Biomedical subjects

K E Kim

Publications and source records attributed to K E Kim.

At least 19 recordsLinked to original sources

PubMed2001

Pharmacokinetics of a new antitumor 3-arylisoquinoline derivative, CWJ-a-5.

1-(4-Methylpiperazinyl)-3-phenylisoquinoline hydrochloride (CWJ-a-5) is a newly developed from benzo[c]phenanthridine alkaloids and derivative and has exhibited potent antitumor activities, in vitro and in vivo. The pharmacokinetics of this novel antitumor 3-arylisoquinoline derivative was studied after intravenous (i.v.), oral (p.o.) and hepatoportal (p.v.) administration in rats. A simple high performance liquid chromatographic method was developed to determine the concentrations of CWJ-a-5 in plasma, bile and urine. Plasma concentration profiles of CWJ-a-5 were best fitted by the two-compartment model after i.v. administration and showed a linear pharmacokinetic behavior up to 20 mg/kg doses. The half-life of CWJ-a-5 in the post-distributive phase (t1/2beta), total-body plasma clearance (CLt), and volume of distribution at steady-state (Vdss) were 86.9 min, 5.72 l/h per kilogram and 9.79 l/kg, respectively, after i.v. administration of 10 mg/kg. Biliary and urinary excretion of CWJ-a-5 was < 1% after i.v. injection of 10 mg/kg. The bioavailability of CWJ-a-5 after p.o. and p.v. administration (50 and 10 mg/kg, respectively) was 52.9 and 72.2%, respectively. Gastrointestinal bioavailability was calculated to be 73.3%. The apparent partition coefficient (log P) of CWJ-a-5 between n-octanol and water was 2.64. Plasma protein binding of CWJ-a-5 measured by the ultrafiltration method was > 95%.

Animals↗

PubMed2001

Dimethylbenzanthracene carcinogenesis in Gadd45a-null mice is associated with decreased DNA repair and increased mutation frequency.

Mice lacking the Gadd45a gene are susceptible to ionizing radiation-induced tumors. Increased levels of Gadd45a transcript and protein are seen after treatment of cells with ionizing radiation as well as many other agents and treatments that damage DNA. Because cells deficient in Gadd45a were shown to have a partial defect in the global genomic repair component of the nucleotide excision repair pathway of UV-induced photoproducts, dimethylbenzanthracene (DMBA) carcinogenesis was investigated because this agent produces bulky adducts in DNA that are also repaired by nucleotide excision repair. Wild-type mice and mice deficient for Gadd45a were injected with a single i.p. dose of DMBA at 10-14 days of age. The latency for spontaneous deaths was slightly decreased for Gadd45a-null mice compared with wild-type mice. At 17 months, all surviving animals were killed, and similar percentages of each genotype were found to have tumors. However, nearly twice as many Gadd45a-null than wild-type mice had multiple tumors, and three times as many had multiple malignant tumors. The predominant tumor types in wild-type mice were lymphoma and tumors of the intestines and liver. In Gadd45a-null mice, there was a dramatic increase in female ovarian tumors, male hepatocellular tumors, and in vascular tumors in both sexes. In wild-type mice, this dose of DMBA induced a >5-fold increase in Gadd45a transcript in the spleen and ovary, whereas the increase in liver was >20-fold. Nucleotide excision repair, which repairs both UV- and DMBA-induced DNA lesions, was substantially reduced in Gadd45a-null lymphoblasts. Mutation frequency after DMBA treatment was threefold higher in Gadd45a-null liver compared with wild-type liver. Therefore, lack of basal and DMBA-induced Gadd45a may result in enhanced tumorigenesis because of decreased DNA repair and increased mutation frequency. Genomic instability, decreased cell cycle checkpoints, and partial loss of normal growth control in cells from Gadd45a-null mice may also contribute to this process.

9,10-Dimethyl-1,2-benzanthracene↗

PubMed2001

Expression of recombinant human granulocyte macrophage-colony stimulating factor (hGM-CSF) in mouse urine.

We have generated transgenic mice expressing human granulocyte macrophage-colony stimulating factor (hGM-CSF) in urine. In particular, the expression plasmid DNA containing mouse uroplakin II promoter was used to direct uroepithelium-specific transcription of transgene. In this study, hGM-CSF transcript was detected only in bladder uroepithelium as determined by northern blot analysis. Furthermore, hGM-CSF protein was detected in the suprabasal layer of the uroepithelium and ureter by immunohistochemistry. The hGM-CSF was secreted into urine at high level (up to 180 ng/ml), and enhanced proliferation of hGM-CSF-dependent human acute monocyte leukemic cells, suggesting that transgenic urine-derived hGM-CSF was bioactive. This is the first case of demonstrating biological activity of a cytokine produced in the urine of a transgenic animal. Our results demonstrate that bladder can be used as a bioreactor to produce biologically important substances. In addition, it suggests a potential application of bladder expression system to livestock for high-yield production of pharmaceuticals.

Animals↗

PubMed2001

Identification of cDNA encoding a serine protease homologous to human complement C1r precursor from grafted mouse skin.

We isolated a cDNA clone from grafted mouse skin that encodes a serine protease homologous to human C1r. The C1r protease is involved in the activation of the first component of the classical pathway in the complement system. In order to identify novel transcripts whose expression is regulated in grafted mouse skin, we first performed differential display reverse transcription polymerase chain reaction analysis and obtained 18 partial cDNA clones whose protein products are likely to play an important role in allograft rejection. One of these showed significant sequence homology with human complement C1r precursor. The other clones displayed no homology to any known sequences, however. Northern blot analysis demonstrated that the level of this transcript was upregulated in day 8 postgrafted skin. The full-length cDNA 2121 nucleotides in length obtained from screening a mouse skin cDNA library contained a single open reading frame encoding 707 amino acid residues with a calculated molecular weight of 80,732 Da. Its deduced amino acid sequence revealed an 81% identity and 89% similarity to the human C1r counterpart. In particular, mouse C1r contained His501, Asp559, and Ser656, which were conserved among this group of serine proteases. This protein was thus designated as mouse C1r. We have expressed a truncated fragment of C1r protein without the N-terminal hydrophobic sequence in Escherichia coli and generated a polyclonal antibody against it. Subsequent immunohistochemical analysis confirmed that mouse C1r was significantly expressed 8 d after the skin graft in both allografted and autografted skins, compared with normal skins. These collective data suggest that a component of the complement system, C1r, might contribute to the graft versus host immune responses in mice.

Animals↗

PubMed2001

The role of vitamin D in normal and pathologic processes in the colon.

Vitamin D(3) metabolites and analogues have recently been shown to play an important role in the regulation of a number of important cellular processes, including proliferation, differentiation, and apoptosis, in addition to their established roles in mineral homeostasis. The actions of these secosteroids involve both rapid, nongenomic effects and genomic effects; the latter mediated via the vitamin D receptor and other transcription factors. Their effects have been described in a variety of cell types, including normal and malignant colonocytes. This article summarizes the rapid and genomic actions of vitamin D(3) metabolites and analogues on normal and pathologic processes in the colon, with particular emphasis on the potential of these secosteroids to prevent colon cancer.

Journal Article↗

PubMed2001

Prevalences of symptoms of asthma and other allergic diseases in korean children: a nationwide questionnaire survey.

The purpose of this study was to estimate the national prevalence of childhood asthma and other allergic diseases in Korea, and to determine potential risk factors for the diseases. Stratified random samples of 42,886 were selected from 34 elementary (6-12 yr olds) and 34 middle schools (12-15 yr olds) nationwide, and 38,955 were in the final analysis. The Korean-translated modified version of the International Study of Asthma and Allergies in Childhood questionnaire was used in this cross-sectional survey. Twelve-month prevalences of the symptoms of asthma, rhinoconjunctivitis, and flexural eczema were 8.7%, 10.5%, 7.3% in 6-12 yr olds, and 8.2%, 10.0%, 3.9% in 12-15 yr olds, respectively. For allergic conjunctivitis, food allergy, and drug allergy, the prevalences in 6-12 yr olds were 11.2%, 6.5%, and 1.5%, respectively. Asthma and flexural eczema decreased significantly with age. Other significant risk factors were also noted. For 6-12 yr-old asthma, adjusted odds ratio (aOR) of body mass index was 1.21 with 95% confidence interval (CI) 1.0-1.48, aOR of passive smoking was 1.37 with 95%CI 1.24-1.51, aOR of carpet use was 1.28 with 95%CI 1.10-1.49. For 6-12 yr-old eczema, aOR of affluence was 1.22 with 95%CI 1.07-1.39. The control of obesity and passive smoking would be the most important preventive measures of allergic diseases.

Adolescent↗

PubMed2000

Distribution of the native strain in human alpha 1-antitrypsin and its association with protease inhibitor function.

Serine protease inhibitors (serpins) are metastable in their native state. This strain, which is released upon binding to target proteases, is essential for the inhibitory activity of serpins. To understand the structural basis of the native strain, we previously characterized stabilizing mutations of alpha(1)-antitrypsin, a prototypical inhibitory serpin, in regions such as the hydrophobic core. The present study evaluates the effects of single point mutations throughout the molecule on stability and protease inhibitory activity. We identified stabilizing mutations in most secondary structures, suggesting that the native strain is distributed throughout the molecule. Examination of the substitution patterns and the structures of the mutation sites revealed surface hydrophobic pockets as a component of the native strain in alpha(1)-antitrypsin, in addition to the previously identified unusual interactions such as side chain overpacking and cavities. Interestingly, many of the stabilizing substitutions did not affect the inhibitory activity significantly. Those that affected the activity were confined in the regions that are mobilized during the complex formation with a target enzyme. The results of our study should be useful for designing proteins with strain and for regulating the stability and functions of serpins.

Enzyme Stability↗

PubMed2000

Cloning and characterization of the UDP-sugar hydrolase gene (ushA) of Enterobacter aerogenes IFO 12010.

A bacterial alkaline phosphatase (BAP, the phoA gene product) is primarily responsible for the hydrolysis of the substrates 5-bromo-4-chloro-3-indolylphosphate-p-toluidine (XP) and p-nitrophenyl phosphate (pNPP). Using these substrates and an E. coli phoA mutant, we have cloned Enterobacter aerogenes genes conferring an XP(+) phenotype. Two types of clones were identified based on phenotypic tests and DNA sequences. One of them is a E. aerogenes phoA gene (XP(+), pNPP(+)) as expected; surprisingly the other one was found to be a ushA gene (XP(+), pNPP(-)), which encodes an UDP (uridine 5'-diphosphate)-sugar hydrolase. The E. aerogenes ushA gene shares high sequence identity with ushA of E. coli and the mutationally silent ushA0 gene of Salmonella typhimurium at both the nucleotide (over 79%) and amino acid (over 93%) levels. Expression of the E. aerogenes ushA gene in E. coli produced high level of UDP-sugar hydrolase, as confirmed by TLC (thin layer chromatography) analysis together with a presence of a strong band due to a XP hydrolysis on a polyacrylamide gel.

Amino Acid Sequence↗

PubMed2000

Transcriptional regulatory effects of lymphoma-associated NFKB2/lyt10 protooncogenes.

C-terminal truncations of the NFKB2 p100 gene product have been observed in a number of cases of human cutaneous T cell lymphomas, as well as human B-cell lymphomas and myelomas. The contribution of these alterations to lymphomagenesis is not understood; however, truncation at amino acid 666 to generate 80 - 85 kD proteins in the HUT78 cell line is associated with addition of a short (serine-alanine-serine) fusion at the 3' end of p80HT, as well as with increased expression of NFKB2 mRNA. We therefore examined the effects of p80HT on the regulation of NFKB2 expression, as well as the properties of a series of other tumor-associated, and site directed mutations of NFKB2. While p80HT had not itself acquired novel transcriptional activation properties with respect to the NFKB2 P1 or P2 promoters or the IL-6 kappaB promoter, p80HT had lost the potent inhibitory (IkappaB-like) activity associated with the wild-type, p100 gene product. Loss of the inhibitory property depended on the SAS residues in the fusion protein, direct truncation at aa666 was fully inhibitory, as was a substitution of three alanines for the SAS residues. The presence of as few as two C-terminal ankyrin motifs was sufficient for inhibition of NF-kappaB-mediated transcriptional activation. Assays of a series of additional lymphoma-associated NF-kappaB-2 truncation suggested that the C-terminal truncation associated with these proteins was also associated with a loss of the IkappaB-like activities of p100 NF-kappaB-2, for at least some NF-kappaB target promoters. Thus, the loss of IkappaB-like activity of lymphoma-associated NFKB2 mutations may play an important role in the genesis of a subset of human lymphomas.

Gene Expression Regulation, Neoplastic↗

PubMed2000

Identification and characterization of the major allergens of buckwheat.

BACKGROUND: Buckwheat (BW) has been recognized as a common food allergen in Korea, Japan, and other countries. Until now, serologic findings of BW food-allergic patients and its major allergenic components have not been clarified. In this study, we analyzed the serologic findings of BW food allergy and characterized its major allergenic components. METHODS: Nineteen BW-allergic subjects with symptoms after BW ingestion and 15 asymptomatic control subjects with positive skin prick test to BW were recruited. BW-specific IgE was measured with the Pharmacia CAP kit. Allergenic components of BW were analyzed by IgE immunoblotting, periodate oxidation, two-dimensonal PAGE, and sequencing of N-terminal amino acids. RESULTS: From the BW-allergic patients and asymptomatic controls, the sensitivity (100%), specificity (53%), and negative (100%) and positive predictive values (73%) of Pharmacia CAP specific IgE for diagnosis were estimated. The prevalence of IgE binding to 24-kDa (pI 8.3), 16-kDa (pI 5.6), and 9-kDa (pI 5.0/ 6.0) allergens was higher than 50% in BW-allergic and asymptomatic subjects. However, the specific IgE to split 19-kDa (pI 6.5/7.0) allergens were more specifically found in BW-allergic patients than in asymptomatic subjects (78% vs 7%). N-terminal amino-acid sequences of 19-kDa and 16-kDa allergens showed moderate and weak homology to the 19-kDa globulin protein of rice and alpha-amylase/trypsin inhibitor of millet, respectively. The N-terminus of the 9-kDa isoallergens were not different from each other and were identified as the reported trypsin inhibitors of BW. Attenuation of the IgE binding to the 9-kDa allergen was found with periodate oxidation. CONCLUSIONS: The allergens of 24, 19, 16, and 9 kDa are strong candidates to be major allergens, and the 19-kDa allergen was relatively specific for BW-allergic patients. Moreover, measurement of BW-specific IgE and the features of immunoblotting should be very useful tools in the diagnosis of BW allergy.

Adolescent↗

PubMed1999

TCR internalization induced by peptide/MHC ligands requires the transmembrane domains of alphabeta chains of TCR, but not the expression of CD8 and Thy-1 molecules.

T-cell receptor (TCR) internalization occurs via TCR recognition of the peptide/MHC molecule complex on antigen presenting cell (APC). In this study, the requirements for inducing the internalization of TCR molecules on Ld major histocompatibility complex (MHC) class I-restricted T-cells were investigated with 2C cytotoxic T-lymphocyte (CTL) clones with defined peptides as the antigen. To evaluate the function of the transmembrane region of TCR alphabeta chains in TCR internalization, we generated T-cell transfectants expressing the wild type and glycosylphosphatidyl inositol (GPI)-linked form of 2C TCR. Among all peptides forming proper ligands to 2C TCR, only the Qp2Ca peptide induced TCR internalization, which was known to have the highest affinity to both Ld MHC class I molecules and TCR in association with Ld molecules. Such TCR internalization was not observed in cells expressing the GPI-linked form of 2C TCR. Furthermore, the expression of CD8 coreceptor and Thy-1 accessory molecules were both not required for Qp2Ca-induced TCR internalization, and these molecules did not accompany TCR internalization. Altogether, these results suggest that TCR internalization on CTL is not a prerequisite for CTL function.

Animals↗

PubMed1999

Genomic instability in Gadd45a-deficient mice.

Gadd45a-null mice generated by gene targeting exhibited several of the phenotypes characteristic of p53-deficient mice, including genomic instability, increased radiation carcinogenesis and a low frequency of exencephaly. Genomic instability was exemplified by aneuploidy, chromosome aberrations, gene amplification and centrosome amplification, and was accompanied by abnormalities in mitosis, cytokinesis and growth control. Unequal segregation of chromosomes due to multiple spindle poles during mitosis occurred in several Gadd45a -/- cell lineages and may contribute to the aneuploidy. Our results indicate that Gadd45a is one component of the p53 pathway that contributes to the maintenance of genomic stability.

Animals↗

PubMed1999

Regulation of glycoprotein hormone alpha-subunit gene expression.

The appropriate, regulated expression of the glycoprotein hormone subunit genes is required to enable the biosynthesis of luteinizing hormone, follicle-stimulating hormone, thyroid-stimulating hormone, and chorionic gonadotropin. We have focused our attention on mechanisms mediating regulated transcription of the common alpha-subunit gene. Our studies have examined both the signaling mechanisms and the DNA elements and transcription factors that are important for alpha-subunit expression. Our initial efforts involved characterization of DNA elements of the alpha-subunit gene important for basal and GnRH-stimulated expression. Clustered point mutation analysis identified two different, unrelated sequences that play a role in alpha-subunit transcription. When tested as multiple copies on a minimal promoter, one of these elements was sufficient to permit a response to GnRH, while the other enhanced basal expression. Therefore, we designated these DNA elements as the GnRH-response element (GnRH-RE) and the pituitary glycoprotein hormone basal element (PGBE), respectively. The GnRH-RE contains a consensus binding site for the Ets family of transcription factors. As several Ets factors have been shown to mediate transcriptional responses to the mitogen-activated protein kinase (MAPK) pathway, we investigated the possibility that GnRH effects on alpha-subunit transcription may involve the MAPK cascade. We found that GnRH can indeed activate MAPK and that MAPK activation is sufficient and necessary for transcriptional activation of the alpha-subunit gene. Efforts to further characterize proteins that interact with the PGBE led to the cloning of a LIM-homeodomain transcription factor designated LH-2. Recombinant LH-2 selectively binds to the PGBE in vitro. Transfection experiments have shown that an expression vector for LH-2 can activate the alpha-subunit promoter in heterologous cells. LH-2 appears to be a component of the endogenous factors that bind to the PGBE. Thus, LH-2 appears to be an excellent candidate as a factor responsible for basal expression of the alpha-subunit gene. Overall, these studies have contributed to identification of molecular components important for regulated expression of the glycoprotein hormone alpha-subunit gene.

Animals↗

PubMed1998

Examination of antisense RNA and oligodeoxynucleotides as potential inhibitors of avian leukosis virus replication in RP30 cells.

Avian leukosis virus (ALV) is an economically important pathogen of chickens. Both antisense RNA and antisense oligodeoxynucleotides (ODN) have been used to diminish the replication and spread of other retroviruses. The use of antisense RNA and ODN to inhibit ALV replication has been examined in cultured RP30 cells. Using an expression system that constitutively transcribes antisense ALV RNA, one transfected cell clone showed a significant reduction in virus growth. However, this effect was not reproducibly observed in other transfected cell lines or in cells in which the antisense transcript was expressed from a regulatable promoter, even though a substantial amount of antisense transcript was generated. Antisense ODN complementary to several different target sites near the 5' end of the ALV genome were also tested for antiviral activity, by comparison of antisense ODN effects to those of randomized sequence controls. An antisense ODN complementary to the ALV primer binding site demonstrated a reproducible reduction in viral replication. However, when the corresponding region was specifically employed as a target for intracellular antisense RNA expression, there again was no significant inhibition of ALV. These results suggest that in vivo expression of antisense RNA is unlikely to be an effective way to generate transgenic poultry that are resistant to field strains of ALV.

Animals↗

PubMed1998

IgE binding patterns to German cockroach whole body extract in Korean atopic asthmatic children.

It is widely known that the cockroach is an inhalant allergen in atopic asthma and allergic rhinitis. Even though Bla g I and Bla g II are considered as the major allergens, several relatively high-molecular weight (MW) cockroach allergens have also been recently identified by IgE-immunoblot in western countries. However, the environmental control and diagnostic tests mainly focussed on Bla g I and Bla g II. Furthermore there is no data about major IgE-binding cockroach antigens in Korea. We performed this study to identify the major German cockroach allergens in Korean atopic children. By the results of allergy skin tests, 14 children with atopic asthma (9 were cockroach-sensitive and 5 were cockroach-nonsensitive atopics) were enrolled in this study. We conducted IgE immunoblot and autoradiographic analysis using Yonsei-extract of German cockroach antigen produced in our laboratory, individual sera from 9 cockroach-sensitive children, and the pooled sera of 5 house-dust-mites-only-sensitive children. We performed an allergic skin test to cockroach mix, and a radioallergosorbent test (RAST) using German cockroach crude extract on all subjects. German cockroach-specific IgE was detected in 6 out of 9 subjects by RAST. We identified at least 15 IgE-binding protein bands, and among them, the components of MWs of 76, 64, 50, 38, and < 14 kilodaltons (kDa) were the major German cockroach allergens in study subjects. Therefore, Bla g I (25-30 kDa) and Bla g II (36 kDa) could not be the absolute indicators of German cockroach sensitization and parameters of environmental control.

Adolescent↗

PubMed1998

Effect of rebamipide on liver damage and increased tumor necrosis factor in a rat model of endotoxin shock.

We investigated the effect of rebamipide, a novel antiinflammatory agent, on liver damage in a rat model of circulatory shock induced by bacterial endotoxin (E. coli lipopolysaccharide, LPS). Endotoxemia for 6 hr resulted in a 5.9-fold rise in the serum levels of nitrite (P < 0.05) with a significant rise in the serum levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST), and lactic dehydrogenase (LDH), suggestive of liver dysfunction. The increased activities of serum ALT, AST, and LDH, but not serum nitrite were significantly inhibited by rebamipide (100 mg/kg, orally for five days). Myeloperoxidase activity in the liver was significantly elevated in the rats with endotoxemia by 2.4-fold (P < 0.05), which was also significantly inhibited by rebamipide. Upon LPS injection, serum TNF-alpha levels peaked at 1 hr after LPS (from 167.4 +/- 20.0 to 1570.0 +/- 100.0 pg/ml) and thereafter rapidly declined. The increased TNF-alpha level measured at 1 hr was significantly inhibited by pretreatment with rebamipide (100 mg/kg for five days). It is suggested that rebamipide exerts a strong protective effect on the LPS-induced liver damage through inhibition of activation of neutrophils and TNF-alpha production.

Alanine↗

PubMed1997

B-cell epitopes recognized by IgE from patients sensitive to Amb a 5.

BACKGROUND: The response to allergens characterized by IgE-mediated hypersensitivity is selective. The search for the inherited contribution to atopy has among other things, focused on the linkage of sensitivity to the presence of specific alleles in the DR and DQ locus. More than 90% of the responders to Amb a 5, an allergen from ambrosia artemisifolia, are DR-2 positive. This relationship is logically linked to the T-cell epitope presentation by the HLA complex. OBJECTIVES: This study aims to investigate a possible relationship between T-cell epitopes, B-cell epitopes and the alleles of the DR and DQ loci in Amb a 5 sensitive DR-2+ and DR-2- individuals. METHODS: Inhibition of solid state Elisa assays by IgE-enriched and IgG-depleted, heated sera. The inhibition was carried out in checkerboard pattern, bidirectionally; A inhibits B and B inhibits A. RESULTS: The B-cell epitopes defined by the inhibition pattern were all found to be conformational. Three different epitope patterns (A, B, C) were recognized. The IgE and IgG complexes were found in only one responder. The DR and DQ locus alleles were all sequenced. Although all the individuals studied responding to Amb a 5 show presence of alleles such as 1501, associated with DR-2, our data indicates no correlation between the B-cell epitopes recognized and the DR and DQ locus alleles. A well known, general T-cell motif was recognized in the known sequence of Amb a 5. CONCLUSIONS: Our investigation suggests that the choice of B-cell recognition is regulated independently of a putative link between T-cell epitope recognition and the D locus.

Adult↗

PubMed1996

The affinity of allergen specific IgE and the competition between IgE and IgG for the allergen in Amb a V sensitive individuals.

We have calibrated a solid state RAST assay with affinity purified allergen-specific IgE. We then utilized the calibrated assay to measure the average affinity of individual IgE-containing sera in terms of the average association constant < K > for purified allergen Amb a V. The binding data yielded linear reciprocal plots indicating that the range of affinities of the responding clones was narrow. The range of the average association constant for the IgE-Amb a V complex was 0.9-26 x 10(10) M-1. The average affinity of the corresponding IgG response in the same individual, estimated by inhibition studies of IgE binding, was 10(7) M-1 in one case and lower than 10(6) M-1 in all the other cases.

Allergens↗