Biomedical subjects

Tae Soo Kim

Publications and source records attributed to Tae Soo Kim.

At least 19 recordsLinked to original sources

PubMed2006

Effects of lactate dehydrogenase suppression and glycerol-3-phosphate dehydrogenase overexpression on cellular metabolism.

In order to conduct a physiological functional study of lactate dehydrogenase (LDH) and glycerol-3-phosphate dehydrogenase (GPDH), we engineered a CHO dhfr(-) cell, by overexpressing either the anti-sense LDH-A RNA (anti-LDH cells) or GPDH (GP3 cells), or both (GP3/anti-LDH cells). LDH activity in the cell cytosol, and lactate content and pHe change in the growth media were found to decrease according to the order: cell lines GP3/anti-LDH > anti-LDH > GP3 > CHO. Intracellular ATP contents, representing the extent of respiration rate, also decreased, according to a rank order as follows: GP3 > CHO > GP3/anti-LDH > anti-LDH. We also attempted to identify and characterize any physiological changes occurring in the cells which harbored diverse metabolic pathways. First, anti-LDH cells with heightened respiration rates were found to display a higher degree of sensitivity to the prooxidant tert-butyl hydroperoxide (tBOOH), and the mitochondrial complex III inhibitor, antimycin A, than the GPDH-expressing cells (GP3 and GP3/anti-LDH), which have a lower respiration rate. Second, the anti-sense LDH-A RNA-expressing cells (anti-LDH and GP3/anti-LDH) evidenced a higher degree of resistance to apoptosis by cell-cell contact inhibition, and a faster doubling time ( approximately 19 h compared with approximately 26 h) than the CHO and GP3 cells. Additionally, cell growth in an extended culture under HCO(3) (-)-free conditions to induce a steep acidification could be maintained with the anti-sense LDH-A RNA-expressing cells, but could not be maintained with the CHO and GP3 cells. Third, we observed that the most appropriate cell line for the optical production of a certain therapeutic protein (Tissue-Plasminogen Activator) was the GP3/anti-LDH cells. Collectively, our data indicate a variety of physiological roles for LDH and GPDH, including cellular acidosis, oxidoresistance, apoptosis by both acidosis and cell-cell contact inhibition, cell growth, and the generation of recombinant proteins.

Animals↗

PubMed2006

Ribonucleotide reductase M1 (RRM1) 2464G>A polymorphism shows an association with gemcitabine chemosensitivity in cancer cell lines.

OBJECTIVES: Significant variability in the efficacy and toxicity of an anticancer drug is observed in cancer patients. Currently, there are no standard tools for prediction of a patient's tumor response or his risk of adverse events to chemotherapy. METHODS: We investigate an association between polymorphisms of gemcitabine metabolism-related genes and its chemosensitivity in vitro using 62 human cancer cell lines of various origins. Polymorphisms of gemcitabine metabolism-related genes of deoxycytidine monophosphate deaminase (DCTD), deoxycytidine kinase (DCK) and ribonucleotide reductase M1 (RRM1) were evaluated using the CEQ8000 Genetic analysis system and GeneDoc software. Chemosensitivity of gemcitabine was expressed as an IC50 using MTT assay. RESULTS: The frequency of the polymorphisms was 21% in DCTD 315T>C, 45.2% in RRM1 1082C>A, 59.7% in RRM1 2455A>G, and 79% in RRM1 2464G>A. When examining the association between these polymorphisms and IC50, only the RRM1 2464G>A showed the tendency to be more chemosensitive to gemcitabine (P=0.011), and haplotypes containing 2464G>A polymorphism also showed the association with chemosensitivity when compared to wild-type RRM1 (G2464G). We could not see the significant differences of mRNA expression level with real-time PCR between cell lines according to G2464A polymorphism. In oligonucleotide microarray 73 GenBank Accession Number (69 genes) were selected which expressed differently between RRM1 wild-type and the G2464A polymorphism. CONCLUSIONS: RRM1 2464G>A polymorphism demonstrated an association with gemcitabine sensitivity, which needs functional studies with co-expressing genes and prospective clinical studies for the clinical application as a predictive bio-marker.

Antimetabolites, Antineoplastic↗

PubMed2006

Systematic analysis of cDNA microarray-based CGH.

cDNA microarray-based CGH (Microarray-CGH) is a useful technique for detecting genomic aberrations with a high resolution. However, the criteria for determining a genomic alteration have not been determined. We evaluated the genome-wide measurement of copy number of each gene in normal gastric and placenta tissues with both sex-matched, direct and sex-mismatched, indirect designs using 17K cDNA microarray. The results revealed the range of genomic copy number of normal tissues to be +/-0.3 of the log(2) ratio (gain >0.3, loss <-0.3) in the autosomal genes with direct and indirect designs. The copy number at a gene level from the X chromosomal genes using the direct and indirect sex-mismatched designs was +/-0.68 of the log(2) ratio (amplification >0.68, deletion <-0.68). In summary, the suggested method can be used as a guideline for analysis of genomic aberration using a Microarray-CGH in both direct and indirect designs.

Chromosomes, Human↗

PubMed2005

Selenite-induced apoptosis of osteoclasts mediated by the mitochondrial pathway.

The possible effects of sodium selenite on mature osteoclasts were investigated. Incubation of osteoclast-like cells differentiated from RAW 264.7 cells with sodium selenite induced apoptosis as revealed by morphological changes, internucleosomal DNA fragmentation, and activation of caspase-3. Selenite also induced generation of the superoxide anion and reduced the number of free thiol groups in the osteoclast-like cells, suggestive of a shift to a more oxidizing intracellular environment. In addition, selenite induced protein aggregation by thiol cross-linking, loss of the mitochondrial membrane potential, and cytochrome c release in mitochondria isolated from the osteoclast-like cells. Finally, selenite-induced DNA fragmentation in osteoclasts was inhibited both by cyclosporin A, a blocker of the mitochondrial permeability transition pore, and by DEVD-CHO, a cell-permeable inhibitor of caspase-3. These results thus suggest that selenite induces apoptosis mediated by the mitochondrial pathway in mature osteoclasts.

Animals↗

PubMed2005

Determination of genes related to gastrointestinal tract origin cancer cells using a cDNA microarray.

PURPOSE: We evaluated the genome-wide gene expression profiles of various cancer cell lines to identify the gastrointestinal tract cancer cell-related genes. EXPERIMENTAL DESIGN: Gene expression profilings of 27 cancer cell lines and 9 tissues using 7.5K human cDNA microarrays in indirect design with Yonsei reference RNA composed of 11 cancer cell line RNAs were done. The significant genes were selected using significant analysis of microarray in various sets of data. The selected genes were validated using real-time PCR analysis. RESULTS: After intensity-dependent, within-print-tip normalization by loess method, we observed that expression patterns of cell lines and tissues were substantially different, divided in two discrete clusters. Next, we selected 115 genes that discriminate gastrointestinal cancer cell lines from others using significant analysis of microarray. Among the expression profiles of five gastric cancer cell lines, 66 genes were identified as differentially expressed genes related to metastatic phenotype. YCC-16, which was established from the peripheral blood of one advanced gastric cancer patient, produced a unique gene expression pattern resembling the profiles of lymphoid cell lines. Quantitative real-time reverse transcription-PCR results of selected genes, including PXN, KRT8, and ITGB5, were correlated to microarray data and successfully discriminate the gastrointestinal tract cancer cell lines from hematologic malignant cell lines. CONCLUSIONS: A gene expression database could serve as a useful source for the further investigation of cancer biology using the cell lines.

Cell Line, Tumor↗

PubMed2005

Destiny of autologous bone marrow-derived stromal cells implanted in the vocal fold.

OBJECTIVES: The aim of this study was to investigate the destiny of implanted autologous bone marrow-derived stromal cells (BSCs) containing mesenchymal stem cells. We previously reported the successful regeneration of an injured vocal fold through implantation of BSCs in a canine model. However, the fate of the implanted BSCs was not examined. In this study, implanted BSCs were traced in order to determine the type of tissues resulting at the injected site of the vocal fold. METHODS: After harvest of bone marrow from the femurs of green fluorescent transgenic mice, adherent cells were cultured and selectively amplified. By means of a fluorescence-activated cell sorter, it was confirmed that some cells were strongly positive for mesenchymal stem cell markers, including CD29, CD44, CD49e, and Sca-1. These cells were then injected into the injured vocal fold of a nude rat. Immunohistologic examination of the resected vocal folds was performed 8 weeks after treatment. RESULTS: The implanted cells were alive in the host tissues and showed positive expression for keratin and desmin, markers for epithelial tissue and muscle, respectively. The implanted BSCs differentiated into more than one tissue type in vivo. CONCLUSIONS: Cell-based tissue engineering using BSCs may improve the quality of the healing process in vocal fold injuries.

Animals↗

PubMed2005

Quantitative detection of telomerase activity by real-time TRAP assay in the body fluids of cancer patients.

Real-time TRAP assay was developed to improve the sensitivity and quantitative detection of telomerase activity in the body fluids of cancer patients. The sensitivity and clinical significance of the real-time TRAP assay was investigated. Real-time PCR protocol was modified by using ACX primer and SYBR green mixture from the process of TS primer extension. Real-time TRAP showed high correlation (r2=0.843, p=0.001) and sensitivity (25 times higher) compared to conventional TRAP. Of 164 samples, there were 8 positives in cytology (4.9%), 7 (4.3%) using the conventional TRAP, and 41 (25%) using real-time TRAP. In cytology positive samples, real-time TRAP showed a higher positivity than conventional TRAP (75% vs 63%) suggesting a higher sensitivity in the body fluids. There was a tendency towards a longer progression-free duration in telomerase negative patients than in positive patients, as determined by conventional and real-time TRAP. The diagnostic interval between the first positivity documentation and clinical progression was short in the order of real-time TRAP, conventional TRAP and cytology. In conclusion, real-time TRAP assay can detect telomerase activity at an earlier phase of cancer progression and can replace conventional TRAP assay for detecting the telomerase activity in body fluids.

Body Fluids↗

PubMed2004

Different distributions of selenoprotein W and thioredoxin during postnatal brain development and embryogenesis.

Whereas the levels of other selenoproteins in the brain decrease when selenium is deficient, the level of selenoprotein W (Se-W) is maintained, suggesting that it has a critical role in the brain. Previously, we reported that Se-W is a GSH-dependent antioxidant [Jeong et al. (2002)]. In this study, the expression of Se-W and thioredoxin (Trx) in the brain and during embrynic development was analyzed by an in situ hybridization technique. Se-W mRNA was highly expressed in the cortex, dentate gyrus, and hippocampus of postnatal rat brains, and in the spinal cord and brain of developing embryos. In contrast, Trx mRNA was highly expressed in the cerebellum, olfactory bulb, and dentate gyrus of postnatal rat brains, and in the liver, telencephalon, and back muscle of developing embryos. Thus these two antioxidant proteins have different and non-overlapping expression patterns. The distribution of Se-W suggests that it plays an important role as an antioxidant in the developing brain and embryo.

Amino Acid Sequence↗

PubMed2004

Role of reactive radicals in degeneration of the auditory system of mice following cisplatin treatment.

OBJECTIVE: It has been suggested that reactive radical species are involved in the mechanism of cisplatin-induced hearing loss. However, the nature of the free radicals involved is not fully understood. We examined the effects of two highly reactive species, hydroxyl radicals and peroxynitrite, on the auditory system of mice following cisplatin treatment. MATERIAL AND METHODS: Expression of 4-hydroxynonenal (HNE), a marker of lipid peroxidation by the hydroxyl radical, and nitrotyrosine (NT), a marker for protein peroxidation by peroxynitrite, was examined immunohistochemically in mouse cochleae injured by means of local application of cisplatin. RESULTS: Loss of outer hair cells (OHCs) and spiral ganglia was found in cochleae affected by cisplatin. Both HNE and NT were detected in auditory epithelia and neurons damaged by cisplatin. Interestingly, auditory hair cells produced HNE, but not NT. Our findings indicate contributions by both HNE and NT to the degeneration of the auditory system due to cisplatin, and a crucial role of the hydroxyl radical in degeneration of OHCs. CONCLUSION: The hydroxyl radical may be a critical target for a strategy aimed at protecting auditory function from cisplatin toxicity.

Aldehydes↗

PubMed2004

Signaling pathway for apoptosis of vestibular hair cells of mice due to aminoglycosides.

Previous studies on regeneration of mammalian vestibular hair cells have indicated the potential for self-repair of damaged hair cells. The rescue of damaged hair cells from cell death may therefore increase regenerated hair cells in affected vestibular epithelia. The role of apoptosis in the degradation of vestibular hair cells following aminoglycoside treatment has been elucidated. To seek a method of protecting vestibular hair cells from aminoglycoside toxicity, we examined the apoptosis signaling pathway of vestibular hair cells due to aminoglycoside toxicity. Induction of apoptosis in hair cells of mouse ampullar cristae damaged by local application of neomycin was evaluated by the terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) method and transmission electron microscopy (TEM). Immunohistochemistry for apoptosis-related proteins was employed to determine the signaling pathway of apoptosis of hair cells. The occurrence of apoptosis in hair cells was demonstrated by TUNEL staining and TEM. In apoptotic hair cells, activation of caspase-3 and -9, and redistribution of cytochrome c was identified, while there was no expression of activated caspase-8 or apoptosis-inducing factor. In conclusion, these findings indicate that the mitochondria-mediated pathway of apoptosis may play a role in inducing the apoptosis of vestibular hair cells due to aminoglycoside toxicity. Stabilization of the mitochondrial membrane may therefore rescue vestibular hair cells from apoptosis, leading to an increase in self-repaired hair cells in affected vestibular epithelia.

Animals↗

PubMed2004

Glucose repression of STA1 expression is mediated by the Nrg1 and Sfl1 repressors and the Srb8-11 complex.

In the yeast Saccharomyces diastaticus, expression of the STA1 gene, which encodes an extracellular glucoamylase, is negatively regulated by glucose. Here we demonstrate that glucose-dependent repression of STA1 expression is imposed by both Sfl1 and Nrg1, which serve as direct transcriptional repressors. We show that Nrg1 acts only on UAS1, and Sfl1 acts only on UAS2, in the STA1 promoter. When bound to its specific site, Sfl1 (but not Nrg1) prevents the binding to UAS2 of two transcriptional activators, Ste12 and Tec1, required for STA1 expression. We also found that Sfl1 contributes to STA1 repression by binding to the promoter and inhibiting the expression of FLO8, a gene that encodes a third transcriptional activator involved in STA1 expression. In addition, we show that the levels of Nrg1 and Sfl1 increase in glucose-grown cells, suggesting that the effects of glucose are mediated, at least in part, through an increase in the abundance of these repressors. NRG1 and SFL1 expression requires the Srb8-11 complex, and correspondingly, the Srb8-11 complex is also necessary for STA1 repression. However, our evidence indicates that the Srb8-11 complex does not associate with either the SFL1 or the NRG1 promoter and thus plays an indirect role in activating NRG1 and SFL1 expression.

DNA-Binding Proteins↗

PubMed2004

Recruitment of the Swi/Snf complex by Ste12-Tec1 promotes Flo8-Mss11-mediated activation of STA1 expression.

In the yeast Saccharomyces diastaticus, expression of the STA1 gene, which encodes an extracellular glucoamylase, is activated by the specific DNA-binding activators Flo8, Mss11, Ste12, and Tec1 and the Swi/Snf chromatin-remodeling complex. Here we show that Flo8 interacts physically and functionally with Mss11. Flo8 and Mss11 bind cooperatively to the inverted repeat sequence TTTGC-n-GCAAA (n = 97) in UAS1-2 of the STA1 promoter. In addition, Flo8 and Mss11 bind indirectly to UAS2-1 of the STA1 promoter by interacting with Ste12 and Tec1, which bind to the filamentation and invasion response element (FRE) in UAS2-1. Furthermore, our findings indicate that the Ste12, Tec1, Flo8, and Mss11 activators and the Swi/Snf complex bind sequentially to the STA1 promoter, as follows: Ste12 and Tec1 bind first to the FRE, whereby they recruit the Swi/Snf complex to the STA1 promoter. Next, the Swi/Snf complex enhances Flo8 and Mss11 binding to UAS1-2. In the final step, Flo8 and Mss11 directly promote association of RNA polymerase II with the STA1 promoter to activate STA1 expression. In the absence of glucose, the levels of Flo8 and Tec1 are greatly increased, whereas the abundances of two repressors, Nrg1 and Sfl1, are reduced, suggesting that the balance of transcriptional regulators may be important for determining activation or repression of STA1 expression.

DNA, Fungal↗

PubMed2004

Mechanisms of apoptosis induced by cisplatin in marginal cells in mouse stria vascularis.

Degeneration of the stria vascularis (SV) is amongst the major causes of cisplatin (CDDP)-induced hearing impairment. The pathways of apoptosis occurring in the SV due to CDDP were examined using a mouse experimental model. Temporal bones of adult C57BL/6 mice were collected on days 3, 7 and 14 after the local application of CDDP. Terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling assay and immunostaining for apoptosis-related proteins or reactive radical species were employed for analysis. Local application of CDDP caused apoptotic cell death of marginal cells 3 days after CDDP treatment. Immunohistochemical analyses demonstrated activation of caspase-3 and -9, but not -8, and redistribution of cytochrome c in affected marginal cells, indicating a caspase-dependent, mitochondrion-mediated apoptotic pathway in marginal cells. Temporary expression of hydroxynonenal, nitrotyrosine and inducible nitric oxide synthase in the SV was observed at the induction of apoptosis in marginal cells. CDDP toxicity generates reactive radical species in the SV, which causes mitochondrial membrane permeabilization leading to apoptosis of marginal cells.

Aldehydes↗

PubMed2004

Fabrication of high quality cDNA microarray using a small amount of cDNA.

DNA microarray technology has become an essential part of biological research. It enables the genome-scale analysis of gene expression in various types of model systems. Manufacturing high quality cDNA microarrays of microdeposition type depends on some key factors including a printing device, spotting pins, glass slides, spotting solution, and humidity during spotting. UsingEthe Microgrid II TAS model printing device, this study defined the optimal conditions for producing high density, high quality cDNA microarrays with the least amount of cDNA product. It was observed that aminosilane-modified slides were superior to other types of surface modified-slides. A humidity of 30+/-3% in a closed environment and the overnight drying of the spotted slides gave the best conditions for arraying. In addition, the cDNA dissolved in 30% DMSO gave the optimal conditions for spotting compared to the 1X ArrayIt, 3X SSC and 50% DMSO. Lastly, cDNA in the concentration range of 100-300 ng/ micro l was determined to be best for arraying and post-processing. Currently, the printing system in this study yields reproducible 9000 spots with a spot size 150 mm diameter, and a 200 nm spot spacing.

DNA, Complementary↗

PubMed2003

STA10 repression of STA gene expression is caused by a defective activator, flo8, in Saccharomyces cerevisiae.

The expression of STA genes that encode extracellular glucoamylase isozymes is repressed in most laboratory Saccharomyces cerevisiae strains, which are believed to contain an undefined repressor, designated STA10. To identify the regulator involved in STA10 repression, we investigate the FLO8, MSN1, MSS11, STE12, and TEC1 genes. The Deltaflo8 or Deltamss11 deletion mutants in the sta10 genetic background exhibit both a loss of flocculation ability and a reduction in extracellular glucoamylase activity, as in the STA10 strain. Moreover, the STA10 repression is suppressed completely or partially by the introduction of a single copy of the FLO8 or MSS11 genes. Sequence analysis and complementation testing of the STA10 strain reveal that it has an inactive, mutated flo8-1 allele. A random spore analysis and transplacement (allele replacement) experiment confirms that the repressive phenotype of STA10 is due to the amber mutation of the transcriptional activator, FLO8.

Alleles↗

PubMed2003

Fate of neural stem cells grafted into injured inner ears of mice.

Loss of sensory hair cells in the inner ear is a major cause of permanent hearing loss, since regeneration of hair cells rarely occurs in mammals. The aim of this study was to examine the potential of neural stem cell transplantation to restore inner ear hair cells in mice. Fetal neural stem cells were transplanted into the mouse inner ear after drug-induced injury. Histological analysis demonstrates that the majority of grafted cells differentiated into glial or neural cells in the inner ear. Strikingly, however, we show that grafted cells integrate in vestibular sensory epithelia and express specific markers for hair cells. This finding suggests that transplanted neural stem cells have the potential to differentiate and restore inner ear hair cells.

Animals↗

PubMed2003

Trophic support of mouse inner ear by neural stem cell transplantation.

In the auditory system, efforts to reduce degeneration of spiral ganglion neurons have the immediate objective of improving clinical benefits of cochlear implants, which are small devices designed to stimulate spiral ganglion neurons electronically. Recent studies have indicated several neurotrophins can enhance survival of spiral ganglion neurons. However, the strategy for application of neurotrophins in inner ear is still a matter of debate. In this study, we examined the potential of cell therapy as a strategy for application of neurotrophins in the inner ear. Neural stem cells obtained from green fluorescent protein-transgenic mice were used as donor cells. Medium containing neural stem cells was injected into mouse inner ear. Histological analysis 4 weeks later revealed that transplant-derived cells survived in inner ear and that most transplant-derived cells in the cochlea had differentiated into glial cells. Moreover, expression of glial cell line-derived neurotrophic factor and brain-derived neurotrophic factor was observed in transplant-derived cells. These findings indicate that transplantation of neural stem cells can be a useful strategy for application of neurotrophins in inner ear.

Animals↗

PubMed2003

A novel technique for inducing local inner ear damage.

With significant development of mouse genomics and the availability of transgenic and knockout mice, the mouse will be the preferred animal model for inner ear research. However, few studies have used mice as experimental animals for examination of hair cell degeneration, because of their relative resistance to ototoxic agents and difficulties in surgical treatment. This study presents a model for induction of apoptotic cell death in sensory epithelia of the mouse inner ear using injection of neomycin into the posterior semicircular canal. Terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) assay revealed that local application of neomycin produced sufficient induction of apoptotic cell death in both auditory and vestibular epithelia over a definite time course. Supplementation of the general caspase-inhibitor significantly reduced induction of TUNEL-positive cells, indicating caspase-dependency of apoptotic cell death observed in the present model. In addition, the approach to the posterior semicircular canal was an easy technique, and sham-operation induced no significant induction of TUNEL-positive cells. This model, hence, enables the use of various genetic tools in studies for mechanisms of hair cell apoptosis.

Amino Acid Chloromethyl Ketones↗