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Y W Chen

Publications and source records attributed to Y W Chen.

At least 37 records · Page 2Linked to original sources
PubMed2000

Enantiomeric separation of some pharmaceutical intermediates and reversal of elution orders by high-performance liquid chromatography using cellulose and amylose tris(3,5-dimethylphenylcarbamate) derivatives as stationary phases.

Several pairs of enantiomers of pharmaceutical intermediates were separated by HPLC directly on cellulose and amylose tris(3,5-dimethylphenylcarbamate) derivatives (Chiralcel OD and Chiralpak AD) using hexane as mobile phase with 2-propanol or ethanol as modifier. The separation and elution order of the enantiomers on the two columns using different alcohol modifiers were compared. Reversal of the elution order of some enantiomeric pairs associated with increased retention of many of these solutes upon changing the mobile phase modifier from 2-propanol to ethanol was observed. The effect of structural variation of two pairs of enantiomers on their k' and separation factor alpha was noted. Chiralcel OD and Chiralpak AD columns provided different retention, separation and elution order of some of the enantiomeric pairs.

Amylose↗
PubMed2000

Expression profiling in the muscular dystrophies: identification of novel aspects of molecular pathophysiology.

We used expression profiling to define the pathophysiological cascades involved in the progression of two muscular dystrophies with known primary biochemical defects, dystrophin deficiency (Duchenne muscular dystrophy) and alpha-sarcoglycan deficiency (a dystrophin-associated protein). We employed a novel protocol for expression profiling in human tissues using mixed samples of multiple patients and iterative comparisons of duplicate datasets. We found evidence for both incomplete differentiation of patient muscle, and for dedifferentiation of myofibers to alternative lineages with advancing age. One developmentally regulated gene characterized in detail, alpha-cardiac actin, showed abnormal persistent expression after birth in 60% of Duchenne dystrophy myofibers. The majority of myofibers ( approximately 80%) remained strongly positive for this protein throughout the course of the disease. Other developmentally regulated genes that showed widespread overexpression in these muscular dystrophies included embryonic myosin heavy chain, versican, acetylcholine receptor alpha-1, secreted protein, acidic and rich in cysteine/osteonectin, and thrombospondin 4. We hypothesize that the abnormal Ca(2)+ influx in dystrophin- and alpha-sarcoglycan-deficient myofibers leads to altered developmental programming of developing and regenerating myofibers. The finding of upregulation of HLA-DR and factor XIIIa led to the novel identification of activated dendritic cell infiltration in dystrophic muscle; these cells mediate immune responses and likely induce microenvironmental changes in muscle. We also document a general metabolic crisis in dystrophic muscle, with large scale downregulation of nuclear-encoded mitochondrial gene expression. Finally, our expression profiling results show that primary genetic defects can be identified by a reduction in the corresponding RNA.

Actins↗
PubMed2000

CD44 interaction with c-Src kinase promotes cortactin-mediated cytoskeleton function and hyaluronic acid-dependent ovarian tumor cell migration.

In this study we have demonstrated that both CD44 (the hyaluronan (HA) receptor) and c-Src kinase are expressed in human ovarian tumor cells (SK-OV-3.ipl cell line), and that these two proteins are physically associated as a complex in vivo. Using a recombinant cytoplasmic domain of CD44 and an in vitro binding assay, we have detected a specific interaction between CD44 and c-Src kinase. Furthermore, the binding of HA to SK-OV-3.ipl cells promotes c-Src kinase recruitment to CD44 and stimulates c-Src kinase activity, which, in turn, increases tyrosine phosphorylation of the cytoskeletal protein, cortactin. Subsequently, tyrosine phosphorylation of cortactin attenuates its ability to cross-link filamentous actin in vitro. In addition, transfection of SK-OV-3.ipl cells with a dominant active form of c-Src (Y527F)cDNA promotes CD44 and c-Src association with cortactin in membrane projections, and stimulates HA-dependent/CD44-specific ovarian tumor cell migration. Finally, overexpression of a dominant-negative mutant of c-Src kinase (K295R) in SK-OV-3.ipl cells impairs the tumor cell-specific phenotype. Taken together, these findings strongly suggest that CD44 interaction with c-Src kinase plays a pivotal role in initiating cortactin-regulated cytoskeleton function and HA-dependent tumor cell migration, which may be required for human ovarian cancer progression.

Animals↗
PubMed2000

3,5-Bis(trifluoromethyl)pyrazoles: a novel class of NFAT transcription factor regulator.

A series of bis(trifluoromethyl)pyrazoles (BTPs) has been found to be a novel inhibitor of cytokine production. Identified initially as inhibitors of IL-2 synthesis, the BTPs have been optimized in this regard and even inhibit IL-2 production with a 10-fold enhancement over cyclosporine in an ex vivo assay. Additionally, the BTPs show inhibition of IL-4, IL-5, IL-8, and eotaxin production. Unlike the IL-2 inhibitors, cyclosporine and FK506, the BTPs do not directly inhibit the dephosphorylation of NFAT by calcineurin.

Animals↗
PubMed2000

Ankyrin-Tiam1 interaction promotes Rac1 signaling and metastatic breast tumor cell invasion and migration.

Tiam1 (T-lymphoma invasion and metastasis 1) is one of the known guanine nucleotide (GDP/GTP) exchange factors (GEFs) for Rho GTPases (e.g., Rac1) and is expressed in breast tumor cells (e.g., SP-1 cell line). Immunoprecipitation and immunoblot analyses indicate that Tiam1 and the cytoskeletal protein, ankyrin, are physically associated as a complex in vivo. In particular, the ankyrin repeat domain (ARD) of ankyrin is responsible for Tiam1 binding. Biochemical studies and deletion mutation analyses indicate that the 11-amino acid sequence between amino acids 717 and 727 of Tiam1 ((717)GEGTDAVKRS(727)L) is the ankyrin-binding domain. Most importantly, ankyrin binding to Tiam1 activates GDP/GTP exchange on Rho GTPases (e.g., Rac1). Using an Escherichia coli-derived calmodulin-binding peptide (CBP)-tagged recombinant Tiam1 (amino acids 393-728) fragment that contains the ankyrin-binding domain, we have detected a specific binding interaction between the Tiam1 (amino acids 393-738) fragment and ankyrin in vitro. This Tiam1 fragment also acts as a potent competitive inhibitor for Tiam1 binding to ankyrin. Transfection of SP-1 cell with Tiam1 cDNAs stimulates all of the following: (1) Tiam1-ankyrin association in the membrane projection; (2) Rac1 activation; and (3) breast tumor cell invasion and migration. Cotransfection of SP1 cells with green fluorescent protein (GFP)-tagged Tiam1 fragment cDNA and Tiam1 cDNA effectively blocks Tiam1-ankyrin colocalization in the cell membrane, and inhibits GDP/GTP exchange on Rac1 by ankyrin-associated Tiam1 and tumor-specific phenotypes. These findings suggest that ankyrin-Tiam1 interaction plays a pivotal role in regulating Rac1 signaling and cytoskeleton function required for oncogenic signaling and metastatic breast tumor cell progression.

Animals↗
PubMed2000

Isolation and characterization of proteins that bind to galactose, lipopolysaccharide of Escherichia coli, and protein A of Staphylococcus aureus from the hemolymph of Tachypleus tridentatus.

In this study, we report the isolation and characterization of three novel hemolymph proteins that are believed to be involved in the innate immune response of horseshoe crabs, Tachypleus tridentatus. They include two closely related proteins, one that binds to the protein A of Staphylococcus aureus (PAP) and another that binds to the lipopolysaccharide of Escherichia coli (LBP). PAP binds specifically to staphylococcal protein A (SpA) with a K(D) of 3.86 x 10(-5) M, whereas LBP binds to lipopolysaccharide (LPS) with a K(D) of 1.03 x 10(-6) M. Both PAP and LBP are glycoproteins with an apparent molecular mass of about 40 kDa. N-terminal sequences of PAP and LBP showed 61.9 and 72.2% identity, respectively, to tachylectin-3, a lectin isolated from the amebocyte of T. tridentatus, previously characterized by its affinity to the O-antigen of LPS and blood group A antigen (Muta, T., and Iwanaga, S. (1996) Curr. Opin. Immunol. 8, 41-47). The third protein, a galactose-binding protein (GBP), was found to bind tightly to Sepharose CL-4B and could only be eluted from the column matrix with chaotropic agents, such as 4 M urea or 2 M guanidine hydrochloride. Further analysis indicated that GBP binds to D(+)-galactose with a K(D) of 2.47 x 10(-7) M. N-terminal sequence analysis showed that GBP shared a 50% identity with lectin L-6, identified in the granules of amebocyte of T. tridentatus. (Gokudan, S., Muta, T., Tsuda, R., Koori, K., Kawahara, T., Seki, N., Mizunoe, Y., Wai, S. N. , Iwanaga, S., and Kawabata, S. (1999) Proc. Natl. Acad. Sci. U. S. A. 96, 10086-10091). Lectin-L6 and tachylectin-3 are nonglycosylated intracellular proteins with about half the molecular mass of PAP, LBP, and GBP. GBP also binds to PAP and LBP with K(D) values of 1.25 x 10(-7) and 1.43 x 10(-8) M, respectively, and this binding is enhanced about 10-fold upon the addition of SpA and LPS to form the GBP.PAP.SpA and GBP.LBP.LPS complexes, respectively.

Amino Acid Sequence↗
PubMed2000

CD44 interaction with tiam1 promotes Rac1 signaling and hyaluronic acid-mediated breast tumor cell migration.

In this study we have explored the interaction between CD44 (the hyaluronic acid (HA)-binding receptor) and Tiam1 (a guanine nucleotide exchange factor) in metastatic breast tumor cells (SP1 cell line). Immunoprecipitation and immunoblot analyses indicate that both the CD44v3 isoform and the Tiam1 protein are expressed in SP1 cells and that these two proteins are physically associated as a complex in vivo. Using an Escherichia coli-derived calmodulin-binding peptide-tagged Tiam1 fragment (i.e. the NH(2)-terminal pleckstrin homology (PHn) domain and an adjacent protein interaction domain designated as PHn-CC-Ex, amino acids 393-738 of Tiam1) and an in vitro binding assay, we have detected a specific binding interaction between the Tiam1 PHn-CC-Ex domain and CD44. Scatchard plot analysis indicates that there is a single high affinity CD44 binding site in the PHn-CC-Ex domain of Tiam1 with an apparent dissociation constant (K(d)) of 0.2 nM, which is comparable with CD44 binding (K(d) = approximately 0.13 nM) to intact Tiam1. These findings suggest that the PHn-CC-Ex domain is the primary Tiam1-binding region for CD44. Most importantly, the binding of HA to CD44v3 of SP1 cells stimulates Tiam1-catalyzed Rac1 signaling and cytoskeleton-mediated tumor cell migration. Transfection of SP1 cells with Tiam1cDNA promotes Tiam1 association with CD44v3 and up-regulates Rac1 signaling as well as HA/CD44v3-mediated breast tumor cell migration. Co-transfection of SP1 cells with PHn-CC-Ex cDNA and Tiam1 cDNA effectively inhibits Tiam1 association with CD44 and efficiently blocks tumor behaviors. Taken together, we believe that the linkage between CD44v3 isoform and the PHn-CC-EX domain of Tiam1 is required for HA stimulated Rac1 signaling and cytoskeleton-mediated tumor cell migration during breast cancer progression.

Adjuvants, Immunologic↗
PubMed2000

Postero-medio-anterior approach of the ankle for the pilon fracture.

A good view of the operative field is important for better reduction and fixation in surgical treatment of fractures. The exposure of the ankle joint for the pilon fracture is commonly through the anterior approach, or combined with the medical approach. But sometimes it is still difficult to have complete viewing of the articular surface and to apply internal fixation by that approach. In recent years, we developed a "postero-medio-anterior" approach of the ankle joint by one incision. This approach provides an excellent exposure of the anterior, medial and posterior aspects of the ankle joint with a clear view of the articular surface. In our 45 cases of pilon fracture during 1991 to 1995, there was no incisional injury to the neurovascular bundle. Superficial wound edge necrosis was noted in two cases which healed later without further procedure. Therefore, we recommend this approach as a simple and reliable incision for open reduction of pilon fractures.

Adult↗
PubMed2000

Crystallization and preliminary crystallographic studies of a SAM domain at the C-terminus of human p73alpha.

p73 is a recently discovered homologue of the tumour suppressor p53 and contains all three functional domains of p53. The alpha-splice variant of p73 (p73alpha) contains an additional structural domain near its C--terminus that has sequence homology with the sterile alpha-motif (SAM) domain. This domain is considered to be responsible for mediating protein-protein interactions. Pyramidal crystals of human p73alpha SAM domain were obtained by the hanging-drop vapour-diffusion method with ammonium dihydrogen orthophosphate as the precipitant. The crystals diffract to 2.54 A resolution and belong to the tetragonal space group P4(1)2(1)2, with unit-cell parameters a = b = 32.02, c = 133.84 A. The structure was solved by molecular replacement using the NMR structure of the same protein as the search model.

Amino Acid Motifs↗
PubMed2000

A systematic case study on using NMR models for molecular replacement: p53 tetramerization domain revisited.

Molecular replacement using search models derived from nuclear magnetic resonance (NMR) spectroscopy has often proved problematic. It has been known for some time that the overall differences in atomic positions (r.m.s.d.) between the crystalline and the solution states of the same protein are of the order of 1-2 A and approach the limit of molecular replacement. In most cases, this structural difference is a result of calculating the NMR structure with insufficient data, yielding an NMR structure of limited accuracy. A systematic case study was performed to investigate the use of NMR models for molecular replacement on the p53 tetramerization domain: NMR search models of varying degrees of accuracy were employed to solve phases for the 1.5 A X-ray diffraction data. An approximate correlation was found between the accuracy of the NMR search model and the clarity and quality of the molecular-replacement solution. It was found that ensemble models perform better than single averaged models and have a larger tolerance in model inaccuracy. Also, distance-derived B factors can improve the performance of single models.

Crystallography, X-Ray↗
PubMed2000

Carotid and transcranial color-coded duplex sonography in different types of carotid-cavernous fistula.

BACKGROUND AND PURPOSE: Patients with carotid-cavernous fistula (CCF) may undergo direct or indirect shunting. Ultrasonography has value that is complementary to angiography in the assessment and follow-up of these patients. The aim of this study was to characterize findings provided by carotid duplex sonography (CDS) and transcranial color-coded duplex sonography (TCCD) in patients with different types of CCF. METHODS: CDS and TCCD were independently performed by technologists and neurologists. Digital subtraction or MR angiography was interpreted by a neuroradiologist. Ultrasonographic studies were categorized into 4 types: I, direct shunting only; II, direct shunting with a carotid aneurysm; III, indirect shunting only; and IV, mixed (direct and indirect) shunting. In addition to carotid and intracranial flow velocities, volume, and pulsatility, other direct and indirect ultrasound signs of shunting were evaluated. The direct sign of CCF was a mosaic flash detected by TCCD. Alteration of hemodynamic parameters on CDS and demonstration of draining veins with the use of TCCD were considered indirect signs. RESULTS: Fifteen patients (8 men, 7 women) were included in the study. According to angiographic results, patients in ultrasonographic classification types I (n=7) and II (n=3) corresponded to type A of Barrow's classification. Patients with type III (n=8) were Barrow's type C. Type IV (n=1) had a combination of Barrow's types A and C. On ultrasound, both direct and indirect signs were seen in types I, II, and IV CCF. The presence of a 2-colored oval mass divided by a zone of separation without turbulence differentiated type I from type II CCF. All patients with type III CCF had indirect signs, and only 1 patient had direct signs on TCCD. Abnormal TCCD findings were most commonly seen through the transorbital window (100%), followed by the transtemporal window (63%) and transforaminal window (40%). CONCLUSIONS: If only indirect ultrasonographic signs of CCF are present, TCCD can be used to predict an indirect CCF type on the basis of the origin of the fistula. With direct communication between carotid artery and cavernous sinus, both direct and indirect ultrasonographic signs can be found. The combination of CDS/TCCD may provide a noninvasive and reliable way to classify patients with CCF.

Adult↗
PubMed2000

The interference of uptake of thallium-201 in cultured rat myocardial cells with existence of potassium related pharmaceuticals--a preliminary report.

Thallium-201 myocardial perfusion imaging is wildly used to detect and assess the extent of jeopardized myocardial ischemia in the coronary artery disease and the viability of myocardium post infarction. In recent years, there has been a great deal of pharmacological development of blockers and openers of potassium channel. In this study, we will discuss the interference of uptake of thallium-201 ion in cultured neonatal rat myocytes with existence of a variety of pharmacological agents. The cultures of neonatal rat myocardial cells were incubated with different agents such as potassium chloride, sodium-potassium ATPase pump inhibitor (ouabain), cesium compound, variable potassium channel blockers (4 AP, TEA and glibenclamide) and their openers (minoxidil, and cromakalim). The radioactivity of intracellular thallium-201 that could enter rat myocardial cells was detected by gamma counter sixty minutes after thallium-201 was added. In this study we found that thallium and potassium ions behave in an analogous manner in cultured rat myocardial cells. Both 2.5 mM and 5 mM concentration of extracellular potassium ion significantly result in reduction of thallium-201 ion influx in rat myocardial cells. 0.5 mM ouabain, an inhibitor of sodium-potassium ATPase pump, reduced about 40% of influx of thallium-201 ion in cultured rat myocardial cells via active transport. Combination of both potassium ion and ouabain inhibit most of thallium-201 ions influx in myocardial cells, but it is not completely inhibited. Cesium, a potassium antagonist, also interferes with the uptake of thallium-201 in cultured rat myocytes in our study. The most interesting finding in our investigation is that potassium channel blockers such as TEA and glibenclamide, inhibit the influx of thallium-201 in myocytes. However, potassium channel openers have no overt effect on influx of thallium-201 in cultured rat myocytes. We indirectly observe about 60% of influx of thallium-201 ion into cultured rat myocardial cells via active sodium-potassium ATPase pump. Potassium, cesium and potassium channel blockers, such as TEA and glibenclamide, inhibited the different percentage of influx of thallium-201 in cultured rat myocardial cells in this study.

Adenosine Triphosphate↗
PubMed2000

Tc99m sulfur colloid scintigraphy for detection of intrathoracic extramedullary hematopoiesis in patient with beta-thalassemia major--a case report.

Extramedullary hematopoiesis (EMH) refers to the production of blood cells outside the bone marrow and is a compensatory mechanism for bone marrow dysfunction. A 34 year-old female patient with beta thalassemia major was found to have multiple large, well-circumscribed radiopaque paravertibral mass lesions in chest radiography. Magnetic resonance imaging (MRI) of the thorax disclosed a right upper apical and two lower thoracic paraspinal mass lesions with heterogeneous isointensity on T1-weighted images and hypointensity on T2-weighted images. Because intrathoracic EMH is suspected in our case, which had obvious bone marrow dysfunction, radionuclide bone marrow scintigraphy is helpful in supporting the diagnosis. Tc99m sulfur colloid scintigraphy demonstrated three intense radioactive thoracic paraspinal mass lesions corresponding to the lesions seen on MRI. We believe whole body bone marrow scintigraphy with Tc99m sulfur colloid is the best convenient noninvasive method for supporting the diagnosis of EMH.

Adult↗
PubMed1999

Application and comparison of derivatized cellulose and amylose chiral stationary phases for the separation of enantiomers of pharmaceutical compounds by high-performance liquid chromatography.

The direct high-performance liquid chromatographic separation of three pairs of structurally related enantiomers on derivatized cellulose and amylose chiral stationary phases (Chiralcel OD, Chiralpak AD and Chiralpak AS) was studied using hexane as the mobile phase with 2-propanol or ethanol as modifiers. The separation, retention and elution order of the enantiomers on the different columns using different alcohol modifiers were compared. The effect of structural variation of the solutes on their k' was noted. A reversal of elution order of one enantiomeric pair upon changing the mobile-phase modifier was observed. Chiralcel OD and Chiralpak AD columns provided different elution orders of the enantiomers, including a fourth pair of enantiomers that were not structurally related to the other three pairs.

Amylose↗
PubMed1999

Inhibition of p56(lck) tyrosine kinase by isothiazolones.

Lck encodes a 56-kDa protein-tyrosine kinase, predominantly expressed in T lymphocytes, crucial for initiating T cell antigen receptor (TCR) signal transduction pathways, culminating in T cell cytokine gene expression and effector functions. As a consequence of a high-throughput screen for selective, novel inhibitors of p56(lck), an isothiazolone compound was identified, methyl-3-(N-isothiazolone)-2-thiophenecarboxylate(A-125800), which inhibits p56(lck) kinase activity with IC50 = 1-7 microM. Under similar assay conditions, the isothiazolone compound was equipotent in blocking the ZAP-70 tyrosine kinase activity but was 50 to 100 times less potent against the catalytic activities of p38 MAP kinase and c-Jun N-terminal kinase 2alpha. A-125800 blocked activation-dependent TCR tyrosine phosphorylation and intracellular calcium mobilization in Jurkat T cells (IC50 = 35 microM) and blocked T cell proliferation in response to alloantigen (IC50 = 14 microM) and CD3/CD28-induced IL-2 secretion (IC50 = 2.2 microM) in primary T cell cultures. Inhibition of p56(lck )by A-125800 was dose- and time-dependent and was irreversible. A substitution of methylene for the sulfur atom in the isothiazolone ring of the compound completely abrogated the ability to inhibit p56(lck) kinase activity and TCR-dependent signal transduction. Incubation with thiols such as beta-ME or DTT also blocked the ability of the isothiazolone to inhibit p56(lck) kinase activity. LC/MS analysis established the covalent modification of p56(lck) at cysteine residues 378, 465, and 476. Together these data support an inhibitory mechanism, whereby cysteine -SH groups within the p56(lck) catalytic domain react with the isothiazolone ring, leading to ring opening and disulfide bond formation with the p56(lck) enzyme. Loss of p56(lck) activity due to -SH oxidation has been suggested to play a role in the pathology of AIDS. Consequently, a similar mechanism of sulfhydryl oxidation leading to p56(lck) inhibition, described in this report, may occur in the intact T cell and may underlie certain T cell pathologies.

Adenosine Triphosphate↗
PubMed1999

TWEAK induces angiogenesis and proliferation of endothelial cells.

TWEAK is a recently described member of the Tumor Necrosis Factor (TNF) ligand family whose transcripts are present in a wide variety of human tissues (Chicheportiche, Y., Bourdon, P. R., Xu, H., Hsu Y. M., Scott, H., Hession, C., Garcia, I., and Browning, J. L. (1997) J. Biol. Chem. 272, 32401-32410). TWEAK is a weak inducer of apoptosis in transformed cells when administered with interferon-gamma or cycloheximide (Chicheportiche, Y., Bourdon, P. R., Xu, H., Hsu Y. M., Scott, H., Hession, C., Garcia, I., and Browning, J. L. (1997) J. Biol. Chem. 272, 32401-32410; Masters, S. A., Sheridan, J. P., Pitti, R. M., Brush, A. G., and Ashkenazi, A. (1998) Curr. Biol. 8, 525-528) and also promotes IL-8 secretion in cultured cells. We report here that picomolar concentrations of recombinant soluble TWEAK induce proliferation in a variety of normal human endothelial cells and in aortic smooth muscle cells and reduce culture requirements for serum and growth factors. Blocking antibodies to Vascular Endothelial Growth Factor (VEGF) do not significantly inhibit TWEAK-induced proliferation, indicating that TWEAK does not function indirectly through up-regulation of VEGF. Pellets containing TWEAK induce a strong angiogenic response when implanted in rat corneas, suggesting a role for TWEAK in vasculature formation in vivo.

Animals↗
PubMed1999

Crystal structure of a dimeric chymotrypsin inhibitor 2 mutant containing an inserted glutamine repeat.

We have constructed mutants of chymotrypsin inhibitor 2 with short glutamine repeats inserted into its inhibitory loop. These mutants oligomerize when expressed in Escherichia coli. The dimer of a mutant with four glutamines now has been crystallized, and its structure has been solved by molecular replacement by using the wild-type monomer as a search model. The structure of each half of the dimer is found to be the same as that of the wild-type monomer, except around the glutamine insertion. It was proposed that the components of the oligomers are held together by hydrogen bonds between the main-chain and side-chain amides of the glutamine repeats. Instead, they appear to form by swapping domains on folding in E. coli, and the glutamine repeats connecting the components of the dimers are disordered.

Amino Acid Sequence↗