar patoc. We concluded that the SphH is a novel hemolysin of a pathogenic Leptospira species, which may be a putative pore-forming protein.

Amino Acid Sequence↗
PubMed2000

Direct interaction of SOS1 Ras exchange protein with the SH3 domain of phospholipase C-gamma1.

A recent report that microinjection of the SH3 domain of PLC-gamma1 could induce DNA synthesis raised the functional importance of the SH3 domain of PLC-gamma1 in mitogenic signaling. In this report, we provide evidence that SOS1, a p21Ras-specific guanine nucleotide exchange factor, directly binds to the SH3 domain of PLC-gamma1, and that the SH3 domain of PLC-gamma1 is involved in SOS1-mediated p21Ras activation. SOS1 was coprecipitated with the GST-fused SH3 domain of PLC-gamma1 in vitro. The interaction between SOS1 and the PLC-gamma1 SH3 domain is mediated by direct physical interaction. The carboxyl-terminal proline-rich domain of SOS1 is involved in the interaction with the PLC-gamma1 SH3 domain. Moreover, PLC-gamma1 could be co-immunoprecipitated with SOS1 antibody in cell lysates. From transient expression studies, we could demonstrate that the SH3 domain of PLC-gamma1 is necessary for the association with SOS1 in vivo. Intriguingly, overexpression of the SH3 domain of PLC-gamma1, lipase-inactive PLC-gamma1, or wild-type PLC-gamma1 elevated p21Ras activity and ERK activity when compared with vector transfected cells. The PLC-gamma1 mutant lacking the SH3 domain could not activate p21Ras. p21Ras activities in cell lines overexpressing either PLC-gamma1 or the SH2-SH2-SH3 domain of PLC-gamma1 were elevated about 2-fold compared to vector transfected cells. This study is the first to demonstrate that the PLC-gamma1 SH3 domain enhances p21Ras activity, and that the SH3 domain of PLC-gamma1 may be involved in the SOS1-mediated signaling pathway.

Animals↗
PubMed2000

Cellular distribution of isozymes of protein kinase C in septal olfactory epithelium of mice.

To determine the presence of protein kinase C (PKC) isozymes in the septal olfactory epithelium of mice (mSOE), western blotting and immunohistochemistry were performed using antibodies against PKC isozymes. With the exception of PKC-betaI, all of the PKC isozymes were detected in the whole lysate of septal tissue layer and apparent molecular weights for each isoform were found. PKC-alpha, PKC-gamma and PKC-epsilon were detected in the olfactory glandular cells of the lamina propria, and PKC-betaI and PKC-betaII were located in the microvillar cells. Neither novel PKC nor atypical PKC was detected in olfactory glandular cells or microvillar cells, except for PKC-epsilon. PKC-lambda was localized in the mucous layer of the mSOE. Meanwhile, PKC-delta and PKC-xi were distributed in the receptor cells in the mSOE. These data demonstrate the isoform-specific expression of PKC in mSOE and suggest a role for the novel and atypical types of PKC in olfactory transduction.

Animals↗
PubMed2000

Overexpression of phospholipase Cbeta-1 protects NIH3T3 cells from oxidative stress-induced cell death.

Oxidative stress has been implicated in a wide range of cellular damage which includes DNA oxidation, membrane lipid peroxidation, and apoptosis. In our study, we found that overexpression of PLC-beta1 in NIH3T3 fibroblasts protected them from cell death occuring in response to oxidative stress. Cell death caused by treatment with prooxidant tert-butylhydroperoxide (TBH), H2O2, or CdCl2 was considerably suppressed in PLC-beta1 overexpressed NIH/beta1-14 cells in comparison to control NIH/neo cells. However, overexpression of PLC-beta1 failed to protect the cells from toxicity by diamide or KCN. In addition, while accumulation of c-fos mRNA was observed within 30 min of TBH treatment in vector transfected NIH/neo cells, TBH-induced c-fos mRNA generation was completely suppressed in NIH/beta1-14 cells, while that of c-jun and GAPDH was not affected. These findings suggest that PLC-beta1 may play a role in process that can protect cells from oxidative stress-induced cell death.

3T3 Cells↗
PubMed2000

Rb protein down-regulates the stress-activated signals through inhibiting c-Jun N-terminal kinase/stress-activated protein kinase.

The Rb protein is the product of the retinoblastoma susceptibility gene and loss of Rb function is detected in many types of human cancers. Rb plays important roles in the regulation of cell proliferation, differentiation, senescence, and apoptotic cell death. Here we show that Rb can physically interact with c-Jun NH(2)-terminal kinase/stress-activated protein kinase (JNK/SAPK), thereby inhibiting intracellular signals mediated by JNK/SAPK. Both in vitro binding and in vitro kinase studies suggest that a carboxyl-terminal domain of Rb containing amino acids 768-928 might be crucial for inhibiting JNK/SAPK. In comparison, Rb did not affect enzymatic activity of either extracellular signal-regulated kinase 1 or p38. Ectopically expressed Rb also abrogated the apoptotic cell death induced by ultraviolet radiation or the activation of MEKK1, an upstream kinase that can stimulate the JNK/SAPK cascade. JNK/SAPK inhibition highlights a novel function of Rb, which may provide a new mechanism by which Rb regulates cell death. JNK/SAPK is a major protein kinase that can be stimulated in response to a variety of cellular stresses. Our results, therefore, suggest that Rb, by inhibiting JNK/SAPK, may act as a negative regulator in stress-activated intracellular signaling cascades.

Apoptosis↗
PubMed2000

Emissions of lead and zinc from candles with metal-core wicks.

We measured the amount of lead released from 14 different brands of candles with metal-core wicks sold in Michigan. The emissions of lead were found to range from 0.5 to 66 microg/h, and the rates for zinc were from 1.2 to 124 microg/h. It is estimated that burning four of the candles bought in Michigan for 2 h can result in airborne lead concentrations that can pose a threat to human health. In addition to inhalation of lead in the air, children get exposed to lead in candle fumes deposited on the floor, furniture and walls through their hand-to-mouth activity. Burning candles with leaded-core wick may be an important exposure route for lead that has generally been ignored.

Air Pollution, Indoor↗