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Biomedical subjects

F Wang

Publications and source records attributed to F Wang.

At least 199 records · Page 11Linked to original sources
PubMed2000

Leukocytes navigate by compass: roles of PI3Kgamma and its lipid products.

Morphologic polarity is necessary for the motility of mammalian cells. In leukocytes responding to a chemoattractant, this polarity is regulated by activities of small Rho guanosine triphosphatases (Rho GTPases) and the phosphoinositide 3-kinases (PI3Ks). Moreover, in neutrophils, lipid products of PI3Ks appear to regulate activation of Rho GTPases, are required for cell motility and accumulate asymmetrically to the plasma membrane at the leading edge of polarized cells. By spatially regulating Rho GTPases and organizing the leading edge of the cell, PI3Ks and their lipid products could play pivotal roles not only in establishing leukocyte polarity but also as compass molecules that tell the cell where to crawl.

Actins↗
PubMed2000

Pre-mRNA splicing imprints mRNA in the nucleus with a novel RNA-binding protein that persists in the cytoplasm.

We describe a novel RNA binding protein, Y14, a predominantly nuclear nucleocytoplasmic shuttling protein. Interestingly, Y14 associates preferentially with mRNAs produced by splicing but not with pre-mRNAs, introns, or mRNAs produced from intronless cDNAs. Y14 associates with both nuclear mRNAs and newly exported cytoplasmic mRNAs. Splicing of a single intron is sufficient for Y14 association. Y14-containing nuclear complexes are different from general hnRNP complexes. They contain hnRNP proteins and several unique proteins including the mRNA export factor TAP. Thus, Y14 defines novel intermediates in the pathway of gene expression, postsplicing nuclear preexport mRNPs, and newly exported cytoplasmic mRNPs, whose composition is established by splicing. These findings suggest that pre-mRNA splicing imprints mRNA with a unique set of proteins that persists in the cytoplasm and thereby communicates the history of the transcript.

Animals↗
PubMed2000

Analysis of neurotoxin 3-N-oxalyl-L-2,3-diaminopropionic acid and its alpha-isomer in Lathyrus sativus by high-performance liquid chromatography with 6-aminoquinolyl-N-hydroxysuccinimidyl carbamate (AQC) derivatization.

A method was developed for the quantitative determination of the neurotoxic nonprotein amino acid, 3-N-oxalyl-L-2,3-diaminopropionic acid (beta-ODAP), and its nontoxic alpha-isomer, 2-N-oxalyl-L-2, 3-diaminopropionic acid (alpha-ODAP), in the plant samples of Lathyrus sativus after derivatization with 6-aminoquinolyl-N-hydroxysuccinimidyl carbamate (AQC) by reversed-phase high-performance liquid chromatography (HPLC). Hippuric acid was used as an internal standard. A linear response was recorded in the concentration rang 0.32-32 nmol with r > 0.999. The RP HPLC detection limit for both isomers is 1.8 ng. According to different experimental needs, a ternary gradient system can be used to determine toxin and other nonprotein amino acids. The RP HPLC method and a colorimetric method were compared for measuring ODAP.

Amino Acids, Diamino↗
PubMed2000

Carbon disulfide at a Chinese viscose factory external and internal exposure assessment.

This article presents the results of carbon disulfide exposure measurements in a Chinese viscose rayon factory. The objectives of the study were to identify the external exposure levels at a large factory and to investigate the 2-thiothiazolidine-4-carboxylic acid (TTCA) concentrations in the urine of the subjects who were exposed to carbon disulfide in the working place atmosphere. The metabolism of carbon disulfide in the exposed subjects was also studied in order to demonstrate the best points in time for the internal exposure sampling. The measurement of the amount of personal exposure to carbon disulfide in the air of the workplace was performed by GC-FPD; the presence of TTCA in the workers urine was analyzed by use of a modified HPLC method. The kinetics of TTCA excretion was studied by analyses at different time-points both during and after exposure to carbon disulfide in the subjects. A total of 155 personal samples were obtained. The carbon disulfide concentration in the staple viscose hall was 13.72 +/- 1.12 mg m-3 in terms of the geometric mean +/- geometric standard deviation, and was 20.05 +/- 1.33 mg m-3 in the filament spinning hall. The TTCA values in the subjects who worked in the staple spinning hall were 1.18 +/- 0.43 mg g-1 creatinine and 1.07 +/- 0.38 mg g-1 creatinine for subjects working in the filament spinning hall. The best time for TTCA sampling is at the end of the working shift, the TTCA excretion was stable for a period of 4-12 h after exposure of the subjects to the carbon disulfide. It might be that the Chinese have different anthropometric characteristics; a sampling bias may therefore appear among different races.

Adult↗
PubMed2000

Preconditioning with cortical spreading depression decreases intraischemic cerebral glutamate levels and down-regulates excitatory amino acid transporters EAAT1 and EAAT2 from rat cerebal cortex plasma membranes.

We previously reported a 50% reduction in cortical infarct volume following transient focal cerebral ischemia in rats preconditioned 3 days earlier with cortical spreading depression (CSD). The mechanism of the protective effect of prior CSD remains unknown. Recent studies demonstrate reversal of excitatory amino acid transporters (EAATs) to be a principal cause for elevated extracellular glutamate levels during cerebral ischemia. The present study measured the effect of CSD preconditioning on (a) intraischemic glutamate levels and (b) regulation of glutamate transporters within the ischemic cortex of the rat. Three days following either CSD or sham preconditioning, rats were subjected to 200 min of focal cerebral ischemia, and extracellular glutamate concentration was measured by in vivo microdialysis. Cortical glutamate exposure decreased 70% from 1,772.4 +/- 1,469.2 microM-min in sham-treated (n = 8) to 569.0 +/- 707.8 microM-min in CSD-treated (n = 13) rats (p <0.05). The effect of CSD preconditioning on glutamate transporter levels in plasma membranes (PMs) prepared from rat cerebral cortex was assessed by western blot analysis. Down-regulation of the glial glutamate transporter isoforms EAAT2 and EAAT1 from the PM fraction was observed at 1, 3, and 7 days but not at 0 or 21 days after CSD. Semiquantitative lane analysis showed a maximal decrease of 90% for EAAT2 and 50% for EAAT1 at 3 days post-CSD. The neuronal isoform EAAT3 was unaffected by CSD. This period of down-regulation coincides with the time frame reported for induced ischemic tolerance. These data are consistent with reversal of glutamate transporter function contributing to glutamate release during ischemia and suggest that down-regulation of these transporters may contribute to ischemic tolerance induced by CSD.

ATP-Binding Cassette Transporters↗
PubMed2000

Islet amyloid polypeptide regulates multiple steps in stimulus-secretion coupling of beta cells in rat pancreatic islets.

Islet amyloid polypeptide (IAPP) is produced in pancreatic beta cells. Intraislet function of IAPP is still uncertain. In the present study, we investigated effects of IAPP and somatostatin on stimulus-secretion coupling of beta cells in isolated rat pancreatic islets. Insulin secretion induced by 22.2 mM glucose was increased by an IAPP antiserum (0.1%) or an IAPP antagonist (IAPP8-37, 10 microM). Pretreatment of islets with pertussis toxin (PTX) abolished the stimulating effect of IAPP8-37 on glucose-induced insulin secretion. In contrast, IAPP antiserum and IAPP8-37 did not change insulin secretion induced by 30 mM KCl. Somatostatin (1 nM) inhibited insulin secretion induced by 22.2 mM glucose, 10 mM L-arginine, 25 microM forskolin, and 200 microM carbachol. IAPP (10 microM) enhanced the inhibitory effect of somatostatin on insulin secretion induced by L-arginine or forskolin. PTX pretreatment abolished the effects of somatostatin and IAPP on arginine-induced insulin secretion. In conclusion, IAPP regulates multiple steps in signal transductions of beta cells. The effects of IAPP on beta cells are mediated by PTX-sensitive regulatory G proteins.

Amyloid↗
PubMed2000

Assembly, secretion, and vacuolar delivery of a hybrid immunoglobulin in plants.

Secretory immunoglobulin (Ig) A is a decameric Ig composed of four alpha-heavy chains, four light chains, a joining (J) chain, and a secretory component (SC). The heavy and light chains form two tetrameric Ig molecules that are joined by the J chain and associate with the SC. Expression of a secretory monoclonal antibody in tobacco (Nicotiana tabacum) has been described: this molecule (secretory IgA/G [SIgA/G]) was modified by having a hybrid heavy chain sequence consisting of IgG gamma-chain domains linked to constant region domains of an IgA alpha-chain. In tobacco, about 70% of the protein assembles to its final, decameric structure. We show here that SIgA/G assembly and secretion are slow, with only approximately 10% of the newly synthesized molecules being secreted after 24 h and the bulk probably remaining in the endoplasmic reticulum. In addition, a proportion of SIgA/G is delivered to the vacuole as at least partially assembled molecules by a process that is blocked by the membrane traffic inhibitor brefeldin A. Neither the SC nor the J chain are responsible for vacuolar delivery, because IgA/G tetramers have the same fate. The parent IgG tetrameric molecule, containing wild-type gamma-heavy chains, is instead secreted rapidly and efficiently. This strongly suggests that intracellular retention and vacuolar delivery of IgA/G is due to the alpha-domains present in the hybrid alpha/gamma-heavy chains and indicates that the plant secretory system may partially deliver to the vacuole recombinant proteins expected to be secreted.

Brefeldin A↗
PubMed2000

Immucillin-H binding to purine nucleoside phosphorylase reduces dynamic solvent exchange.

The rate and extent of hydrogen/deuterium (H/D) exchange into purine nucleoside phosphorylase (PNP) was monitored by electrospray ionization mass spectrometry (ESI-MS) to probe protein conformational and dynamic changes induced by a substrate analogue, products, and a transition state analogue. The genetic deficiency of PNP in humans is associated with severe T-cell immunodeficiency, while B-cell immunity remains functional. Inhibitors of PNP have been proposed for treatment of T-cell leukemia, to suppress the graft-vs.-host response, or to counter type IV autoimmune diseases without destroying humoral immunity. Calf spleen PNP is a homotrimer of polypeptide chains with 284 amino residues, molecular weight 31,541. Immucillin-H inhibits PNP with a Kd of 23 pM when only one of the three catalytic sites is occupied. Deuterium exchange occurs at 167 slow-exchange sites in 2 h when no catalytic site ligands are present. The substrate analogue and product prevented H/D exchange at 10 of the sites. Immucillin-H protected 32 protons from exchange at full saturation. When one of the three subunits of the homotrimer is filled with immucillin-H, and 27 protons are protected from exchange in all three subunits. Deuterium incorporation in peptides from residues 132-152 decreased in all complexes of PNP. The rate and/or extent of deuterium incorporation in peptides from residues 29-49, 50-70, 81-98, and 112-124 decreased only in the complex with the transition state analogue. The peptide-specific H/D exchange demonstrates that (1) the enzyme is most compact in the complex with immucillin-H, and (2) filling a single catalytic site of the trimer reduces H/D exchange in the same peptides in adjacent subunits. The peptides most highly influenced by the inhibitor surround the catalytic site, providing evidence for reduced protein dynamic motion caused by the transition state analogue.

Amino Acid Sequence↗
PubMed2000

Cloning of the rhesus lymphocryptovirus viral capsid antigen and Epstein-Barr virus-encoded small RNA homologues and use in diagnosis of acute and persistent infections.

Epstein-Barr virus (EBV) is the most common cause of infectious mononucleosis and is associated with the development of several human malignancies. A closely related herpesvirus in the same lymphocryptovirus (LCV) genera as EBV naturally infects rhesus monkeys and provides an important animal model for studying EBV pathogenesis. We cloned the small viral capsid antigen (sVCA) homologue from the rhesus LCV and developed a peptide enzyme-linked immunosorbent assay (ELISA) to determine whether epitopes in the rhesus LCV sVCA are a reliable indicator of rhesus LCV infection. In order to define a "gold standard" for rhesus LCV infection, we also cloned the EBV-encoded small RNA 1 (EBER1) and EBER2 homologues from rhesus LCV and developed a reverse transcription (RT)-PCR assay to detect persistent LCV infection in rhesus monkey peripheral blood lymphocytes. Animals from a conventional and a hand-reared colony were studied to compare the prevalence of rhesus LCV infection in the two groups. There was a 100% correlation between the peptide ELISA and EBER RT-PCR results for rhesus LCV infection. In addition, specificity for LCV infection and exclusion of potential cross-reactivity to the rhesus rhadinovirus sVCA homologue could be demonstrated using sera from experimentally infected animals. These studies establish two novel assays for reliable diagnosis of acute and persistent rhesus LCV infections. The rhesus LCV sVCA peptide ELISA provides a sensitive and reliable assay for routine screening, and these studies of the hand-reared colony confirm the feasibility of raising rhesus LCV-naive animals.

Amino Acid Sequence↗
PubMed2000

Sequence and functional analysis of EBNA-LP and EBNA2 proteins from nonhuman primate lymphocryptoviruses.

The Epstein-Barr virus (EBV) EBNA-LP and EBNA2 proteins are the first to be synthesized during establishment of latent infection in B lymphocytes. EBNA2 is a key transcriptional regulator of both viral and cellular gene expression and is essential for EBV-induced immortalization of B lymphocytes. EBNA-LP is also important for EBV-induced immortalization of B lymphocytes, but far less is known about the functional domains and cellular cofactors that mediate EBNA-LP function. While recent studies suggest that serine phosphorylation of EBNA-LP and coactivation of EBNA2-mediated transactivation are important, more detailed mutational and genetic studies are complicated by the repeat regions that comprise the majority of the EBNA-LP sequence. Therefore, we have used a comparative approach by studying the EBNA-LP homologues from baboon and rhesus macaque lymphocryptoviruses (LCVs) (baboon LCV and rhesus LCV). The predicted baboon and rhesus LCV EBNA-LP amino acid sequences are 61 and 64% identical to the EBV EBNA-LP W1 and W2 exons and 51% identical to the EBV EBNA-LP Y1 and Y2 exons. Five evolutionarily conserved regions can be defined, and four of eight potential serine residues are conserved among all three EBNA-LPs. The major internal repeat sequence also revealed a highly conserved Wp EBNA promoter with strong conservation of upstream activating sequences important for Wp transcriptional regulation. To test whether transcriptional coactivating properties were common to the rhesus LCV EBNA-LP, a rhesus LCV EBNA2 homologue was cloned and expressed. The rhesus LCV EBNA2 transcriptionally transactivates EBNA2-responsive promoters through a CBF1-dependent mechanism. The rhesus LCV EBNA-LP was able to further enhance rhesus LCV or EBV EBNA2 transactivation 5- to 12-fold. Thus, there is strong structural and functional conservation among the simian EBNA-LP homologues. Identification of evolutionarily conserved serine residues and regions in EBNA-LP homologues provides important clues for identifying the cellular cofactors and molecular mechanisms mediating these conserved viral functions.

Amino Acid Sequence↗
PubMed2000

Structural, functional, and genetic comparisons of Epstein-Barr virus nuclear antigen 3A, 3B, and 3C homologues encoded by the rhesus lymphocryptovirus.

EBNA-3A, -3B, and -3C are three latent infection nuclear proteins important for Epstein-Barr virus (EBV)-induced B-cell immortalization and the immune response to EBV infection. All three are hypothesized to function as transcriptional transactivators, but little is known about their precise mechanism of action or their role in EBV pathogenesis. We have cloned and studied the three EBNA-3 homologues from a closely related lymphocryptovirus (LCV) which naturally infects rhesus monkeys. The rhesus LCV EBNA-3A, -3B, and -3C homologues have 37, 40, and 36% amino acid identity with the EBV genes, respectively. Function, as measured by in vitro assays, also appears to be conserved with the EBV genes, since the rhesus LCV EBNA-3s can interact with the transcription factor RBP-Jkappa and the rhesus LCV EBNA-3C encodes a Q/P-rich domain with transcriptional activation properties. In order to better understand the relationship between these EBV and rhesus LCV latent infection genes, we asked if the rhesus LCV EBNA-3 locus could be recombined into the EBV genome and if it could substitute for the EBV EBNA-3s when assayed for human B-cell immortalization. Recombination between the EBV genome and rhesus LCV DNA was reasonably efficient. However, these studies suggest that the rhesus LCV EBNA-3 locus was not completely interchangeable with the EBV EBNA-3 locus for B-cell immortalization and that at least one determinant of the species restriction for LCV-induced B-cell immortalization maps to the EBNA-3 locus. The overall conservation of EBNA-3 structure and function between EBV and rhesus LCV indicates that rhesus LCV infection of rhesus monkeys can provide an important animal model for studying the role of the EBNA-3 genes in LCV pathogenesis.

Amino Acid Sequence↗
PubMed2000

Distribution of IGFBP-rP1 in normal human tissues.

IGFBP-rP1/mac25 is a recently described member of the insulin-like growth factor binding protein (IGFBP) family. It has structural homology to the other members of the IGFBP family but has a lower affinity for insulin-like growth factors (IGFs). In previous studies using RNA blot hybridization, it was shown that the expression of IGFBP-rP1/mac25 was ubiquitous in normal human tissues. In this report we show by immunohistochemistry that the expression of IGFBP-rP1/mac25 is actually restricted to certain organs and specific cell types. We used an antibody raised against a decapeptide of the C-terminal part of the protein that recognizes a approximately 37-kD protein under reduced conditions. The immunohistochemistry performed on normal human tissues showed a ubiquitous intense staining of peripheral nerves and a variable degree of positive staining in smooth muscle cells, including those from blood vessel walls, gut, bladder, and prostate. Cilia from the respiratory system, epididymis, and fallopian tube showed intense immunoreactivity. Most endothelial cells showed some positivity, whereas fat cells, plasma cells, and lymphocytes were negative. There was specific expression limited to certain cell types in the kidney, adrenal gland, and skeletal muscle, indicating a possible specialized function of IGFBP-rP1/mac25 in these organs. We further noted an opposite pattern of staining in the lining epithelium of breast (typically positive) and prostate glands (largely negative). The specific localization of IGFBP-rP1/mac25 as described implies a function of the protein. However, its regulation within the IGF axis or a possible direct action of IGFBP-rP1/mac25 remains to be demonstrated.

Antibodies↗
PubMed2000

Transcriptional activation of cathepsin D gene expression by growth factors.

Insulin-like growth factor-I (IGF-I), transforming growth factor alpha (TGFalpha) and epidermal growth factor (EGF) induced cathepsin D gene expression and reporter gene activity in MCF-7 human breast cancer cells transiently transfected with a construct (pCD1) containing a -2576 to -124 cathepsin D gene promoter insert. In contrast, IGF-I, but not TGFalpha or EGF, induced reporter gene activity in cells cotransfected with wild-type estrogen receptor (ER) expression plasmid and a construct (pCD2) containing estrogen-responsive downstream elements from -208 to -101. Promoter deletion and mutational analysis experiments identified four GC-rich sites and an imperfect palindromic estrogen responsive element required for IGF-I activation of the ER (ligand-independent). Subsequent studies with the mitogen-activated protein kinase (MAPK) inhibitor, PD98059, and a serine(118(-ER mutant confirmed the role of the MAPK pathway for IGF-I activation of the ER in MCF-7 cells. Thus, growth factor activation of ER can mediate transactivation vs ER/Sp1 binding to GC-rich sites and represents a novel pathway for ligand-independent ER action. The divergent pathways for IGF-I and TGFalpha/EGF activation of the ER observed in MCF-7 cells contrast with previous data indicating that pathways for growth factor activation of the ER are dependent on the gene and/or gene promoter and on cell context.

Base Sequence↗
PubMed2000

Apoptosis during goitre involution - the role of Bcl-2.

Goitrogenesis is accompanied by hyperplasia and hypertrophy and involves tissue remodelling and angiogenesis. During the involution of the goitre there must be removal of this increased thyroid volume, in addition to further remodelling, which may involve apoptosis. We investigated apoptosis in the involuting rat thyroid using male Fisher rats that were on a goitrogenic regimen for 14 days and then returned to a normal diet. Thyroid weights increased fourfold with the goitrogenic regimen. During involution, the largest decrease in weight was between day 2 and day 4 after withdrawal of treatment. After 34 days of involution, the thyroid weight plateaued, but had not returned to control values. High levels of Bcl-2 immunoreactivity were observed in normal and goitrous rat thyroids. These high levels were significantly reduced at 2 days of involution, after which high levels of Bcl-2 immunoreactivity returned. In situ end-labelling of apoptotic cells showed that there was an increase in the number of cells undergoing DNA fragmentation during goitrogenesis (1.0+/-0.8 cells/100 cells, n=9) compared with controls, in which no positive staining was observed. After 2 days of goitrogen withdrawal, there was a further fourfold increase in the number of in situ end-labelled cells (day 16: 4.1+/-1.7, n=9). Numbers of positive cells returned to low levels after 4 days of involution (day 18: 0.3+/-0.8, n=9). Using antiserum to apoptosis-specific protein, we found increased immunoreactivity during goitrogenesis and after 2 days of involution that was localised predominantly with the stromal and vascular tissue at both time points. The data show that rapid downregulation of Bcl-2 accompanies thyroid involution, which involves increased levels of apoptosis within the stromal compartment.

Animals↗
PubMed2000

Preparation of biodegradable microspheres of testosterone with poly(D,L-lactide-co-glycolide) and test of drug release in vitro.

Biodegradable microspheres formulation of testosterone (T) can be used as a new physiological approach for androgen replacement in hypogonadal men. In this study, poly(D,L-lactide-co-glycolide) (PLGA) microspheres containing T were prepared by a solvent-evaporation/solvent-diffusion process and the drug release tests of the microspheres were carried out in vitro. T/PLGA microspheres with good yield, desired size and satisfied drug loading were obtained. A significant testosterone sustained release was shown in the drug release tests in vitro. Since PLGA microspheres preparations are normally sterilized by colbat-60 irradiation, the effects of 25 kGy colbat-60 irradiation on physicochemical properties and in vitro drug release profile of T/PLGA microsphere were investigated. The results showed that the irradiation didn't have any effects on the physicochemical properties of T. Though about one-third decrease in molecular weight of PLGA was caused by the irradiation, no significant changes were observed on the drug release profile in vitro.

Biodegradation, Environmental↗