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S Kim

Publications and source records attributed to S Kim.

At least 901 records · Page 50Linked to original sources
PubMed1995

The effect of post-asphyxial reoxygenation with 21% vs. 100% oxygen on Na+,K(+)-ATPase activity in striatum of newborn piglets.

To compare the effect of 21% vs. 100% oxygen during post-asphyxial reoxygenation on brain cell membrane function in the striatum, 20 anesthetized, ventilated newborn piglets were studied: group 1 (normoxia, n = 5), group 2 (asphyxia, no reoxygenation, n = 5), group 3 (asphyxia followed by reoxygenation with 21% O2, n = 5), and group 4 (asphyxia followed by reoxygenation with 100% O2, n = 5). Asphyxia was induced by a stepwise reduction in FiO2 at 20 min intervals from 21% to 14%, 11%, and 8%. Following a total 60 min of asphyxia, piglets in groups 3 and 4 were recovered for 2 h with either 21% or 100% O2. Na+,K(+)-ATPase activity (mumol Pi/mg protein/h) in striatal cell membranes was 31 +/- 1, 22 +/- 2, 32 +/- 2 and 26 +/- 1 in groups 1, 2, 3 and 4, respectively. Na+,K(+)-ATPase activities in groups 2 and 4 were significantly lower than in groups 1 and 3 (p < 0.01). Piglets recovered post-asphyxia for 2 h with 21% O2 had restoration of Na+,K(+)-ATPase activity to baseline levels, while those treated with 100% O2 during recovery had persistent Na+,K(+)-ATPase inhibition of 16%. This could result from increased free radical production during reoxygenation with 100% O2 which could contribute to post-asphyxial cellular injury in the striatum.

Animals↗
PubMed1995

Crystal structure of a complement factor D mutant expressing enhanced catalytic activity.

Complement factor D is a serine protease regulated by a novel mechanism that depends on conformational changes rather than cleavage of a zymogen for expression of proteolytic activity. The conformational changes are presumed to be induced by the single natural substrate, C3bB, and to result in reversible reorientation of the catalytic center and of the substrate binding site of factor D, both of which have atypical conformations. Here we report that replacement of Ser94, Thr214, and Ser215 of factor D (chymotrypsinogen numbering has been used for comparison purposes) with the corresponding residues of trypsin, Tyr, Ser, and Trp, is sufficient to induce substantially higher catalytic activity associated with a typical serine protease alignment of the catalytic triad residues His57, Asp102, and Ser195. These results provide a partial structural explanation for the low reactivity of "resting-state" factor D toward synthetic substrates.

Amino Acid Sequence↗
PubMed1995

Blockage of the early events of mitogenic signaling by interferon-gamma in macrophages in response to colony-stimulating factor-1.

Inhibition of cell proliferation is an important biologic function of interferons (IFNs), which has been exploited in therapeutic treatment of certain hematologic malignancies. However, the molecular mechanism was not clear. We have recently shown that IFNs (alpha/beta and gamma) inhibit protein kinase C (PKC)-dependent (such as PDGF and phorbol ester) but not PKC-independent (such as epidermal growth factor) activation of Raf-1 and mitogen-activated protein kinases (MAPK/ERKs) in fibroblasts (Xu et al, Mol Cell Biol 14:8018, 1994), suggesting a novel mechanism by which IFNs execute their antiproliferative function. Monocytes/macrophages are primary targets in vivo for IFN-gamma, the major activity of macrophage-activating factor. In the present study, mechanism of IFN-gamma-induced antiproliferative action in macrophages in response to colony-stimulating factor-1 (CSF-1) has been investigated. Our results show that antiproliferative effect of IFN-gamma overrode mitogenic effect of CSF-1 and phorbol ester, as measured by early gene expression, DNA synthesis and cell proliferation. Although activation, phosphorylation, and turnover of the CSF-1 receptor and CSF-1-induced increase in diacylglycerol production remained normal, IFN-gamma blocked CSF-1-stimulated activation of mitogen-activated protein kinases, Raf-1 kinase, increase in GTP-bound Ras and tyrosine phosphorylation, and activation of protein kinase C delta (PKC-delta). PKC-delta was required for CSF-1-induced mitogenic signaling and a primary target for IFN-gamma-induced inhibition. Interestingly, although phorbol myristate acetate stimulated Ras activation, PKC-delta did not appear to be an upstream activator of Ras. These studies clearly indicated that IFN-gamma specifically inhibits PKC-delta activation, resulting in blockage of the early events of mitogenesis in macrophages in response to CSF-1.

Animals↗
PubMed1995

Solution structure of the C-terminal core domain of human TFIIB: similarity to cyclin A and interaction with TATA-binding protein.

TFIIB is an essential component of the machinery that transcribes protein-coding genes. The three-dimensional structure of the human TFIIB core domain (TFIIBc) has been determined using multidimensional heteronuclear magnetic resonance spectroscopy. The molecule consists of two direct repeats that adopt similar alpha-helical folds, conferring pseudo-twofold symmetry. An extensive, central basic surface including an amphipathic alpha helix is critical to the function of TFIIB as a bridge between the TBP-promoter complex and RNA polymerase II and associated general and regulatory transcription factors. Similarities between the TFIIBc and cyclin A folds indicate that elements of the eukaryotic cell cycle control apparatus evolved from more fundamental transcriptional control components, demonstrating a link between the transcription and cell cycle molecular machineries.

Amino Acid Sequence↗
PubMed1995

Angiotensin II type 1 receptor blockade inhibits the expression of immediate-early genes and fibronectin in rat injured artery.

BACKGROUND: Vascular injury activates various kinds of genes, including proto-oncogenes, growth factors, and extracellular matrix proteins. However, the significance of activation of these genes in neointimal formation is poorly understood. Angiotensin II type 1 (AT1) receptor antagonist is shown to prevent neointimal formation after vascular injury, although the mechanism is unclear. To understand the molecular mechanism of vascular thickening, we examined the effects of AT1 receptor blockade on the gene expression of proto-oncogenes, transforming growth factor-beta 1 (TGF-beta 1), and extracellular matrix proteins after vascular injury. METHODS AND RESULTS: Endothelial denudation of the left common carotid artery in Sprague-Dawley rats was performed with a Fogarty 2F balloon catheter. TCV-116 (10 mg.kg-1.d-1), a selective nonpeptide AT1 receptor antagonist, or vehicle was administered orally to rats from 1 day before to 14 days after balloon injury. Injured left and uninjured right common carotid arteries were removed from rats at 1, 6, and 24 hours and 3, 7, and 14 days after balloon injury. Tissue mRNA levels were measured with Northern blot analysis using specific cDNA probes and corrected for 18S ribosomal RNA value. Arterial mRNAs for c-fos, c-jun, jun B, jun D, and Egr-1 increased significantly at 1 hour after balloon injury and decreased rapidly. At 6 hours, ornithine decarboxylase (ODC) mRNA expression reached the maximal levels. TGF-beta 1 and fibronectin mRNA levels started to increase at 6 hours after injury and remained enhanced until 7 days after injury. On the other hand, collagen types I, III, and IV and laminin mRNA levels were not significantly increased over 7 days. Treatment with TCV-116 significantly inhibited the induction of mRNAs for c-fos, c-jun, Egr-1, ODC, and fibronectin in injured artery, whereas the increase in TGF-beta 1 gene expression after injury was not prevented by TCV-116. Immunohistological studies indicated that TCV-116 decreased not only the intimal thickening but also the amount of these extracellular matrix proteins in the intima. CONCLUSIONS: The results indicate that AT1 receptor blockade inhibits the induction of immediate-early genes, ODC, and fibronectin in rat injured artery. Thus, inhibition of intimal thickening by AT1 receptor blockade may be mediated at least in part by suppression of multiple genes related to cell growth and migration in the very early phase after vascular injury.

Angioplasty, Balloon↗
PubMed1995

Multiple subtypes of phospholipase C are encoded by the norpA gene of Drosophila melanogaster.

The norpA gene of Drosophila melanogaster encodes a phosphatidylinositol-specific phospholipase C that is essential for phototransduction. Besides being found abundantly in retina, norpA gene products are expressed in a variety of tissues that do not contain phototransduction machinery, implying that norpA is involved in signaling pathways in addition to phototransduction. We have identified a second subtype of norpA protein that is generated by alternative splicing of norpA RNA. The alternative splicing occurs at a single exon that is excluded from mature norpA transcripts when a substitute exon of equal size is retained. The net difference between the two subtypes of norpA protein is 14 amino acid substitutions occurring between amino acid positions 130 and 155 of the enzyme. Results from Northern analyses suggest that norpA subtype I transcripts are most abundantly expressed in adult retina, while subtype II transcripts are most abundant in adult body. Moreover, norpA subtype I RNA can be detected by the reverse transcription-polymerase chain reaction in extracts of adult head tissue but not adult body nor at earlier stages of Drosophila development. Conversely, norpA subtype II RNA can be detected by reverse transcription-polymerase chain reaction throughout development as well as in heads and bodies of adults. Furthermore, norpA subtype I RNA is easily detected in retina using tissue in situ hybridization analysis, while subtype II RNA is not detectable in retina but is found in brain. Since only norpA subtype I RNA is found in retina, we conclude that subtype I protein is utilized in phototransduction. Since norpA subtype II RNA is not found in retina but is expressed in a variety of tissues not known to contain phototransduction machinery, subtype II protein is likely to be utilized in signaling pathways other than phototransduction. The amino acid differences between the two subtypes of norpA protein may reflect the need for each subtype to interact with signaling components of different signal-generating pathways.

Alternative Splicing↗
PubMed1995

Scanning electron microscopy of the human esophagus: application to Barrett's esophagus, a precancerous lesion.

In Barrett's esophagus, metaplastic columnar epithelium replaces the normal squamous epithelium. The importance of this lesion lies in the increased incidence of adenocarcinoma of the esophagus occurring in patients with Barrett's esophagus. We characterized the surface epithelial cells of Barrett's esophagus using quantitative scanning electron microscopy. Three distinct surface cell types, in addition to the globlet cell, were recognized in Barrett's epithelium: the gastric-like cell and the intestinal-like cell, both of which were similar to normal gastric and small intestinal surface cells, respectively, by quantitative scanning electron microscopy, and the variant cell which had a range of surface features. In four biopsy specimens from the squamo-Barrett's junction in three patients, we found the distinctive cell that had features intermediate between those of squamous and columnar epithelium. On the distinctive cell's surface there are two disparate structures not normally present on the same cell in the gastrointestinal tract: microvilli (a scanning electron microscopy feature of glandular epithelium) and intercellular ridges (a scanning electron microscopy feature of squamous epithelium). The surface characteristics of this cell were almost identical to those of cells found in the transformation zone of the uterine cervix, an area in which squamous epithelium physiologically replaces columnar epithelium. Scanning electron microscopy of Barrett's esophagus has increased our understanding of this precancerous lesion by showing striking cellular heterogeneity. It has also identified the distinctive cell which may represent an intermediate step in the development of Barrett's epithelium during which the surface characteristics of two different cell types, columnar and squamous, coexist in the same cell.

Barrett Esophagus↗
PubMed1995

Phospholipase C rescues visual defect in norpA mutant of Drosophila melanogaster.

Mutations in the norpA gene of Drosophila melanogaster severely affect the light-evoked photoreceptor potential with strong mutations rendering the fly blind. The norpA gene has been proposed to encode phosphatidylinositol-specific phospholipase C (PLC), which enzymes play a pivotal role in one of the largest classes of signaling pathways known. A chimeric norpA minigene was constructed by placing the norpA cDNA behind an R1-6 photoreceptor cell-specific rhodopsin promoter. This minigene was transferred into norpAP24 mutant by P-element-mediated germline transformation to determine whether it could rescue the phototransduction defect concomitant with restoring PLC activity. Western blots of head homogenates stained with norpA antiserum show that norpA protein is restored in heads of transformed mutants. Moreover, transformants exhibit a large amount of measurable PLC activity in heads, whereas heads of norpAP24 mutant exhibit very little to none. Immunohistochemical staining of tissue sections using norpA antiserum confirm that expression of norpA protein in transformants localizes in the retina, more specifically in rhabdomeres of R1-6 photoreceptor cells, but not R7 or R8 photoreceptor cells. Furthermore, electrophysiological analyses reveal that transformants exhibit a restoration of light-evoked photoreceptor responses in R1-6 photoreceptor cells, but not in R7 or R8 photoreceptor cells. This is the strongest evidence thus far supporting the hypothesis that the norpA gene encodes phospholipase C that is utilized in phototransduction.

Animals↗
PubMed1995

Structure and expression of a smooth muscle cell-specific gene, SM22 alpha.

SM22 alpha is expressed exclusively in smooth muscle-containing tissues of adult animals and is one of the earliest markers of differentiated smooth muscle cells (SMCs). To examine the molecular mechanisms that regulate SMC-specific gene expression, we have isolated and structurally characterized the murine SM22 alpha gene. SM22 alpha is a 6.2-kilobase single copy gene composed of five exons. SM22 alpha mRNA is expressed at high levels in the aorta, uterus, lung, and intestine, and in primary cultures of rat aortic SMCs, and the SMC line, A7r5. In contrast to genes encoding SMC contractile proteins, SM22 alpha gene expression is not decreased in proliferating SMCs. Transient transfection experiments demonstrated that 441 base pairs of SM22 alpha 5'-flanking sequence was necessary and sufficient to program high level transcription of a luciferase reporter gene in both primary rat aortic SMCs and A7r5 cells. DNA sequence analyses revealed that the 441-base pair promoter contains two CArG/SRF boxes, a CACC box, and one potential MEF-2 binding site, cis-acting elements which are each important regulators of striated muscle transcription. Taken together, these studies have identified the murine SM22 alpha promoter as an excellent model system for studies of developmentally regulated, lineage-specific gene expression in SMCs.

Amino Acid Sequence↗
PubMed1995

Catalytic role of a surface loop of the complement serine protease factor D.

We have investigated the structural determinants of the unique functional properties of complement factor D by constructing and testing a series of trypsin-like mutants of the enzyme. Mutational replacement of the primary substrate-binding pocket of factor D with that of trypsin resulted in a mutant (M1) with greatly reduced proteolytic activity and slightly reduced reactivity toward small thioester substrates. Combining the M1 mutations with substitution of Tyr for Ser94, previously shown to enhance substantially both the proteolytic and esterolytic activities of factor D, produced a mutant (M2) with reactivities similar to M1. Replacement of the surface loop formed by residues 184-188 of M1 and M2 with the corresponding loop of trypsin produced mutants exhibiting one and two orders of magnitude higher esterolytic activity, respectively, than native factor D. However, the proteolytic activity of both mutants was similar to that of M1 and M2. We conclude that loop184-188 is an important determinant of the geometry of the primary specificity pocket of factor D. The low proteolytic activity of these mutants supports the proposal that the proteolytically active conformation of factor D is induced by its natural substrate, C3bB.

Amino Acid Sequence↗
PubMed1995

A fiber-optic cocaine biosensor.

A fiber-optic biosensor was developed for detection of cocaine, its metabolites, and other coca alkaloids, using a monoclonal antibody (mAb) against a derivatized benzoylecgonine (BE). The mAb was immobilized noncovalently on quartz fibers and a flow fluorometer was used to detect changes in evanescent wave fluorescence. A fluorescein (FL) conjugate of BE bound to the mAb specifically in a saturable manner and with high affinity (Kd = 7.6 nM). Cocaine or other test compounds competed with FL-BE for binding to the mAb in a concentration-dependent manner, thereby reducing the initial rate or steady-state fluorescence. Addition of cocaine to the flow buffer after reaching steady-state fluorescence enhanced the dissociation of bound FL-BE, and cocaine removal allowed fiber regeneration for multiple measurements. The detection limits for cocaine, cocaethylene, norcocaine, and BE were 5, 5, 29, and 30 ng/ml, respectively, but for ecgonine it was 4600 ng/ml and for methylecgonine it was 2000 ng/ml. Tropacocaine was detected at 10 ng/ml, but atropine was detected at 2900 ng/ml. The biosensor discriminated by 833-fold between cocaine and its stereoisomer pseudococaine. Structural features necessary for high-affinity recognition by this mAb are benzoate and 3 beta configuration, both of which are found in BE, cocaine, norcocaine, and cocaethylene.

Antibodies, Monoclonal↗