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Biomedical subjects

K J Hwang

Publications and source records attributed to K J Hwang.

At least 19 recordsLinked to original sources
PubMed2007

Human amniotic fluid-derived stem cells have characteristics of multipotent stem cells.

OBJECTIVES: To characterize mesenchymal stem cell-like cells isolated from human amniotic fluid for a new source of therapeutic cells. MATERIALS: Fibroblastoid-type cells obtained from amniotic fluid at the time of birth. METHODS: The ability of ex vivo expansion was investigated until senescence, and stem cell-like characteristics were analyzed by examining differentiation potential, messenger RNA expression and immunophenotypes. RESULTS AND CONCLUSIONS: A morphologically homogenous population of fibroblastoid-type (HAFFTs) cells, similar to mesenchymal stem cells from bone marrow (BM-MSCs), was obtained at the third passage. The cells became senescent after 27 passages over a period of 8 months while undergoing 66 population doublings. Under appropriate culture conditions, by the 8th passage they differentiated into adipocytes, osteocytes, chondrocytes and neuronal cells, as revealed by oil red O, von Kossa, Alcian blue and anti-NeuN antibody staining, respectively. Immunophenotype analyses at the 17th passage demonstrated the presence of TRA-1-60; SSEA-3 and-4; collagen types I, II, III, IV and XII; fibronectin; alpha-SMA; vimentin; desmin; CK18; CD44; CD54; CD106; FSP; vWF; CD31; and HLA ABC. Reverse transcriptase-polymerase chain reaction analysis of the HAFFTs from passages 6-20 showed consistent expression of Rex-1, SCF, GATA-4, vimentin, CK18, FGF-5 and HLA ABC genes. Oct-4 gene expression was observed up to the 19th passage but not at the 20th passage. HAFFTs showed telomerase activity at the 5th passage with a decreased level by the 21st passage. Interestingly, BMP-4, AFP, nestin and HNF-4alpha genes showed differential gene expression during ex vivo expansion. Taken together, these observations suggest that HAFFTs are pluripotent stem cells that are less differentiated than BM-MSCs, and that their gene expression profiles vary with passage number during ex vivo expansion.

Amniotic Fluid↗
PubMed2006

Progesterone-dependent release of transforming growth factor-beta1 from epithelial cells enhances the endometrial decidualization by turning on the Smad signalling in stromal cells.

Endometrial decidualization results from the differentiation of stromal cells in an ovarian steroid-sensitive manner. Human endometrial tissues obtained from fertile women at various stages of the menstrual cycle were subjected to immunohistochemistry to localize the components of the transforming growth factor-beta (TGF-beta) system. TGF-beta receptor-I and -II expression was higher in stromal cells than in epithelial cells during the secretory phase while no such variation was observed during the proliferative phase. The expression of phosphorylated Smad3 (pSmad2/3), an activated form of a component of the TGF-beta signalling pathway, and translocation of pSmad2/3 from the cytoplasm to the nucleus were more pronounced in secretory endometrium. In coculture of human endometrial epithelial with stromal cells, each isolated from the proliferative endometrium, administration of progesterone stimulated decidualization as well as TGF-beta signalling activation in stromal cells. Progesterone also significantly elevated the concentration of TGF-beta1 in the coculture medium. Careful manipulation of the coculture, i.e. selective addition and omission of the cellular components, showed that this progesterone-induced increase in secretion of TGF-beta1 come mainly from epithelial cells. Moreover, administration of TGF-beta1 (10 ng/ml) directly to cultured stromal cells enhanced the expression of prolactin as well as pSamd2/3 even without progesterone. Taken together, our present data support the notion that progesterone induces stromal decidualization indirectly, i.e. by enhancing the expression and secretion of TGF-beta1 from epithelial cells. The secreted, epithelial-derived TGF-beta1 then acts on adjacent stromal cells, at least in part, to turn on Smad signalling that could lead to stromal decidualization.

Adenomyoma↗
PubMed2001

Backbone dynamics for the wild type and a double H52R/T56W mutant of the vnd/NK-2 homeodomain from Drosophila melanogaster.

The (15)N relaxation behavior and heteronuclear Overhauser effect data for the wild type and an H52R/T56W double mutant protein that encompasses the vnd/NK-2 homeodomain from Drosophila melanogaster were used to characterize and describe the protein backbone dynamics. This investigation, which includes a description of a model structure for the H52R/T56W double mutant vnd/NK-2 homeodomain, was carried out for the two proteins in both the free and DNA-bound states. The double residue replacement at positions 52 and 56 within the DNA recognition helix of vnd/NK-2 has been shown to lead to a significant secondary structural modification resulting in an increase in the length of the recognition helix for the unbound protein. These structural changes are accompanied by corresponding changes in the T(1) and T(1)(rho) relaxation times as well as in the heteronuclear Overhauser effect (XNOE) values that show that the structural stability of the protein is enhanced by the two residue replacements. The values of the rotational anisotropy, D(parallel)/D(perpendicular), derived from analysis of the (15)N T(1) and T(1)(rho) relaxation values are small (1.189 for the unbound homeodomain and 1.110 for the bound homeodomain; both analyzed as prolate ellipsoids of revolution). A comparison of the T(2) values of the wild type and double mutant homeodomain reveals the presence of a low-frequency exchange contribution for the wild type analogue. These relaxation studies show that the motional behavior of the protein primarily reflects the tertiary structure and stability of the homeodomain backbone as well as the respective changes induced upon site-directed residue replacement or DNA binding.

Animals↗
PubMed2001

4-Hydroxy-6-oxo-6,7-dihydro-thieno[2,3-b] pyrimidine derivatives: synthesis and their biological evaluation for the glycine site acting on the N-methyl-D-aspartate (NMDA) receptor.

Bioisostere approach has been shown to be useful to augment potency or to modify certain physiological properties of a lead compound. Based upon well documented bioisosterism, an isosteric replacement of benzene ring of 4-hydroxy-2-quinolone compound (L-695902) with a thiophene moiety was carried out to prepare the title compounds, 4-hydroxy-6-oxo-6,7-dihydro-thieno[2,3-b] pyrimidines 15. The resulting bioisosteric compounds 15 were evaluated for their antagonistic activity (binding assay) for NMDA receptor glycine site.

Animals↗
PubMed2001

Effect of particle size on the performance of batchwise centrifugal filtration.

The effect of particle size distribution on the performance of batchwise centrifugal filtration is studied. By analyzing the velocity of particles in a filter, a numerical program is designed for simulating the migration and deposition of particles. The particle size distributions and the average specific filtration resistances of cake are then estimated under various rotating speeds of the centrifuge. A large deviation of particle concentration profiles in the filter chamber will occur if the particle size distribution is not taken into consideration. A more heterogeneous cake will form under a lower rotating speed due to the sedimentation effect of particles. The predicted results of particle size distribution and average specific filtration resistance of cake agree well with the available experimental data.

Centrifugation↗
PubMed2000

Detection of antifungal activity in Portulaca oleracea by a single-cell bioassay system.

The antifungal activity of Portulaca oleracea extracts against hyphal growth of various fungi was evaluated in real time using an automatic single-cell bioassay system. Target organisms were the filamentous fungi Aspergillus and Trichophyton and the yeast Candida. A colony of test fungi was in contact with the assay medium, or assay medium containing plant extract, in sequence. The antifungal activity of each fraction of P. oleracea was evaluated based on the dynamic hyphal growth response curves of test fungi. A crude sample obtained by EtOAc extract showed a specific and marked activity against dermatophytes of the genera Trichophyton.

Antifungal Agents↗
PubMed2000

Synthesis of 7,8-dichloro-6-nitro-1H-1,5-benzodiazepine-2,4-(3H, 5H)-dione as a potential NMDA receptor glycine site antagonist.

An efficient procedure for the preparation of 7,8-dichloro-6-nitro-1H-1,5-benzodiazepine-2,4-(3H, 5H)-dione(7) as a potential lead compound for the NMDA receptor glycine binding site antagonist, starting from readily available 4,5-dichloro-2-nitroaniline(8), is described. The key step in the synthesis involves the cyclization of malonic ester amide 10 to compound 11.

Benzazepines↗
PubMed1999

Phospholipid-based microemulsions of flurbiprofen by the spontaneous emulsification process.

The purpose of this study was to investigate the possibility for parenteral delivery of flurbiprofen without chemical modification using a phospholipid-based microemulsion system. Microemulsions composed of ethyl oleate, lecithin and distearoylphosphatidyl-ethanolamine-N-poly(ethyleneglycol) 2000 (DSPE-PEG) were prepared using ethanol as a cosolvent. The effect of formulation variables on the particle size of the microemulsion was investigated. Flurbiprofen concentrations in plasma and various organs after the intravenous administration of flurbiprofen-loaded microemulsion were measured and compared with those after the intravenous administration of flurbiprofen axetil-entrapped emulsion (Lipfen(R), 50 mg/5 ml as flurbiprofen axetil) and flurbiprofen solution. Phospholipid-based microemulsions could solubilize more than 10 mg ml-1 of flurbiprofen at the ratio of vehicle to drug at least 10:1, if the oil contents (10 or 20%) of common parenteral emulsions were used. The half-life, AUC and MRT of flurbiprofen loaded in microemulsion (ethyl oleate:lecithin:DSPE-PEG:flurbiprofen=8:3:1:1.2) increased significantly. The biodistribution of flurbiprofen loaded in this microemulsion was quite different from others. Reticuloendothelial uptake of flurbiprofen loaded in microemulsion decreased compared with that in solution or Lipfen(R). It is concluded that the current microemulsion system might be applicable to formulate the parenteral dosage form of poorly water-soluble flurbiprofen without chemical modification.

Animals↗
PubMed1999

Effects of low oxygen condition on the generation of reactive oxygen species and the development in mouse embryos cultured in vitro.

OBJECTIVE(S): To elucidate the relationship between intracellular H2O2 production and embryo development in different oxygen culture conditions. METHODS: Pronuclear stage embryos were obtained from C57BL/CBA F1 and ICR mice. Measurement of H2O2 level was performed with 2',7'-dichlorodihydroflourescein diacetate and the number of blastomeres was counted after staining with 4',6'-diamidino-2-phenylindole. RESULTS: Regardless of strains, H2O2 level reached a peak at the 2-cell stage in 20% O2. But in embryos cultured in 5% O2 it was significantly lower at the 2-cell and 4-cell stages compared to those from 20% O2. The embryos cultured in 20% O2 showed developmental delay or block, but in 5% O2 these phenomena were overcome and the development was significantly increased with an infrequent fragmentation. CONCLUSIONS: Our data suggest that the 5% O2 decreases the relative concentration of H2O2 and results in improved embryo development in terms of quantity and quality without regard to type of strains.

Animals↗
PubMed1998

Conservative therapy of adnexal torsion employing color Doppler sonography.

STUDY OBJECTIVE: To ascertain if color Doppler sonography (CDS) can detect viability of adnexa in torsion. DESIGN: Concurrent, nonrandomized study (Canadian Task Force classification II-2). SETTING: Tertiary medical care center in a university hospital. PATIENTS: Twenty-seven women suspected of having torsion of the ovary in which a benign tumor was present. INTERVENTIONS: Color Doppler sonography of the ovarian pedicle suggested torsion. Laparoscopy or laparotomy was performed in 19 patients and confirmed the diagnosis. MEASUREMENTS AND MAIN RESULTS: We identified a twisted vascular pedicle of the ovarian tumor by CDS in 24 (88%) of 27 women. Pedicle arterial and venous blood flow was observed in 10 women; pathology specimens revealed normal tissue, edema with congestion, or early hemorrhage. In all nine in whom only arterial blood flow or no blood flow was observed, pathology revealed hemorrhagic necrosis. In five women with arterial and venous blood flow the tumor was managed conservatively, either by detorsion or cystectomy, after which no embolism or tumor recurrence was seen at follow-up ultrasonography. Normal follicular development and ovulation occurred in these patients. CONCLUSION: For women of childbearing age in whom torsion of benign adnexal tumors is suspected, CDS should be performed to confirm the diagnosis and ascertain whether or not pedicle arterial and venous blood flow is present. If such blood flow is detected, the adnexa is considered to be viable and detorsion or cystectomy may be performed, thus preserving the ovary. It is recommended that further studies with a larger group be undertaken to provide a basis for statistical analysis.

Abdomen, Acute↗
PubMed1998

Detection of reactive oxygen species (ROS) and apoptosis in human fragmented embryos.

In human in-vitro fertilization (IVF)-embryo transfer, the in-vitro culture environment differs from in-vivo conditions in that the oxygen concentration is higher, and in such conditions the mouse embryos show a higher concentration of reactive oxygen species (ROS) in simple culture media. ROS are believed to cause damage to cell membranes and DNA fragmentation in somatic cells. This study was conducted to ascertain the level of H2O2 concentration within embryos and the morphological features of cell damage induced by H2O2. A total of 62 human oocytes and embryos (31 fragmented, 15 non-fragmented embryos, 16 unfertilized oocytes) was obtained from the IVF-embryo transfer programme. The relative intensity of H2O2 concentrations within embryos was measured using 2',7'-dichlorodihydrofluorescein diacetate by Quanti cell 500 fluorescence imaging and DNA fragmentation was observed with transmission electron microscopy and an in-situ apoptosis detection kit. The H2O2 concentrations were significantly higher in fragmented embryos (72.21 +/- 9.62, mean +/- SEM) compared to non-fragmented embryos (31.30 +/- 3.50, P < 0.05) and unfertilized oocytes (30.75 +/- 2.67, P < 0.05). Apoptosis was observed only in fragmented embryos, and was absent in non-fragmented embryos. Electron microscopic findings confirmed apoptotic bodies and cytoplasmic condensation in the fragmented blastomeres. We conclude that there is a direct relationship between increased H2O2 concentration and apoptosis, and that further studies should be undertaken to confirm these findings.

Apoptosis↗
PubMed1997

NMR structure determination of a novel conotoxin, [Pro 7,13] alpha A-conotoxin PIVA.

A high-resolution solution conformation of a novel conotoxin, [Pro 7,13] alpha A-conotoxin PIVA, GCCGSYPNAACHPCSCKDROSYCGQ-NH2, has been determined by two-dimensional 1H NMR methods and distance geometry calculations. The total of 324 NOE-derived interproton distance restraints including 33 long-range NOE restraints as well as 11 phi and 7 chi 1 torsion angle restraints was used for computation of structures. Back-calculation from the experimental NOE spectrum has provided 49 new NOE restraints and yielded the final R-factors of Ra = 0.641 and Rb = 0.157. The final RMSD values are 0.90 and 1.16 A for the backbone and the heavy atoms, respectively. The C-terminal half of the molecule involving the residues 12-24 is extremely well-defined with a backbone RMSD value of 0.56 A, whereas the N-terminal 3-11 disulfide loop is relatively flexible, possessing a backbone RMSD value of 1.09 A. The [Pro 7,13] alpha A-conotoxin PIVA does not contain any significant secondary structure although the 21S-24G nearly completes one turn of a 3(10) helix. The overall protein fold is largely maintained by the three disulfide bridges of 2-16, 3-11, and 14-23. The presence of the three disulfide bridges imposes geometric constraints that force the molecule to form six continuous bends involving the following residues: 3C-5S, 7P-10A, 12H-14C, 15S-17K, 17K-19R, and 21S-25Q. The overall shape of the [Pro 7,13] alpha A-conotoxin PIVA can be described as an "iron". Residues 15S-19R form a loop that protrudes out of the "bottom plate" formed by the rest of the protein and constitute the handle of the iron. The N-terminal tip of the molecule is relatively immobile due to attractive electrostatic interactions between the gamma-hydroxyl group of 20 Hyp and the phenolic hydroxyl group of 22Y. The flexible 3-11 disulfide loop consists mostly of hydrophobic residues, while the best-defined 14-23 disulfide loop contains the highly charged hydrophilic 15S-19R "handle" domain exposed to the exterior of the protein. Binding to nicotinic acetylcholine receptor can be mediated through two different types of interactions: one involving the aromatic hydrophobic residues such as 6Y and 12H and the other involving the positively charged hydrophilic side chain of the 19R. The side chain of the 19R in the [Pro 7, 13] alpha A-conotoxin PIVA and that of the 9R of the alpha-conotoxin G1, and also the side chains of the 12H and 6Y in the former and those of 10H and 11Y in the latter can be aligned to point to the same direction when the corresponding backbone atoms are superimposed to an RMSD value of 2.5 A.

Amino Acid Sequence↗
PubMed1997

Synthesis and purification of NB1-palmitoyl insulin.

A procedure for synthesizing NB1-palmitoyl insulin for incorporation into liposomes for targeting to hepatocytes was developed. The amino group of the first amino acid phenylalanine on the B chain (B1) of insulin was selected for conjugation with palmitic acid in anticipation that its binding to the insulin receptor would be preserved. Two other free amino groups present in insulin, the first amino acid glycine on the A chain (A1) and the 29th amino acid lysine on the B chain (B29), were first protected with a t-butoxycarbonyloxy (t-Boc) group to yield NA1, B29-di-(t-Boc) insulin. The identity of this di-(t-Boc) insulin was confirmed by amino acid analysis as well as by enzyme hydrolysis coupled with matrix-assisted laser-desorption time of flight mass spectrometry (MALDI-TOF MS). NA1,B29-Di-(t-Boc) insulin was then reacted with the N-hydroxysuccinimide ester of palmitic acid, followed by deblocking the t-Boc groups, to yield NB1-palmitoyl insulin, the structure of which was further confirmed by MALDI-TOF MS analysis. NB1-palmitoyl insulin was found to interact with the insulin receptor on fat cells, thereby catalyzing the conversion of [14C]glucose into lipids, at reduced efficiency (30-40%).

Animals↗
PubMed1995

Studies using fluorescent tetrahydrochrysene estrogens for in situ visualization of the estrogen receptor in living cells.

We have analyzed four fluorescent nonsteroidal estrogens for their potential to serve as vital cytological stains to visualize the estrogen receptor (ER) in a model receptor expression system. The novel estrogen fluorophores are based on the rigidified stilbene-like structure of 5,6,11,12-tetrahydrochrysene (THC), and they embody electron-donor (hydroxyl) and electron-acceptor groups (nitrile, amide, ester, or ketone) that afford efficient, long wavelength, and environment-sensitive fluorescence. These probes bind with high affinity to human ER (relative binding affinity, 22-85 vs. estradiol, 100), and they stimulate the transcriptional activity of this receptor. The strong fluorescence of the estrogenic THCs permits visualization, using conventional epifluorescence microscopy, of ER in transfected Cos-7 cells that express elevated levels of receptor. Cell staining by the donor-acceptor THCs characteristically displays a nonuniform pattern of nuclear fluorescence that can be fully inhibited by nonfluorescent estrogens such as estradiol or diethylstilbestrol. Additionally, this staining appears to be specific for ER, since it coincides with the distribution of receptor as determined by indirect immunofluorescence analysis using an ER-specific monoclonal antibody. Using these probes, we have analyzed the intracellular distribution of ER mutants containing a variety of deletions. Evidence is presented to show that removal of amino-terminal sequences within the ER polypeptide results in an altered pattern of intranuclear distribution with preferential accumulation of receptor protein within the nucleolus. These THC fluorophores therefore represent excellent probes for cytological studies involving ER expressed in cultured cells and represent an important advance toward the goal of exploiting fluorescence technology to analyze the expression and distribution of ER within tissue samples.

Animals↗
PubMed1994

Identification of Borrelia burgdorferi isolated in Korea using outer surface protein A (OspA) serotyping system.

Two characteristic strains (935T, 934U) of B. burgdorferi isolated from Ixodes persulcatus and a wild rodent (Apodemus agrarius) in Korea were selected and analyzed by an immunoblot method using the monoclonal antibodies directed to different epitopes of outer surface protein A (OspA). The reactive pattern of strain 934U with these monoclonal antibodies was identical to that of strains belonging to B. afzelii and that of strain 935T was different from other isolates. Monoclonal antibody (5TEE3) which is specific to strain 935T did not react with any other Western and Japanese isolates. So, it was suggested that there exist at least two groups of B. burgdorferi in Korea. One could be classified as B. afzelii and the other is a divergent group from three known species of B. burgdorferi sensu stricto, B. garinii and B. afzelii.

Animals↗
PubMed1993

Use of the gamma-ray perturbed angular correlation (PAC) technique for monitoring liposomal phospholipid bilayer integrity.

A membrane labeling method based on the principle of gamma-ray perturbed angular correction (PAC) was developed to monitor the structural integrity of liposomal membranes. The reporter group was 111In(III) complexed with the lipophilic diethylenetriaminepentaacetic acid (DTPA) derivative of dipalmitoylphosphatidylethanolamine (DPPE) embedded in the phospholipid bilayers of small unilamellar liposomes. Using this method, complete chemical digestion of the constituent phospholipids in these DTPA-conjugated liposomes by phospholipase A2 or phospholipase C in the presence of Ca2+ was found not to be followed by an immediate disruption of the liposomal membrane. Compared with other methods, the method developed permits the continuous noninvasive monitoring of the microenvironment of the lipid bilayer at the molecular level. It may potentially be applicable to evaluate liposomal fusion, screen for penetration enhancers under development for enhancement in mucosal drug penetration, and monitor liposomal degradation within the living animal.

Gamma Rays↗
PubMed1993

A fluorescence quenching method for estimating chelating groups in chelate-conjugated macromolecules.

A terbium-dipicolinic acid (Tb-DPA) fluorescence quenching method for estimating free chelating groups conjugated to protein molecules was developed. This method was based on competitive displacement of DPA from binding to terbium by stronger chelating groups such as diethylenetriaminepentaacetic acid (DTPA), EDTA, nitrilotriacetic acid (NTA), DTPA-conjugated bovine serum albumin (BSA-DTPA), or DTPA-conjugated immunoglobulin G (IgG-DTPA), resulting in a significant reduction in terbium fluorescence. The chelating ability of the tested reagent, from high to low, was in the following order: BSA-DTPA > DTPA > IgG-DTPA > EDTA, NTA. At low terbium concentrations, the reduction was linear for DTPA. This fluorescence quenching method was not only rapid, simple, and as accurate as conventional radiosotopic or chromatographic methods, but also sensitive and reproductible. The detection limit was 10 nM for DTPA. The interrun coefficient of variation was at most 8%. The advantage of this method over other indirect methods is that it reveals the actual chelating ability of the tested macromolecule, unencumbered by complicating factors such as trace metal contamination and dimer/polymer formation during conjugation.

Chelating Agents↗