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Biomedical subjects

Q Xu

Publications and source records attributed to Q Xu.

At least 199 records · Page 11Linked to original sources
PubMed1999

Eph receptors and ephrins restrict cell intermingling and communication.

Eph proteins are receptors with tyrosine-kinase activity which, with their ephrin ligands, mediate contact-dependent cell interactions that are implicated in the repulsion mechanisms that guide migrating cells and neuronal growth cones to specific destinations. Ephrin-B proteins have conserved cytoplasmic tyrosine residues that are phosphorylated upon interaction with an EphB receptor, and may transduce signals that regulate a cellular response. Because Eph receptors and ephrins have complementary expression in many tissues during embryogenesis, bidirectional activation of Eph receptors and ephrin-B proteins could occur at interfaces of their expression domains, for example at segment boundaries in the vertebrate hindbrain. Previous work has implicated Eph receptors and ephrin-B proteins in the restriction of cell intermingling between hindbrain segments. We therefore analysed whether complementary expression of Eph receptors and ephrins restricts cell intermingling, and whether this requires bidirectional or unidirectional signalling. Here we report that bidirectional but not unidirectional signalling restricts the intermingling of adjacent cell populations, whereas unidirectional activation is sufficient to restrict cell communication through gap junctions. These results reveal that Eph receptors and ephrins regulate two aspects of cell behaviour that can stabilize a distinct identity of adjacent cell populations.

Animals↗
PubMed1999

In vivo cell sorting in complementary segmental domains mediated by Eph receptors and ephrins.

The restriction of intermingling between specific cell populations is crucial for the maintenance of organized patterns during development. A striking example is the restriction of cell mixing between segments in the insect epidermis and the vertebrate hindbrain that may enable each segment to maintain a distinct identity. In the hindbrain, this is a result of different adhesive properties of odd- and even-numbered segments (rhombomeres), but an adhesion molecule with alternating segmental expression has not been found. However, blocking experiments suggest that Eph-receptor tyrosine kinases may be required for the segmental restriction of cells. Eph receptors and their membrane-bound ligands, ephrins, are expressed in complementary rhombomeres and, by analogy with their roles in axon pathfinding, could mediate cell repulsion at boundaries. Remarkably, transmembrane ephrins can themselves transduce signals, raising the possibility that bi-directional signalling occurs between adjacent ephrin- and Eph-receptor-expressing cells. We report here that mosaic activation of Eph receptors leads to sorting of cells to boundaries in odd-numbered rhombomeres, whereas mosaic activation of ephrins results in sorting to boundaries in even-numbered rhombomeres. These data implicate Eph receptors and ephrins in the segmental restriction of cell intermingling.

Animals↗
PubMed1999

Genes for calcineurin B-like proteins in Arabidopsis are differentially regulated by stress signals.

An important effector of Ca2+ signaling in animals and yeast is the Ca2+/calmodulin-dependent protein phosphatase calcineurin. However, the biochemical identity of plant calcineurin remained elusive. Here we report the molecular characterization of AtCBL (Arabidopsis thaliana calcineurin B-like protein) from Arabidopsis. The protein is most similar to mammalian calcineurin B, the regulatory subunit of the phosphatase. AtCBL also shows significant similarity with another Ca2+-binding protein, the neuronal calcium sensor in animals. It contains typical EF-hand motifs with Ca2+-binding capability, as confirmed by in vitro Ca2+-binding assays, and it interacts in vivo with rat calcineurin A in the yeast two-hybrid system. Interaction of AtCBL1 and rat calcineurin A complemented the salt-sensitive phenotype in a yeast calcineurin B mutant. Cloning of cDNAs revealed that AtCBL proteins are encoded by a family of at least six genes in Arabidopsis. Genes for three isoforms were identified in this study. AtCBL1 mRNA was preferentially expressed in stems and roots and its mRNA levels strongly increased in response to specific stress signals such as drought, cold, and wounding. In contrast, AtCBL2 and AtCBL3 are constitutively expressed under all conditions investigated. Our data suggest that AtCBL1 may act as a regulatory subunit of a plant calcineurin-like activity mediating calcium signaling under certain stress conditions.

Amino Acid Sequence↗
PubMed1999

[Rapid detection of non-deletional alpha-thalassemia mutations by PCR-LIS-SSCP]

OBJECTIVE: To establish a rapid and convenient single strand conformation polymorphism (SSCP) analysis method for detecting the point mutation of non-deletional alpha-thalassemia. METHODS: The 543bp DNA fragment spanning the hot spot region mainly responsible for non-deletional alpha-thalassemia was nested amplified using the selective amplification of alpha2 globin gene as a template and was denatured with low ionic strength(LIS) solution followed by SSCP analysis. RESULTS: LIS buffer was more efficient for ssDNA formation than formamide buffer was and the formation of ssDNA was very stable. In addition to a normal electrophoresis pattern, at least three SSCP profiles can be detected by the present method when the DNA samples bearing non-deletional genes of Hb H disease were screened. Confirmed by DNA sequencing analysis, the DNAs represented these profiles have turned out to be the different three mutants, i.e., the alphaCS mutation, the alpha QS mutation and the alphaWestmead mutation, respectively. Only 3 hours were needed to complete the electrophoresis procedure of this method. CONCLUSION: PCR-LIS-SSCP can be used as a tool in rapid screening for the alterations in human alpha-globin gene.

Journal Article↗
PubMed1999

Endothelial cytotoxicity mediated by serum antibodies to heat shock proteins of Escherichia coli and Chlamydia pneumoniae: immune reactions to heat shock proteins as a possible link between infection and atherosclerosis.

BACKGROUND: Growing evidence suggests that an immunological reaction against heat shock proteins (HSPs) may be involved in atherogenesis. Because HSPs show a high degree of amino acid sequence homology between different species, from prokaryotes to humans, we investigated the possibility of "antigenic mimicry" caused by an immunological cross-reaction between microorganisms and autoantigens. METHODS AND RESULTS: Serum antibodies against the Escherichia coli HSP (GroEL) and the 60-kDa chlamydial HSP (cHSP60) from subjects with atherosclerosis were purified by use of affinity chromatography. Western blot analyses and competitive ELISAs confirmed the cross-reaction of the eluted antibodies with human HSP60 and the bacterial counterparts. The cytotoxicity of anti-GroEL and anti-cHSP60 antibodies was determined on human endothelial cells labeled with 51Cr. A significant difference (40% versus 8%) was observed in the specific 51Cr release of heat-treated (42 degrees C for 30 minutes) and untreated cells, respectively, in the presence of these anti-HSP antibodies and complement. This effect was blocked by addition of 100 microg/mL recombinant GroEL. In addition, seropositivity against specific non-HSP60 Chlamydia pneumoniae antigens is more prominent among high-anti-HSP titer sera than low-titer sera. CONCLUSIONS: Serum antibodies against HSP65/60 cross-react with human HSP60, cHSP60, and GroEL; correlate with the presence of antibodies to C pneumoniae and endotoxin; and mediate endothelial cytotoxicity. These findings suggest that humoral immune reactions to bacterial HSPs, such as cHSP60 and GroEL, may play an important role in the process of vascular endothelial injury, which is believed to be a key event in the pathogenesis of atherosclerosis.

Adult↗
PubMed1999

[Healthy life years(HeaLY): a new indicator for measuring the burden of diseases].

Healthy life years(HeaLY) is a new indicator for measuring the burden of diseases. The data of Beijing MONICA were analyzed and reevaluated by using this indicator, and were compared with another indicator, disability adjusted life years (DALY), for the burden of diseases provided by WHO and World Bank several years ago.

Cost of Illness↗
PubMed1999

Flexibility of interdomain contacts revealed by topological isomers of bivalent consolidated ligands to the dual Src homology domain SH(32) of abelson.

Src homology (SH)2 and SH3 domains are found in a variety of proteins involved in the control of cellular signaling and architecture. The possible interrelationships between domains are not easily investigated, even though several cases of multiple domain-containing constructs have been studied structurally. As a complement to direct structural methods, we have developed consolidated ligands and tested their binding to the Abl SH(32) complex. Consolidated ligands combine in the same molecule peptide sequences recognized by SH2 and SH3 domains, i.e., Pro-Val-pTyr-Glu-Asn-Val and Pro-Pro-Ala-Tyr-Pro-Pro-Pro-Pro-Val-Pro, respectively; these are joined by oligoglycyl linkers. Four types of ligands were chemically synthesized, representing all the possible relative orientations of ligands. Their affinities were found to vary with binding portion topologies and linker lengths. Two of these types were shown to bind to both SH2 and SH3 dual domains with high affinities and specificities, showing increases of one order of magnitude, as compared to the most strongly bound monovalent equivalent. These results suggest that the relative orientation of SH2 and SH3 in Abl SH(32) is not fixed, and this synthetic approach may be generally useful for determining the structures of ligated complexes and for developing reagents with high affinities and specificities.

Amino Acid Sequence↗
PubMed1999

Purification, crystallization and preliminary X-ray diffraction analysis of the yeast phosphorelay protein YPD1.

YPD1 is a yeast osmoregulatory protein that functions in a phosphorelay signal-transduction pathway. YPD1 has been expressed in Escherichia coli, purified to homogeneity and crystallized. The crystals were obtained by hanging-drop vapor-diffusion using PEG 4000 as a precipitant. Preliminary X-ray diffraction analysis indicates that the crystals belong to tetragonal space group P43212 or P41212 with unit-cell dimensions a = b = 52.71, c = 244.02 A. X-ray data to 2.7 and 3.0 A have been collected from native crystals and a heavy-atom derivative, respectively. Positions for two Hg atoms have been located by analysis of difference Patterson maps.

Base Sequence↗
PubMed1999

Application of capillary nongel sieving electrophoresis for gene analysis.

Capillary electrophoresis (CE) has proved to be a strong tool for DNA analysis and has found abundant applications in the fields of restriction fragment sizing, mutation screening, polymerase chain reaction (PCR) product characterizing and forensic identifying. CE may be the main alternative to slab gel electrophoresis. Capillary nongel electrophoresis is the most favorable mode when aiming for this purpose because of its advantages of long lifetime, easy operation, good reproducibility, and low expense. In this paper, a new kind of sieving matrix, with mannitol as the additive for capillary electrophoresis, as well as related methods and their application for gene analysis were reported. Nine DNA fragments amplified by multiplex PCR from a normal dystrophin gene were well separated by this system. Three different deletions were found in Duchenne muscular dystrophy (DMD) patients. Three to four copies of the sex-determination region of the Y chromosome (SRY) gene, as well as the phenylalanine hydroxylase (PAH) gene, could be detected in mixed samples. The frequencies of short tandem repeats (STR) in PAH genes was analyzed in 61 normal Chinese individuals and 6 phenylketonuria families. One case of prenatal gene diagnosis was performed. By using this matrix, CE coupled with reverse transcription PCR (RT-PCR), the analysis of the alternative splicing expression pattern of the fragile X mental retardation 1 (FMR1) gene in adult lung tissue was achieved.

Adult↗
PubMed1999

Methods of assaying Bcl-2 and Bax family proteins in yeast.

Bcl-2 family proteins play an evolutionarily conserved role in regulating the life and death of the cell. Certain proapoptotic members of the Bcl-2 family, Bax and Bak, have intrinsic cytotoxic activities in that they not only induce or sensitize mammalian cells to undergo apoptosis but also display a lethal phenotype when ectopically expressed in two yeast species Saccharomyces cerevisiae and Schizosaccharomyces pombe. Furthermore, the antiapoptotic Bcl-2 and Bcl-XL proteins can protect yeast against Bax-mediated lethality, suggesting that the death-regulatory functions of these Bcl-2 family proteins are well preserved in yeast. These observations provide the opportunity to study the function of Bcl-2 family proteins in genetically tractable yeast and to apply classical yeast genetics and functional cloning approaches to the dissection of programmed cell death pathway regulated by Bcl-2 family proteins. We describe here methods used in our laboratory to express and to study the functions of Bcl-2 family proteins in both the budding yeast S. cerevisiae and the fission yeast S. pombe.

Apoptosis↗
PubMed1999

Effects of several drugs on the liver injury induced by delayed-type hypersensitivity to picryl chloride by regulating suppressor or helper T cells.

The effects of histamine, cimetidine, and diphenhydramine on picryl chloride (PCl)-induced ear contact sensitivity, as well as liver injury, were examined in mice. Histamine was found to produce less response in mice to PCl. In contrast, cimetidine, a selective antagonist of histamine type 2 receptor, significantly enhanced the response, while diphenhydramine, a selective antagonist of histamine type 1 receptor showed no effect. The pre-treatment of 2,4, 6-trinitrobenzene sulphonic acid (TNBS) significantly caused a tolerance to the formation of the liver injury induced by delayed-type hypersensitivity (DTH) to PCl. Against the tolerance, the single intravenous administration of 150 mg kg-1 cyclophosphamide (Cy) at 3 days before the TNBS-treatment recovered the response and induced a remarkable elevation of serum transaminases. On the other hand, cyclosporin A protected the liver injury. These observations revealed that the development of acute PCl-DTH liver injury was regulated by the functional state of suppressor and helper T cells.

Animals↗
PubMed1999

Myocardial injury leads to a release of heat shock protein (hsp) 60 and a suppression of the anti-hsp65 immune response.

OBJECTIVE: While atherosclerosis is associated with high titers of autoantibodies to bacterial hsp65 crossreacting with human hsp60 (anti-hsp60 autoantibodies), myocardial infarction entails decreased humoral immune response to hsp65. We previously hypothesized that myocardial ischemia and subsequent infarction not only induce myocardial hsp60 expression, but also trigger release of myocardial hsp60 into the circulation, influencing the systemic hsp immune response via immune complex formation. METHODS: In the present study, organ culture of rat hearts under circulatory arrest provided a model of myocardiocyte injury due to ischemia. RESULTS: Reperfusion of ischemic hearts confirmed the occurrence of myocardial injury by a rise of heart enzymes. Myocardial hsp60 expression was induced up to threefold in response to ischemia, and most of hsp60 expression was localized to the muscle fibers. Analysis of coronary eluate revealed release of hsp60 from myocardium. In addition, hsp60-containing, but not hsp60-free, coronary eluate was recognized by anti-hsp65 serum antibodies and induced proliferation of hsp65-specific T cells. When hsp60-containing coronary eluate was reinjected into an hsp65-primed rat, both humoral and cellular hsp65-immune responses were strongly downregulated. CONCLUSION: Our findings demonstrate the release of highly immunogenic and crossreactive hsp60 into the circulation in response to myocardial ischemia and myocardiocyte injury.

Animals↗
PubMed1999

Benzylisoquinoline alkaloids from gnetum parvifolium

The investigation of the chemical constituents from the lianas of Gnetum parvifolium resulted in the isolation of three new benzylisoquinoline alkaloids, (+/-)-N-methylhigenamine (1), (-)-N-methylhigenamine N-oxide (2), and (+/-)-8-(p-hydroxybenzyl)-2, 3,10,11-tetrahydroxyprotoberberine (3), together with two known alkaloids, higenamine and trigonelline. The structures of 1-3 were determined by chemical methods and spectral analysis including 2D NMR and NOE studies.

Journal Article↗
PubMed1999

Characterization and analysis of a novel glycoprotein from snake venom using liquid chromatography-electrospray mass spectrometry and Edman degradation.

An N-linked glycosylation in a novel C-lectin protein from snake venom was observed by Edman degradation and liquid chromatography-electrospray mass spectrometry. The peptides obtained by trypsin cleavage were analyzed to confirm the amino acid sequence and Asn5 was found to be the N-glycosylation site. The result was further confirmed by N-glycosidase digestion. In addition, the protein and tryptic peptides with and without glycan chain were characterized by mass spectrometry according to the mass difference. The glycopeptide obtained from proteolytic digestion was analyzed and the glycoforms were identified as high-mannose type by tandem MS coupled with alpha-mannosidase digestion. An oxidized Met residue was detected and located in the protein by mass spectrometry.

Amidohydrolases↗
PubMed1999

Integrin alphavbeta3-RGDS interaction mediates fibrin-induced morphological changes of glomerular endothelial cells.

BACKGROUND: In our previous studies, we found that intraglomerular deposition of fibrin and its metabolites was related to glomerular sclerosis and reduced renal function. It has been reported that both overlying and underlying fibrin may induce specific morphological changes of cultured endothelial cells from large blood vessels. The dependency of these morphological changes on the integrin alphavbeta3-arginyl-glycyl-aspartyl-serine (RGDS) interaction is still controversial. We hypothesized that glomerular endothelial cells (GECs) stimulated by fibrin might undergo morphological changes through an integrin alphavbeta3-RGDS interaction. Methods. In vitro studies were performed to examine the growing status of GECs stimulated by overlying and underlying fibrin gels in the presence or absence of the following: 50 microg/ml anti-alphavbeta3 integrin monoclonal antibody 23C6 or nonimmune mouse IgG, 1 mg/ml synthetic RGDS or arginyl-glycyl-glycyl-serine (RGGS) peptide, 10 mg/ml sodium heparin, 100 microg/ml cycloheximide, and 10 microM actinomycin D. Fast protein liquid chromatography (FPLC)-purified fibrinogen and the third to fifth passages of human GECs were also used in this study. RESULTS: GECs developed capillary tube structure after 60 hours of culturing on fibrin gels, and GECs cultured on gelatin-coated plates displayed a monolayer of cobblestone-like cells in the presence or absence of 23C6 and synthetic RGDS peptide. Fibrin-induced capillary tube formation was promoted by 23C6 and inhibited by RGDS peptide, cycloheximide, and actinomycin D. Disorganization of the GEC monolayer was induced by overlying fibrin, but was not induced by overlying agarose gels and glass cover slips or culturing in fibrinogen, 0.05 NIH U/ml thrombin, fibrin supernatants, as well as in fibrin degradation products. Disorganization of GEC monolayer can be induced by both des-AA-fibrin and des-AABB-fibrin and was unaffected by heparin. Furthermore, both 23C6 and synthetic RGDS peptide prevented disorganization of GECs induced by overlying fibrin, whereas nonimmune mouse IgG, synthetic RGGS peptide, cycloheximide, and actinomycin D had no similar effect. CONCLUSIONS: GECs cultured on fibrin gels may develop capillary structure spontaneously, and GECs covered by fibrin gels may undergo disorganization. Our data suggest that these GEC morphological changes are mediated by an integrin alphavbeta3-RGDS interaction.

Antibodies, Monoclonal↗
PubMed1999

A new flavanone isolated from rhizoma smilacis glabrae and the structural requirements of its derivatives for preventing immunological hepatocyte damage.

From the rhizome of Smilax glabra Roxb., a new flavanone was isolated and named as smitilbin (1), together with 6 known compounds, engeletin (2), astilbin (3), dihydroquercetin (4), eurryphin (5), resveratrol (6), and 5-O-caffeoylshikimic acid (7). These compounds were applied to the assay of liver nonparenchymal cells (NPC) against hepatocytes (HC) isolated from mice with an immunological liver injury. Against the NPC-caused elevation of ALT (alanine transminase) in culture supernatant from HC, the pretreatment of NPC with flavanoids (1-3) dose-dependently blocked the ALT release while 4, the aglycone of 3, did not. The chromone 5 showed a much stronger inhibition. Compound 6 also showed the activity. However, 1-7 did not show any suppression of NPC or CCl4-induced ALT release when they were used to pretreat HC. These results suggest that compounds 1-3, 5, and 6 could protect the hepatocyte damage from NPC through selectively producing the dysfunction of NPC with an essential requirement of rhamnose, and the chromone part in their structures may be critical for exhibiting the activity rather than through protecting the hepatocyte membranes.

Animals↗
PubMed1999

Recombinant fusion protein and DNA vaccines against foot and mouth disease virus infection in guinea pig and swine.

In this study, we provide evidence that a recombinant fusion protein containing beta-galactosidase and a tandem repeat peptide of immunogenic dominant epitope of foot-and-mouth disease virus (FMDV) VP1 protein elicits high levels of neutralizing antibody and protects both guinea pigs and swine against infection. Vaccination with this fusion protein induced a FMDV-specific proliferative T-cell response and a neutralizing antibody response. The immunized guinea pigs and swine were protected against FMD type O virus infection. Two DNA plasmids expressing genes of foot-and-mouth disease were constructed. Both plasmids pBO1 and pCO1 contain a signal sequence of the swine immunoglobulin G (IgG) gene and fusion protein gene of pXZ84. The signal sequence and fusion protein gene were under the control of a metallothionein promoter in the case of the pBO1 plasmid and under the control of a cytomegalovirus immediate early promoter in the case of pCO1 plasmid. When pBO1 and pCO1 were inoculated intramuscularly into guinea pigs, both plasmids elicited a neutralizing antibody response and spleen cell proliferation increased following stimulation with FMDV antigen, but animals were not protected from viral challenge.

Animals↗