Biomedical subjects

Y Cheng

Publications and source records attributed to Y Cheng.

At least 91 records · Page 5Linked to original sources

PubMed2000

Transient suppression of cortactin ectopically induces large telencephalic neurons towards a GABAergic phenotype.

Excitatory and inhibitory neuronal cell fates require specific expression of both neurotransmitter and morphological phenotypes. The role of the F-actin cytoskeleton in morphological phenotypes has been well documented, but its role in neurotransmitter phenotype expression remains unknown. Here we present evidence that the F-actin binding protein cortactin participates in determining both aspects of cell fate in large telencephalic neurons. We show that the expression of cortactin was upregulated early in development just prior to appearance of GABAergic neurons in the chick telencephalon at embryonic day 6. This program was faithfully maintained in primary neuronal cultures derived from E6 telencephalon, where immature neurons differentiate either to large pyramidal and large stellate excitatory neurons or to small inhibitory GABAergic neurons. Immunostaining revealed that cortactin was enriched in areas of membrane budding, growth cones, and in the cell cortex of immature neurons. With differentiation, intense punctate staining was also observed in an extraction-resistant cytosolic compartment of the soma and processes. More importantly, suppression of cortactin by inhibition of cortactin mRNA translation with antisense oligonucleotides caused permanent phenotypic changes. Specifically, a transient suppression of cortactin was achieved in immature neurons with a single exposure to antisense oligonucleotides. This inhibition first induced both the expression of mRNA and the enzymatic activity of GAD significantly earlier than in control neurons. Second, cortactin-suppressed large projectional neurons exhibited significantly shorter processes and growth cones with protrusive filopodia and an enlarged lamellipodia veil. Most importantly, this remodeling of neuritic outgrowth in projectional somata was accompanied by the ectopic induction of GABA (*-aminobutyric acid) expression. Considering this data altogether, it appears that cortactin may function to suppress concurrently several parameters of the GABAergic program in large developing neurons.

Animals↗

PubMed2000

Lanthanides enhance pulmonary absorption of insulin.

In an effort to investigate the enhancement effect of lanthanide ions (Ln3+) on the absorption of larger molecules from the pulmonary pathway, insulin (mol. wt. = 5730) was chosen as a model peptide. The absorption of insulin preadministered or coadministered with Ln3+ from the lung was investigated by means of an in situ pulmonary absorption experiment. The enhancement absorption of insulin by Ln3+ ions was evaluated by calculating the various bioavailabilities (Fr) of insulin from pulmonary absorption. Moreover, the temporal change of Gd content in serum was also investigated. Results showed that the promoting effect of Ln3+ on the bioavailability of insulin is closely related to its species, concentration, and delivery order. The effect of the median Ln3+ series was remarkably greater than that of light and heavy Ln3+. The anionic form of Gadolinium (Fr = 68.4%) seemed to be more effective compared with its cationic form (Fr = 59.5%). Coadministration of Gd3+ with insulin (Fr = 80.1%) was the most effective in increasing insulin absorption from the lung. Gd3+ was rapidly absorbed and metabolized to a normal level after 4 h. It was suggested that lanthanides in a very low concentration might become potent absorption enhancers to improve absorption of larger molecules via the pulmonary pathway.

Absorption↗

PubMed2000

Biclustering of expression data.

An efficient node-deletion algorithm is introduced to find submatrices in expression data that have low mean squared residue scores and it is shown to perform well in finding co-regulation patterns in yeast and human. This introduces "biclustering", or simultaneous clustering of both genes and conditions, to knowledge discovery from expression data. This approach overcomes some problems associated with traditional clustering methods, by allowing automatic discovery of similarity based on a subset of attributes, simultaneous clustering of genes and conditions, and overlapped grouping that provides a better representation for genes with multiple functions or regulated by many factors.

Algorithms↗

PubMed2000

[Glucose isomerase gene knock-out by denatured double-stranded DNA].

After Genetic background analysis of Streptomyces diastaticus No. 7 strain M1033, the modified conditions of M1033 protoplasts and transformation were established. Replacement plasmid for homologous recombination was also constructed by inserting tsr gene into glucose isomerase gene. The homologous recombination of GI gene in M1033 chromosomes was achieved by using denatured linearized DNA fragments and glucose isomerase deficient strain M1033LJ was obtained. It is basic for introducing mutation into M1033 chromosome and realizing site-directed molecular reformation.

Aldose-Ketose Isomerases↗

PubMed2000

Review of pre-eclampsia in Taiwan: a multi-institutional study.

BACKGROUND: Because the reported frequency of pre-eclampsia in Taiwan varies significantly, the aims of this study were to measure the current incidence of pre-eclampsia and its correlated morbidity and mortality for both mothers and fetuses in Taiwan. METHODS: We retrospectively studied all reported cases of pre-eclampsia and eclampsia from January 1, 1993 to December 31, 1997 in the 14 tertiary medical centers and regional hospitals in Taiwan. Recruiting criteria were pregnancy-induced hypertension (systolic blood pressure > or = 140 mmHg or diastolic blood pressure > or = 90 mmHg) with proteinuria (> or = 300 mg of urinary protein per 24 hours) and independent part edema. RESULTS: There were 4,193 patients with pre-eclampsia and eclampsia for a frequency of 2.03% of 206,551 deliveries during the study period. Of these, 58.9% of patients were classified as having mild pre-eclampsia while 38.4% had severe pre-eclampsia. Advanced maternal age (> 35 years) (odds ratio [OR] = 4.56; 95% confidence interval [CI] = 4.23-4.90; p < 0.001), primiparity (OR = 1.71; 95% CI = 1.61-1.82; p = 0.02) and twin pregnancy (OR = 1.92; 95% CI = 1.64-2.25; p = 0.01) were significant risk factors for developing pre-eclampsia. However, multivariate analysis showed that only advanced maternal age was a significant risk factor for pre-eclampsia (OR = 3.21; 95% CI = 2.95-3.50; p < 0.001). In contrast to mild pre-eclampsia, severe pre-eclampsia resulted in significantly worse outcomes for both mothers and fetuses. Complications in patients with severe pre-eclampsia included placental abruption, acute renal failure, pulmonary edema, postpartum hemorrhage, pleural effusion, preterm labor, intrauterine growth retardation, stillbirth, neonatal mortality and low birth weight infants, all of which occurred significantly more frequently than in patients with mild pre-eclampsia (p < 0.001). CONCLUSIONS: Pre-eclampsia remains a big challenge in modern obstetrics in Taiwan. Early diagnosis and management of patients with pre-eclampsia to prevent progression would significantly improve outcomes for mothers and fetuses.

Adult↗

PubMed2000

Cyclo-oxygenase products and atherothrombosis.

The advent of selective inhibitors of the cyclo-oxygenase (COX)-2 enzyme has afforded the opportunity to reduce the incidence of gastrointestinal complications of traditional nonsteroidal anti-inflammatory drugs (NSAIDs). The widespread use of these drugs has increased interest in their role in the cardiovascular system. Although deletion of the prostacyclin receptor (the IP) accelerates atherogenesis in the mouse, retention of one copy of the IP is atheroprotective. This is consistent with the failure of biochemically defined, selective doses of a COX-2 inhibitor to accelerate atherogenesis in the mouse, despite suppressing prostacyclin biosynthesis by roughly 60%. Inhibition of both COX isozymes, by contrast, markedly retards atherogenesis. Consistent with these observations, antagonism of the thromboxane receptor (the TP) retards atherogenesis and diminishes the proliferative response to vascular injury in the mouse. Even partial suppression of prostacyclin (without coincident inhibition of platelet COX-1-dependent thromboxane formation) by COX-2 inhibitors may be undesirable in acute vascular occlusive syndromes. However, these drugs are unlikely to accelerate progression of the underlying vascular disease. By contrast, the effects of TP antagonists, aspirin, and even traditional NSAIDs on atherosclerotic plaque progression merit further evaluation in humans.

Animals↗

PubMed2000

[The regulating mechanism of anti-fungicides on mouse oocyte development].

AIM: To study the mechanism of the effect of gonadotropin-induced oocyte maturation. METHODS: Mouse oocytes were cultured in HX-medium, and the effects of amphotericin B and ketoconazole on resumption of meiosis of mouse oocyte were examined. RESULTS: 1. FSH(10-200 IU/L) induced a dose-dependent manner of oocytes maturation in CEO. A maximum increase in GVBD was observed with 25-50 IU/L FSH. 2. Amphotericin tericim B (0.025-2.5 micrograms/L) caused significant increases in GVBD in CEO, which mimicked the function of FSH. 3. Ketoconazole (10(-7)-10(-3) mol/L) inhibited the effect of FSH on resumption of meiosis, but no effect on oocyte spontaneous maturation. CONCLUSION: Amphotericin B and ketoconazole are able to affect mouse oocyte maturation, and indicates that they have a regulation on FSH-induced synthesis of meiosis-activating sterol.

Amphotericin B↗

PubMed2000

[Cloning and expression of TTV gene fragment in E. coil and identification of the expressed protein].

OBJECTIVE: To construct a recombinant expression vector containing TTV gene fragment and identify the expressed protein. METHODS: Based on computer-aided studies, TTV ORF12 gene fragment coding protein of antigenicity was amplified by PCR from the patient serum with non-A-G hepatitis. Then the gene fragment was integrated into the multiple cloning site of pQE-30 and expressed in E coil. The expressed protein was identified by ELISA. RESULTS: The relative molecular weight of the expressed protein matches the predication and its yield was up to 22.33% of total bacterial proteins of E. coli The expressed protein was detected by ELISA. CONCLUSIONS: The recombinant vector has been constructed successfully and the expressed protein possesses antigenicity.

Adolescent↗

PubMed2000

Molecular cloning and protein expression of EC1-2 and EC3-4 epitopes of pemphigus vulgaris antigen.

OBJECTIVE: To clone and express EC1-2 and EC3-4 epitopes of PVA (pemphigus vulgaris antigen, desmoglein-3) in order to diagnose pemphigus and study the relationship between epitopes of PVA and anti-PVA antibody. METHODS: RNA was extracted from keratinocytes and the cDNA of epitopes EC1-2 and EC3-4 was synthesized by reverse transcription. Amplified genes of EC1-2 and EC3-4 were inserted into the expression plasmid, PGEX-4T-1, and transformed into E. coli BL21 by electric transduction. Recombinant fusion proteins of EC1-2 and EC3-4 epitopes were expressed by IPTG induction. These proteins were separated on SDS-PAGE gels and electroblotted to nitrocellulose to detect the anti-PVA antibody. RESULTS: The sequences of cloned EC1-2 and EC3-4 genes were identical to the sequence registered in PC/GENE. Expressed recombinant proteins reacted only to sera from patients with pemphigus vulgaris, not to sera from patients with bullous pemphigoid, systemic lupus erythematosus or normal persons. CONCLUSIONS: These recombinant proteins are very specific in antigenicity. This may provide a new method for the diagnosis of pemphigus vulgaris (PV) or the differential diagnosis of other bullous cutaneous diseases via patient sera. It is also helpful in understanding the relationship between adhesion molecules and the pathogenic mechanism of pemphigus vulgaris.

Autoantibodies↗

PubMed1999

Virtual electrode-induced reexcitation: A mechanism of defibrillation.

Mechanisms of defibrillation remain poorly understood. Defibrillation success depends on the elimination of fibrillation without shock-induced arrhythmogenesis. We optically mapped selected epicardial regions of rabbit hearts (n=20) during shocks applied with the use of implantable defibrillator electrodes during the refractory period. Monophasic shocks resulted in virtual electrode polarization (VEP). Positive values of VEP resulted in a prolongation of the action potential duration, whereas negative polarization shortened the action potential duration, resulting in partial or complete recovery of the excitability. After a shock, new propagated wavefronts emerged at the boundary between the 2 regions and reexcited negatively polarized regions. Conduction velocity and maximum action potential upstroke rate of rise dV/dt (max) of shock-induced activation depended on the transmembrane potential at the end of the shock. Linear regression analysis showed that dV/dt(max) of postshock activation reached 50% of that of normal action potential at a V(m) value of -56.7+/-0.6 mV postshock voltage (n=9257). Less negative potentials resulted in slow conduction and blocks, whereas more negative potentials resulted in faster conduction. Although wavebreaks were produced in either condition, they degenerated into arrhythmias only when conduction was slow. Shock-induced VEP is essential in extinguishing fibrillation but can reinduce arrhythmias by producing excitable gaps. Reexcitation of these gaps through progressive increase in shock strength may provide the basis for the lower and upper limits of vulnerability. The former may correspond to the origination of slow wavefronts of reexcitation and phase singularities. The latter corresponds to fast conduction during which wavebreaks no longer produce sustained arrhythmias.

Animals↗

PubMed1999

Murine Siva-1 and Siva-2, alternate splice forms of the mouse Siva gene, both bind to CD27 but differentially transduce apoptosis.

CD27, a member of the TNFR family known to provide essential co-stimulatory signals for T cell growth and B cell Ig synthesis, can also mediate cell death. Using the CD27 cytoplasmic tail as the bait in yeast two hybrid assay, we previously cloned human Siva, a pro-apoptotic molecule. Here we report the characterization of the mouse Siva gene as a 4 kb sequence containing 4 exons and 3 introns. RT-PCR has revealed the presence of two forms of mouse Siva mRNA, the longer full length form Siva-1 and the shorter Siva-2 lacking the sequence coded by exon 2. Immunoblotting with anti-Siva (human) antibodies clearly demonstrate the presence of both Siva-1 and Siva-2. Cotransfection experiments in 293T cells reveal that mouse CD27 receptor can interact with both forms of Siva. Although mouse Siva-1 can trigger apoptosis in Rat-1 cells and in some of the mouse cell lines in transient transfection experiments, similar to the observation made with human Siva, intriguingly its alternate splice form, Siva-2 appears to be much less toxic. It is therefore likely that Siva-2 could regulate the function of Siva-1.

3T3 Cells↗

PubMed1999

Detection of IgM to hepatitis B core antigen in a reductant containing, chemiluminescence assay.

The Abbott PRISM(R) hepatitis B core (HBc) antigen assay is an automatic in vitro competitive chemiluminescence immunoassay for the detection of total antibody to HBc (anti-HBc) antigen in human serum or plasma. The assay utilizes cysteine solution as a reducing reagent in order to maximize specificity. To help understand the effect of cysteine on detection of anti-HBc antigen, we separated and purified anti-HBc IgM and IgG from human plasma using size exclusion, protein A/G, and affinity chromatography techniques. We showed that cysteine affected the reactivity of anti-HBc IgM with recombinant HBc (rHBc) antigen but not the reactivity of anti-HBc IgG. Anti-HBc IgM treated with cysteine yielded byproducts which were reactive in the PRISM HBcore assay. Reduction-sensitive factor (RSF) - IgM fraction from serum known to be non-specific for anti-HBc activity, similarly treated with cysteine, was no longer reactive in the PRISM HBcore assay. We showed that cysteine treatment is effective against non-specific IgM in human blood. Also, the inclusion of cysteine in the PRISM HBcore assay does not compromise the detection of HBc specific antibodies.

Antibody Specificity↗

PubMed1999

Gadolinium induces domain and pore formation of human erythrocyte membrane: an atomic force microscopic study.

Lanthanide cations bind to human erythrocyte membranes and enhance cell permeability. It was postulated that this effect is due to their likeness with calcium ions, which have been used to induce perforation of cells. However, the nature and mechanism of the perforation are still not clear. In the present work, the change in surface topography of erythrocyte membranes exposed to various gadolinium species was imaged with an atomic force microscope (AFM) in order to get direct evidence of perforation. The images of the whole cell and regions in nanometer scale showed that the normal surface is featured by closely packed nanometer size particles. The AFM images showed that Gd(3+) binding to erythrocytes led to domain structure at low concentration and pore formation at higher concentration. The domain structures that appeared after incubation with 1.0x10(-6)-1.0x10(-5) mol/l Gd(3+) solution for 30 min are featured by the particles aggregated to form ranges and the separations among them enlarged to gorges. With a higher concentration, 2.5x10(-5) mol/l Gd(3+), the further aggregation developed into crater-shaped 'pores'. By washing with EDTA the 'pores' can be resealed but the domain structure remained. The anionic complex of Gd(3+), [Gd(Cit)(2)](3-) of this concentration, can only induce the domain structure formation. The domain and 'pore' structures mediated by Gd(3+) concentrations might be responsible for both enhanced permeability and perforation. The mechanism of Gd-induced domain formation and perforation is discussed on the basis of aggregation of membrane proteins and the coexistence of different phases of membrane lipids resulting from Gd(3+) binding.

Animals↗

PubMed1999

The events relating to lanthanide ions enhanced permeability of human erythrocyte membrane: binding, conformational change, phase transition, perforation and ion transport.

The binding and uptake of Gd3+ ions by human erythrocytes in vitro were studied by determining the Gd contents in membrane and in cytosol by means of particle-induced X-ray emission (PIXE) spectrometry. Results obtained from varied incubation time revealed that the Gd3+ ions bind to the membrane proteins and lipids at first. Gd3+ binding to the membrane lipids and proteins lasts 0 approximately 20 and 20 approximately 100 ms respectively, as shown by the stopped-flow studies. Then a fraction of Gd3+ ions diffuses through the membrane. The kinetics of Gd3+ binding indicates that the binding to phospholipids is prior to that to the membrane proteins, but a portion of the lipid-bound Gd3+ redistributed later to the proteins. PIXE studies showed that the entry of Gd3+ increased the influx of Ca2+ and Cl-. By monitoring the changes in fluorescence of proteins and that of the Ln3+, the uptake of La3+, Eu3+, Gd3+ and Tb3+ was shown to be a process comprising a series of events. Binding to the membrane molecules induces the phase transition of lipid bilayer and conformational changes and aggregation of membrane proteins. Conformational changes of the proteins were characterized by Fourier transform IR spectroscopy (FT-IR) deconvolved spectra, i.e. alpha-helix content decreases while beta-sheet increases. ESR spectra of MSL-labeled proteins reflect the aggregation state related with the conformational change. [31P]NMR spectra of membrane lipid bilayer revealed the Ln3+ ions induced hexagonal (H(II)) phase formation. Phase transition and aggregation of membrane proteins cause the formation of domain structure and perforation in the membrane. These alterations in membrane structure are responsible for the Ln3+ enhanced membrane permeability. Thus the previous Ln3+ binding will facilitate the across-membrane transport of other Ln3+ ions through the membrane.

Blood Proteins↗

PubMed1999

The projection structure of the membrane protein microsomal glutathione transferase at 3 A resolution as determined from two-dimensional hexagonal crystals.

The formation of two-dimensional crystals of the membrane-bound enzyme microsomal glutathione transferase is sensitive to fractional changes in the lipid-to-protein ratio. Variation of this parameter results in crystal polymorphism. The projection structure of a p6 crystal form of the enzyme has been determined by the use of electron crystallography. The unit cell at 3 A resolution is comprised of two trimers. The hexagonal p6 and the orthorhombic p21212 crystal types have common elements in the packing arrangement which imply dominant crystal contacts. An overall structural similarity between the protein molecules in the two crystal forms is suggested by the projection maps. Furthermore, a comparison of the p6 and p21212 projection maps identifies additional corresponding protein densities which could not be assigned to the microsomal glutathione transferase trimer previously. Surprisingly, an ambiguity of the rotational orientation was found for trimers interspersed at certain positions within the crystal lattice.

Animals↗

PubMed1999

The 69 bp circadian regulatory sequence (CRS) mediates per-like developmental, spatial, and circadian expression and behavioral rescue in Drosophila.

The period (per) gene is an essential component of the circadian timekeeping mechanism in Drosophila. This gene is expressed in a circadian manner, giving rise to a protein that feeds-back to regulate its own transcription. A 69 bp clock regulatory sequence (CRS) has been identified previously upstream of the period gene. The CRS confers wild-type mRNA cycling when used to drive a lacZ reporter gene in transgenic flies. To determine whether the CRS also mediates proper developmental and spatial expression and behavioral rescue, we used the CRS to drive either lacZ or per in transgenic flies. The results show that the CRS is able to activate expression in pacemaker neuron precursors in larvae and essentially all tissues that normally express per in pupae and adults. The CRS is sufficient to rescue circadian feedback loop function and behavioral rhythms in per01 flies. However, the period of locomotor activity rhythms shortens if a stronger basal promoter is used. This study shows that regulatory elements sufficient for clock-dependent and tissue-specific per expression in larvae, pupae, and adults are present in the CRS and that the period of adult locomotor activity rhythms is dependent, in part, on the overall level of per transcripts.

Animals↗

PubMed1999

Influence of various nucleotides on the in situ crystallization of Ca2+-ATPase.

A reproducible in situ crystallization of the Ca2+-ATPase in isolated sarcoplasmic reticulum (SR) membranes was studied. The addition of various nucleotides to the washing buffer allowed the formation of tubular crystals, which is induced by vanadate. SR membranes washed with nucleotide-free buffer could not form tubular crystals upon subsequent incubation with vanadate.

Adenosine Triphosphate↗

PubMed1999

The transport kinetics of lanthanide species in a single erythrocyte probed by confocal laser scanning microscopy.

A novel method has been developed to visualize and follow the temporal course of lanthanide transport across the membrane into a single living erythrocyte. By means of confocal scanning microscopy and the optical section technique, the entry of lanthanide ions was followed by the fluorescence quenching of fluorescein isothiocyanate (FITC)-labeled membrane and cytosol. From the difference of the quenching kinetics of the whole section and the central area, the time for diffusion through the membrane and the diffusion in the extracellular and intracellular media can be deduced. To clarify the mechanism of lanthanide-induced fluorescence quenching of FITC-labeled erythrocytes and to ensure that this reaction can be used in this method, the reaction was investigated by steady-state fluorescence techniques. The results showed that the lanthanides strongly quenched the florescence emitted by FITC covalently bound to membrane proteins and cytosolic proteins. The static quenching mechanism is responsible for the fluorescence quenching of FITC-labeled proteins by Ln species. The quenching mechanism is discussed on the basis of complex formation. The dependence of fluorescence quenching on both ion size and the total orbital angular momentum L supports the complexation mechanism. The transport time across the membrane is strikingly correlated with Ln species and extracellular concentration. For a given concentration, the transport time of [Ln(cit)2]3- is much shorter than that of Ln3+, since they enter the cells via the anion channel. This is supported by the inhibition effect of 4,4'-diisothiocyanato-2,2'-stilbenendisulfonate on the transport of [Ln(cit)2]3-. On the other hand, the transport of free Ln3+ might be attributed to the enhanced permeability of erythrocytes owing to Ln3+ binding. These findings strongly demonstrate the existence of the non-internalization mechanism of Ln species uptake by erythrocytes.

Animals↗