Search PubMed⌕ Search

Biomedical subjects

Z H Lu

Publications and source records attributed to Z H Lu.

At least 37 records · Page 2Linked to original sources
PubMed1998

The role of GM1 ganglioside in regulating excitatory opioid effects.

Our studies with cultured cells have provided new insight into the particular role of GM1 in regulating excitatory opioid responses. GM1 is significantly elevated in chronic opioid-treated cells via Gs/adenylyl cyclase activation. Such GM1 elevation promotes coupling of opioid receptor with Gs, resulting in attenuation of inhibitory opioid effects and induction of a sustained excitatory response. Application of exogenous GM1, but not other gangliosides, induces excitatory opioid responses not only in neurons and neuroblastoma cells that bear intrinsic opioid receptors but also in nonneuronal cells that are transfected with delta-opioid receptor. The latter system provides evidence that allosteric binding of GM1 changes receptor conformation from a Gi-coupled to a Gs-coupled mode. This is supported by preliminary experiments with a mutated delta-opioid receptor.

Adenylyl Cyclases↗
PubMed1998

The role of GM1 and other gangliosides in neuronal differentiation. Overview and new finding.

The pronounced increases in gangliosides belonging to the gangliotetraose family during the neurite outgrowth phase of neuronal differentiation have suggested a functional requirement for these substances related to process extension, arborization, and possibly synaptogenesis. Support for this hypothesis has come from a variety of experimental paradigms utilizing neuroblastoma cell lines, primary neuronal cultures, and observations on the developing nervous system. We have recently observed that differentiation of both primary neurons and neuroblastoma cells by Ca(2+)-elevating stimulants is characterized by upregulation of GM1 in the nuclear membrane. Immunostaining revealed these Ca(2+)-induced neurites to have axonal characteristics. Recent work has indicated that nuclear GM1 facilitates efflux of nuclear Ca2+, thereby contributing to the reduced level of nuclear Ca2+ that characterizes the differentiated neuron. Thus, while GM1 is generally recognized as a pluripotent molecule with several modulatory roles in the plasma membrane of developing and mature neurons, regulation of Ca2+ flux across the nuclear membrane is proposed as another critical function of this ganglioside in neuronal development, with special relevance to axonogenesis.

Animals↗
PubMed1998

CXCR4 sequences involved in coreceptor determination of human immunodeficiency virus type-1 tropism. Unmasking of activity with M-tropic Env glycoproteins.

The interaction of human immunodeficiency virus type 1 (HIV-1) with CD4 and one of a cadre of chemokine receptors triggers conformational changes in the HIV-1 envelope (Env) glycoprotein that lead to membrane fusion. The coreceptor activity of the second extracellular loop of CXCR4, which is restricted to dual tropic and T-tropic strains, was insensitive to the removal of charged residues either singly or in combinations by alanine scanning mutagenesis or to the conversion of acidic residues to lysine. Conversion of Asp-187 to a neutral residue exclusively unmasked activity with M-tropic Env in fusion and infection experiments. Insertion of the D187V mutation into chimeras containing extracellular loop 2 of CXCR4 in a CXCR2 framework also resulted in the acquisition of M-tropic coreceptor activity. The independence of CXCR4 coreceptor activity from charged residues and the extension of its repertoire by removing Asp-187 suggest that this interaction is not electrostatic and that coreceptors have the potential to be utilized by a spectrum of Env, which may be masked by charged amino acids in extracellular domains. These findings indicate that the primary structural determinants of coreceptors that program reactivity with M-, dual, and T-tropic Env are surprisingly subtle and that relatively insignificant changes in CXCR4 can dramatically alter utilization by Env of varying tropism.

Flow Cytometry↗
PubMed1998

Hematological effects of atypical and Cameroon beta-globin gene haplotypes in adult sickle cell anemia.

To examine the effects of unusual or atypical beta-globin gene cluster haplotypes on the hematological features and Hb F levels of sickle cell anemia, we studied African Americans who had an atypical or Cameroon haplotype chromosome in association with a typical haplotype. We identified over 20 atypical haplotypes. The distribution of 5' sub-haplotypes of the atypical chromosomes mirrored the distribution of common haplotypes in African Americans with sickle cell anemia. Neither 5' nor 3' subhaplotypes of the atypical chromosomes affected Hb F levels, packed cell volume, or mean corpuscular volume in individuals with a Benin chromosome. That the 5' subhaplotype is unaffected might be a consequence of the small numbers of Senegal 5' subhaplotypes in our sample, the need for linkage of both 5' and 3' subhaplotypes of any haplotype for an effect on Hb F to be present, or the likelihood that a normal beta-globin gene contributed the 5' subhaplotypes of some atypical haplotypes.

Adult↗
PubMed1998

Induction of axon-like and dendrite-like processes in neuroblastoma cells.

Neuroblastoma cells are widely utilized models for the study of the neuritic outgrowth phase of neuronal differentiation, but relatively few such studies have attempted to identify the nature of the process outgrowths. This identification will be necessary in developing strategies for utilizing these models to distinguish the underlying mechanisms involved in axonogenesis vs dendritogenesis. In an effort to identify procedures for inducing specific types of neurite outgrowth, and for distinguishing axon- from dendrite-like processes, we have subjected two neuroblastoma cell lines to a variety of stimuli previously shown to induce neurite outgrowth in these cells. These include neuraminidase, ionomycin, KCl+dibutyryl cAMP, cholera toxin B subunit, retinoic acid, dibutyryl cAMP (alone), GM1 ganglioside, and low serum. The first four of these (group 1) gave rise to neurites with axon-like characteristics, including immunostaining that was positive for phosphorylated high molecular weight neurofilament protein (NF-H) and synaptic vesicle protein-2 (SV2), but negative for microtubule-associated protein-2 (MAP2). The next three treatments (group 2) resulted in dendrite-like processes, as evidenced in immunostaining that was positive for MAP2 and negative for NF-H and SV2. Neurites produced by low serum had mixed properties. These cytoskeletal differences were supported by immunoblot analysis with antisera to the above cytoskeletal proteins. Striking morphological differences were also noted, group 2-induced neurites being significantly shorter with more branch points than those generated by group 1 stimulants. Time of exposure to stimulatory agent was crucial in determining expression of the neuritic phenotype. Correlation with previous studies suggests that axon-like neurites result from stimulants which elevate intracellular Ca2+, a dependence not previously reported to our knowledge. Dendrite-like process outgrowth, on the other hand, does not appear to depend on altered intracellular Ca2+.

Animals↗
PubMed1998

Histone H1 reduces the frequency of initiation in Xenopus egg extract by limiting the assembly of prereplication complexes on sperm chromatin.

Somatic histone H1 reduces both the rate and extent of DNA replication in Xenopus egg extract. We show here that H1 inhibits replication directly by reducing the number of replication forks, but not the rate of fork progression, in Xenopus sperm nuclei. Density substitution experiments demonstrate that those forks that are active in H1 nuclei elongate to form large tracts of fully replicated DNA, indicating that inhibition is due to a reduction in the frequency of initiation and not the rate or extent of elongation. The observation that H1 dramatically reduces the number of replication foci in sperm nuclei supports this view. The establishment of replication competent DNA in egg extract requires the assembly of prereplication complexes (pre-RCs) on sperm chromatin. H1 reduces binding of the pre-RC proteins, XOrc2, XCdc6, and XMcm3, to chromatin. Replication competence can be restored in these nuclei, however, only under conditions that promote the loss of H1 from chromatin and licensing of the DNA. Thus, H1 inhibits replication in egg extract by preventing the assembly of pre-RCs on sperm chromatin, thereby reducing the frequency of initiation. These data raise the interesting possibility that H1 plays a role in regulating replication origin use during Xenopus development.

Animals↗
PubMed1998

An animal model to study erythrocyte senescence with a narrow time window of erythrocyte production: alterations in osmotic fragility and deformability of erythrocytes during their life span.

Using the model in which the entire RBC population was nearly synchronously produced following the induction of spherocytic anemia in the rabbit with antibody serum, we determined the changes of RBC osmotic fragility and deformability with aging. The results showed that the osmotic fragility increased with the RBC aging process in a nonlinear manner, being much more profound in the later part of the RBC life span. The RBC deformation index (DI) was measured by an ektacytometry. It is found that the DI decreased with RBC aging in a nonlinear fashion, with increasingly greater changes in the later part of the RBC life span. The alterations of RBC mechanical properties with aging may be attributable to a number of factors, including changes of RBC size and shape, and the viscoelasticity of the cytoplasm and membrane.

Anemia, Hemolytic↗
PubMed1997

Differential utilization of CCR5 by macrophage and T cell tropic simian immunodeficiency virus strains.

Certain chemokine receptors serve as cofactors for HIV type 1 envelope (env)-mediated cell-cell fusion and virus infection of CD4-positive cells. Macrophage tropic (M-tropic) HIV-1 isolates use CCR5, and T cell tropic (T-tropic) strains use CXCR4. To investigate the cofactors used by simian immunodeficiency viruses (SIV), we tested four T-tropic and two M-tropic SIV env proteins for their ability to mediate cell-cell fusion with cells expressing CD4 and either human or nonhuman primate chemokine receptors. Unlike HIV-1, both M- and T-tropic SIV envs used CCR5 but not CXCR4 or the other chemokine receptors tested. However, by testing a panel of CCR5/CCR2b chimeras, we found that the structural requirements for CCR5 utilization by M-tropic and T-tropic SIV strains were different. T-tropic SIV strains required the second extracellular loop of CCR5 whereas a closely related M-tropic SIV strain could, like M-tropic HIV-1 strains, use the amino-terminal domain of CCR5. As few as two amino acid changes in the SIV env V3 domain affected the regions of CCR5 that were critical for fusogenic activity. Receptor signaling was not required for either fusion or infection. Our results suggest that viral tropism may be influenced not only by the coreceptors used by a given virus strain but also by how a given coreceptor is used.

Animals↗
PubMed1997

Fetal hemoglobin in sickle cell anemia: determinants of response to hydroxyurea. Multicenter Study of Hydroxyurea.

Hydroxyurea (HU) can increase fetal hemoglobin (HbF) in sickle cell anemia (HbSS). To identify determinants of the HbF response, we studied 150 HU-treated patients grouped by quartiles of change in HbF from baseline to 2 years. Half of the HU-assigned patients had long-term increments in HbF. In the top two quartiles, HbF increased to 18.1% and 8.8%. These patients had the highest baseline neutrophil and reticulocyte counts, and largest treatment-associated decrements in these counts. In the lower two quartiles, 2-year HbF levels (4.2% and 3.9%) and blood counts changed little from baseline. In the highest HbF response quartile, myelosuppression developed in less than 6 months, compliance was best, and final doses of HU were 15 to 22.5 mg/kg. All four quartiles had substantial increases of F cells in the first year. This was maintained for 2 years only in the top three quartiles. Leukocyte and reticulocyte counts decreased initially in all quartiles, but drifted back toward baseline levels in the lowest HbF response quartile. Initial HbF level and phenotype of the F-cell production (FCP) locus were not associated with HbF response, but absence of a Central African Republic (CAR) haplotype was. Bone marrow ability to withstand HU treatment may be important for sustained HbF increases during HU treatment of HbSS.

Adult↗
PubMed1997

Interaction of the delta-opioid receptor with GM1 ganglioside: conversion from inhibitory to excitatory mode.

Previous studies have shown GM1 ganglioside to play a crucial role in regulating excitatory opioid receptor function, which may underlie some aspects of opioid dependence, tolerance, and supersensitivity. To study the mechanism of this receptor modulation we have employed CHO cells containing a single, transfected opioid receptor of the delta-type. When forskolin was employed to elevate cAMP the reduction affected by 10 microM DADLE was counteracted by preincubation of the cells with GM1. No effect was observed with GD1a, GD1b, GT1b GM3, or the GM1 derivative, GM1-OH. In pertussis toxin-treated cells 10 nM DADLE increased basal levels of cAMP after preincubation with as little as 10 nM GM1. The results suggest conformational alteration of the opioid receptor from a form coupled primarily to G(i)/G(o) to one also capable of interacting with G(s).

Adenylyl Cyclases↗
PubMed1997

Opioid receptor and calcium channel regulation of adenylyl cyclase, modulated by GM1, in NG108-15 cells: competitive interactions.

GM1 ganglioside was previously shown to function as a specific regulator of excitatory opioid activity in dorsal root ganglion neurons and F11 hybrid cells, as seen in its facilitation of opioid-induced activation of adenylyl cyclase and its ability to dramatically reduce the threshold opioid concentration required to prolong the action potential duration. The elevated levels of GM1 resulting from chronic opioid exposure of F11 cells were postulated to cause the ensuing opioid excitatory supersensitivity. We now show that GM1 promotes opioid (DADLE)-induced activation of adenylyl cyclase in NG108-15 cells which possess the delta-type of receptor. In keeping with previous studies of other systems, this can be envisioned as conformational interaction of GM1 with the receptor that results in uncoupling of the receptor from Gi and facilitated coupling to Gs. This would also account for the observation that DADLE-induced attenuation of forskolin-stimulated adenylyl cyclase was reversed by GM1, provided the cells were not pretreated with pertussis toxin. When the cells were so pretreated, GM1 evoked an unexpected attenuation of forskolin-stimulated adenylyl cyclase attributed to GM1-promoted influx of calcium which was postulated to inhibit a calcium-sensitive form of adenylyl cyclase. This is concordant with several studies showing GM1 to be a potent modulator of calcium flux. Pertussis toxin in these experiments exerted dual effects, one being to promote interaction of the delta-opioid receptor with Gs through inactivation of Gi, and the other to enhance the GM1-promoted influx of calcium by inactivation of Go; the latter is postulated to function as constitutive inhibitor of the relevant calcium channel. NG108-15 cells thus provide an interesting example of competitive interaction between two GM1-regulated systems involving enhancement of both opioid receptor excitatory activity and calcium influx.

Adenylate Cyclase Toxin↗
PubMed1997

Two distinct CCR5 domains can mediate coreceptor usage by human immunodeficiency virus type 1.

The chemokine receptor CCR5 is the major fusion coreceptor for macrophage-tropic strains of human immunodeficiency virus type 1 (HIV-1). To define the structures of CCR5 that can support envelope (Env)-mediated membrane fusion, we analyzed the activity of homologs, chimeras, and mutants of human CCR5 in a sensitive gene reporter cell-cell fusion assay. Simian, but not murine, homologs of CCR5 were fully active as HIV-1 fusion coreceptors. Chimeras between CCR5 and divergent chemokine receptors demonstrated the existence of two distinct regions of CCR5 that could be utilized for Env-mediated fusion, the amino-terminal domain and the extracellular loops. Dual-tropic Env proteins were particularly sensitive to alterations in the CCR5 amino-terminal domain, suggesting that this domain may play a pivotal role in the evolution of coreceptor usage in vivo. We identified individual residues in both functional regions, Asp-11, Lys-197, and Asp-276, that contribute to coreceptor function. Deletion of a highly conserved cytoplasmic motif rendered CCR5 incapable of signaling but did not abrogate its ability to function as a coreceptor, implying the independence of fusion and G-protein-mediated chemokine receptor signaling. Finally, we developed a novel monoclonal antibody to CCR5 to assist in future studies of CCR5 expression.

Amino Acid Sequence↗
PubMed1997

Histone H1 modulates DNA replication through multiple pathways in Xenopus egg extract.

We investigated the effects of histone H1s on DNA replication using Xenopus egg extract. Mouse variants H1c and H10 were assembled onto Xenopus sperm chromatin by the extract during the remodeling that accompanies nuclear decondensation. The association of H1 with chromatin was rapid and concentration dependent. H1-associated chromatin displayed a typical nucleosomal repeat pattern indicating that linker histones are properly positioned along the DNA. The presence of H1 on sperm chromatin reduced both the rate and extent of DNA replication in egg extract. This reduction in rate is due, in part, to a delay in initiation of replication within individual nuclei. Initiation in extract is dependent upon nuclear assembly. Analysis of the assembly process revealed that H1 does not inhibit nuclear membrane formation or the import of nuclear protein, however, it does slow the rate of nuclear lamina formation. This H1-induced delay in lamina assembly is responsible for the delay in initiation as pre-assembled H1-containing nuclei initiate replication at the same time as control nuclei. However, H1 inhibits replication even when lamina assembly is complete suggesting that H1 also affects replication directly. These data indicate that H1 modulates DNA replication through multiple pathways in egg extract.

Animals↗
PubMed1996

Trophic effect of cholera toxin B subunit in cultured cerebellar granule neurons: modulation of intracellular calcium by GM1 ganglioside.

Survival of cerebellar granule cells (CGC) in culture was significantly improved in the presence of cholera toxin B subunit (Ctx B), a ligand which binds to GM1 with specificity and high affinity. This trophic effect was linked to elevation of intracellular calcium ([Ca2+]i), and was additive to that of high K+. Survival was optimized when Ctx B was present for several days during the early culture period. 45Ca2+ and cell survival studies indicated the mechanism to involve enhanced influx of Ca2+ through L-type voltage-sensitive channels, since the trophic effect was blocked by antagonists specific for that channel type. Inhibitors of N-methyl-D-aspartate receptor/channels were without effect. During the early stage of culture Ctx B, together with 25 mM K+, caused [Ca2+]i to rise to 0.2-0.7 microM in a higher proportion of cells than 25 mM K+ alone. A significant change in the nature of GM1 modulation of Ca2+ flux occurred after 7 days in culture, at which time Ctx B ceased to elevate and instead reduced [Ca2+]i below the level attained with 25 mM K+. GM1 thus appears to serve as intrinsic inhibitor of one or more L-type Ca2+ channels during the first 7 days in vitro, and then as intrinsic activator of (possibly other) L-type channels after that period. This is the first demonstration of a modulatory role for GM1 ganglioside affecting Ca2+ homeostasis in cultured neurons of the CNS.

Animals↗
PubMed1996

Fetal hemoglobin in sickle cell anemia: relation to regulatory sequences cis to the beta-globin gene. Multicenter Study of Hydroxyurea.

Very different fetal hemoglobin levels among adult sickle cell anemia patients suggest genetic modulation of gamma-globin gene expression. In sickle cell anemia, different fetal hemoglobin levels are associated with distinct beta-globin gene haplotypes. Haplotype may be a marker for linked DNA that modulates gamma-globin gene expression. From 295 individuals with sickle cell anemia, we chose for detailed studies 53 patients who had the highest or the lowest fetal hemoglobin levels and 7 patients whose fetal hemoglobin levels were atypical of their haplotype. In these individuals, we examined portions of the beta-globin gene locus control region hypersensitive sites two and three, an (AT)x(T)y repeat 5' to the beta-globin gene, a 4-bp deletion 5' to the A gamma T gene, promoters of both gamma-globin genes, 5' flanking region of the G gamma-globin gene, and A gamma-globin gene IVS-II. Of the regions we studied all polymorphisms were always haplotype-linked and no additional mutations were present. This suggested that variations in these areas are uncommon mechanisms of fetal hemoglobin modulation in sickle cell anemia. Whereas unexamined cis-acting sequences may regulate gamma-globin gene transcription, trans-acting factors may play a more important role.

Anemia, Sickle Cell↗
PubMed1996

Beta-globin gene haplotype in Hb SC disease.

We asked the question, is the haplotype found with the sickle hemoglobin gene associated with different hematological characteristics in patients who were combined heterozygotes for sickle hemoglobin and hemoglobin C (Hb SC disease)? In 73 adults with Hb SC disease, a Benin haplotype chromosome was present in 56%, and Bantu (or Central African Republic; CAR), Senegal, and atypical haplotype chromosomes were found in 25%, 6%, and 12%, respectively. No significant difference were found in hematological characteristics or fetal hemoglobin levels of patients with Benin/C, CAR/C, Senegal/C, and atypical/C haplotypes. There were 71% C I, 18% C II, and 11% other beta(c) haplotypes. Fetal hemoglobin levels are lower in Hb SC disease than in sickle-cell anemia. Perhaps because haplotype has no discernible effect on fetal hemoglobin level in Hb SC disease, it does not modulate its hematological features.

Adult↗