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Biomedical subjects

J S Shin

Publications and source records attributed to J S Shin.

At least 55 records · Page 3Linked to original sources
PubMed1999

Cloning and characterization of the chloroplast elongation factor EF-Tu cDNA of Oryza sativa L.

From the rice leaf cDNA library, we have cloned a cDNA encoding rice chloroplast translational elongation factor EF-Tu (tufA). The rice tufA cDNA clone contains 1678 nucleotides and codes for a 467 amino acid protein including a putative chloroplast transit peptide of 59 amino acid residues. The predicted molecular mass of the mature protein is approximately 45 kDa. This cDNA clone contains the 61 nucleotides of the 5' untranslated region (UTR) and the 213 nucleotides of 3' UTR. Amino acid sequence identity of the rice tufA with the mature chloroplast EF-Tu proteins of tobacco, pea, arabidopsis, and soybean ranges from 83% to 86%. The deduced polypeptide of the rice tufA cDNA contains GTP binding domains in its N-terminal region and chloroplast EF-Tu signature regions in the C-terminal region. The rice tufA appears to exist as a single copy gene, although its homologues of maize and oat exist as multiple copy genes. The rice tufA gene is located in chromosome 1 and is more highly expressed in the leaf than in root tissue.

Amino Acid Sequence↗
PubMed1999

Asymmetric synthesis of chiral amines with omega-transaminase.

The asymmetric synthesis of chiral amines using prochiral ketones was carried out with (S)-specific omega-transaminase (omega-TA) from Vibrio fluvialis JS17. This reaction is inhibited severely by both products, (S)-amine and deaminated ketone. In addition, thermodynamic equilibrium strongly favored the reverse reaction. L-Alanine proved to be the best amino donor based on easy removal of the products. Optimal pH of the reactions with both whole cells and cell-free extract was 7. Amino acceptor reactivities of ketone substrates and reaction profiles of the asymmetric synthesis showed that the initial rate as well as the reaction yield were lower when the resulting (S)-amine from a prochiral ketone substrate was a more reactive amino donor. The yield could be increased dramatically by removing pyruvate, which is a more inhibitory product than (S)-alpha-methylbenzylamine [(S)-alpha-MBA] when acetophenone and L-alanine are used as an amino acceptor and donor, respectively. The removal of pyruvate was carried out by incorporating lactate dehydrogenase (LDH) in cell-free extract or by using whole cells. The whole cell reaction yielded a much better result. When 25 mM benzylacetone and 30 mM acetophenone were used as an amino acceptor with 300 mM L-alanine, 90.2% and 92.1% of the reaction yields after 1 day were obtained with whole cells, respectively. Enantiomeric excesses of both (S)-alpha-MBA and (S)-1-methyl-3-phenylpropylamine [(S)-MPPA] were all above 99%.

Alanine↗
PubMed1999

Gene silencing-mediated resistance in transgenic tobacco plants carrying potato virus Y coat protein gene.

Unlike other pathogens, plant viruses are hardly controlled by chemical agents. Potato virus Y (PVY) is distributed around the world, and causes a great loss economically. In an attempt to minimize the damage by viruses, the PVY coat protein (CP) gene was introduced into tobacco by Agrobacterium-mediated transformation. A significant proportion of the transgenic plants displayed resistance to PVY and showed substantially decreased CP transgene expression at both protein and steady-state mRNA levels compared to susceptible transgenic or nontransgenic plants. A resistant plant was selected and self-fertilized for several generations until T4 progenitor lines were obtained. Most of these T4 plants accumulated extremely low levels of CP protein and steady-state mRNA, and exhibited almost complete resistance to PVY. DNA gel blot analysis revealed that the transgenic plants typically had two or three copies of the transgene. These results are characteristic of pathogen-derived resistance, in which the resistance against virus is the consequence of post-transcriptional gene silencing directed by homologous transgenes. To uncover factors that may play roles in gene silencing, sequences in the 3' part of the transcribed region of the CP gene were transcribed in vitro and the RNA fragments were incubated with cell extracts from transgenic plants. A ribonuclease activity was detected that appeared to be specific for this transcript in the PVY-resistant transgenic plants.

Capsid↗
PubMed1999

Isolation and characterization of a cDNA encoding the cysteine proteinase inhibitor, induced upon flower maturation in carnation using suppression subtractive hybridization.

The suppression subtractive hybridization (SSH) method was used to isolate differentially expressed genes during carnation flower maturation. Five cDNA clones, designated as carnation flower maturation-induced (CFMI), were verified as flower maturation-induced cDNAs. Sequence analysis of five CFMI (CFMI-5, CFMI-6, CFMI-7, CFMI-9, and CFMI-10) clones revealed that one of the clones, CFMI-5, showed high sequence similarity to the cysteine proteinase inhibitor gene, predicted to be involved in flower maturation. The full length cDNA clone CFMI-5 was 531 nucleotides (nts) long and consisted of an open reading frame of 294 nucleotides, encoding a 98 amino acid protein, 12 nucleotides of 5'-untranslated region and 3'-untranslated region (225 nts) with a poly(A)+ tail. The predicted CFMI-5 amino acid sequence had a conserved sequence Gln-Val-Val-Ala-Gly, which corresponds to the active site of proteinase inhibition. Northern blot analysis revealed tissue-specific expression of CFMI-5 transcripts, as the transcripts were expressed preferentially in petals and styles. A PCR-based cDNA subtraction method, termed suppression subtractive hybridization, was identified as a rapid method to screen differentially expressed genes in a short time.

Amino Acid Sequence↗
PubMed1999

Expression of fas ligand in human hepatoma cell lines: role of hepatitis-B virus X (HBX) in induction of Fas ligand.

It has been postulated that tumor cells expressing Fas ligand (FasL) can evade immune surveillance by inducing apoptosis in T cells expressing Fas. In this study, we investigated FasL expression in 13 human hepatoma cell lines. Strong FasL expression was detected by reverse transcription-polymerase chain reaction or immunofluorescence in Hep G2.2.15, in which the hepatitis-B-virus (HBV) genome was transfected, and in SNU-354, which showed HBx transcripts. To determine the biological activity of FasL, Hep G2.2. 15 was co-cultured with MOLT-4, T-cell-leukemia cells. Hep G2.2.15 induced apoptosis in MOLT-4 and this was inhibited by the antagonistic anti-Fas antibody, ZB4. For further analysis of the role of HBx in the induction of FasL, PLC/PRF/5 cells were transfected transiently with the HBV genome, or HBx, or the frameshift mutant of HBx. In PLC/PRF/5 cells transfected with the HBV genome or HBx but not in cells transfected with the frameshift mutant of HBx, FasL expression was detected. Our data suggest that HBx plays a role in the induction of FasL in hepatoma cells and in the escape from immune surveillance.

Apoptosis↗
PubMed1999

Synthesis and hypoglycemic effect of chrysin derivatives.

A series of 18 chrysin derivatives, prepared by alkylation and condensation, were fully characterized by NMR and other techniques and tested in vivo against the diabetes mellitus. Several modified compounds especially those with propyl, butyl, octyl and tolyl groups were found to have hypoglycemic effect on diabetec mice in spite of the fact that chrysin itself had inhibited insulin release by 40-60%. None of the test animals died at the maximum dose 500mg/kg and did not cause any significant change in general feature, water and food consumption, body weight and organ weight when we examined the acute oral toxicity of those compounds having significant hypoglycemic effect.

Animals↗
PubMed1999

Particle bombardment mediated transformation and GFP expression in the moss Physcomitrella patens.

There are few plants facilitated for the study of development, morphogenesis and gene expression at the cellular level. The moss Physcomitrella patens can be a very useful plant with several advantages: simple life cycle containing a major haploid gametophyte stage, easy manipulation, small genome size (6 x 10(8) bp) and high similarities with higher plants. To establish the transformation system of mosses as a model for basic plant research, a series of experiments were performed. Mosses were cultured in cellophane overlaid BCD media, transformed by particle bombardment and selected by the choice of appropriate antibiotics. Initial transformants appeared 8 d or 14 d after selection, showing different sensitivities toward the antibiotics used. Heat treatment during the preparation of particles revealed that denaturing the DNA enabled a more efficient way to deliver a transgene into the chromosome. This was proven by the increase in the number of transformants by five times in the plants with denatured DNA. In the test for the repairing capacity of mosses, 154 and 195 transformants survived from 1 d and 3 d incubations, respectively, indicating that a longer period of incubation seemed to be recommendable for better survival. The selected transformants were further analyzed at the DNA and expression level. Transformed genes were confirmed by PCR where all the transformants showed the expected size of amplification. Histochemical beta-glucuronidase (GUS) and green fluorescent protein (GFP) expression also confirmed the integration of exogenous DNA. In a comparison of the two different forms of GFP, soluble-modified GFP (smGFP) expressed stronger signals than modified GFP (mGFP) due to its improved solubility. Confirmation of the transgene in the chloroplast transformation has improved the applicability of moss as a model system for the study of basic biological researches.

Blotting, Southern↗
PubMed1999

Fertilization and in vitro development of porcine oocytes following intracytoplasmic injection of round spermatid or round spermatid nuclei.

The objective of this study was to determine fertilization rates and developmental ability of porcine oocytes following injection of round spermatid and round spermatid nucleus with artificial activation either 2 h before or immediately after injection. Electrical stimulation at 2 h before spermatid injection significantly increased the incidence of normal fertilization compared with that following injection without stimulation or with stimulation immediately after injection. Incidences of formation of 2 pronuclei and of apposition were not different in oocytes following intracytoplasmic spermatid and spermatid nucleus injection. Chromosome analysis revealed that most oocytes were diploid either following round spermatid or round spermatid nucleus injection. There was no diploid set of chromatin in oocytes at 20 h following sham injection. At 6 d following injection blastocoele formation was seen in the oocytes following round spermatid (25%) and round spermatid nucleus injection (27%). However, none of the oocytes developed to the blastocyst stage 6 d following sham injection. The average cell numbers of blastocysts 8 d after injection of spermatid and spermatid nucleus were 99 and 87, respectively. These results suggest that electrical stimulation before injection enhances the incidence of fertilization following round spermatid injection in the pig. Our study also indicates that either the round spermatid or it's nucleus can be used to produce viable embryos by injection into unfertilized porcine oocytes.

Animals↗
PubMed1999

Bacteria-host cell interaction mediated by cellular cholesterol/glycolipid-enriched microdomains.

Gram negative bacterial infection is a leading cause of fatality and is attributed, at least in part, to the bacteria's capacity to persist in the host in spite of appropriate antibiotic therapy. It has been suggested that bacteria evade antibiotics by hiding within host cells. We sought to investigate this important aspect of infections in mast cells, which are inflammatory cells found in close proximity to the host-environment interface and which have recently been reported to play a crucial role in the early innate immune response to bacteria. We examined mast cell interactions with FimH-expressing E. coli, one of the major opportunistic pathogens of humans. We determined that in serum free conditions, these bacteria were able to trigger mast cell uptake without loss of bacterial viability. CD48, a mannose containing GPI (glycosylphosphatidylinositol)-linked molecule was found to be the receptor of FimH-expressing E. coli in mouse mast cells. We found that the internalization via CD48 was blocked by filipin, a cholesterol binding drug known to disrupt cholesterol/glycolipid-enriched microdomains and the bacteria-encasing vacuoles were rich in cholesterol inside cells. Interestingly, we found that mast cells subsequently expelled majority of the intracellular bacteria in 24 hours. This expulsion process was blocked by lovastatin/cyclodextrin treatment, which is known to inhibit cellular trafficking of cholesterol/glycolipid-enriched microdomains. Thus, the bacterial entry into and expulsion from mast cells were critically dependent on cholesterol/glycolipid-enriched microdomains, which represents a novel mode of tussle between the pathogen and the mast cell occurring in opsonin deficient sites in the body or even at other sites in naive or immunocompromised hosts which have low systemic levels of E. coli specific antibody.

Adhesins, Bacterial↗
PubMed1999

Inhibitory effects of higenamine on dopamine content in PC12 cells.

The inhibitory effects of (+/-)-higenamine, a benzylisoquinoline alkaloid, on dopamine biosynthesis in PC12 cells were investigated. Higenamine decreased the intracellular dopamine content dose-dependently and showed 55.2% inhibition of dopamine content in PC12 cells at a concentration of 20 microM with 24 h incubation. The IC50 value of higenamine was 18.2 microM. Dopamine content was lowered and reached minimal level at 12-24 h after exposure to higenamine. In this condition, tyrosine hydroxylase, the rate-limiting enzyme of catecholamine biosynthesis, was also inhibited by the treatment of higenamine in PC12 cells (21.9% inhibition at 20 microM). Higenamine at 20 microM lowered the intracellular Ca2+ concentration by 33.1% inhibition relative to control in PC12 cells. These results suggest that the inhibition of tyrosine hydroxylase activity by higenamine might partially contribute to the decrease in dopamine content in PC12 cells.

Alkaloids↗
PubMed1999

Monoclonal antibodies with various reactivity to p58 killer inhibitory receptors.

Human natural killer (NK) cells have receptors that bind to HLA class I molecules. Among these receptors, p58 and p50 bind to HLA-C. P58 belongs to the killer inhibitory receptor (KIR) and p50 belongs to the killer activatory receptor (KAR). Previously, we obtained three recombinant p58 KIR or p50 KAR proteins, KAR-K1 (KIR2DS4), KIR-K6 (KIR2DL1), and KIR-K7 (KIR2DL3). In this study, we produced and characterized seven monoclonal antibodies (MAbs) to the p58 KIR and p50 KAR proteins. The MAbs were classified in three groups according to their antigen-binding specificity: 5IB103 and 26ID707 were KAR-K1-specific; A809, 190IIC311, and 197IIC611 were KIR-K7-specific; while A210 and A803g bound to all three recombinant proteins. The MAbs reactive to KIR-K7 bound to the gamma3 domain among two immunoglobulin (Ig) domains of KIR-K7. Immunofluorescence staining and flow cytometric analysis with A803g showed reactivity to about 10% of peripheral blood mononuclear cells and 35% of purified natural killer cells. Double immunofluorescence staining with A803g and anti-CD56 Ab showed that CD56 and p58 or p50 were expressed on NK cells in a mutually exclusive way. We also investigated T-cell markers in A803g+ cells. A803g+ T cells were almost CD8+ cells. MAbs produced in this study can be utilized practically in the investigation of biological characteristics of p58 KIR and p50 KAR.

Animals↗
PubMed1999

Management of pancreatic pseudocysts by endoscopic cystogastrostomy.

Open surgical drainage is currently the treatment of choice for pancreatic pseudocysts, but endoscopic transmural drainage is another minimally invasive surgical alternative. In this report, we describe two patients with symptomatic pancreatic pseudocysts treated with endoscopic cystogastrostomy. The first patient, a 15-year-old boy, had an episode of traumatic pancreatitis after abdominal injury from a car accident, and complained of postprandial vomiting and abdominal distention 4 weeks later. A large pancreatic pseudocyst, about 10 cm x 6 cm, was noted. The second patient, a 44-year-old man, had a 1-year history of recurrent alcoholic pancreatitis prior to this admission. He suffered from abdominal distention for several weeks. Sonography revealed a large pancreatic pseudocyst, about 18 cm x 9 cm in size. Both patients underwent successful endoscopic cyst-drainage without recurrence. These cases illustrate that endoscopic transmural drainage provides a minimally invasive and effective approach to the management of pancreatic pseudocysts.

Adolescent↗
PubMed1998

Expression of Fas-related genes in human hepatocellular carcinomas.

Hepatocellular carcinoma (HCC) is one of the tumors known to be resistant to Fas-mediated apoptosis. To elucidate the possible mechanisms of this resistance, we examined the mRNA transcripts of genes related to Fas in 10 hepatitis B virus (HBV)-associated HCC cell lines and six HBV-associated HCC tissues. Most of the HBV-associated HCC cell lines showed markedly decreased Fas expression. Some cell lines expressed FAP, a possible inhibitor of apoptosis, or showed downregulated expression of FADD or FLICE, molecules essential to Fas-mediated apoptosis. All six HBV-associated HCC tissues also showed decreased Fas expression compared with their non-malignant counterparts. Our data demonstrate that HBV-associated HCC showed a common defect in the expression of Fas, upregulation of FAP and downregulation of downstream molecules such as FADD and FLICE.

Apoptosis↗
PubMed1998

Kinetic modeling of omega-transamination for enzymatic kinetic resolution of alpha-methylbenzylamine.

A kinetic model for omega-transaminase from Bacillus thuringiensis JS64 was developed by using the King-Altman method to simulate the kinetic resolution of alpha-methylbenzylamine (alpha-MBA). Starting from a ping-pong bi-bi mechanism, a complete kinetic model including substrate inhibition only in the reverse reaction (i.e., transamination between acetophenone and L-alanine) was developed. The asymmetric synthesis of (S)-alpha-MBA proved to be difficult due to a much lower maximum reverse reaction rate than the maximum forward reaction rate, thermodynamically exergonic forward reaction (i.e., transamination between (S)-alpha-MBA and pyruvate), and the severe product and substrate inhibition of the reverse reaction. Experimental values for kinetic parameters show that the product inhibition constant of (S)-alpha-MBA is the most important parameter on determining the resolution reaction rate, suggesting that the resolution reaction rate will be very low unless (S)-alpha-MBA strongly inhibits the reverse reaction. Using the kinetic model, the kinetic resolution of alpha-MBA in aqueous buffer was simulated, and the simulation results showed a high degree of consistency with experimental data over a range of reaction conditions. Various simulation results suggest that the crucial bottleneck in the kinetic resolution of alpha-MBA lies mainly in the accumulation of acetophenone in reaction media as the reaction proceeds, whereas L-alanine exerts a little inhibitory effect on the reaction. The model predicts that removing acetophenone produced during the reaction can enhance the reaction rate dramatically. Indeed, the biphasic reaction system is capable of extracting acetophenone from the aqueous phase, showing a much higher reaction rate compared to a monophasic reaction system. The kinetic model was also useful in predicting the properties of other, better enzymes as well as the optimal concentrations of amino acceptor and enzyme in the resolution reaction.

Acetophenones↗
PubMed1998

Inhibitory effects of bulbocapnine on dopamine biosynthesis in PC12 cells.

The effects of bulbocapnine, an aporphine isoquinoline alkaloid, on dopamine biosynthesis in PC12 cells were investigated. Bulbocapnine showed 45.2% inhibition on dopamine content in PC12 cells at a concentration of 20 microM for 12 h. The IC50 value of bulbocapnine was 26.7 microM. Bulbocapnine at concentrations up to 80 microM was not cytotoxic towards PC12 cells. Tyrosine hydroxylase (TH) activity was inhibited by the treatment of bulbocapnine in PC12 cells (24.4% inhibition at 20 microM). Bulbocapnine at 20 microM also decreased the intracellular Ca2+ concentration by 12.9% inhibition relative to control in PC12 cells. However, TH mRNA level was not altered by bulbocapnine treatment. These results suggest that the inhibition of TH activity by bulbocapnine might be involved in at least one component of the reduction of dopamine biosynthesis in PC12 cells.

Animals↗
PubMed1998

Rehabilitation of a triple amputee including a hip disarticulation.

A multiple amputee more severe than a triple amputee is uncommon. There have been no reports on the rehabilitation outcome of a triple amputee, including hip disarticulation and transtibial amputation. The authors report the rehabilitation of a patient with left hip disarticulation, right trans-tibial amputation, and left trans-humeral amputation due to a train accident. He has successfully completed the rehabilitation programme and has become independent in prosthetic ambulation, activities of daily living, and driving.

Adult↗
PubMed1998

[On the traditional surgery of Choson period (II)].

In this paper, authors discussed the classification of swelling (Chong), swelling-treatment methods (Chi-Chong) in Choson period, traditional notions about the natural history, treatment principles, popular healing methods of swelling, and depicted specially designed acupuncture instruments applied to it. Among the six professional guide books at that time, A Secret Recipe of Swelling Treatment (16th century) introduced the invasive surgical method into the narrow disease category i.e. carbuncle and furuncle, cellulitis, erysipelae, and gravitation abscess. The writer named these diseases as 'fire, stone, water, hemp, silk carbuncle', following each specific characteristics. Another surgical book, Orientation to Swelling Treatment (16th century) extended the surgical method to 'non-external' but 'looks-like swelling' diseases, such as pleurisy, tympanitis, testicular swelling, sequestrum of osteomyelitis. It is natural that some researchers doubted whether the book was used in real practice. However, the content of the book is too detailed to be regarded as an imaginary product. From these books and other materials, we found that the traditional notion of swelling was closely related with the notions of 'knotted' or 'pent in'. So 'pent-up rancor' or grudge was thought to aggravate any kind of swelling, and was tabooed or contraindicated in swelling treatment. 'Knotting in mind' was regarded as one of the principal etiologies of 'swelling and abscess formation in the body'. 'Fire in mind' was also regarded as one of the causes of phlegm aggregation resulting in swelling.

Abscess↗