Biomedical subjects

Y Wu

Publications and source records attributed to Y Wu.

At least 613 records · Page 34Linked to original sources

PubMed1998

Interleukin (IL)-1 receptor-associated kinase (IRAK) requirement for optimal induction of multiple IL-1 signaling pathways and IL-6 production.

Interleukin (IL)-1 is a proinflammatory cytokine with pleiotropic effects in inflammation. IL-1 binding to its receptor triggers a cascade of signaling events, including activation of the stress-activated mitogen-activated protein (MAP) kinases, c-Jun NH2-terminal kinase (JNK) and p38 MAP kinase, as well as transcription factor nuclear factor kappaB (NF-kappaB). IL-1 signaling results in cellular responses through induction of inflammatory gene products such as IL-6. One of the earliest events in IL-1 signaling is the rapid interaction of IL-1 receptor-associated kinases, IRAK and IRAK-2, with the receptor complex. The relative roles of IRAK and IRAK-2 in IL-1 signaling pathways and subsequent cellular responses have not been previously determined. To evaluate the importance of IRAK in IL-1 signaling, IRAK-deficient mouse fibroblast cells were prepared and studied. Here we report that IL-1-mediated activation of JNK, p38, and NF-kappaB were all reduced in embryonic fibroblasts deficient in IRAK expression. In addition, IL-6 production in response to IL-1 was also dramatically reduced in IRAK-deficient embryonic fibroblasts and in skin fibroblasts prepared from IRAK-deficient mice. Our results demonstrate that IRAK plays an essential proximal role in coordinating multiple IL-1 signaling pathways for optimal induction of cellular responses.

Animals↗

PubMed1998

B7-CTLA4 interaction enhances both production of antitumor cytotoxic T lymphocytes and resistance to tumor challenge.

Expression of B7-family costimulatory molecules CD80 (B7-1) and CD86 (B7-2) on tumor cells enhances host immunity. However, the role of the two B7 receptors, CD28 and CTLA4 (CD152), on T cells in antitumor immune response has not been clearly elucidated. Based on the effects of anti-CD28 and anti-CTLA4 mAbs on T cell response, it was proposed that CD28-B7 interaction promotes antitumor immunity, whereas B7-CTLA4 interaction down-regulates it. A critical test for the hypothesis is whether selective engagement of CTLA4 receptors by their natural ligands CD80 and CD86 enhances or reduces antitumor immunity. Here we used tumors expressing wild-type and mutant CD80, as well as mice with targeted mutation of CD28, to address this issue. We report that in syngeneic wild-type mice, B7W (W88>A), a CD80 mutant that has lost binding to CD28 but retained binding to CTLA4, can enhance the induction of antitumor cytotoxic T lymphocytes (CTL); B7Y (Y201>A), which binds neither CD28 nor CTLA4, fails to do so. Consistent with these observations, B7W-transfected J558 plasmocytoma and EL4 thymoma grow significantly more slowly than those transfected with either vector alone or with B7Y. Optimal tumor rejection requires wild-type CD80. Moreover, expression of a high level of CD80 on thymoma EL4 cells conveys immunity in mice with a targeted mutation of CD28 gene. Taken together, our results demonstrate that B7-CTLA4 interaction enhances production of antitumor CTL and resistance to tumor challenge and that optimal enhancement of antitumor immunity by CD80 requires its engagement of both CD28 and CTLA4.

Abatacept↗

PubMed1998

Influence of proximal side mutations on the molecular and electronic structure of cyanomet myoglobin: an 1H NMR study.

A series of proximal side mutants of sperm whale metmyoglobin (metMb) that involves residues which provide hydrogen bonds to the axial His and heme have been prepared, and the CO binding and solution molecular and electronic structure has been investigated by 1H NMR. These include Ser92(F7), whose O gamma serves as a hydrogen-bond acceptor to the axial His ring NdeltaH and whose O gamma H serves as hydrogen-bond donor to the 7-propionate carboxylate, and His97(FG3) whose ring provides the other hydrogen-bond donor to the 7-propionate carboxylate. 2D NMR data on the S92A-metMbCN, S92P-metMbCN and H97F-metMbCN show that the distal structure is completely conserved and that proximal side structural changes are highly localized. For the S92A-metMbCN, altered dipolar contacts to the F-helix backbone show that the axial His imidazole has rotated clockwise by approximately 10 degrees relative to a stationary heme, while in H97F-metMbCN, the altered heme-E helix backbone contacts reveal that the heme has rotated counterclockwise by approximately 3 degrees relative to a conserved axial His. The pattern of axial His rotation was qualitatively predicted by energy minimization calculations. The assignments and conserved structural elements allow the determination of a set of magnetic axes whose major magnetic axis is unchanged with respect to WT and confirms that local distal, and not proximal, interactions control the orientation of the major magnetic axis and, by inference, the degree and direction of tilt of the Fe-CN from the heme normal. The rhombic magnetic axes in S92A-metMbCN are rotated approximately 10 degrees in the opposite direction from the established approximately 10 degrees rotation for the axial His ring as expected. It is shown, moreover, that the pairwise alpha-, gamma-meso vs beta-, delta-meso-H hyperfine shift differences are well predicted by the change in the location of the rhombic magnetic axes. Carbon monoxide ligation rates experience minor but systematic perturbation for the S92A substitutions which confirms an influence (albeit very small) for axial His orientation on ligand affinity.

Animals↗

PubMed1998

The bis-azo compound FP-21399 inhibits HIV-1 replication by preventing viral entry.

The bis-azo compound FP-21399 inhibits HIV-1 infection. We now show that FP-21399 acts on the HIV-1 envelope glycoprotein to prevent viral replication. This compound targets the entry step of the HIV-1 replication cycle as demonstrated by time-of-addition and single cycle viral entry assays. The entry of SIVmac239, which uses the same coreceptors (CD4/CCR5) as HIV-1, was not inhibited by FP-21399, indicating that the antiviral effect of FP-21399 is specific for the HIV-1 envelope glycoprotein and is not dependent upon the cellular receptors CD4 and CCR5. FP-21399 inhibits neither the activity of HIV-1 reverse transcriptase nor the expression of HIV-1 early mRNA. Finally, this compound inhibits gp120 shedding of the T-tropic virus. Our results suggest that the anti-HIV activity of FP-21399 is due to its interaction with HIV-1 gp120/41 complex during viral entry.

Animals↗

PubMed1998

The autocrine derivation of the opioid growth factor, [Met5]-enkephalin, in ocular surface epithelium.

Endogenous opioid peptides serve as growth factors in developing, renewing, healing, and neoplastic cells and tissues. A native opioid peptide, [Met5]-enkephalin, termed opioid growth factor (OGF), has been discovered to regulate DNA synthesis in the epithelium of the ocular surface. OGF and its receptor zeta have been localized in both the basal and suprabasal cells of the epithelium. This study examined the hypothesis that OGF is an autocrine growth factor. Using probe for preproenkephalin (PPE) mRNA that encodes OGF, and in situ hybridization techniques, silver grains related to PPE mRNA were detected in both basal and suprabasal cells of the central and peripheral cornea, limbus, and conjunctiva. No distinct regional differences in the presence or location of message, as reflected by the density and distribution of PPE mRNA signal, were noted. These results demonstrate that a growth factor known to serve as a tonic, inhibitory, and receptor-mediated influence on the epithelium of the ocular surface is derived in an autocrine manner, thereby permitting local control of homeostatic cellular replication.

Animals↗

PubMed1998

CD28-independent induction of T helper cells and immunoglobulin class switches requires costimulation by the heat-stable antigen.

It is well established that B7-CD28/CTLA4 interactions play an important role in the induction of T helper cells for T-dependent antibody responses. However, targeted mutation of CD28 does not significantly affect production of IgG and activation of CD4 T helper cells in response to infections by some viruses and nematode parasites. To test whether the CD28-independent induction of Ig class switches requires costimulation by the heat-stable antigen (HSA), we compared T helper cell induction and antibody response in mice deficient for either HSA, CD28, or both genes. We found that after immunization with KLH-DNP, mice deficient for both CD28 and HSA lack DNP-specific IgA and all subtypes of IgG. This deficiency corresponds to a reduced number of effector helper T cells that rapidly produce IL-2, IL-4, and IFN-gamma after in vitro stimulation with carrier antigen KLH. In contrast, priming of T helper cells and Ig class switch are normal in mice deficient with either HSA or CD28 alone. IgM responses are not affected by any of these targeted mutations. These results demonstrate that CD28-independent induction of T helper cells and Ig class-switches requires costimulation by the HSA.

Animals↗

PubMed1998

A fifty percent reduction of platelet surface glycoprotein Ib does not affect platelet adhesion under flow conditions.

Glycoprotein (GP) Ib is an adhesion receptor on the platelet surface that binds to von Willebrand Factor (vWF). vWF becomes attached to collagens and other adhesive proteins that become exposed when the vessel wall is damaged. Several investigators have shown that during cardiopulmonary bypass (CPB) surgery and also during platelet activation in vitro by thrombin or thrombin receptor activating peptide (TRAP) GPIb disappears from the platelet surface. Such a disappearance is presumed to lead to a decreased adhesive capacity. In the present study, we show that a 65% decrease in platelet surface expression of GPIb, due to stimulation of platelets in Orgaran anticoagulated whole blood with 15 micromol/L TRAP, had no effect on platelet adhesion to both collagen type III and the extracellular matrix (ECM) of human umbilical vein endothelial cells under flow conditions in a single-pass perfusion system. In contrast to adhesion, ristocetin-induced platelet agglutination was highly dependent on the presence of GPIb. Immunoelectron microscopic studies showed that GPIb almost immediately returned to the platelet surface once platelets had attached to collagen. In a subsequent series of experiments, we showed that when less than 50% of GPIb was blocked by an inhibitory monoclonal antibody against GPIb (6D1), platelet adhesion under flow conditions remained unaffected.

Blood Platelets↗

PubMed1998

Differential transcriptional regulation of the human thrombin receptor gene by the Sp family of transcription factors in human endothelial cells.

The mitogenic effects of thrombin are mediated by a G-protein-coupled receptor. Because the effects of thrombin are strongly influenced by the expression of its receptor, an understanding of its regulatory mechanisms is essential. To identify mechanisms of human thrombin receptor (HTR) gene regulation, a series of HTR-promoter-luciferase constructs were made and transfected into human microvascular endothelial cells for analysis. Deletion from bp -303 to -164 abolished reporter gene expression. Dimethyl sulphate treatment in vivo and DNase I footprinting in vitro demonstrated that a cluster of three GC box consensus sites was occupied, and electrophoretic mobility-shift assays established that Sp1 and Sp3 both bind to this 3' GC box cluster. We mutated each of the three GC boxes individually and all three collectively within this 3' cluster. Basal promoter activity was decreased to 46%, 78% and 29% of control for each of the GC boxes mutated individually, and to 6% when the three were mutated collectively. To test the individual abilities of Sp1 and Sp3 to activate or repress HTR transcription, we conducted co-transfection experiments with wild-type or mutated HTR-promoter-luciferase constructs. Co-transfection with Sp1 significantly augmented wild-type HTR promoter activity. Sp3 alone did not affect activity, and inhibited Sp1-mediated activation. Competition for shared binding sites by Sp1 and Sp3 might differentially regulate HTR expression in vascular endothelial cells.

Aorta↗

PubMed1998

Tyrosine phosphorylation regulates the SH3-mediated binding of the Wiskott-Aldrich syndrome protein to PSTPIP, a cytoskeletal-associated protein.

Wiskott-Aldrich syndrome is an X-linked hematopoietic disease that manifests itself in platelet deficiency and a compromised immune system. Analysis of hematopoietic cells from affected individuals reveals that mutations in the Wiskott-Aldrich syndrome protein (WASP) result in structural and functional abnormalities in the cell cortex, consistent with the suggestion that WASP is involved with regulation of the actin-rich cortical cytoskeleton. Here we report that WASP interacts with a recently described cytoskeletal-associated protein, PSTPIP, a molecule that is related to the Schizosaccharomyces pombe cleavage furrow regulatory protein, CDC15p. This association is mediated by an interaction between the PSTPIP SH3 domain and two polyproline-rich regions in WASP. Co-expression of PSTPIP with WASP in vivo results in a loss of WASP-induced actin bundling activity and co-localization of the two proteins, which requires the PSTPIP SH3 domain. Analysis of tyrosine phosphorylation of PSTPIP reveals that two sites are modified in response to v-Src co-transfection or pervanadate incubation. One of these tyrosines is found in the SH3 domain poly-proline recognition site, and mutation of this tyrosine to aspartate or glutamate to mimic this phosphorylation state results in a loss of WASP binding in vitro and a dissolution of co-localization in vivo. In addition, PSTPIP that is tyrosine phosphorylated in the SH3 domain interacts poorly with WASP in vitro. These data suggest that the PSTPIP and WASP interaction is regulated by tyrosine phosphorylation of the PSTPIP SH3 domain, and this binding event may control aspects of the actin cytoskeleton.

Adaptor Proteins, Signal Transducing↗

PubMed1998

Evidence for a Cytoskeleton-Associated Binding Site Involved in Prolamine mRNA Localization to the Protein Bodies in Rice Endosperm Tissue

Previous studies have demonstrated that the mRNAs encoding the prolamine and glutelin storage proteins are localized to morphologically distinct membranes of the endoplasmic reticulum (ER) complex in developing rice (Oryza sativa L.) endosperm cells. To gain insight about this mRNA localization process, we investigated the association of prolamine polysomes on the ER that delimit the prolamine protein bodies (PBs). The bulk of the prolamine polysomes were resistant to extraction by 1% Triton X-100 either alone or together with puromycin, which suggests that these translation complexes are anchored to the PB surface through a second binding site in addition to the well-characterized ribosome-binding site of the ER-localized protein translocation complex. Suppression of translation initiation shows that these polysomes are bound through the mRNA, as shown by the simultaneous increase in the amounts of ribosome-free prolamine mRNAs and decrease in prolamine polysome content associated with the membrane-stripped PB fraction. The prolamine polysome-binding activity is likely to be associated with the cytoskeleton, based on the association of actin and tubulin with the prolamine polysomes and PBs after sucrose-density centrifugation.

Journal Article↗

PubMed1998

Requirement of two specific tyrosine residues for the catalytic activity of Bcr serine/threonine kinase.

Bcr is a novel serine/threonine protein kinase that is believed to require two cysteine pairs for activity (Maru and Witte, Cell, 67, 459, 1991). Tyrosine phosphorylated Bcr has dramatically reduced kinase activity, and tyrosine 360 of Bcr, which is one of the sites of phosphorylation by the Bcr-Abl oncoprotein, is required for transkinase activity (Liu et al., Mol. Cell Biol., 16, 998, 1996). Results presented here indicate that Bcr tyrosine 328 is also phosphorylated within Bcr-Abl expressing cells and is required for Bcr's serine/threonine kinase activity. Bcr Y328F, like Bcr Y360F, had defective transkinase activity but can autophosphorylate. However, the Y328F/Y360F double mutant of Bcr is defective in both trans- and autokinase activities. Taken together with the kinase inhibitory effects of tyrosine phosphorylation of Bcr by Bcr-Abl, our studies with tyrosine to phenylalanine Bcr mutants indicate that the hydroxyl residues of tyrosines 328 and 360 play crucial roles in Bcr's kinase activity.

Amino Acid Sequence↗

PubMed1998

Opioid gene expression in the developing and adult rat heart.

Opioid peptides are known to play a role in the function and growth of the mammalian heart. Although some information about gene expression of opioids in the heart is available, there is no data on the cellular location of opioid gene expression during development or in the adult. Using in situ hybridization and rat heart ranging from embryonic day 14 (E14) to adulthood, we have evaluated the distribution of gene expression for proenkephalin, proopiomelanocortin, and prodynorphin. With respect to preproenkephalin mRNA (PPE mRNA), message in the ventricle was abundant from E14 (the first time point examined) until shortly after birth, with a marked reduction noted on postnatal days 5, 10, and 21. Adults displayed considerable message, though less than in preparations of embryonic and neonatal heart. PPE mRNA was detected in epicardial, myocardial, and endocardial cells, as well as the walls of blood vessels, capillaries, and fibroblasts. Preproopiomelanocortin (POMC) mRNA was only found in adults, and was localized to the myocardium. Message for preprodynorphin could not be observed in the ventricles of developing or adult rats. These results are the first to define the temporal and spatial ontogeny of opioid gene expression with regard to the emergence of cardiac architecture. The data suggest that gene expression for proenkephalin is especially prevalent in embryonic and neonatal rats and may be related to the modulatory activity of the opioid growth factor, [Met5]-enkephalin, on cell proliferation and differentiation. The role of PPE and POMC mRNA in adult rat heart requires elucidation.

Animals↗

PubMed1998

Ontogeny of the opioid growth factor, [Met5]-enkephalin, preproenkephalin gene expression, and the zeta opioid receptor in the developing and adult aorta of rat.

Opioid peptides are known to play a role in the function of the mammalian cardiovascular system in the newborn. To learn about mechanisms underlying the morphogenesis of the developing vasculature and the principles contributing to the organization and maintenance of adult blood vessels, we have investigated the expression of the opioid growth factor (OGF), [Met5]-enkephalin, and its receptor, zeta. Moreover, gene expression for preproenkephalin mRNA, which encodes OGF, was studied to determine the source(s) of this inhibitory growth factor. By using immunocytochemistry, both OGF and the zeta opioid receptor were detected at embryonic day (E) 16 in the mesenchymal cells of the aortic wall. Staining appeared to be abundant in endothelial cells, smooth muscle cells, and fibroblasts at E20 and in the neonate. Immunoreactivity was noted to decrease progressively from day 5 to 10, but by weaning (day 21) and continuing into adulthood intense staining for both the peptide and receptor were observed. Preproenkephalin mRNA was detected throughout the aortic wall at E16, and the number of silver grains increased up today. Message was progressively reduced at days 5, 10, and 21, but signal in the adult aorta was comparable to that observed at day 5. These results indicate that components related to an endogenous opioid system regulating growth are present in the embryo, display a distinct spatial and temporal pattern of ontogeny, and persist into adulthood. In addition, these data indicate that OGF is an autocrine produced growth factor that is related to the emergence of vascular architecture and the maintenance of homeostasis in blood vessels.

Animals↗

PubMed1998

Fibrinogen adsorption and host tissue responses to plasma functionalized surfaces.

The physical and chemical characteristics of material surfaces are thought to play important roles in biomaterial-mediated tissue responses. To understand the importance of discrete biomaterial chemical characteristics in modifying host tissue responses, we constructed surfaces bearing different functional groups using radio frequency glow discharge plasma polymerization. Surfaces evaluated included those having high concentrations of -OH, -NH2, -CF3, and siloxyl groups. These surfaces and polyethylene terephthalate controls were used to assess the importance of particular physicochemical characteristics in surface:protein:cell interactions both in vitro and in vivo. The results obtained show that surface functionalities do significantly affect both the adsorption and "denaturation" of adsorbed fibrinogen (which is an important mediator of inflammatory responses to biomaterial implants). In addition, these surfaces provoke different degrees of acute inflammatory responses. Interestingly, the amounts of "denatured" fibrinogen that spontaneously accumulate on the individual surfaces correlate closely with the extent of biomaterial-mediated inflammation. These results suggest that surfaces that tend to "irreversibly" bind fibrinogen prompt greater acute inflammatory responses. Unexpectedly, all test surfaces except those bearing a siloxyl group engender relatively similar biomaterial-mediated fibrotic responses. Thus surface functionalities alone may not be sufficient to affect subsequent fibrotic responses.

Adsorption↗

PubMed1998

Altered expression of inhibitory guanine nucleotide regulatory proteins (Gi-proteins) in experimental hepatocellular carcinoma.

Guanine nucleotide regulatory proteins (G-proteins) play an important role in the onset and progression of malignancy. We hypothesized that alterations in inhibitory G-protein (Gi) expression and/or function may contribute to cellular invasion and formation of hepatocellular carcinoma (HCC). H4IIE hepatoma cells were inoculated directly into the liver parenchyma of ACI strain rats, and membranes were prepared from HCC livers and adjacent nonneoplastic livers 12 days following the initial inoculation. Expression of inhibitory Gialpha proteins was determined by Western blot analysis and changes in the functional activity of these proteins confirmed by pertussis toxin catalyzed ADP ribosylation and adenylyl cyclase activity. Inhibitory Gialpha1, Gialpha1/2, and Gialpha3 protein expression was significantly elevated in HCC when compared to adjacent nonneoplastic liver and sham-operated hepatic tissue. Pertussis toxin catalyzed ADP ribosylation of Gialpha substrates was significantly enhanced in HCC concomitant with increased basal and stimulated adenylyl cyclase activity following uncoupling of Gi-proteins with manganese ions. The role of Gi-proteins in cellular proliferation was confirmed using cultured H4IIE cells and normal hepatocytes. In quiescent H4IIE cells, mastoparan (Gialpha activator) increased [3H] thymidine incorporation and cell growth in a dose-dependent manner, whereas both pertussis toxin (a Gi-protein inhibitor) and 8-bromo-cAMP inhibited mitogenesis. In contrast, in isolated cultured hepatocytes, mastoparan inhibited [3H] thymidine incorporation, while pertussis toxin and 8-bromo-cAMP were mitogenic. We conclude that HCC is associated with marked changes in Gialpha-protein expression in vivo and in vitro, direct activation of which leads to increased mitogenesis in H4IIE cells in vitro.

8-Bromo Cyclic Adenosine Monophosphate↗

PubMed1998

Alignment echo of spin-3/2 9Be nuclei: detection of ultraslow motion

It is demonstrated that the alignment echo of 9Be is quite useful for detecting ultraslow atomic motions in the metallic glass Zr46. 75Ti8.25Cu7.5Ni10Be27.5. The time scale of detectable atomic motion is between T2 = 1.5 ms and T1 of a few seconds. Similar to previous works of 2H NMR, the Jeener-Broekaert sequence is used to create quadrupolar order for spin-32 nuclei upon nonselective excitations. Since the chemical and Knight shift distributions are not negligible for 9Be, the proper choice of the dephasing time between the first and the second pulses is essential for achieving pure quadrupolar order. It is demonstrated experimentally that slow atomic motions contribute significantly to the decay of the alignment echo near the glass transition temperature. Copyright 1998 Academic Press.

Journal Article↗

PubMed1998

Arterial patterns in the thoracic and abdominal segments of the esophagus: anatomy and clinical significance.

Cadaver studies were made to obtain more details about the arterial patterns in the thoracic and abdominal segments of the esophagus, with emphasis on the arterial supply to the lower esophageal sphincter (LES). The results showed that the inferior thoracic segment of the esophagus usually received blood supply from a single artery, with an average diameter of 1.42 +/- 0.49 mm (X +/- s), whereas the abdominal segment was commonly supplied by two arteries. The abdominal esophageal arteries coming from the left gastric artery were semi-circular in shape with an average diameter of 2.06 +/- 0.70 mm. From these semi-circular esophageal arteries, 3 to 8 smaller arteries branched off and penetrated into the muscle of the abdominal segment of the esophagus or anastomosed with arterial branches from the inferior thoracic segment. Thus, the blood supply of the abdominal segment was richer than that of the inferior thoracic segment. This richness of blood supply, characterized by semicircular esophageal arteries in the abdominal segment, may be important for the function of the LES.

Adult↗

PubMed1998

Study on amino acid composition of HSP70 and the level of plasma free amino acids of workers with long-term exposure to harmful factors.

In order to know the practical value of heat stress protein 70 (HSP70) and to know the changes of plasma free amino acids of workers with the induction of HSP70 by harmful factors, the amino acid composition of major HSP, HSP70 purified from the heated cultured human leukemia cancer cell line K562, rabbit liver, rat liver and heart, and mouse liver with two-step procedures of DE52-cellulose ion exchange chromatography and affinity chromatography on ATP-agarose was examined. The level of plasma free amino acids of workers with long-term exposure to heat, carbon monoxide and the combined effect of both heat and carbon monoxide was also investigated. The results showed that the three richest amino acids in HSP70 of all origins were Gly, Glu and Asp, except that of rat heart which was rich in Gly, Phe and Glu. Additionally, Lys, Val, Leu and Ala were also found very rich in HSP70 of all origins. Compared with controls, the most of plasma free amino acids tended to increase and free methionine and tryptophan were increased significantly (P < 0.05) after a long-term exposure to heat, carbon monoxide, and both. These findings suggested that further studies need to be done to find the substances or drugs which induce the synthesis of HSP70 and reduce the inhibition of synthesis of normal proteins for the purpose of protecting people with exposure to harmful factors against the damage of the factors.

Amino Acids↗