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Y Xu

Publications and source records attributed to Y Xu.

At least 631 records · Page 35Linked to original sources
PubMed1998

Crystallization and preliminary X-ray analysis of Escherichia coli GlnK.

The trimeric signal-transduction protein GlnK, from Escherichia coli, has been over-expressed, purified to homogeneity and crystallized. The crystals belong to space group P213 with a = 85.53 A and have two subunits in the asymmetric unit. The complex of GlnK with ATP crystallized in space group P63 with a = 57.45 and c = 54.79 A. These crystals have a single subunit in the asymmetric unit. High-quality diffraction data from crystals of GlnK and the GlnK complex have been collected to 2.0 A.

Bacterial Proteins↗
PubMed1998

Mice lacking transcription factor NF-E2 provide in vivo validation of the proplatelet model of thrombocytopoiesis and show a platelet production defect that is intrinsic to megakaryocytes.

Mechanisms of platelet production and release by mammalian megakaryocytes are poorly understood. We used thrombocytopenic knockout mice to better understand these processes. Proplatelets are filamentous extensions of terminally differentiated megakaryocytes that are thought to represent one mechanism of platelet release; however, these structures have largely been recognized in cultured cells and there has been no correlation between thrombocytopoiesis in vivo and proplatelet formation. Mice lacking transcription factor NF-E2 have a late arrest in megakaryocyte maturation, resulting in profound thrombocytopenia. In contrast to normal megakaryocytes, which generate abundant proplatelets, cells from these mice never produce proplatelets, even after prolonged stimulation with c-Mpl ligand. Similarly, megakaryocytes from thrombocytopenic mice with lineage-selective loss of transcription factor GATA-1 produce proplatelets very rarely. These findings establish a significant correlation between thrombocytopoiesis and proplatelet formation and suggest that the latter represents a physiologic mechanism of platelet release. We further show that proplatelet formation by normal megakaryocytes and its absence in cells lacking NF-E2 are independent of interactions with adherent (stromal) cells. Similarly, thrombocytopenia in NF-E2(-/-) mice reflects intrinsic defects in the megakaryocyte lineage. These observations improve our understanding of platelet production and validate the study of proplatelets in probing the underlying mechanisms.

Animals↗
PubMed1998

Lysophosphatidic acid as a potential biomarker for ovarian and other gynecologic cancers.

CONTEXT: Lysophosphatidic acid (LPA) has been shown to stimulate proliferation of ovarian cancer cells and is present in the ascitic fluid of patients with ovarian cancer. OBJECTIVES: To determine whether elevated levels of LPA are present in plasma from patients with ovarian cancer and other gynecologic malignancies compared with healthy controls and to evaluate whether an elevated LPA plasma level may be a biomarker for these diseases. DESIGN: A research assay was used to measure total LPA levels in plasma from healthy women and women with different diseases. All LPA assays and comparison of LPA levels and CA125 (an ovarian cancer biomarker) levels were performed by observers blinded to patient status or group. SETTING: The Cleveland Clinic Foundation. PARTICIPANTS: A convenience sample of 48 healthy control women, 48 women with ovarian cancer, 36 women with other gynecologic cancers, 17 women with benign gynecologic diseases, 11 women with breast cancer, and 5 women with leukemias. MAIN OUTCOME MEASURES: Total LPA levels in plasma samples from patients and controls. RESULTS: Patients in the ovarian cancer group had significantly higher plasma LPA levels (mean, 8.6 micromol/L; range, 1.0-43.1 micromol/L) compared with the healthy control group (mean, 0.6 micromol/L; range, <0.1-6.3 micromol/L) (P<.001). Elevated plasma LPA levels were detected in 9 of 10 patients with stage I ovarian cancer, 24 of 24 patients with stage II, III, and IV ovarian cancer, and 14 of 14 patients with recurrent ovarian cancer. Of 36 patients with other gynecologic cancers, 33 also showed higher LPA levels(mean, 14.9 micromol/L; range, <0.1-63.2 pmol/L), compared with healthy controls (P<.001). Elevated plasma LPA levels were detected in 5 of 48 controls and 4 of 17 patients with benign gynecologic diseases and in no women with breast cancer or leukemia. In comparison, among a subset of patients with ovarian cancer, 28 of 47 had elevated CA125 levels, including 2 of 9 patients with stage I disease. CONCLUSIONS: Plasma LPA levels may represent a potential biomarker for ovarian cancer and other gynecologic cancers. However, these findings are preliminary and require confirmation in larger studies.

Adult↗
PubMed1998

A 29-kilodalton Golgi soluble N-ethylmaleimide-sensitive factor attachment protein receptor (Vti1-rp2) implicated in protein trafficking in the secretory pathway.

Expressed sequence tags coding for a potential SNARE (soluble N-ethylmaleimide-sensitive factor attachment protein receptor) were revealed during data base searches. The deduced amino acid sequence of the complete coding region predicts a 217-residue protein with a COOH-terminal hydrophobic membrane anchor. Affinity-purified antibodies raised against the cytoplasmic region of this protein specifically detect a 29-kilodalton integral membrane protein enriched in the Golgi membrane. Indirect immunofluorescence microscopy reveals that this protein is mainly associated with the Golgi apparatus. When detergent extracts of the Golgi membrane are incubated with immobilized glutathione S-transferase alpha soluble N-ethylmaleimide-sensitive factor attachment protein (GST-alpha-SNAP), this protein was specifically retained. This protein has been independently identified and termed Vti1-rp2, and it is homologous to Vti1p, a yeast Golgi SNARE. We further show that Vti1-rp2 can be qualitatively coimmunoprecipitated with Golgi syntaxin 5 and syntaxin 6, suggesting that Vti1-rp2 exists in at least two distinct Golgi SNARE complexes. In cells microinjected with antibodies against Vti1-rp2, transport of the envelope protein (G-protein) of vesicular stomatitis virus from the endoplasmic reticulum to the plasma membrane was specifically arrested at the Golgi apparatus, providing further evidence for functional importance of Vti1-rp2 in protein trafficking in the secretory pathway.

Amino Acid Sequence↗
PubMed1998

Anti-Gal(alpha)1-3Gal antibody response to porcine bone marrow in unmodified baboons and baboons conditioned for tolerance induction.

BACKGROUND: Mixed lymphohematopoietic chimerism can provide an effective means of inducing longterm immunological tolerance and has been documented in a monkey allograft model. A conditioning regimen including nonmyeloablative or myeloablative irradiation and splenectomy has been used to induce chimerism in a pig-to-primate transplantation model. Since the presence of anti-Gal(alpha)1-3Gal (alphaGal) natural antibodies leads to the hyperacute rejection of pig organs transplanted into primates, extracorporeal immunoaffinity adsorption (EIA) of anti-alphaGal antibodies is also included in the regimen. The effect of the tolerance induction protocol on the anti-alphaGal antibody response has been assessed. METHODS: Anti-alphaGal antibody was measured after the EIA of plasma through an alphaGal immunoaffinity column in baseline studies involving two unmodified baboons, one splenectomized baboon, and one baboon that received a challenge with porcine bone marrow (BM), and in three groups of baboons (n=2 in each group) that received different conditioning regimens for tolerance induction. Group 1 received a nonmyeloablative conditioning regimen without porcine BM transplantation. Group 2 received nonmyeloablative conditioning with pig BM transplantation and pig cytokine therapy. Group 3 received myeloablative conditioning, an autologous BM transplant (with BM depleted of CD2+ or CD2+/CD20+ cells), and pig BM transplantation. RESULTS: In the baseline studies, a single EIA of anti-alphaGal antibodies in an unmodified animal initially depleted anti-alphaGal antibody, followed by a mild rebound. Nonmyeloablative conditioning (group 1) in the absence of pig cell exposure reduced the rate of anti-alphaGal antibody return. Pig BM cells markedly stimulated anti-alphaGal antibody production in an unmodified baboon (alphaGal IgM and IgG levels increased 40- and 220-fold, respectively). This response was significantly reduced (to an only 2- to 5.5-fold increase of IgM and IgG) in baboons undergoing nonmyeloablative conditioning (group 2). A myeloablative conditioning regimen (group 3) prevented the antibody response to pig BM, with the reduction in response being greater in the baboon that received autologous BM depleted of both CD2+ and CD20+ cells. No new antibody directed against pig non-aGal antigens was detected in any baboon during the 1 month follow-up period. CONCLUSIONS: (i) EIA of anti-alphaGal antibody in unmodified baboons results in a transient depletion followed by a mild rebound of antibody; (ii) exposure to pig BM cells results in a substantial increase in anti-alphaGal antibody production; (iii) a nonmyeloablative conditioning regimen reduces the rate of antibody return and (iv) markedly reduces the response to pig BM cells; (v) the anti-alphaGal response is completely suppressed by a myeloablative regimen if CD2+ and CD20+ cells are eliminated from the autologous BM inoculum. Furthermore, (vi) challenge with pig BM cells appears to stimulate only an anti-alphaGal antibody response without the development of other (non-alphaGal) anti-pig antibodies. We conclude that regimens used for T-cell tolerance induction can be beneficial in reducing the anti-alphaGal antibody response to porcine BM.

Animals↗
PubMed1998

Chemical and enzymatic properties of bridging 5'-S-phosphorothioester linkages in DNA.

We describe physicochemical and enzymatic properties of 5' bridging phosphorothioester linkages at specific sites in DNA oligonucleotides. The susceptibility to hydrolysis at various pH values is examined and no measurable hydrolysis is observed at pH 5-9 after 4 days at 25 degrees C. The abilities of three 3'- and 5'-exonuclease enzymes to hydrolyze the DNA past this linkage are examined and it is found that the linkage causes significant pauses at the sulfur linkage for T4 DNA polymerase and calf spleen phosphodiesterase, but not for snake venom phosphodiesterase. Restriction endonuclease (Nsi I) cleavage is also attempted at a 5'-thioester junction and strong resistance to cleavage is observed. Also tested is the ability of polymerase enzymes to utilize templates containing single 5'-S-thioester linkages; both Klenow DNA polymerase and T7 RNA polymerase are found to synthesize complementary strands successfully without any apparent pause at the sulfur linkage. Finally, the thermal stabilities of duplexes containing such linkages are measured; results show that T m values are lowered by a small amount (2 degrees C) when one or two thioester linkages are present in an otherwise unmodified duplex. The chemical stability and surprisingly small perturbation by the 5' bridging sulfur make it a good candidate as a physical and mechanistic probe for specific protein or metal interactions involving this position in DNA.

Animals↗
PubMed1998

Leukocystatin, a new Class II cystatin expressed selectively by hematopoietic cells.

We describe a new cystatin in both mice and humans, which we termed leukocystatin. This protein has all the features of a Class II secreted inhibitory cystatin but contains lysine residues in the normally hydrophobic binding regions. As determined by cDNA library Southern blots, this cystatin is expressed selectively in hematopoietic cells, although fine details of the distribution among these cell types differ between the human and mouse mRNAs. In addition, we have determined the genomic organization of mouse leukocystatin, and we found that in contrast to most cystatins, the leukocystatin gene contains three introns. The recombinant proteins corresponding to these cystatins were expressed in Escherichia coli as N-terminal glutathione S-transferase or FLAGTM fusions, and studies showed that they inhibited papain and cathepsin L but with affinities lower than other cystatins. The unique features of leukocystatin suggests that this cystatin plays a role in immune regulation through inhibition of a unique target in the hematopoietic system.

Amino Acid Sequence↗
PubMed1998

Synthesis and characterization of long chain alkyl acyl carnitine esters. Potentially biodegradable cationic lipids for use in gene delivery.

A series of alkyl acyl carnitine esters (alkyl 3-acyloxy-4-trimethylammonium butyrate chloride) were synthesized as potential biocompatible cationic lipids for use in gene transfer. The physicochemical properties of the lipids, liposomes prepared from them, and their complexes with DNA were characterized by differential scanning calorimetry (DSC), particle size, zeta potential, and surface monolayer measurements. The transition temperatures and behavior at an air-water interface for this series are similar to phosphatidylcholines with the same hydrocarbon chain length. The physical properties of the l derivatives were not significantly different from the dl derivatives. At 70 degrees C, the acyl chains were readily hydrolyzed at pH 7. The influence of the aliphatic chain length (n = 12-18) on transfection efficiency in vitro was determined using cationic liposomes prepared from these lipids or their mixtures with the helper lipids, dioleoylphosphatidylethanolamine (DOPE), dioleoylphosphatidylcholine, monooleoylglycerol, and cholesterol (Chol). The mixture of myristyl 3-myristoyloxy-4-trimethylammonium butyrate chloride (MMCE, 4d) with DOPE at a 1:1 molar ratio mediated the highest transfection efficiency in cell culture. The mixture of oleyl 3-oleoyloxy-4-trimethylammonium butyrate chloride (OOCE, 4f) with Chol at a 1:1 molar ratio gave the highest transfection efficiency after intravenous administration in mice. In vivo gene expression using 4f was comparable to values obtained with the best cationic lipids reported to date.

Animals↗
PubMed1998

[Chromosome assignment of a novel microsatellite polymorphic marker (D14S1435) by fluorescence in situ hybridization].

OBJECTIVE: Chromosome assignment of a novel microsatellite marker (D14S1435) containing CA repeats and showing 11 alleles and 0.85 of PIC value in Chinese population) isolated from microdissected human chromosome 14q24.3 DNA library by fluorescence in situ hybridization(FISH). METHODS: The polymorphic marker was used as probe to screen a human Lambda/DASH genome library, a positive clone was isolated and the recombinant phage DNA was extracted as primary material. The insert was recovered through BamH I digestion to complete and low melting point agarose gel electro- phoresis, after processed with Sau3A I the probe was prepared by linker- catch PCR method and the following fluorescence in situ hybridization(FISH) was conducted to metaphase chromosomes. RESULTS: this novel STR was precisely mapped to human chromosome 14q24.3, and this demonstrated the validity of isolating chromosome band-specific genetic markers from probe pool made by chromosome microdissection. CONCLUSION: Confirmed by chromosome fluorescence in situ hybridization, the novel STR obtained by chromosome microdissection, PCR and subcloning method was really from the micro- dissected region. This not only increases the numbers of genetic markers used for linkage analysis in the region,but also provides a novel valuable genetic marker for the gene diagnosis and gene cloning of the possible disease-causing genes located in the region of 14q24.3.

Chromosome Mapping↗
PubMed1998

Electrical conductivity of olivine, wadsleyite, and ringwoodite under upper-mantle conditions

Geophysical models show that electrical conductivity in Earth's mantle rises about two orders of magnitude through the transition zone in the depth range 410 to 660 kilometers. Impedance measurements obtained on Mg1.8Fe0.2SiO4 olivine, wadsleyite, and ringwoodite at up to 20 gigapascals and 1400 degreesC show that the electrical conductivities of wadsleyite and ringwoodite are similar and are almost two orders of magnitude higher than that of olivine. A conductivity-depth profile to 660 kilometers, based on these laboratory data, shows a conductivity increase of almost two orders of magnitude across the 410-kilometer discontinuity; such a profile favors a two-layer model for the upper mantle. Activation enthalpies of 1.2 to 1.7 electron volts permit appreciable lateral variations of conductivity with lateral temperature variations.

Journal Article↗
PubMed1998

A cell surface receptor defined by a mAb mediates a unique type of cell death similar to oncosis.

Cell death is mediated by distinct pathways including apoptosis and oncosis in response to various death signals. To characterize molecules involved in cell death, a panel of mAbs was raised by immunizing mice with apoptotic cells. One of these antibodies, designated anti-Porimin (for pro-oncosis receptor inducing membrane injury), was found to directly induce a unique type of cell death in Jurkat cells. Anti-Porimin defines a 110-kDa cell surface receptor on Jurkat cells. Functionally, anti-Porimin alone rapidly mediates pore formation on the plasma membrane and induces cell death without participation of complement. Both the cellular expression and functional characteristics of the Porimin antigen indicate that it is distinct from the CD95 (Fas/Apo-1) and other cell receptors known to induce apoptosis. Anti-Porimin-mediated cell death was preceded by cell aggregation, formation of plasma membrane pores, and the appearance of membrane blebs. More important, these cells show neither DNA fragmentation nor apoptotic bodies, but display lethal damage of the cell membrane. Cell death by anti-Porimin is distinct from complement-dependent cytolysis or complement-independent apoptosis but is similar to that described for oncosis, a form of cell death accompanied by the membrane damage followed by karyolysis. The induction of cell death by anti-Porimin may represent a unique cell surface receptor-mediated pathway of cell death in the human lymphoid system.

Animals↗
PubMed1998

Characterization of the promoter region of the human somatostatin receptor subtype 2 gene and localization of sequences required for estrogen-responsiveness.

Somatostatin receptor (sst) subtype 2 mRNA is potently regulated by 17beta-estradiol (E2) in a time and dose-dependent manner in the estrogen receptor (+) human breast cancer cell line T47D. To explore the mechanism involved in E2 regulated expression of sst2, we isolated and characterized a genomic clone containing over 5.3 kilobase pairs (kb) of the 5' flanking region of the human sst2 gene. The 5'-flanking region lacks both TATA and CCAAT boxes. Primer extension and RNAase protection analysis revealed the existence of two transcriptional start sites, located within an initiator-like sequence, 85 and 82 bp upstream of the translational initiation methionine. The 5.3-kb 5'-flanking region was an active promoter in T47D cells but was inactive in MDA MB 435s cells, a human breast cancer cell line that does not express sst2. Deletion analysis identified both positive and negative regulatory elements within the 5.3-kb fragment. Furthermore, transcriptional regulation by E2 was mediated by a distal 1.5-kb fragment located 3.8 kb from the transcriptional start sites. Determining the precise ERE will provide important insights into the complex regulation of sst2 expression in normal and neoplastic tissue.

Base Sequence↗
PubMed1998

Tolerization of anti-Galalpha1-3Gal natural antibody-forming B cells by induction of mixed chimerism.

Xenotransplantation could overcome the severe shortage of allogeneic organs, a major factor limiting organ transplantation. Unfortunately, transplantation of organs from pigs, the most suitable potential donor species, results in hyperacute rejection in primate recipients, due to the presence of anti-Galalpha1-3Gal (Gal) natural antibodies (NAbs) in their sera. We evaluated the ability to tolerize anti-Gal NAb-producing B cells in alpha1,3-galactosyltransferase knockout (GalT KO) mice using bone marrow transplantation (BMT) from GalT+/+ wild-type (WT) mice. Lasting mixed chimerism was achieved in KO mice by cotransplantation of GalT KO and WT marrow after lethal irradiation. The levels of anti-Gal NAb in sera of mixed chimeras were reduced markedly 2 wk after BMT, and became undetectable at later time points. Immunization with Gal+/+ xenogeneic cells failed to stimulate anti-Gal antibody production in mixed chimeras, whereas the production of non-Gal-specific antixenoantigen antibodies was stimulated. An absence of anti-Gal-producing B cells was demonstrated by enzyme-linked immunospot assays in mixed KO + WT --> KO chimeras. Thus, mixed chimerism efficiently induces anti-Gal-specific B cell tolerance in addition to T cell tolerance, providing a single approach to overcoming both the humoral and the cellular immune barriers to discordant xenotransplantation.

Animals↗
PubMed1998

A specific, nonproliferative role for E2F-5 in choroid plexus function revealed by gene targeting.

Homozygous E2F-5 knockout embryos and mice have been generated. Although embryonic development appeared normal, newborn mice developed nonobstructive hydrocephalus, suggesting excessive cerebrospinal fluid (CSF) production. Although the CSF-producing choroid plexus displayed normal cellular organization, it contained abundant electron-lucent epithelial cells, consistent with excessive CSF secretory activity. Moreover, E2F-5 CNS expression in normal animals was largely confined to the choroid plexus. Cell cycle kinetics were not perturbed in homozygous knockout embryo fibroblasts. Thus, E2F-5 is not essential for cell proliferation. Rather, it affects the secretory behavior of a differentiated neural tissue.

Alleles↗
PubMed1998

Functional role for the c-Abl tyrosine kinase in meiosis I.

The c-Abl tyrosine kinase is activated by ionizing radiation and certain other DNA-damaging agents. The DNA-dependent protein kinase (DNA-PK) and the ataxia telangiectasia mutated (ATM) gene product, effectors in the DNA damage response, contribute to the induction of c-Abl activity. The present study demonstrates that c-Abl is expressed in mouse and rat testes, and predominantly in pachytene spermatocytes of meiosis I. The results also demonstrate that c-Abl interacts directly with meiotic chromosomes. In concert with a requirement for c-Abl at the pachytene stage, we show that, in contrast to wild-type mice, testes from Abl-/- mice exhibit defects in spermatogenesis. These findings provide the first demonstration that c-Abl plays a functional role in meiosis.

Animals↗
PubMed1998

Vitamin E modulation of dieldrin-induced hepatic focal lesion growth in mice.

The effect of vitamin E on dieldrin-induced hepatic focal lesion growth in male B6C3F1 mice previously treated with diethylnitrosamine (DEN) was investigated. After hepatic focal lesions were formed, mice were placed into one of the following treatment groups: Group 1, 50 mg vitamin E/kg diet (control NIH-07 diet); Group 2, 10 mg dieldrin/kg NIH-07 diet; Group 3, 10 mg dieldrin and 450 mg vitamin E/kg NIH-07 diet; and Group 4, 450 mg vitamin E/kg NIH-07 diet. Mice were killed and necropsied after 30 and 60 d of dietary treatment. The effect of treatment on lesion growth was examined by measuring the number of focal lesions per liver and the relative hepatic focal lesion volume. In addition, the possible cellular mechanism of focal hepatocyte growth was investigated by examining both focal DNA synthesis and apoptosis. Dieldrin treatment alone (Group 2) increased the focal lesion volume, focal lesion number, and focal lesion labeling index. Supplementation with vitamin E (Group 3) blocked this effect. Vitamin E supplementation to the diet alone (Group 4) also enhanced focal lesion growth and increased the number of lesions per liver, the relative focal volume, and the labeling index in hepatic focal lesions. Interestingly, vitamin E supplementation inhibited apoptosis in normal liver but did not produce an observable decrease in apoptosis in hepatic focal lesions. The present study showed that dieldrin (Group 2) or vitamin E supplementation alone (Group 4) promoted the growth of hepatic focal lesions in mice. However, when vitamin E is supplemented to dieldrin-fed mice (Group 3), there is an inhibition of hepatic focal lesion growth.

Animals↗
PubMed1998

The duration of viral suppression during protease inhibitor therapy for HIV-1 infection is predicted by plasma HIV-1 RNA at the nadir.

OBJECTIVE: To determine markers that are associated with the durability of virologic response to therapy with HIV protease inhibitors in HIV-infected individuals. DESIGN: This study encompassed two retrospective analyses of the duration of virologic response to protease inhibitor therapy. The first analysis included 29 patients receiving either monotherapy or combination therapy with the protease inhibitor ritonavir whose plasma HIV RNA levels rebounded from the point of greatest decline with mutations associated with resistance to ritonavir. The second analysis included a cohort of 102 patients who initially responded to randomized treatment with either monotherapy with ritonavir or combination therapy with ritonavir and zidovudine. METHODS: Durability of response was defined as the time from the initiation of therapy to the point at which plasma HIV RNA displayed a sustained increase of at least 0.6 log10 copies/ml from the nadir value. In the first analysis, durability of response was analyzed with respect to baseline HIV RNA, HIV RNA at the nadir, and the drop in HIV RNA from baseline to the nadir. In the second analysis, time to rebound was examined using Kaplan-Meier analysis, stratifying by either baseline HIV RNA or HIV RNA at the nadir. RESULTS: In both analyses, the durability of response was not highly associated with either baseline RNA or the magnitude of RNA decline from baseline. Instead, a strong relationship was observed between the durability of response and the nadir plasma HIV-1 RNA value (P < 0.01). The nadir in viral load was generally reached after 12 weeks of randomized therapy. CONCLUSIONS: Viral RNA determinations at intermediate timepoints may be prognostic of impending virologic failure of protease inhibitor therapy. Therapeutic strategies that allow intensification of initial antiretroviral regimens in the subset of patients with incomplete virological response before the emergence of high level resistance should be investigated.

Anti-HIV Agents↗