Biomedical subjects

D H Kim

Publications and source records attributed to D H Kim.

At least 91 records · Page 5Linked to original sources

PubMed2001

Murine Xist RNA isoforms are different at their 3' ends: a role for differential polyadenylation.

Murine Xist is an essential transcript for X chromosome inactivation (X inactivation). According to recently revised structure, Xist is at least 17.8 kb long. It consists of seven exons and there are two major transcripts in female somatic cells. In this study we further defined the molecular structures of the two isoforms, namely short (S) and long (L) forms by northern blot and RNAse protection assay (RPA). The following lines of evidences suggest that mouse Xist depends on differential polyadenylation, not alternative splicing, to generate the two RNA isoforms: (1) only one band was detectable with the northern probes spanning the 3' end of Xist. (2) RPA showed the 3' termini of both S and L forms, and there are putative polyadenylation signals and hairpin structures close to these ends. (3) Analyses by splice site prediction program did not show any evidence of splice motifs in the sequence of L form. (4) Alignments between Xist 3' end (ESTs) and genomic sequence support the absence of splicing event in the region. The newly revised structure of Xist isoforms may have different stability and roles in the process of X inactivation.

Animals↗

PubMed2001

Solution structure of the antiapoptotic protein bcl-2.

The structures of two isoforms of Bcl-2 that differ by two amino acids have been determined by NMR spectroscopy. Because wild-type Bcl-2 behaved poorly in solution, the structures were determined by using Bcl-2/Bcl-x(L) chimeras in which part of the putative unstructured loop of Bcl-2 was replaced with a shortened loop from Bcl-x(L). These chimeric proteins have a low pI compared with the wild-type protein and are soluble. The structures of the two Bcl-2 isoforms consist of 6 alpha-helices with a hydrophobic groove on the surface similar to that observed for the homologous protein, Bcl-x(L). Comparison of the Bcl-2 structures to that of Bcl-x(L) shows that although the overall fold is the same, there are differences in the structural topology and electrostatic potential of the binding groove. Although the structures of the two isoforms of Bcl-2 are virtually identical, differences were observed in the ability of the proteins to bind to a 25-residue peptide from the proapoptotic Bad protein and a 16-residue peptide from the proapoptotic Bak protein. These results suggest that there are subtle differences in the hydrophobic binding groove in Bcl-2 that may translate into differences in antiapoptotic activity for the two isoforms.

Amino Acid Sequence↗

PubMed2001

Long-term preservation of rat pancreatic islets under physiological conditions.

In this study, we found that islet cells treated with polyphenol could be preserved for over 2 months under physiological conditions retaining their original function and maintaining their spherical shapes without any insulin secretion. When islets were treated at higher concentration than 250 microg ml(-1), these islets could retain their compact spherical shape over 65 days whereas non-treated islets were scattered ease to break within 2 weeks. The secretional capacity from treated islets in the initial stage is also lower than untreated islets. However, in the case of untreated islets, insulin release rapidly lowered with the progress in the culture time and secretion completely disappeared after 9 days. On the contrary, islets treated with polyphenol (250 microg ml(-1)) in RPMI culture medium showed significant enhancement of insulin secretion on 40th day. The secretional capacity of islets was greatly dependent on the treating concentration. Polyphenol treatment may be a useful method for preservation of mammalian islet cells. By changing the concentration of polyphenol, it is possible to control the preservation duration and insulin secretion of islets.

Animals↗

PubMed2001

Identification of a negative Cis-regulatory element and multiple DNA binding proteins that inhibit transcription of the transforming growth factor-beta type II receptor gene.

Expression of the transforming growth factor-beta type II receptor (TGF-beta RII) is highly regulated and is a critical determinant of the cellular response to TGF-beta. Previous analysis of the promoter region for the TGF-beta RII gene introduced the possible existence of a negative regulatory element (NRE) upstream adjacent to the core promoter region (Bae et al., 1995. J. Biol. Chem. 270, 29460-29468). We have confirmed the presence of a strong NRE located between base pairs -100 and -67 relative to the transcription start site. Utilizing DNA transfection techniques and a series of synthesized oligonucleotide promoter fragments, we have shown that this NRE is active in a variety of cell lines. Electrophoretic mobility shift assays have revealed the presence of multiple DNA binding proteins specifically interacting with the NRE. At least three distinct protein complexes are variably present depending on the specific cell line examined, and mutational analysis of the NRE has identified a ten-base pair recognition sequence which is shared by all three complexes. This palindromic sequence has not been previously reported and does not share homology with any known transcription factor consensus sequences. When inserted into an E4Delta heterologous promoter construct, the NRE binding sequence failed to inhibit either basal or activated transcription of the target gene, indicating that the NRE does not act as a general repressor but may specifically operate within the context of the TGF-beta RII core promoter.

Animals↗

PubMed2001

Differential regulation of protein tyrosine kinase on free radical production, granule enzyme release, and cytokine synthesis by activated murine peritoneal macrophages.

The present study examined the regulatory effect of tyrosine kinase inhibitors (genistein, tyrphostin, and 2,5-dihydroxycinnamate) on the free radical production, granule enzyme release, and synthesis of interleukin (IL)-8 and granulocyte macrophage-colony stimulating factor (GM-CSF) in murine peritoneal macrophages exposed to different stimulators [10 ng/mL of IL-1, 1 microgram/mL of lipopolysaccharide (LPS), and 1 microM N-formyl-methionyl-leucyl-phenylalanine (fMLP)]. Protein tyrosine kinase (PTK) inhibitors attenuated the stimulated superoxide, hydrogen peroxide, and nitric oxide production in macrophages stimulated with IL-1, LPS, or fMLP. N,N-Dimethylsphingosine (DMS) alone stimulated superoxide and hydrogen peroxide production by intact macrophages, but at 45 microM the stimulatory effect on superoxide production was not found. In contrast, DMS attenuated nitric oxide production by macrophages. High concentrations of DMS, tyrphostin, and 2,5-dihydroxycinnamate showed cytotoxic effects. PTK inhibitors did not exhibit a significant effect on granule enzyme release induced by IL-1, whereas they attenuated the effect of LPS and fMLP on degranulation. Genistein and tyrphostin decreased the production of IL-8 and GM-CSF in macrophages activated by IL-1, whereas 2,5-dihydroxycinnamate did not affect it. The results suggest that tyrosine kinases exposed to IL-1, LPS, and fMLP may exert different modulatory actions on macrophage responses. The IL-1-activated macrophage responses, particularly degranulation, appear to be differently regulated by tyrosine kinases compared with the responses activated by LPS and fMLP.

Animals↗

PubMed2001

Contact and fumigant activities of constituents of Foeniculum vulgare fruit against three coleopteran stored-product insects.

The insecticidal activities of materials derived from the fruit of fennel, Foeniculum vulgare, against adults of Sitophilus oryzae, Callosobruchus chinensis and Lasioderma serricorne were examined using direct contact application and fumigation methods. The biologically active constituents of the Foeniculum fruits were characterized as the phenylpropenes (E)-anethole and estragole, and the monoterpene (+)-fenchone, by spectroscopic analysis. Responses varied with insect species, compound, dose and exposure time. In a filter paper diffusion test, estragole at 0.168 mg cm-2 caused 91% mortality to S oryzae adults within 1 day after treatment (DAT), whereas (+)-fenchone and (E)-anethole gave over 90% mortality at 2 and 4 DAT, respectively. Against C chinensis adults, all test compounds revealed potent insecticidal activities at 0.021 mg cm-2 at 2 DAT. Against L serricorne adults at 0.105 mg cm-2, (E)-anethole gave 100% mortality at 1 DAT, whereas 90 and 60% mortality at 4 DAT was achieved with estragole and (+)-fenchone, respectively. In a fumigation test, the compounds were much more effective against adults of S oryzae, C chinensis and L serricorne in closed cups than in open ones, indicating that the insecticidal activity of test compounds was largely attributable to fumigant action. As naturally occurring insect-control agents, the F vulgare fruit-derived materials described could be useful for managing field populations of S oryzae, C chinensis and L serricorne.

Allylbenzene Derivatives↗

PubMed2001

Quantitative determination of medroxyprogesterone acetate in plasma by liquid chromatography/electrospray ion trap mass spectrometry.

A sensitive and rapid liquid chromatography/electrospray ion trap mass spectrometry (LC/MS/MS) method has been developed for the quantitative determination of medroxyprogesterone acetate (MPA) in human plasma. Plasma samples (1.0 mL) were simply extracted with pentane and the extracts were analyzed by HPLC with the detection of the analyte in the selective reaction monitoring (SRM) mode. The determination of MPA was accurate and reproducible, with a limit of quantitation of 0.05 ng/mL in plasma. The standard calibration curve for MPA was linear (r = 0.998) over the concentration range 0.05-6.0 ng/mL in human plasma. Analysis precision over the concentration range of MPA was lower than 18.8% (relative standard deviation, RSD) and accuracy was between 96.2 and 108.7%.

Calibration↗

PubMed2001

In vitro anti-Helicobacter pylori activity of panaxytriol isolated from ginseng.

This study investigated the effect that some polyacetylenes and protopanaxatriol, which were isolated from heated ginseng (family Araliaceae), have on inhibiting Helicobacter pylori (HP) growth. Among the compounds tested, panaxytriol was quite effective in inhibiting HP growth with an MIC of 50 microg/ml. Ginsenoside Rh1 and protopanaxatriol weakly inhibited H+/K+-ATPase from a rat stomach.

Alkynes↗

PubMed2001

Annomocherin, annonacin and annomontacin: a novel and two known bioactive mono-tetrahydrofuran annonaceous acetogenins from Annona cherimolia seeds.

A novel and two known bioactive mono-tetrahydrofuran (THF) annonaceous acetogenins, annomocherin (1), annonacin (2) and annomontacin (3), have been isolated from the fractionated ethanolic extracts of the seeds of Annona cherimolia, guided by the brine shrimp lethality test (BST). Their structures were elucidated on the basis of spectroscopic and chemical methods. All compounds have a relative stereochemistry of threo/trans/threo for the mono-THF ring with two flanking hydroxyls. Compound 1 has a double bond at C-23/24 of aliphatic chain. Compound 1 was isolated from natural sources for the first time, and was named annomocherin. Two known Compounds 2 and 3 which have never been isolated from this species before, were obtained. Compound 1 exhibited potent and selective cytotoxicities against the breast carcinoma (MCF-7) and kidney carcinoma (A-498) cell lines with 100 to 1,000 times the potency of adriamycin. In brine shrimp lethality test (BST), 1-3 exhibited cytotoxicity.

4-Butyrolactone↗

PubMed2001

Cloning, sequence analysis, and characterization of the astA gene encoding an arylsulfate sulfotransferase from Citrobacter freundii.

Arylsulfate sulfotransferase (ASST) transfers a sulfate group from a phenolic sulfate ester to a phenolic acceptor substrate. In the present study, the gene encoding ASST was cloned from a genomic library copy of Citrobacter freundii, subcloned into the vector pGEM3Zf(-) and sequenced. Sequencing revealed two contiguous open reading frames (ORF1 and ORF2) on the same strand and based on amino acid sequence homology, they were designated as astA and dsbA, respectively. The amino acid sequence of astA deduced from C. freundii was highly similar to that of the Salmonella typhimurium, Enterobacter amnigenus, Klebsiella, Pseudomonas putida, and Campylobacter jejuni, encoded by the astA genes. However, the ASST activity assay revealed different acceptor specificities. Using p-nitrophenyl sulfate (PNS) as a donor substrate, alpha-naphthol was found to be the best acceptor substrate, followed by phenol, resorcinol, p-acetaminophen, tyramine and tyrosine.

Amino Acid Sequence↗

PubMed2001

Intestinal bacterial beta-glucuronidase activity of patients with colon cancer.

The fecal beta-glucuronidase activity of patients with colon cancer and healthy controls were measured to determine the relationship between the fluctuation of intestinal bacterial beta-glucuronidase and colon cancer. The fecal beta-glucuronidase activity of patients with colon cancer was 1.7 times higher than that of the healthy controls. However, when these fecal specimens were sonicated, the enzyme activity of patients with colon cancer was 12.1 times higher than that of the healthy controls. The fecal beta-glucuronidase activity of human intestinal bacteria was drastically induced by its substrate or the bile secreted after a subcutaneous injection of 1,2-dimethylhydrazine (DMH) and benzo[a]pyrene into rats. DMH- and benzo[a]pyrene-treated biles induced beta-glucuronidase activity in the human intestinal microflora by approximately 1.5- and 2.3-fold, respectively. They also induced beta-glucuronidase in E. coli HGU-3, which is a beta-glucuronidase-producing bacterium from the human intestine. D-saccharic acid 1,4-lactone similarly inhibited fecal beta-glucuronidase in several patients with colon cancer in addition to the healthy controls. This suggests that potent beta-glucuronidase activity is a prime factor in the etiology of colon cancer.

Adult↗

PubMed2001

Constituents and the antitumor principle of Allium victorialis var. platyphyllum.

To search for cytotoxic components from Allium victorialis, MTT assays on each extract and an isolated component, gitogenin 3-O-lycotetroside, were performed against cancer cell lines. Cytotoxicities of most extract were shown to be comparatively weak, though IC50 values of CHCl3 fraction was found to be <31.3-368.4 microg/ml. From the incubated methanol extract at 36 degrees C, eleven kinds of organosulfuric flavours were predictable by GC-MS performance. The most abundant peak was revealed to be 2-vinyl-4H-1,3-dithiin (1) by its mass spectrum. Further, this extract showed significant cytotoxicities toward cancer cell lies. Silica gel column chromatography of the n-butanol fraction led to the isolation of gitogenin 3-O-lycotetroside (3) along with astragalin (4) and kaempferol 3, 4'-di-O-beta-D-glucoside (5). This steroidal saponin exhibited significant cytotoxic activities (IC50, 6.51-36.5 microg/ml) over several cancer cell lines. When compound 3 was incubated for 24 h with human intestinal bacteria, a major metabolite was produced and then isolated by silica gel column chromatography. By examining parent- and prominent ion peak in FAB-MS spectrum of the metabolite, the structure was speculated not to be any of prosapogenins of 3, suggesting that spiroketal ring were labile to the bacterial reaction. These suggest that disulfides produced secondarily are the antitumor principles.

Allium↗

PubMed2001

Albumin release from biodegradable hydrogels composed of dextran and poly(ethylene glycol) macromer.

Biodegradable hydrogels based on glycidyl methacrylate dextran (GMD) and dimethacrylate poly(ethylene glycol) (DMP) were proposed for colon-specific drug delivery. GMD was synthesized by coupling of glycidyl methacrylate with dextran in the presence of 4-(N,N-dimethyl-amino)pyridine (DMAP) using dimethylsulfoxide as a solvent. Methacrylate-terminated poly (ethylene glycol) (PEG) macromer was prepared by the reaction of PEG with methacryloyl chloride. GMD/DMP hydrogels were prepared by radical polymerization of phosphate buffer solution (0.1M, pH 7.4) of GMD and DMP, using ammonium peroxydisulfate (APS) and UV as initiating system. The synthetic GMD, DMP, and GMD/DMP hydrogels were characterized by fourier transform infrared (FT-IR) spectroscopy. The FITC-albumin loaded hydrogels were prepared by adding FITC-albumin solution before UV irradiation. Swelling capacity of GMD/DMP hydrogels was controlled not only by molecular weight of dextran, but also by incorporation ratio of DMP Degradation of the hydrogels has been studied in vitro with dextranase. FITC-albumin release from the GMD/DMP hydrogels was affected by molecular weight of dextran and the presence of dextranase in the release medium.

Acrylates↗

PubMed2001

Two kinds of body representation are used to control hand movements following tactile stimulation.

The task of reaching with a hand toward a target given in space is generally described as an ill-posed problem. It is often assumed that some kind of internal body model is required to solve this problem. This article provides information concerning the nature of this representation. Experiments were carried out in which blindfolded subjects were stimulated mechanically at one of eight possible stimulation sites on the legs and then asked to move one hand as quickly as possible to the stimulated site. In nine different postures of legs and hands, the frequency was recorded of the use of either hand. In addition, reaction times (RT) were measured and in two choice reaction tasks RTs were measured for different conditions (morphologically compatible vs morphologically incompatible, hands parallel vs hands crossed): The results support the hypothesis that the representation of the body is based on at least two systems, one which provides spatial information concerning the body position and a second one which is based on a morphological representation. According to this hypothesis, our results could be described by the following schema. Following the application of the stimulus, two processes were started in parallel. One concerned the activation of the spatial representation of body position, including the position of stimulus site and the possible response sites. This was more difficult, i.e., took more time, when the hands were crossed. Within this spatial representation, the distances between stimulus site and response sites were determined, and it was found that the smaller the distance, the more strongly the hands were activated. Simultaneously, in a second process the response site which was morphologically ipsilateral to the stimulus was excited. This schema could explain our results: In the behavioral experiments that hand which was most strongly excited, and which therefore exceeded a threshold sooner, won the decision and showed smaller RT values. In the choice reaction experiments, the winning hand was compared with the externally given task. The more strongly, according to the given task, the wrong hand was excited, the longer was the RT of the response.

Adult↗

PubMed2001

Biomechanical properties after pre-twist of canine patellar tendon.

We studied what effect a rotational pre-twist of the patellar tendon had on its mechanical properties. We used the central strip of canine patellar tendons. The length, width and thickness of each specimen were measured and the specimens were mounted in custom-made aluminum pots. Five groups of 10 specimens each were measured with neutral rotation, 90 degrees external rotation, 180 degrees external rotation, 270 degrees external rotation and 360 degrees external rotation. The ultimate stress, ultimate strain, average elastic modulus, and strain energy density were measured. There were no statistically significant differences among the five groups. Twisting the graft up to 360 degrees did not decrease the biomechanical properties of the graft.

Animals↗

PubMed2001

N-(Hydroxyaminocarbonyl)phenylalanine: a novel class of inhibitor for carboxypeptidase A.

N-(Hydroxyaminocarbonyl)phenylalanine (1) was designed rationally as a new type of inhibitor for carboxypeptidase A (CPA). The designed inhibitor was readily prepared from phenylalnine benzyl ester in two steps and evaluated to find that rac-1 inhibits CPA in a competitive fashion with the Ki value of 2.09 microM. Surprisingly, inhibitor 1 having the D-configuration is more potent (Ki = 1.54 microM) than its antipode by about 3-fold. A possible explanation for the stereochemistry observed in the inhibition of CPA with 1 is presented.

Binding Sites↗

PubMed2001

A new inhibitor design strategy for carboxypeptidase A as exemplified by N-(2-chloroethyl)-N-methylphenylalanine.

N-(2-Chloroethyl)-N-methylphenylalanine was designed and synthesized in an optically active form as a novel class of mechanism-based inactivator for carboxypeptidase A (CPA). It was anticipated that the chloroethylamino moiety of the CPA bound inhibitor undergoes an intramolecular SN2 reaction to generate a chemically reactive species (an aziridinium ion) which is expectedly subjected to a nucleophilic attack by the carboxylate of Glu-270, leading to covalent modification of the carboxylate. The irreversible nature of the inhibition of CPA by the inhibitor was supported by the kinetic data: the enzyme lost its enzymic activity in a time-dependent manner in the presence of the inhibitor and the inactivated CPA failed to regain the activity upon dialysis. Interestingly, the (R)-isomer that belongs to the D-series was more potent than its enantiomer.

Animals↗

PubMed2001

A simple and efficient in vitro method for metabolism studies of radiotracers.

In vitro metabolism of acetylcholinesterase inhibitors containing 3-[(18)F]fluoromethylbenzyl- ([(18)F]1) and 4-[(18)F]fluorobenzyl-piperidine moieties ([(18)F]2) was studied and compared with the in vivo metabolism. Defluorination of the [(18)F]1 mainly occurred to generate [(18)F]fluoride ion both in vitro and in vivo. In contrast, the [(18)F]2 was converted into an unknown polar metabolite in both metabolism methods and another metabolite, 4-[(18)F]fluorobenzoic acid in vitro. These results demonstrated that the in vitro method can be used to predict the in vivo metabolism of both radiotracers.

Animals↗