Biomedical subjects

Y K Shin

Publications and source records attributed to Y K Shin.

At least 19 recordsLinked to original sources

PubMed2002

Effects of rutin and harmaline on rat reflux oesophagitis.

1. This study was aimed at evaluating the effect of rutin and harmaline (1-methyl-7-methoxy-3,4-dihydro-beta-carboline) on the development of the surgically induced reflux oesophagitis, on gastric secretion, lipid peroxidation, polymorphonucleocytes (PMNs) accumulation, superoxide and hydroxyl radical production in PMNs, cytokine [interleukin-1beta (IL-1beta), tumour necrosis factor-alpha (TNF-alpha)] production in blood and [Ca2+]i mobilization in PMNs. 2. Rutin and harmaline significantly prevented the development of reflux oesophagitis and gastric secretion. Treatments of oesophagitis rats with rutin and harmaline inhibited lipid peroxidation, and myeloperoxidase (MPO) in the oesophagus in comparison with untreated rats. 3. Superoxide anion and hydrogen peroxide production in 1 microm formylmethionylleucylphenylalanine (fMLP)- or 0.1 microg ml-1N-phorbol 12-myristate 13-acetate (PMA)-activated PMNs was inhibited by rutin and harmaline in a dose-dependent fashion. Rutin and harmaline effectively scavenged the hydroxyl radical and hydrogen peroxide. Treatments of oesophagitis rats with rutin and harmaline inhibited IL-1beta production in the oesophagus in comparison with untreated rats, but TNF-alpha production was not affected by rutin and harmaline. The fMLP-induced elevation of [Ca2+]i was inhibited by rutin. 4. The results of this study suggest that rutin and harmaline may have beneficial protective effects against reflux oesophagitis by the inhibition of gastric acid secretion, oxidative stress, inflammatory cytokine production (i.e. IL-1beta), and intracellular calcium mobilization in PMNs in rats.

Animals↗

PubMed2001

Effects of ATP on TTX-sensitive and TTX-resistant sodium currents in rat sensory neurons.

Differential effects of ATP on tetrodotoxin-sensitive (TTX-S) and tetrodotoxin-resistant (TTX-R) sodium currents in rat dorsal root ganglion neurons were studied using the whole-cell variation of path-clamp technique. Currents were evoked by step depolarizations to 0 mV from a holding potential of -80 mV. ATP suppressed TTX-S sodium currents while it increased TTX-R sodium currents. The effects were concentration-dependent and were reversible upon washout with ATP-free external solution. ATP-gamma-S, a hydrolysis-resistant ATP analog, also affected two types of sodium currents similarly to ATP, excluding the possibility that the effects were caused by the products of ATP hydrolysis, namely adenosine. ATP by modulating sodium currents may exert profound effects on the transmission of sensory information such as nociception.

Adenosine Triphosphate↗

PubMed2001

Self-assembly of influenza hemagglutinin: studies of ectodomain aggregation by in situ atomic force microscopy.

We have used in situ tapping mode atomic force microscopy (AFM) to study the structural morphology of two fragments of the influenza hemagglutinin protein bound to supported bilayers. The two proteins that we studied are the bromelain-cleaved hemagglutinin (BHA), corresponding to the full ectodomain of the hemagglutinin protein, and FHA2, the 127 amino acid N-terminal fragment of the HA2 subunit of the hemagglutinin protein. While BHA is water soluble at neutral pH and is known to bind to membranes via specific interactions with a viral receptor, FHA2 can only be solubilized in water with an appropriate detergent. Furthermore, FHA2 is known to readily bind to membranes at neutral pH in the absence of a receptor. Our in situ AFM studies demonstrated that, when bound to supported bilayers at neutral pH, both these proteins are self-assembled as single trimeric molecules. In situ acidification resulted in further lateral association of the FHA2 without a large perturbation of the bilayer. In contrast, BHA remained largely unaffected by acidification, except in areas of exposed mica where it is aggregated. Remarkably, these results are consistent with previous observations that FHA2 promotes membrane fusion while BHA only induces liposome leakage at low pH. The results presented here are the first example of in situ imaging of the ectodomain of a viral envelope protein allowing characterization of the real-time self-assembly of a membrane fusion protein.

Bromelains↗

PubMed2001

The 1-127 HA2 construct of influenza virus hemagglutinin induces cell-cell hemifusion.

Conformational changes in the HA2 subunit of influenza hemagglutinin (HA) are coupled to membrane fusion. We investigated the fusogenic activity of the polypeptide FHA2 representing 127 amino-terminal residues of the ectodomain of HA2. While the conformation of FHA2 both at neutral and at low pH is nearly identical to the final low-pH conformation of HA2, FHA2 still induces lipid mixing between liposomes in a low-pH-dependent manner. Here, we found that FHA2 induces lipid mixing between bound cells, indicating that the "spring-loaded" energy is not required for FHA2-mediated membrane merger. Although, unlike HA, FHA2 did not form an expanding fusion pore, both acidic pH and membrane concentrations of FHA2, required for lipid mixing, have been close to those required for HA-mediated fusion. Similar to what is observed for HA, FHA2-induced lipid mixing was reversibly blocked by lysophosphatidylcholine and low temperature, 4 degrees C. The same genetic modification of the fusion peptide inhibits both HA- and FHA2-fusogenic activities. The kink region of FHA2, critical for FHA2-mediated lipid mixing, was exposed in the low-pH conformation of the whole HA prior to fusion. The ability of FHA2 to mediate lipid mixing very similar to HA-mediated lipid mixing is consistent with the hypothesis that hemifusion requires just a portion of the energy released in the conformational change of HA at acidic pH.

Animals↗

PubMed2001

Fungicidal effect of antimicrobial peptide, PMAP-23, isolated from porcine myeloid against Candida albicans.

The antifungal activity and mechanism of a 23-mer peptide, PMAP-23, derived from pig myeloid was investigated. PMAP-23 displayed strong antifungal activity against yeast and mold. To investigate the antifungal mechanism of PMAP-23, fluorescence activated flow cytometry and confocal laser scanning microscopy were performed. Candida albicans treated with PMAP-23 showed higher fluorescence intensity by propidium iodide(PI) staining, which was similar to that of Melittin than untreated cells. Confocal microscopy showed that the peptide was located in the plasma membrane. The action of peptides against fungal cell membranes was examined by treating prepared protoplasts of C. albicans with the peptide and lipid vesicle titration test. The result showed that the peptide prevented the regeneration of fungal cell walls and induced release of the fluorescent dye trapped in the artificial membrane vesicles, indicating that the peptide exerts its antifungal activity by acting on the plasma lipid membrane.

Amino Acid Sequence↗

PubMed2001

CD99 regulates the transport of MHC class I molecules from the Golgi complex to the cell surface.

The down-regulation of surface expression of MHC class I molecules has recently been reported in the CD99-deficient lymphoblastoid B cell line displaying the characteristics of Hodgkin's and Reed-Sternberg phenotype. Here, we demonstrate that the reduction of MHC class I molecules on the cell surface is primarily due to a defect in the transport from the Golgi complex to the plasma membrane. Loss of CD99 did not affect the steady-state expression levels of mRNA and protein of MHC class I molecules. In addition, the assembly of MHC class I molecules and the transport from the endoplasmic reticulum to the cis-Golgi occurred normally in the CD99-deficient cells, and no difference was detected between the CD99-deficient and the control cells in the pattern and degree of endocytosis. Instead, the CD99-deficient cells displayed the delayed transport of newly synthesized MHC class I molecules to the plasma membrane, thus causing accumulation of the molecules within the cells. The accumulated MHC class I molecules in the CD99-deficient cells were colocalized with alpha-mannosidase II and gamma-adaptin in the Golgi compartment. These results suggest that CD99 may be associated with the post-Golgi trafficking machinery by regulating the transport to the plasma membrane rather than the endocytosis of surface MHC class I molecules, providing a novel mechanism of MHC class I down-regulation for immune escape.

12E7 Antigen↗

PubMed2001

Expression of leukemia-associated antigen, JL1, in bone marrow and thymus.

The identification of immunophenotypic markers with restricted expression has long been a critical issue in diagnostic and therapeutic advances for acute leukemias. We previously developed a monoclonal antibody against a new thymocyte surface antigen, JL1, and showed that JL1 is expressed in the majority of acute leukemia cases. In this study, using multiparameter flow cytometric analyses, we found that JL1 was uniquely expressed in subpopulations of normal bone marrow (BM) cells, implying the association of JL1 with the differentiation and maturation process. Although CD34(+) CD10(+) lymphoid precursors and some of maturing myeloid cells express JL1, neither CD34(+) CD38(-/lo) nor CD34(+) AC133(+) noncommitted pluripotent stem cells do. As for the myeloid precursors, CD34(+) CD33(+) cells do not express JL1. During lymphopoiesis, JL1 on the earliest lymphoid precursors disappear in the CD20(+) sIgM(+) stage of B-cell development or after CD1a down-regulation in thymocytes. Despite the highly restricted expression of JL1 in normal BM cells, most of the leukemias express JL1 irrespective of their immunophenotypes. These results indicate that JL1 is not only a novel differentiation antigen of hematopoietic cells, but also a leukemia-associated antigen. Therefore, we suggest that JL1 be a candidate molecule in acute leukemia for the diagnosis and immunotherapy that spares the normal BM stem cells.

Adult↗

PubMed2001

The neuronal t-SNARE complex is a parallel four-helix bundle.

Assembly of the soluble N-ethylmaleimide sensitive factor attachment protein receptor (SNARE) complex is an essential step for neurotransmitter release in synapses. The presynaptic plasma membrane associated proteins (t-SNAREs), SNAP-25 (synaptosome-associated protein of 25,000 Da) and syntaxin 1A may form an intermediate complex that later binds to vesicle-associated membrane protein 2 (VAMP2). Using spin labeling electron paramagnetic resonance (EPR), we found that the two t-SNARE proteins assemble into a parallel four-helix bundle that consists of two identical syntaxin 1A components and the N-terminal and C-terminal domains of SNAP-25. Although the structure is generally similar to that of the final SNARE complex, the middle region of the helical bundle appears more flexible in the t-SNARE complex. Such flexibility might facilitate interactions between VAMP2 and the t-SNARE complex.

Electron Spin Resonance Spectroscopy↗

PubMed2001

Sphingomonas aquatilis sp. nov., Sphingomonas koreensis sp. nov., and Sphingomonas taejonensis sp. nov., yellow-pigmented bacteria isolated from natural mineral water.

The taxonomic status of four strains from several mineral water sources in Taejon, Korea, was examined. The DNA base compositions of these strains ranged from 62 to 66 mol %. All of the strains contained sphingolipids and ubiquinone 10 as the main respiratory quinone. The cellular fatty acids of these strains included octadecenoic acid, hexadecanoic acid and 2-hydroxymyristic acid, without 3-hydroxy fatty acids. On the basis of morphological, physiological and chemotaxonomic characteristics, together with 16S rDNA sequence comparison and DNA-DNA reassociation data, three new species of the genus Sphingomonas are proposed for the strains isolated from natural mineral water: Sphingomonas aquatilis sp. nov. for strain JSS7T (= KCTC 2881T = KCCM 41067T), Sphingomonas koreensis sp. nov. for strains JSS26T (= KCTC 2882T = KCCM 41069T) and JSS28 (= KCTC 2883 = KCCM 41066) and Sphingomonas taejonensis sp. nov. for strain JSS54T (= KCTC 2884T = KCCM 41068T).

Base Composition↗

PubMed2001

Candida thermophila sp. nov., a novel thermophilic yeast isolated from soil.

Yeast strain Y94T, which is capable of growth at high temperature, was isolated from soil in Korea. Characteristics of the strain include asexual reproduction by multilateral budding, the absence of extracellular starch-like compounds, a negative Diazonium blue B colour reaction, and the absence of arthrospores, ballistoconidia and ascospores; the strain can therefore be placed in the genus Candida. A maximum growth temperature of 50-51 degrees C, along with certain other physiological characteristics, and a unique 26S rDNA partial sequence separate this strain from other ascomycetous yeasts. Taken together, these results suggest that the strain is a novel species and the name Candida thermophila sp. nov. (type strain is Y94T = JCM 10994T = KCCM 50661T) is proposed.

Candida↗

PubMed2001

Frequency of acyclovir-resistant herpes simplex virus in clinical specimens and laboratory isolates.

The proportion of acyclovir (ACV)-resistant herpes simplex virus (HSV) isolates in clinical specimens and laboratory isolates was determined. HSV isolates in clinical specimens and laboratory isolates were cultured in the absence or presence of 5 microg of ACV per ml. The frequency of ACV-resistant HSV was calculated as (average virus titer in the wells with ACV)/(average virus titer in the wells without ACV). The mutation frequency of HSV type 1 isolates in clinical samples (directly from patient lesions) was 7.5 x 10(-4) +/- 2.5 x 10(-4) (mean +/- standard error), and that of HSV type 2 isolates was 15.0 x 10(-4) +/- 4.6 x 10(-4). The mutation frequencies of isolates derived in the laboratory from these clinical samples were very similar. Similarly, the 50% inhibitory concentrations for isolates in clinical samples and laboratory isolates were identical. The frequencies of ACV-resistant HSV types 1 and 2 were in a narrow range of 7.5 x 10(-4) to 15.0 x 10(-4) among isolates in clinical specimens and did not change for laboratory-derived pools of viral isolates.

Acyclovir↗

PubMed2001

LMP1-induced downregulation of CD99 molecules in Hodgkin and Reed-Sternberg cells.

Hodgkin's and Reed-Sternberg (H-RS) cells are morphological hallmarks of Hodgkin's disease (HD). So far, several characteristics frequently seen in H-RS cells from different origins have been described, such as the high expression of Epstein-Barr virus latent membrane protein 1 (LMP1), the elevation of NF-kappaB activity, and the aberrant expression of molecules such as CD15, CD30, and CD99. Despite extensive studies on the nature of H-RS cells, the molecular mechanism by which H-RS cells are generated remained elusive. Recently, the forced down-regulation of CD99 was reported to induce typical H-RS phenotypes in vitro in a B cell line. Furthermore, it was revealed that LMP1 markedly reduces the CD99 expression at the transcriptional level. Since the presence of LMP1 is known to be associated with the H-RS cell formation, the data provide a possibility of linkage between LMP1 and HD via CD99, thus suggesting that, at least in part, the loss of CD99 may play a critical role in the pathogenic sequence to the formation of H-RS cells in HD. In this review, the role of CD99 in the generation of H-RS cells and its molecular mechanism will be suggested.

12E7 Antigen↗

PubMed2000

Arachidonic acid and nonsteroidal anti-inflammatory drugs induce conformational changes in the human prostaglandin endoperoxide H2 synthase-2 (cyclooxygenase-2).

By using the technique of site-directed spin labeling combined with EPR spectroscopy, we have observed that binding of arachidonic acid and nonsteroidal anti-inflammatory drugs induces conformational changes in the human prostaglandin endoperoxide H(2) synthase enzyme (PGHS-2). Line shape broadening resulting from spin-spin coupling of nitroxide pairs introduced into the membrane-binding helices of PGHS-2 was used to calculate the inter-helical distances and changes in these distances that occur in response to binding various ligands. The inter-residue distances determined for the PGHS-2 holoenzyme using EPR were 1-7.9 A shorter than those of the crystal structure of the PGHS-2 holoenzyme. However, inter-helical distances calculated and determined by EPR for PGHS-2 complexed with arachidonic acid, flurbiprofen, and SC-58125 were in close agreement with those obtained from the cognate crystal structures. These results indicate that the structure of the solubilized PGHS-2 holoenzyme measured in solution differs from the crystal structure of PGHS-2 holoenzyme obtained by x-ray analysis. Furthermore, binding of ligands induces a conformational change in the holo-PGHS-2, converting it to a structure similar to those obtained by x-ray analysis. Proteolysis protection assays had previously provided circumstantial evidence that binding of heme and non-steroidal anti-inflammatory drugs alters the conformation of PGHS, but the present experiments are the first to directly measure such changes. The finding that arachidonate can also induce a conformational change in PGHS-2 was unexpected, and the magnitude of changes suggests this structural flexibility may be integral to the cyclooxygenase catalytic mechanism.

Animals↗

PubMed2000

Light-induced rotation of a transmembrane alpha-helix in bacteriorhodopsin.

Spin labeling EPR spectroscopy has been used to characterize light-induced conformational changes of bacteriorhodopsin (bR). Pairs of nitroxide spin labels were attached to engineered cysteine residues at strategic positions near the cytoplasmic ends of transmembrane alpha-helices B, F, and G in order to monitor distance changes upon light activation. The EPR analysis of six doubly labeled bR mutants indicates that the cytoplasmic end of helix F not only tilts outwards, but also rotates counter-clockwise during the photocycle. The direction of the rotation of helix F is the opposite of the clockwise rotation previously reported for bovine rhodopsin. The opposite chirality of the F helix rotation in the two systems is perhaps related to the differences in the cis-trans photoisomerization of the retinal in the two proteins. Using time-resolved EPR, we monitored the rotation of helix F also in real time, and found that the signal from the rotation arises concurrently with the reprotonation of the retinal Schiff base.

Animals↗

PubMed2000

Block of sodium currents in rat dorsal root ganglion neurons by diphenhydramine.

To elucidate the local anesthetic mechanism of diphenhydramine, its effects on tetrodotoxin-sensitive (TTX-S) and tetrodotoxin-resistant (TTX-R) sodium currents in rat dorsal root ganglion (DRG) neurons were examined by the whole-cell voltage clamp method. Diphenhydramine blocked TTX-S and TTX-R sodium currents with K(d) values of 48 and 86 microM, respectively, at a holding potential of -80 mV. It shifted the conductance-voltage curve for TTX-S sodium currents in the depolarizing direction but had little effect on that for TTX-R sodium currents. Diphenhydramine caused a shift of the steady-state inactivation curve for both types of sodium currents in the hyperpolarizing direction. The time-dependent inactivation became faster and the recovery from the inactivation was slowed by diphenhydramine in both types of sodium currents. Diphenhydramine produced a profound use-dependent block when the cells were repeatedly stimulated with high-frequency depolarizing pulses. The use-dependent block was more pronounced in TTX-R sodium currents. The results show that diphenhydramine blocks sodium channels of sensory neurons similarly to local anesthetics.

Animals↗

PubMed2000

Amitriptyline modulation of Na(+) channels in rat dorsal root ganglion neurons.

The effects of amitriptyline, a tricyclic antidepressant, on tetrodotoxin-sensitive and tetrodotoxin-resistant Na(+) currents in rat dorsal root ganglion neurons were studied using the whole-cell patch clamp method. Amitriptyline blocked both types of Na(+)currents in a dose-and holding potential-dependent manner. At the holding potential of -80 mV, the apparent dissociation constants (K(d)) for amitriptyline to block tetrodotoxin-sensitive and tetrodotoxin-resistant Na(+) channels were 4.7 and 105 microM, respectively. These values increased to 181 and 193 microM, respectively, when the membrane was held at a potential negative enough to remove the steady-state inactivation. Amitriptyline dose-dependently shifted the steady-state inactivation curves in the hyperpolarizing direction and increased the values of the slope factors for both types of Na(+) channels. The voltage dependence of the activation of both types of Na(+) channels was shifted in the depolarizing direction. It was concluded that amitriptyline blocked the two types of Na(+) channels in rat sensory neurons by modulating the activation and the inactivation kinetics.

Amitriptyline↗

PubMed2000

ATP evokes different currents in TTX-sensitive and TTX-resistant cells of dorsal root ganglia.

The relationship between the level of expression of tetrodotoxin-sensitive (TTX-S) and tetrodotoxin-resistant (TTX-R) sodium currents and the occurrence of two kinetically different ATP-induced currents in rat dorsal root ganglion neurons was studied. ATP evoked two distinct types of currents, one with fast activation and desensitization (I-fast) and the other with slow and persistent development (I-slow). In all TTX-S cells which expressed solely TTX-S sodium currents ATP evoked I-fast. The other cells expressed a variable proportion of TTX-S and TTX-R sodium currents. Only 15% of these TTX-R+S cells responded to ATP with I-fast. I-slow was evoked in both cell types but the magnitude of response was much greater in TTX-R+S cells. This result suggests that a different array of ion channels is equipped in different types of sensory neurons.

Adenosine Triphosphate↗

PubMed2000

Inhibitory action of thimerosal, a sulfhydryl oxidant, on sodium channels in rat sensory neurons.

The effects of thimerosal, a sulfhydryl oxidizing agent, on tetrodotoxin-sensitive (TTX-S) and tetrodotoxin-resistant (TTX-R) sodium channels in rat dorsal root ganglion neurons were studied using the whole-cell patch clamp technique. Thimerosal blocked the two types of sodium channels in a dose-dependent manner. The inhibitory effect of thimerosal was much more pronounced in TTX-R sodium channels than TTX-S sodium channels. The effect of thimerosal was irreversible upon wash-out with thimerosal-free external solution. However, dithiothreitol, a reducing agent, partially reversed it. Thimerosal shifted the steady-state inactivation curves for both types of sodium channels in the hyperpolarizing direction. The voltage dependence of activation of both types of sodium channels was shifted in the depolarizing direction by thimerosal. The inactivation rate in both types of sodium channels increased after thimerosal treatment. All these effects of thimerosal would add up to cause a depression of sodium channel function leading to a diminished neuronal excitability.

Animals↗